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BRAIN FOG
AIR POLLUTION MAY CAUSE COGNITIVE DECLINE
RECONSIDERING
PARKINSON’S
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OCTOBER 2019
Contents
THE SCIENTIST THE-SCIENTIST.COM VOLUME 33 NUMBER 10
© LISA CLARK; © ISTOCK.COM, DEBIBISHOP; © SCIENCESOURCE, SPL
Features ON THE COVER: MODIFIED FROM © ISTOCK.COM, QUICKSHOOTING
30
The Destructive Path
36
Brain Fog
44
NETs: Two-Faced Players
of Parkinson's Air pollution likely has detrimental in Immunity
The disease might not start with effects on human neurodevelopment Webs of nuclear material expelled from
tangles of α-synuclein proteins, but and cognitive function. neutrophils trap invading pathogens, but
with the inability of cells to rid BY CATHERINE OFFORD these newly discovered structures also
themselves of toxic materials. have ties to autoimmunity and cancer.
BY ASHLEY YEAGER BY BORKO AMULIC AND
GABRIEL SOLLBERGER
10. 201 9 | T H E S C IE N T IST 3

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OCTOBER 2019
Department Contents
13 59
16 FROM THE EDITOR
Flesh and Blood and Wonder
LAB TOOLS
Peering into the Brain
The human brain is made of delicate Researchers are developing a variety
tissue. This fragility makes it all the of approaches for clearing neural
more amazing that it can generate tissue to get a better view of the
thoughts and consider its own function. brain’s circuitry.
BY BOB GRANT BY AMBER DANCE
16 NOTEBOOK 63 BIO BUSINESS

Nature as Medicine; Monkey Noise; Breaking the Antidepressant
Overactive Brains; Museum Finds Drought
FDA approvals of the first major new
26 CRITIC AT LARGE classes of antidepressant therapies
Organoids, Not Chimeras in decades open up new possibilities
Scientists are devising human-animal for the development of psychiatric
hybrids for harvesting human organs, drugs.
but lab-derived mini-organs are a less
52 BY BIANCA NOGRADY
ethically fraught solution to meeting
the need for transplantation. 67 READING FRAMES
BY JOHN D. LOIKE AND Warm-Blooded, Warm-Hearted
ROBERT POLLACK How evolution geared the
mammalian brain toward kindness
28 MODUS OPERANDI BY PATRICIA CHURCHLAND
Getting on the Nerves
An implantable wireless device with 72 FOUNDATIONS
UNSPLASH, DAVID MARCU; KELLY FINNAN; © CHRISTOPHER BEAUCHAMP PHOTOGRAPHY
microfluidic and optical components Wine Therapy, Middle Ages

allows remote control of individual BY KERRY GRENS
nerve fibers in a mouse’s extremities.
BY RUTH WILLIAMS IN EVERY ISSUE
10 CONTRIBUTORS
52 THE LITERATURE 15 SPEAKING OF SCIENCE
Stem cell stimulation for stroke 71 THE GUIDE
recovery in mice; the link between
57 paternal age and autism risk; sleep as a
ANSWER
biomarker of Alzheimer’s PUZZLE ON PAGE 15
54 PROFILE P ROO F S C A R L E T
The Cerebellum’s Secrets O M I P M A I
Neuroscientist Kamran Khodakhah S C A NN E R P E C A N
has discovered that the brain I H A H T E
region does more than coordinate T R A NQU I L I Z E R
R U N B A S
movement; it also plays a role in
O RCH I D V I O L E T
reward circuits and in addiction. N H C C A I
BY ANNA AZVOLINSKY J A C KO L A N T E RN
H M S I S G
57 SCIENTIST TO WATCH UMB R A MON S T E R
Johannes Kohl: Circuit Breaker M E N A E E A
BY NICOLETTA LANESE
P A R A DOX WA R T Y
10. 201 9 | T H E S C IE N T IST 5

OCTOBER 2019
Online Contents
THIS MONTH AT THE-SCIENTIST.COM:
VIDEO VIDEO VIDEO

Monkey Music? Cerebellar Surprises NETs on Film
Hear the muted symphony that one Kamran Khodakhah, a researcher at the See neutrophil extracellular traps in
researcher thinks might be a close Albert Einstein College of Medicine, talks antibacterial action.
approximation to how macaques about the cerebellum’s newly described
perceive the performance. role in addictive and social behaviors.
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OCTOBER 2019
Contributors
Patricia Churchland coined the term “neurophilosophy,” a word that refers to the idea that
neurobiological mechanisms underpin all mental processes, including decision-making, prob-
lem-solving, and even emergent properties like consciousness.
Churchland began her graduate studies in both biology and philosophy at the
University of Pittsburgh before transferring to the University of Oxford in 1966. While there, she
“slowly came to the realization that psychologists and neuroscientists were actually the ones who
were really interested in finding out how . . . the mind really works, in as much as the mind is a
product of the way the brain works,” she says. After graduating in 1969, she took a faculty posi-
tion at the University of Manitoba in Winnipeg and began her foray into the world of neurosci-
ence in earnest.
At Manitoba, Churchland attended neuroscience lectures and labs alongside medical stu-
dents, attended clinical rounds in the neurology and neurosurgery departments, and worked
in a lab studying the neural circuitry behind rhythmic walking motions. In 1984, Churchland
became a professor at the University of California, San Diego, and soon befriended neuroscien-
tist Francis Crick, who worked at the Salk Institute for Biological Studies nearby. In 1986, she
published her first book, entitled Neurophilosophy, which explores the relationship between
abstract cognitive theory and neuroscience. Churchland has since written five more texts, each
exploring overlaps between philosophy and brain science.
On page 67 of this issue, Churchland considers questions she says she and Crick discussed
often, such as, “Why do mammals make personal sacrifices for the benefit of others?” Her essay
encapsulates the focus of her most recent work, Conscience: The Origins of Moral Intuition.
“I think the answer slowly came into focus,” she says. “It began to be very clear that there is
COURTESY OF PATRICIA CHURCHLAND; CHRISTOPHER HARBORT; DIANE SCHAD, MAX PLANCK INSTITUTE
a very deep, very rich, very important biologic story here.”
Borko Amulic and Gabriel Sollberger both investigate the function of neutrophils, a type
of white blood cell critical to defending the body against pathogens. The two met as postdocs
in the lab of Arturo Zychlinsky at the Max Planck Institute for Infection Biology. The cellular
microbiologist’s group had previously discovered that neutrophils generate fibrous nets to trap
bacteria, and both Amulic and Sollberger signed on to further explore the mysterious meshes,
known as neutrophil extracellular traps or NETs. The cellular microbiologist’s group had pre-
viously discovered that neutrophils generate fibrous nets to trap bacteria, and both Amulic and
Sollberger signed on to further explore the mysterious meshes, known as neutrophil extracellu-
lar traps or NETs, which they write about on page 44.
Amulic, now an assistant professor at the University of Bristol and a Medical Research
Council fellow, leads a lab that studies how neutrophils are regulated at the molecular level
and throughout the cell cycle. His work suggests that dysregulation of cell cycle proteins
alters the role of neutrophils in immunity and inflammation, which may relate to certain dis-
ease pathologies. “Achieving perfect balance in neutrophil responses seems integral to human
health and it is perhaps something that we can learn to modulate therapeutically,” he writes in
an email to The Scientist.
Sollberger, still a postdoc in Zychlinsky’s lab, completed his doctorate in 2011 at ETH
Zurich, where he studied inflammasome activation in keratinocytes. He took an interest in
NETs because both inflammasome activation and NET production have been connected to
innate immune cell death.
“Many proteins have been described that influence the production of NETs, but we do not
know how many more exist and how they all interact in the process,” Sollberger tells The Sci-
entist in an email. In this issue, Sollberger and Amulic unpack what we know about NETs, and
what we’ve yet to learn.
1 0 T H E SC I EN T I ST | the-scientist.com
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FROM THE EDITOR
Flesh and Blood and Wonder

The human brain is made of delicate tissue. This fragility makes it all the
more amazing that it can generate thoughts and consider its own function.
BY BOB GRANT
T
he most exciting thing about science is that it can dodge or delay at least some of the
ferry humanity into the unknown. The scientific ravages of time, environmental
method, as a mode of observation piloted by humans insult, and biological malfunction.
for generations, has probed outer space, the depths of the In this issue, two feature
oceans, and the inner reaches of cells, molecules, and articles explore emerging
atoms—our amazing brains at the helm. Never satisfied, the ideas poised to reshape our
three-pound, skull-encased lump of flesh strains to know concept of brain physiology.
more, discover more, solve more. And the universe obliges. Our cover story (pg. 36), by
Unimaginably vast swaths of space lie unexplored; most of Catherine Offord, relays the
the ocean floor remains a mystery; and new insights into latest scientific findings on
the functioning of cells and the nature of subatomic mat- the impact of air pollution on
ter emerge on an almost daily basis. neurodevelopment and cogni-
This almost unfathomable potential for discovery and tive function. In the second article (pg. 30), Ashley Yeager
innovation always rockets to the fore of my own three- offers a fresh perspective on the contribution of lysosomal
pound fleshlump when it comes time to edit our annual dysfunction to the development of Parkinson’s disease.
issue on neuroscience. Elsewhere in the magazine, an opinion article by Colum-
Most scientists and science enthusiasts I’ve met are bia University bioethicist John Loike and his biologist col-
intellectually inflamed by the fact that there is so much league Robert Pollack (pg. 26) exposes a potential pitfall of
out there (and in here) that we don’t know—a passion that humanity’s propensity to problem solve. They point out the
transcends disciplines. And is there any mystery more fas- ethical precariousness of creating human-monkey chimeric
cinating than the functioning of the human brain itself? embryos, as an international team of researchers reported
After all, we carry our brains around with us every day doing this summer, with the ultimate goal of using such
and use them to ferret out the patterns and meanings that hybrids to grow organs for transplant into human patients.
throng around us. Neuroscientists, even more than the rest “There have been several studies showing that the expres-
of us, use theirs to think about thinking. sion of human genes in animal brains affects the animals’
Yet, after millennia of intimate interactions with our own behavior,” they write. What a chilling thought—that we
brains and decades of formal study of the organ, “how the might inadvertently bestow something approaching con-
brain works was and still is a complete mystery,” in the words sciousness on an animal that is likely ill-equipped to han-
of Albert Einstein College of Medicine neuroscientist Kamran dle such an unwanted gift—and destined to be sacrificed for
Khodakhah, this month’s profilee (pg. 54). research or xenotransplantation.
How in the name of Ramón y Cajal can cells, amassed But even with the downsides of limitless curiosity, the
in tangled networks and swapping ions across their mem- frailty of the organ from which it emanates, and the evolu-
branes to propagate waves of electrical potential, result in a tionary compromises that come with growing such com-
thought? How does this sequence of physical events form plex and knowledge-thirsty biological computers, I for one
moving pictures, symphonies, emotions, and inspiration? It wouldn’t trade my brain for the world. There are too many
truly boggles . . . well, the brain. things left to learn. 
But despite the black box around the connection between
the human brain and the mind, it is clear that the flesh-and-
blood organ is subject to all the ills that befall any collection of
ANDRZEJ KRAUZE
biological cells and tissues. Our brains become stuck in ruts.

Our brains degenerate. Our brains deceive us.
The fragility of the human brain takes our investigations
into the nuts and bolts of consciousness beyond the realm Editor-in-Chief
of mere curiosity. Studying brain function can help humans eic@the-scientist.com
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References:
• P.A. Jones & S.B. Baylin, “The • M. Berdasco, M. Esteller, “Aberrant • K. Robertson, “DNA methylation and
epigenomics of cancer,” Cell, 128:683- epigenetic landscape in cancer: how human disease,” Nat Rev Genet, 6(8):597-
692, 2007. cellular identity goes awry,” Dev Cell, 610, 2005.
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QUOTES
Speaking of Science
1 2 3 4 5 6 7 The level of neural activity we
are seeing is unprecedented
in vitro. We are one step
8 9 closer to having a model that
can actually generate these
early stages of a sophisticated
10 11 neural network.
—Alysson Muotri, a University of California,
12
San Diego, neuroscientist who recently published a
Note: The answer grid will include every letter of the alphabet.
paper reporting the spontaneous production of fetal

13 14 15 brain-wave-like oscillations by human cortical
organoids grown from neural stem cells in Petri
16 dishes (Physics World, September 3)
17 18
You can’t uncook a brain any
19
more than you can uncook
20 21 22 an egg.
—University of Maryland paleontologist Thomas Holtz
talking to National Geographic about recent findings
suggesting that tissues in the heads of T. rex and
23 24 other dinosaur species helped the animals control the
temperature of their brains and eyes (September 4)
BY EMILY COX AND HENRY RATHVON
ACROSS DOWN
1. Work ending “QED” 1. Antiparticle with a charge of +1 e
4. Descriptor for some pimpernel 2. Tribe of northeastern Nebraska
8. Magnetic resonance imaging tool 3. Appendage on which a frogfish may walk
9. Tree product foraged by 4. Injury commonly treated with RICE
Algonquians 5. Descendant of Devonian
10 Anxiolytic or antipsychotic, sarcopterygian fish
commonly 6. Milk-producing
13 Target of botanical theft in the film 7. Pointed branch of an antler
Adaptation 11. Colloid hydrogel to avoid in bogs
15 One end of a spectrum 12. Whip-tailed, venom-packing kin of a
17 Seasonal use of Cucurbita pepo shark
(hyph.) 14. One of a cardiac quartet
20 Central dark part of a sunspot 16. Stable point in ecological succession
21 Werewolf, vampire, or zombie 18. Compound with a fruity aroma
JONNY HAWKINS
23 Contradiction evoked by Hempel’s 19. Reservoir of fatty tissue, not water

ravens, e.g. 22. Word before moon, math, age, or wave
24 Blemished by papillomaviruses
Answer key on page 5
10.2019 | T H E S C IE N T IST 1 5
FREEZE FRAME
Caught on Camera
Selected Images of the Day from the-scientist.com
MIND ALTERING
»
A study in rats found that tiny doses of the

psychedelic drug DMT (above) may alleviate
anxiety and depression, but there could be
downsides, such as neuronal atrophy or
weight gain.
Posted: March 11, 2019
SMARTPHONE-CONTROLLED BRAINS »
A device (illustrated here, implanted into a mouse's
brain) can target specific neurons with drugs and
stimulating light over long periods.
Posted: August 7, 2019
» CLUSTERED MEMORIES
The signaling protein Fyn, captured here
using super-resolution microscopy in a
mouse hippocampal neuron, may be driven
to clump by the mutant tau proteins that
are characteristic of Alzheimer's disease.
Posted: August 5, 2019
SMELL CIRCUITS
»
A section sliced from a
mouse piriform cortex, a
key brain region for smell
processing.
Posted: July 24, 2019
» SUPER SPEEDY MICROSCOPY

The Scanned Line Angular Projection
(SLAP) microscope system enables
scientists to record neural activity
patterns in the mouse visual cortex,
millisecond-by-millisecond.
Posted: July 24, 2019
Mind Altering: Lindsay Cameron and Lee Dunlap; Clustered Memories: Meunier Lab/University of Queensland; Smell Circuits: Salk Institute;
Smartphone-Controlled Brains: Korea Advanced Institute of Science and Technology; Super Speedy Microscopy: HHMI Janelia Research Campus
NEWS AND ANALYSIS
Notebook OCTOBER 2019
Nature as drops, heart rate decreases, immune

function improves, and the parasympa-
green space within a certain distance
of a person’s home, White says. But “it’s
Medicine thetic nervous system directs the body

to rest and digest.
not so much where you live; it’s whether
you use it or not.”
M
athew White is on a mission As humans increasingly populate The UK government had just the
to garner Mother Nature the urbanized areas, they are spending less data he was looking for. The Depart-
respect he thinks she deserves and less time in natural environments— ment for Environment, Food & Rural
when it comes to human health. For perhaps to the detriment of their health. Affairs (Defra) carries out an annual
decades, scientists and health-care pro- But before doctors can start advising survey called Monitor of Engagement
fessionals have recognized that expo- their patients to head to the nearest with the Natural Environment to gather
sure to green spaces, such as public park, there is an important outstand- data on how UK residents use their local
parks or forests, is linked with lower ing question, says White, an environ- public green spaces to guide policy deci-
risks of all sorts of ailments common mental psychologist at the University of sions regarding land use. In the early
in the developed world—including car- Exeter Medical School in the UK: How 2010s, however, when White wanted to
UNSPLASH, DAVID NARCU
diovascular disease, obesity, diabetes, much time in nature do you need to use the data to estimate what dose of
and mental distress—and even of mor- generate these apparent benefits? Most nature was needed to show benefits to
tality. Experimental work has demon- of the research that has linked health a person’s health, Defra wasn’t gather-
strated myriad physiological responses outcomes with exposure to the natural ing information on health and wellbe-
that occur when people spend time in world didn’t use frequency or duration ing. So he and his colleagues asked the
natural environments: blood pressure of park visits, but rather the amount of government to add a few questions to
the survey, then waited a couple of years tile organic compounds while filtering benefits. “That was kind of the delib-
for the answers to roll in. out pollutants in the air that can have erate medicalization of the language
With responses from nearly 20,000 detrimental effects on human health. around nature and health,” he says.
participants in the 2014/15 and 2015/16 (See “Brain Fog” on page 36.) The University of Washington’s
survey data, White’s group found the Researchers agree that health-care Gregory Bratman, who recently coau-
answer he was after: spending at least recommendations for people to spend thored a paper calling for city planners
two hours in nature per week was time in nature are probably years away, to monitor and consider the mental
strongly correlated with self-reports but “the movement has begun,” Teresa health benefits of natural environments
of being in good health or having high Horton, an evolutionary and ecological (Sci Adv, 5:eaax0903, 2019), agrees
wellbeing. “I was very surprised, to be physiologist at Northwestern Univer- that White’s study is a valuable addition
honest,” says White. “We had no idea” sity, writes in an email to The Scientist. to the literature. “[W]e think that [the]
that such a clear threshold of time per Several organizations around the world study addresses issues of duration/dose
week would emerge from the data (Sci now promote awareness of nature’s in an interesting way!” he writes in an
Rep, 9:7730, 2019). contribution to health. “Researchers email to The Scientist. “[We] hope that
He was further surprised to learn in the area are working to have their this work helps to add to the conver-
that it didn’t seem to matter how many voices heard by policy makers and [to] sation about urban design and plan-
trips to a park people took, so long as organize the available information to ning when it comes to decision making
they got in their two hours per week. make the case for ‘nature as medicine,’” regarding accessible nature.”
It could be a long visit one day, a cou- says Horton. —Jef Akst
ple of hour-long trips, three visits of 40
minutes, or four half-hour excursions.
Humans are spending less
“They were the big categories that we
were able to look at, and we found that and less time in natural envi- Monkey Noise
it was exactly the same,” says White. ronments—perhaps to the If there’s one thing Bevil Conway has
He and his colleagues speculate that, if detriment of their health. learned from studying the visual corti-
nature’s apparent health benefits are a ces of rhesus macaques, it’s that they’re
result of being able to de-stress, then remarkably like those of humans. The
whatever pattern of green space expo- White is now working with the UK visual cortex is anatomically highly sim-
sure fits one’s schedule is probably the government to obtain more longitudi- ilar in the two species, and macaques
best way to achieve that goal. nal data that will yield further insights and humans show comparable behav-
Researchers have yet to completely into the relationship between the nat- ioral and neural responses to colors
describe the physiological mechanisms ural environment and human health— and images. When a macaque opens its
underlying the apparent health benefits and clearer answers to those critical eyes, “I’m pretty sure he’s seeing what
of time in nature, but relief from stress questions of how much time we need I’m seeing,” says Conway, a neuroscien-
is undoubtedly part of the whole pic- to spend in green space, and how often, tist at the National Institutes of Health
ture, says the University of Edinburgh’s in order to achieve the health benefits (NIH) in Bethesda, Maryland. But does
Catharine Ward Thompson, a landscape it can provide. The weekly two-hour the same hold true for what he hears?
architect who studies how the environ- threshold sits well with Ward Thomp- The question came up in 2014 over a
ment affects behavior and health. She son, who got similar results in a small- beer with Sam Norman-Haignere, then
and others have shown, for example, scale study done for BBC2’s television a graduate student with Josh McDer-
that “natural environments appear to program Trust Me, I’m a Doctor, set to mott and Nancy Kanwisher at MIT,
help the body regulate the functioning air over the coming months. But Jules where Conway headed a lab at the time.
of the hypothalamic pituitary adrenal Pretty, a professor of environment and Norman-Haignere told Conway about
axis, which regulates cortisol secretion society at the University of Essex, thinks his groups’ recent collaborative find-
and whose dysregulation is associ- two hours per week sounds “at the low ing that a particular patch of the human
ated with a range of disease outcomes.” end,” speculating that 30 minutes a day auditory cortex is more sensitive to har-
Other proposed mechanisms include might be more beneficial. monic tones—notes that have an easily
natural environments’ positive effect on Nevertheless, efforts to quantify discernible pitch, such as those played
our ability to focus our attention after exposure to natural environments is a on a piano—than to noisy sounds, such
a long, cognitively demanding day, and step in the right direction, says Pretty, as those made by a drum. When a dozen
an abundance of microorganisms that who with colleagues was the first to use people listened to harmonic tones in a
appear to boost immune function. And the term “a dose of nature” to evaluate functional magnetic resonance imaging
plant life can emit antimicrobial vola- the amount of exposure needed to reap (fMRI) scanner, this patch of the audi-
1 0. 201 9 | T H E S C IE N T IST 1 9
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tory cortex flared up with heightened Conway’s surprise, the same brain region quality of those sounds than macaque
neural activity—substantially more than in macaques showed no significant dif- brains were. “I think I owe Sam a beer,”
when the participants listened to noise. ferences in response to the two sets of Conway remarks.
Musing whether there was an equiv- sounds. In fact, if anything, the monkeys’ What makes the finding all the more
alent region in monkeys that would also brains often had a greater response to surprising is that macaque brains are
respond selectively to harmonic tones over noise than to harmonic tones. perfectly capable of detecting differ-
noise, Conway and Norman-Haignere The contrast between macaques ences in audio frequency. Like humans,
made a bet. “I thought, I’m pretty sure and humans couldn’t have been clearer, macaques have a “tonotopic map” in
that monkeys are going to have that,” Conway says, “but then we had to spend their auditory cortex, a pattern of brain
Conway recalls. Given that the visual about two years full-time collecting regions in which each region responds
systems of humans and macaques are data in order to convince ourselves that to a specific range of frequencies,
so similar, he figured that their audi- the result was real.” although the team’s fMRI data, consis-
tory cortices would be as well. Norman- Initially, the team wondered whether tent with earlier research, indicate that
Haignere wasn’t so sure. Humans rely on the monkeys’ lack of selective response this map is arranged slightly differently
harmonic tones not only in music, but to sounds with a specific pitch could be in macaques than it is in humans (Nat
also to articulate vowels in spoken lan- simply due to their unfamiliarity with Neurosci, 22:1057–60, 2019). The mon-
guage. Because monkeys lack these cul- synthetic sounds. So the researchers keys just don’t show a preference for
tural aspects, the animals may not need repeated the same experiment, this time sounds with a specific pitch in the way
a brain region devoted to perceiving har- playing recordings of real macaque calls that humans do.
monic sounds, reasoned Norman-Haig- and computationally engineered, pitch- To Conway, the findings may explain
nere, who is now a postdoc at Columbia less, or noisy, versions of those vocal- why previous researchers have so far
University. izations. Even then, the fMRI results failed to find neurons that respond
Along with McDermott and Kan- revealed that human auditory cortices selectively to sounds with pitch in
wisher, the two researchers set out to were much more sensitive to the pitch macaques, and have had a hard time
repeat Norman-Haignere’s earlier exper-
iments, this time comparing the human
brain’s response to that of the macaque
(Macaca mulatta). To find out if the two
species’ auditory cortices respond selec-
tively to harmonic tones, the research-
ers used computer software to synthesize
two sets of sounds. The first set of stimuli
was harmonic, based on short sequences
of up to a dozen notes differing in mel-
ody and the range of audio frequencies—
that is, the range from the lowest note
to the highest note in the sequence. The
second, control set of stimuli was noisy,
created from a scrambled kind of sound
much like static on the radio or TV. The
team designed the noise to match the
frequency ranges of the harmonic stim-
uli and thus ensure that the sound sets
were comparable.
Three macaques and four people
took their turn in an fMRI machine as
the scientists played the sounds inside
the scanner and monitored each par-
ticipant’s auditory cortex. As observed
KRAUZE
in the earlier experiments, the human

LINE
auditory cortex showed significantly

ANDRZEJ
CREDIT
greater activity in response to the har-

monic stimuli than to the noise. But to
1 0. 201 9 | T H E S C IE N T IST 2 1
NOTEBOOK
training macaques and other monkey sian composer Tchaikovsky’s Fifth Sym- nectivity in the brain. Several functional
species, such as capuchins, to perform phony. “He taped all of the strings on magnetic resonance imaging studies
certain auditory tasks that humans the instruments so they couldn’t make have reported that people with mild
excel at. “The explanation has histori- any pitches, and all that’s recorded is cognitive impairment (MCI), a condi-
cally been that the monkeys’ memory the noise,” Conway says—the swish- tion that often precedes Alzheimer’s,
cortex works differently, but our finding and tapping of bows against tape. appear to have higher brain activity
ings say no, maybe their memory sys- “I think that’s probably what it sounds levels than their age-matched counter-
tems are just fine, but the auditory cor- like to the monkey.” parts. Researchers have also found signs
tex works differently.” —Katarina Zimmer of such changes in healthy people carry-
The heightened responsiveness of the ing the APOE4 allele, as well as in pre-
human brain to pitch may well have to symptomatic stages of Alzheimer’s in
do with the origins of spoken language
as well as music. “Harmonic structures Overactive rodent models of the disease.
Krishna Singh, a physicist and imag-
are very important to humans,” says
Charles Snowdon, a primatologist who Brains ing neuroscientist at the Cardiff Uni-
versity Brain Research Imaging Center
recently retired from the University of There are approximately 5.6 million (CUBRIC) in the UK, and his colleagues
Wisconsin–Madison. “We’re producing people over the age of 65 living with wanted to investigate this theory fur-
very clear harmonics [with] every vowel Alzheimer’s disease in the United States. ther. Previous studies of brain activity in
sound that we use, and we can’t speak With the population aging, that num- young APOE4 carriers were mostly con-
without producing vowels.” ber is projected to grow to 7.1 million by ducted using small sample sizes, accord-
Macaques do use some harmonic 2025. Researchers know that age, a fam- ing to Singh. But by the mid-2010s, his
vocalizations, for friendly calls and ily history of the disease, and carrying a team had access to neuroimaging data
chatter in particular, but they use them genetic variant known as APOE4 are all from close to 200 participants studied
far less frequently than do humans. associated with a higher chance of devel- at CUBRIC as part of an effort to build a
“The macaques make a greater use of oping the condition. But the biological massive dataset of healthy brains. So the
noisy signals and noisy vocalizations in mechanisms leading to Alzheimer’s are researchers decided to use the data to
their own communication and there- still largely a mystery.
fore it makes sense that they would Over the last decade, scientists
have significant amounts of brain tissue have amassed evidence for a hypothe- WIRED: Young carriers of the APOE4 allele have
brains that are more connected (left, red lines
devoted to processing noise,” Snowdon sis that, prior to developing full-blown
illustrate connections between brain areas) and
says. Humans can make vocalizations Alzheimer’s disease, patients experience active (right, yellow indicates activity) than the
that are noisy too—such as growling— a period of hyperactivity and hypercon- brains of those without the allele.
but they are rarely used, “because we
have language instead,” he adds.
But whether humans are unique
among primates in having a preference
for harmonic sounds—what Conway
and his colleagues call “pitch bias”—
will require further research in other
primate species, Snowdon says. Con-
way and his colleagues plan on examin-
ing tamarin monkeys (genus Saguinus)
next, which make many vocalizations
KRISHNA SINGH, ELIFE, 8:E36011, 2019.
that have harmonic structure.

For now, Conway is left to wonder
what a macaque’s experience of the
acoustic world may be. What would
music sound like to a macaque? It’s
impossible to say, but he imagines a
recent performance by Hong Kong
musical artist Samson Young might
offer one indication. Young had an
orchestra play a “muted” version of Rus-
search for signs of unusual brain activ- In rodents, researchers have up getting impaired in Alzheimer’s may
ity and connectivity in people with the found that hyperactivity can be highly active and connected early in
APOE4 allele. life—long before symptoms of the dis-
increase the production and
Using magnetoencephalography ease appear.
(MEG), a neuroimaging technique that spread of amyloid-ß. “This study adds further evidence
records the magnetic fields generated that hyperactivity and hyperconnec-
by electrical activity in the brain, Singh The researchers next compared the tivity may play an influential role in
and his colleagues had measured rest- results to brain activity and connectiv- Alzheimer’s disease,” says Tal Nuriel,
ing-state brain activity in a group of 183 ity data from a previous neuroimaging a professor of pathology and cell biol-
healthy adults, which included 51 indi- study they had conducted, which found ogy at the Columbia University Medi-
viduals who carried at least one copy of that elderly people with early-stage cal Center who wasn’t involved in the
APOE4. The average age of the partic- Alzheimer’s disease had decreased neu- work. Because this was an observational
ipants was 24 years old, although ages ronal activity and connectivity com- study, the findings can only establish a
ranged from 18 to 65 years old. pared with that of age-matched controls correlation between brain activity and
Analysis of the imaging data (Clin Neurophysiol, 128:2347–57, 2017). Alzheimer’s, Nuriel adds, so it’s still
revealed that, compared with controls, The network of brain areas that dis- unclear whether the hyperactivity and
young APOE4 carriers displayed greater played increased connectivity in young hyperconnectivity observed during the
activity in several regions in the right APOE4 carriers, the team found, par- early stages of the disease are a cause or
side of the brain, including parts of tially overlapped with the brain regions a consequence of pathological changes
what’s known as the default mode net- that exhibited a decrease in connectiv- that lead to neurodegeneration.
work, which is active when a person is ity in people with early-stage Alzheim- Scientists used to think that
not focused on a specific task. A simi- er’s (eLife, 8:e36011, 2019). These find- increased activity was simply a compen-
lar set of brain regions also showed an ings are intriguing, Singh says, because satory effect—the brain trying to make
overall increase in connectivity. they suggest that brain areas that end up for a loss of neurons and synapses,
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NOTEBOOK
says Willem de Haan, a neurologist at For the current study, Singh’s team ga’s. But, bizarrely, all of the teeth jut
the Amsterdam University Medical Cen- also trained a machine-learning algo- forward from the jaws, and the bottom
ter who was not involved in the latest rithm to distinguish APOE4 carriers row of teeth are twisted, spiraling left-
study. “But I think there’s overwhelming from non-carriers based on their MEG ward—just as a narwhal’s tusk does.
evidence that this may actually be patho- data and tested whether it would be able One of Lorenzen’s colleagues, narwhal
logical hyperactivity.” to predict cases of Alzheimer’s. They researcher Mads Peter Heide-Jørgensen
found that while the program was able of the Greenland Institute of Natural
to perform above chance, the effect was Resources, discovered the skull on a
not significant. “In a way, that was kind hunter’s roof in the early 1990s and sug-
This study adds further evi- of encouraging,” Singh says. “Because I gested it represented a hybrid of a nar-
dence that hyperactivity don’t think anybody would predict that whal and a beluga (Mar Mammal Sci,
and hyperconnectivity may we could find a signature [for Alzheim- 9:258–68, 1993). Back then, research-
play an influential role in er’s] in 20- and 30-year-olds.” ers didn’t have the tools to test DNA
Alzheimer’s disease. For now, Singh says, his team’s find- from the skull to determine if that was
ings simply shed light on what might be true. By the time Lorenzen came across
—Tal Nuriel,
Columbia University Medical Center going on in the brains of people with the the skull, however, scientists had devel-
APOE4 allele. There are still a number of oped techniques to extract and analyze
unanswered questions—such as when the nucleic acids from animals that lived
transition from hyper- to hypoconnectiv- up to several thousand years ago. (See
Much of that evidence comes from ity and activity happens, what changes “What’s Old Is New Again,” The Scientist,
animal experiments conducted over the occur in the largely understudied middle- June 2015.) Extracting genetic mate-
last decade or so. In rodents, research- aged cohort, and whether there are dif- rial from the teeth and comparing it
ers have found that hyperactivity can ferences between APOE4 carriers who to DNA from the teeth of narwhal and
increase the production and spread of go on to develop Alzheimer’s and those beluga skulls in the Natural History
amyloid-ß, the peptide that accumu- who don’t. Ultimately, to understand Museum of Denmark collection con-
lates into plaques found in the brains how disruptions in neuronal activity lead firmed Heide-Jørgensen’s hypothesis:
of people with Alzheimer’s—and that to behavioral and cognitive deficits, sci- the whale was indeed a hybrid, the son
amyloid-ß can in turn induce neuro- entists need to decipher what’s going on of a narwhal mom and a beluga dad (Sci
nal hyperactivity. These findings have inside a healthy brain, Singh says. “[We] Rep, 9:7729, 2019).
led some scientists to speculate that require a model of how the brain works— “The fact [that they] even managed
there might be a self-amplifying loop, and those are still in their infancy.” to find the skull is incredibly surprising
where a progressive hyperactivity and —Diana Kwon in itself,” says Travis Park, a postdoc who
build-up of amyloid-ß drives pathologi- studies whales at the Natural History
cal changes associated with the neuro- Museum in London and was not involved
degenerative disease.
Research in humans also supports Museum Finds in the new work. “Personally, I like the
fact that the skull is almost exactly what
the idea that hyperactivity could play Every time Eline Lorenzen pushes open you’d imagine you’d get if you mixed a
a causal role in Alzheimer’s disease. the basement door of the Natural His- beluga and a narwhal together. Genetics
In 2012, researchers at Johns Hopkins tory Museum of Denmark at the Uni- usually doesn’t work out so cleanly so it’s
University treated individuals with MCI versity of Copenhagen, a pungent scent nice to have an example of this phenom-
with the anti-epileptic drug levetirace- of dust, whale blubber, and chemi- enon that is so clearly illustrated.”
tam and found that the therapy sup- cals pummels her nose, the biologist The skull offers the first hard evi-
pressed activity in the hippocampus and tells The Scientist. Hundreds of skele- dence that narwhal-beluga hybrids, or
led to improved memory performance tons and skulls are scattered about the narlugas, can occur, though anecdotal
(Neuron, 74:467–74, 2012). The team is room, but there’s one in particular that evidence suggests there may be oth-
currently testing levetiracetam for MCI Lorenzen has been fixated on since she ers. The hunter who shot the whale
in clinical trials. “I think this is one of became the curator of the mammal col- noted that he had seen it with two sim-
the most interesting results,” says de lection at the museum in 2015. At first ilar-looking companions, and natives
Haan. “It seems to show that by correct- glance, the 30-year-old skull has fea- of western Greenland have a common
ing hyperactivity we can actually find tures similar to the face and jaw bones name for a narwhal-beluga hybrid,
some improvements in patients that of toothed whales such as narwhals and itorsaq. The new discovery underscores
might point to a completely new type belugas. It has a row of chompers on the the importance of museum collections,
of therapy for [Alzheimer’s disease].” top and bottom jaws, just like a belu- Lorenzen says. Museums often house
MEET THE NARLUGA: A skull found in the Nat-

other proteins from samples kept in he says. Sequencing one of the proteins,
ural History Museum of Denmark (center) looks
like a cross between the skull of a beluga whale museums all over the world. In June, amelogenin, can even provide informa-
(left) and that of a narwhal (right). the team analyzed collagen while a col- tion about the sex of a specimen, giving
laborating group analyzed ancient DNA scientists with access to multiple individ-
in the bones of living and extinct sloth uals’ samples the ability to study popula-
what she calls “gems”—“specimens that species kept in museums in the US, the tion dynamics and sex ratios.
lead to major insights into our natu- UK, France, and Argentina. The molec- Lorenzen and her colleagues did not
ral world and the natural history of the ular data completely overturned long- analyze the proteins stored in the narlu-
world.” Natalie Cooper, a researcher held assumptions based on anatomical ga’s teeth, but they did run the teeth
focusing on vertebrate ecology and evo- analyses about the evolutionary relation- through isotopic analyses and compared
lution at the Natural History Museum ships among these animals. Extant three- the results to data from narwhals and
in London, agrees. “Museum collections toed sloths, for instance, appear so ana- belugas to see if they could figure out
in general are a treasure trove of infor- tomically different from other species what the hybrid whale ate. The results
mation for all kinds of research,” she that researchers had put them on a com- weren’t at all what the team expected,
writes in an email to The Scientist. pletely separate evolutionary branch. But says study coauthor Mikkel Skovrind,
Such gems often go unnoticed, how- both the mitochondrial and protein evi- who started the project as a graduate
ever, as collections at natural history dence revealed that three-toed sloths are student and is now a postdoc in Loren-
museums are largely untapped, says in fact part of the Megatherioidea super- zen’s lab. Since the whale’s mom was a
Ross MacPhee, curator of the mammal family, a group that includes the extinct, narwhal, the team thought maybe the
collection at the American Museum of elephant-size ground sloth genus Mega- narluga would learn to feed from her,
Natural History in New York City. The therium. Based on the new relation- eating like a narwhal. These whales cre-
newly analyzed narluga skull is a perfect ships among the sloths, the research- ate a vacuum of sorts to suck in their
example, he adds: it sat in the collection ers concluded that the assumption that food and often eat alone. Belugas, on
for a long time before Lorenzen came extant, tree-inhabiting sloths evolved the other hand, hunt together, corral-
along and decided to study it in depth. from a ground-dwelling ancestor was also ling schools of fish to chow down on or
Although museum specimens present wrong. Rather, the ancestral sloth might snagging octopuses, crabs, and snails.
certain challenges for biological analy- have lived both in the trees and on the According to the analysis, however, the
ses—their storage and transport might ground (Nat Ecol Evol, 3:1121–30, 2019; narluga didn’t eat like either parent—
make them prone to contamination, for Curr Biol, 29:2031–2042.e6, 2019). it was a bottom feeder. “[It’s an] even
example, while treatment with chemicals One particularly promising avenue for more unique individual than previously
and changing temperature and humidity unlocking museum specimens’ secrets is thought,” Skovrind says.
conditions could damage genetic mate- extracting proteins from teeth, MacPhee The findings are a reminder of
MIKKEL HØEGH POST
rial—techniques for extracting informa- says. It’s long been known that certain what’s to learn from studying dusty old
tion from such samples are improving, proteins involved in producing enamel bones. “Who knows what else is in these
MacPhee says. in the teeth get sealed into pockets in the collections?” Lorenzen says. “We just
He and his colleagues have been enamel that protect them against deg- need to ask the right questions.”
working to collect ancient collagen and radation like they’re in a “locked vault,” —Ashley Yeager
1 0. 201 9 | T H E S C IE N T IST 2 5
CRITIC AT LARGE
Organoids, Not Chimeras

Scientists are devising human-animal hybrids for harvesting human organs, but lab-derived
mini-organs are a less ethically fraught solution to meeting the need for transplantation.
BY JOHN D. LOIKE AND ROBERT POLLACK
R
ecent studies have shown that even though about
50 percent of adults in the United States have regis-
tered as organ donors, more than 100,000 people in
the United States are waiting for a transplant, and many will
die waiting. The increasing scarcity of human organ donors
has driven research scientists to examine options other than
donation from deceased patients, such as xenotransplanta-
tion, or the use of human organs that were grown in animals.
Xenotransplantation experiments are being conducted in
sheep, pigs, and, most recently, in nonhuman primates. In
July, a group of scientists from Spain and the US claimed to
have successfully created embryos containing both monkey
and human cells as a preliminary step in examining the use
of such human-monkey chimeras as hosts to produce human
organs. According to published reports, the research was con-
ducted in China “to avoid legal issues.”
Xenotransplantation experiments rely on applying gene
editing and stem cell technologies to address the potential
of tissue rejection by the recipient’s immune system and the
possibility of transmitting infectious diseases from donor
animals to human patients. Scientists are years away from
solving technical hurdles, such as developing a human vas-
cular system in these organs, before beginning clinical trials
in human beings.
There are many scientific and ethical issues emerging
from xenotransplantation technologies. For example, will
it be ethical to do the experiments necessary to ensure that
human stem cells introduced into the animal embryo to form
a kidney will not migrate to the animal’s brain to generate
human-like behaviors in such animal chimeras? at birth, there is no need to be ethically concerned that their
There have been several studies showing that the expres- nervous systems might develop any human traits. We believe
sion of human genes in animal brains affects the ani- this argument does not adequately address the ethical issue.
mals’ behavior. In April of 2019, scientists in China pub- By accepting the necessity of their destruction as an element
lished a study in which they claimed to have introduced a of xenotransplantation research, scientists would have had to
DNA sequence expressed in human brains into macaques decide in advance that the potential personhood of human-
and found the monkeys had better short-term memory and animal chimeras could be ignored for the short run, leaving
shorter reaction times than control animals. Even though the open the possibility of a later discovery that such chimeras
monkeys were not chimeras as xenotransplantation hosts might in fact have human traits.
© ISTOCK.COM, SEAMARTINI
would be, it is clear that new ethical boundaries are being Equally important is the trend in many countries, such as
pushed in animal studies. To date, no human-animal hybrid Argentina, that recognize personhood status of nonhuman pri-
embryos have been brought to term, and the US National mates and ban their “incarceration” in zoos. Human-monkey
Institutes of Health has had a moratorium on funding such chimeras would properly have even greater or enhanced per-
work since 2015. sonhood status than normal monkeys.
Some ethicists have argued that because these animal- Bioethicists also may argue that the need to address the
human chimeras will be sacrificed for their human organs organ transplant crisis may override our ethical debates on
Don’t
whether such chimeras are human-like or whether they have
personhood status. That argument has merit, but even if
the ethical debate were to be set aside, the decision to forge
lose
ahead with organ transplants from human-monkey chimeras
to humans would not justify the use of a technology that is
still full of unanswered scientific questions.
There is an alternative that we believe avoids several of
your
these scientific and ethical concerns—the use of stem cell
technology to develop human organoids. We have seen a great
deal of excitement following the development of artificial ova-
ries for women with cancer who had been treated with high-
nerve
dose chemotherapy. The potential fear of being left infertile
from these treatments can be mitigated by developing artifi-
cial ovaries in vitro from their own stem cells and transplant-
over a
ing those lab-grown organs back into the patients after they
are cured of their cancers.
There are other labs that are using 3-D technologies to
develop mini-organs such as livers, placentas, stomachs,
poor
lymph nodes, and even brains. Malfunctioning or diseased
organs can be removed from patients and their cells can be
gently destroyed with detergent, leaving behind a translu-
marker
cent matrix that serves as a platform for organ-directed plu-
ripotent cells to differentiate into new organ-specific cells
in vivo. These stem cells can further develop into organoid
structures within weeks as opposed to the months required to
develop human organs in pigs, sheep, or monkeys. California-
based biotech firm Organovo recently presented new findings
on a stem cell–based approach to developing kidney tissue
Antibodies have been proven to
using its 3-D bioprinting platform. The company reported
that the automated production of complex kidney organoids
help achieve successful identification
could inform in vitro disease modeling and could support the and characterization of neurons. Use
development of treatments for patients with renal disease. our neuronal and glial marker
Most of the scientists in the field are currently using antibodies to correctly identify and
human organoids to study human disease, but these tissues better understand the function and
also have the potential to serve as organ transplants. Using
connectivity of your nerve cells.
human organoids in transplants will avoid the basic problems
of tissue rejection and the possible introduction of pathogens
or oncogenes from animals into human recipients. Equally
important, there is less concern that nonbrain human organ- Learn more at: rockland-inc.com/neuro
oids will generate novel central nervous system neurons that
could alter the behaviors of their recipients.
Without making a final ethical judgment on future
research using human-animal chimeras as a source of human
transplantable organs, we believe that developing human
organoids might offer a faster and sounder platform, both
scientifically and ethically, to address the mounting crisis in
organ transplantation. g
John D. Loike, a professor of biology at Touro College and Uni-

versity Systems, writes a regular column on bioethics for The
Scientist. Robert Pollack is a professor of biological sciences
at Columbia University. A version of this opinion article
appeared on the-scientist.com on August 23, 2019.
MODUS OPERANDI
Getting on the Nerves

An implantable wireless device with microfluidic and optical components allows remote
control of individual nerve fibers in a mouse’s extremities.
BY RUTH WILLIAMS
S
timulating or inhibiting particular neurons
in an animal and observing the effects
is the basis of countless neuroscience Microfluidic
Base station
channels
experiments. To achieve such control, research- Cuff
ers have two key methods at their disposal:
pharmacology and optogenetics.
But methods for targeting drugs or light Sciatic nerve
to specific neurons may interfere with the
behavior or neuronal activity under observation.
Animals generally need to be handled for drug
injections, for example, while light stimulation
often requires illuminating an animal’s skin or
tethering the creature to a fiber optic cable. NEURON REMOTE CONTROL: Peripheral nerves in freely moving mice can be manipulated with
To minimize disturbance to animals during drugs, light, or a combination of both using a new implantable optofluidic device. The base unit of the
device, which houses electronics for wireless control of an LED and microfluidic pumps, is sutured to
experiments, engineers are developing small
the back of an anesthetized mouse. The thin, soft, flexible cuff is inserted into the body and attached
wireless devices containing light-emitting to the nerve of interest (here, the sciatic nerve). Once the mouse recovers from surgery, it can roam
diodes (LEDs) or microfluidic channels (for about an enclosure while a researcher controls nerve activity from afar.
drug delivery) that can be surgically implanted.
But, as yet, such devices are inflexible, bulky,
and only suitable for brain stimulation, for end of the cuff, and collect power transmitted pain neurons, switching on the LED induced
which they can be anchored to the skull, says wirelessly from outside the animal, eliminating discomfort. Conversely, pumping an anesthetic
neuroscientist Robert Gereau of Washington the need for a battery. Running the length of to the sciatic nerve of wild-type mice
University in St. Louis. To stimulate nerves in the the cuff are microfluidic channels through increased the animals’ tolerance to pain.
rest of the body, he adds, you need a device that which the reservoir fluids are pumped, and a The researchers “take two very distinct
“moves with the tissue. It can’t be rigid.” wire connects the base to the LED. The base technologies . . . and combine them into one
To that end, Gereau and colleagues have is sutured onto an animal’s back, with the cuff single device,” says pain researcher Gregory
built a device that consists of a tiny base unit inserted into the body and attached to a nerve Corder of the University of Pennsylvania, who
(5 mm in diameter) and a long, soft, flexible bundle of interest. was not involved in the research. It can be
cuff (approximately 20 mm x 2 mm). The base In proof-of-principle experiments, the used “in an awake, behaving animal without
unit houses microfluidic reservoirs and pumps team hooked up the device to the sciatic an experimenter being present in the room,”
as well as electronics that control the pumps, nerve in mice’s hind legs and showed that in he adds. “It’s extremely impressive.” (Sci Adv,
illuminate a microscale LED sitting at the far animals engineered to have light-sensitive 5:eaaw5296, 2019) g
AT A GLANCE
PERIPHERAL NERVE EASE POTENTIAL FOR BEHAVIORAL TEMPORAL PRECISION SPATIAL PRECISION OF
STIMULATION TECHNIQUE CONFOUNDING OF NERVE ACTIVATION NERVE ACTIVATION
Existing optogenetic and pharmaco- High. Products and protocols for light High. Researcher interaction is often High for light stimulation; low Low
logical methods stimulation and injections are established required, which may cause fear. for drug injections
© GEORGE RETSECK
and readily available.
Implanted optofluidic device Low. Surgery is required to implant Unlikely. The device is remotely High for both light- and drug- High. The device is directly
the device, which is not yet widely controlled. induced activations attached to nerve bundles.
available.
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The Destructive
Path of Parkinson’s
The disease might not start with tangles of α-synuclein proteins,
but with the inability of cells to rid themselves of toxic materials.
BY ASHLEY YEAGER
D
uring her time as a postdoc at antibodies that bound to various pro- did see α-synuclein in the Lewy bodies,
the University of Basel in Swit- teins revealed that the clumps contained as well, but the cores of the structures
zerland, Sarah Shahmoradian a protein called α-synuclein, and after weren’t composed of twisted and tan-
decided to study the abnormal aggregates more work probing Lewy bodies, scien- gled fibrils as researchers had thought.
of protein that develop inside nerve cells tists developed a rough sketch of their Instead, the protein was intermingled
and contribute to Parkinson’s disease. structure—essentially, a dense mass sur- with other cellular material. 1
The protein clumps develop over time in rounded by a halo of twisted filaments The study is one of many that raise
the brains of Parkinson’s patients, lead- of α-synuclein. It’s these filaments, questions about the prevailing idea that
ing some scientists to think they wreak known as fibrils, that Shahmoradian α-synuclein accumulation is the under-
havoc on nerve cells, causing severe dam- and her colleagues were most interested lying cause of the neurodegeneration in
age and hastening their death. A fresh to analyze in postmortem human brains. Parkinson’s disease. Rather, α-synuclein
look at the clumps, called Lewy bodies, Fibrils had been repeatedly produced in buildup may be just one symptom of a more
with cutting-edge microscopy tools could cultured cells and in animal models, but fundamental problem: the cells’ inability
reveal insights that might lead to new no one had ever gotten a clear view of to break down excess lipids and proteins,
treatments for Parkinson’s, Shahmora- them in human brain tissue. including α-synuclein. Some Parkinson’s
dian recalls thinking. “The original goal “We were originally looking for patients carry mutations in genes associ-
was to really find out what the building fibrils,” Shahmoradian says, “but unex- ated with lysosomal function, and studies
blocks of Lewy bodies are, what they are pectedly, we found an abundance of . . . in mice have revealed that natural aging
made of, and what they actually look like.” mitochondria, other organelles, and leads to the build-up of lipids associated
The clumps were first identified in lipid membranes [in the Lewy bod- with Parkinson’s disease.2 These findings
© LISA CLARK
the early 1900s, appearing as abnormal ies].” The researchers also found evi- have led a small but growing set of scien-
material in nerve cells viewed under a dence of lysosomes, organelles that tists to propose that for a vast majority of
microscope. Additional studies using facilitate cellular waste removal. They Parkinson’s patients, the disease is funda-
1 0. 201 9 | T H E S C IE N T IST 3 1
mentally a cellular machinery problem, Based on additional probes into dis- Laboratory investigations in the 1990s
not a protein problem. eased patients’ brains, neurologists found suggested that Lewy bodies were com-
“In this new story, α-synuclein is actually Lewy bodies to be particularly common posed of α-synuclein, while early explora-
a reaction to the root cause of Parkinson’s,” in the substantia nigra, a brain region tions of the genetics of Parkinson’s pub-
Ole Isacson, a neuroscientist at Harvard that sits in the center of the head directly lished around the same time revealed
Medical School, tells The Scientist. behind the eyes. It’s where many of the that patients with an inherited form of
neurons that produce dopamine, a neu- the disease often carried mutations in the
A gut feeling rotransmitter involved in movement and SNCA gene, which encodes α-synuclein.3
In 1912, Fritz Heinrich Lewy, a doctor learning and in regulating mood, orig- Together, the pathology and genetic find-
working in Berlin, studied the brains of inate. These neurons send signals to ings suggested that α-synuclein might be
patients who had died from Parkinson’s another brain region called the striatum, the pathologic protein underlying Parkin-
disease (then known as shaking palsy) forming a neural pathway that facilitates son’s disease, pathologist Kelvin Luk of the
and found odd clumps of proteins in muscle motion; in Parkinson’s disease, it’s Perelman School of Medicine at the Uni-
their nerve cells. Several years later, the dopamine neurons in the substantia versity of Pennsylvania tells The Scientist.
Spanish neurologist Gonzalo Rodrí- nigra that are damaged or destroyed. Peo- In the early 2000s, Goethe Univer-
guez Lafora, who had identified the pro- ple with Parkinson’s typically have trouble sity Frankfurt neuroanatomist Heiko
tein inclusions in the brain of another with balance and walking, and they often Braak built on that work, observing that
patient who had died of shaking palsy, suffer from tremors in the hands or fin- α-synuclein accumulation didn’t just
dubbed them Lewy bodies. gers and other involuntary movements. occur in the brain. Postmortem analy-
FAILURE TO CLEAR HEALTHY CELLS

Many patients with Parkinson’s disease Unneeded proteins, lipids,
and other cellular materials
carry gene variants that lie at the root of
are typically gathered into
problems with cellular waste-clearing vacuoles, which fuse with Lysosome
processes, mediated by the lysosome. lysosomes to clear the cells
One of the proteins that must be cleared of the waste.
from cells is α-synuclein—the protein
that scientists have long-fingered as a
prime pathogenic suspect in Parkinson’s.
When α-synuclein isn’t cleared from
neurons, it can become misfolded and
clump together in Lewy bodies that
prevent these cells from functioning Vacuole
and ultimately cause them to die, lead-
Proteins
ing to telltale symptoms of the disease. α-synuclein
Lipids
But α-synuclein is not the only material
accumulating in the neuron when the
lysosomes aren’t functioning properly;
Lewy bodies are composed of a mix of
cellular material.
Further evidence that Parkinson’s Axon Mitochondrion
disease might be driven by problems
with cellular waste-clearing processes
comes from genes that are related to
mitochondrial dysfunction. Certain
gene variants related to Parkinson’s can
cause the mitochondria to form reactive Presynaptic
© LISA CLARK
terminal
oxygen species and other compounds
that can damage the lysosome, leading
to problems with waste removal.
ses showed that it had accumulated in Staffan Holmqvist, then at Lund Univer- These studies led researchers to pro-
the nasal cavity, in nerves in the throat, sity in Sweden, and colleagues showed pose that Parkinson’s might start in
and in the gut of deceased Parkinson’s that if they injected α-synuclein into the the gut years before the disease mani-
patients. 4 Braak’s postmortem obser- guts of rats, the protein could travel up fested as neurodegeneration in the brain.
vations also showed that aggregations the vagus nerve to their brains.6 And this Despite the growing popularity of this
of the protein appeared in the vagus June, Johns Hopkins University neuro- hypothesis, however, new work is chal-
nerve—a superhighway of nerve-fiber scientist Ted Dawson and an interna- lenging the idea. For example, accord-
bundles running between the brain and tional team of researchers showed that ing to one study, there is no change in
various organs of the body, including the fibrillar, pathological form of the the risk of the disease among patients
the heart, lungs, and gut. He concluded protein can travel in a similar way in who have had their vagal nerves cut to
that some type of pathogen causing the mice and lead to Parkinson’s-like symp- stop the development of gastric ulcers.8
neuronal cell damage seen in Parkin- toms in the rodents. 7 “Not only do the Moreover, in a recent study of more
son’s could invade through the nose or mice have the motor features of Parkin- than 2,000 Parkinson’s patients, only
gut and then travel up to the brain via son’s disease, they also have the nonmo- 0.05 percent had mutations in the SNCA
the vagus nerve.5 tor features,” Dawson told The Scien- gene, leaving scientists questioning how
Researchers then started to wonder tist at the time. “They’ve got cognitive α-synuclein accumulates in the other
if aggregates of α-synuclein might move dysfunction, anxiety, depression, prob- 99.95 percent of cases, and therefore
through the body in a similar way—and lems with smell”—all symptoms seen in if the protein is, in fact, at the root of
studies have shown that it can. In 2014, human patients with Parkinson’s. Parkinson’s disease.
DISEASED CELLS
In the neurons of Parkinson’s patients,
something appears to have gone
wrong with the cellular waste-clearing
Damaged
process. Reactive oxygen species lysosome
(ROS) released from mitochondria
may play a role, damaging lysosomes.
If the lysosomes don’t function
Lewy body
properly, then cellular waste products
are left in the cell to accumulate.
ROS
Axon
Vacuole
α-synuclein
Presynaptic
terminal
Hints that something other than reported that more than 50 percent of is halted, and memory improves in mice.14
α-synuclein might be to blame started Parkinson’s disease patients carry a puta- There are now even a few clinical trials for
to circulate in the late 1990s and early tively damaging mutation in one or more Parkinson’s disease drugs that target faulty
2000s. In 2004, for example, Enza Maria genes that are known to cause lysosomal lysosome function instead of α-synuclein
Valente, then at the Mendel Institute in storage diseases,11 inherited metabolic dis- aggregation, Klein says. Considering all
Rome, and colleagues found that early- orders caused by enzyme deficiencies that the evidence together, “we really had . . .
onset Parkinson’s disease appeared to be allow the buildup of toxic materials inside the guts to [say] that Parkinson’s is a lyso-
caused by mutations in the gene PINK1, cells. That result “sent a signal to the com- somal disease.”
which plays a role in mitochondrial munity that we need to be looking at [the Even as Klein and Mazzulli were
function. 9 (See “Malfunctioning Mito- lysosome] critically to try and understand collating the findings for their review,
chondria” on opposite page.) In 2009, what the mechanism is . . . [that] makes researchers were publishing more data
Ellen Sidransky, a neurogeneticist at dopaminergic neurons so dependent on to support their argument. Isacson and
the National Human Genome Research normal lysosomal function,” Frances Platt, Platt reported in 2018, for example, that
Institute, and colleagues reported results an expert in lysosomal storage diseases at in healthy mice, aging alone causes an
suggesting that Parkinson’s might stem the University of Oxford, tells The Scientist. accumulation of glycolipids also involved
As it turned out, Isacson was already in Parkinson’s disease.2 Later that same
at work on the question, and in 2015 his year, Isacson and another group of col-
In this new story, α-synuclein team found that the enzymatic activity leagues published data showing that
is actually a reaction to the of GBA decreased in mice and in human aging causes α-synuclein and lipids to
root cause of Parkinson’s. dopamine neurons (examined postmor- stick to each other and then to the mem-
tem) with increasing age. This resulted branes of dopamine-containing vesicles
—Ole Isacson, Harvard Medical School
in the accumulation of glycolipids that in neurons.15 These results reveal how
could disrupt neuronal function, suggest- natural aging changes lipids and lyso-
more from a fundamental cellular probing that natural aging alone was enough somes, accelerating neuronal degenera-
lem than from the accumulation of a to reduce GBA activity, leading to lipid tion—a direct challenge to the hypothe-
particular protein. In an analysis of a buildup.12 That same year, Isacson’s group sis that Parkinson’s is primarily a protein
genetic panel taken from more than also showed that blocking the activity of problem, as changes to the lipids and
5,600 Parkinson’s patients and more the GBA enzyme—a proxy for lysosomal lysosomes would precede or provoke
than 4,800 healthy individuals from dysfunction—caused a dramatic accumu- α-synuclein aggregation.
around the world, the team found genes lation of α-synuclein in neurons, spurring Last December, Isacson and col-
associated with Parkinson’s disease that neuroinflammation, which is characteris- leagues found more evidence to chal-
encoded lysosomal components. For tic of Parkinson’s.13 lenge the proteinopathy view of Parkin-
example, 15 percent of Ashkenazi Jewish “The genetics, the biochemistry, and son’s. In the substantia nigra of deceased
patients with the disease and 3 percent the cell biology tell us that the lysosome patients, levels of a glycoprotein called
of non–Ashkenazi Jewish patients had plays a major role in disease pathogen- GPNMB were elevated compared with
a mutation in a gene called GBA, which esis” of Parkinson’s, cell and molecular age-matched controls. 16 Transgenic
encodes a protein active in lysosomes biologist Andres Klein of Universidad mice modeling Parkinson’s with excess
that helps clear cellular waste. 10 The del Desarrollo in Santiago, Chile, tells α-synuclein did not show higher levels
faulty protein made by the mutated GBA The Scientist. of GPNMB, but when wild-type mice
gene prevents the breakdown of an inter- were given lipid-based drugs to simu-
mediary compound in the metabolism of A waste problem late lysosomal dysfunction, their levels of
carbohydrate-containing lipids, or glyco- In an August 2018 review published in GPNMB skyrocketed, mirroring relative
lipids. Other mutations in the same gene Brain, Klein and neuroscientist Joseph levels of the glycoprotein in the Parkin-
can cause the metabolic disorder known Mazzulli of Northwestern’s Feinberg son’s patients’ brains. An accumulation of
as Gaucher disease, which can lead to School of Medicine laid out all of the evi- stray lipids in nerve cells might be enough
brain damage, among numerous other dence for Parkinson’s disease as a lyso- to spur inflammation and cause neuronal
outcomes, strengthening the suspicion somal disorder. In animal models of the damage and death, Isacson says.
that lysosomes play a role in Parkinson’s. disease and in neurons cultured from If Parkinson’s is in fact a lysosomal
Intrigued by the results, Baylor College induced pluripotent stem cells (iPSCs) disorder, it raises the question of whether
of Medicine geneticist Joshua Shulman of Parkinson’s patients, when researchers “some of the treatments that are being
looked into the genomes of Parkinson’s treat the lysosome to correct for the cell developed for lysosomal diseases may
patients for mutations in lysosomal genes clearance problems, the toxic buildup of unexpectedly turn out to be useful in Par-
other than GBA. In 2017, he and colleagues lipids and proteins, including α-synuclein, kinson’s,” Platt notes. Several clinical tri-
als have begun or are being planned to test still think α-synuclein is their major con- 7. S. Kim et al., “Transneuronal propagation of
pathologic α-synuclein from the gut to the brain
whether drugs already used to treat well- stituent. “It’s hard to ignore synuclein,”
models Parkinson’s disease,” Neuron, 103:P627–
characterized lysosomal storage disorders Dawson agrees. But he adds that more 41, 2019.
might also work as Parkinson’s therapeu- researchers are starting to integrate the 8. B. Liu et al., “Vagotomy and Parkinson disease,”
tics. One sponsored by University College data on mitochondrial and lysosomal dys- Neurology, 88:1996–2002, 2017.
London is testing ambroxol, a drug that function into their ideas on the disease. 9. E. Valente et al., “Hereditary early-onset
Parkinson’s disease caused by mutations
reduces mucus production in the respira- They are realizing “it’s all intertwined.” g
in PINK1,” Science, 304:1158–60, 2004.
tory tract, for its ability to increase activ- 10. E. Sidransky et al., “Multicenter analysis of
ity of the lysosomal enzyme GBA and, as a glucocerebrosidase mutations in Parkinson’s
result, reduce the buildup of excess lipids References disease,” N Engl J Med, 361:1651–61, 2009.
and proteins, such as α-synuclein. Another, 1. S.H. Shahmoradian et al., “Lewy pathology 11. L.A. Robak et al., “Excessive burden of lysosomal
in Parkinson’s disease consists of crowded storage disorder gene variants in Parkinson’s
sponsored by Sanofi Genzyme, is recruiting
organelles and lipid membranes,” Nat Neurosci, disease,” Brain, 140:3191–203, 2017.
Parkinson’s patients with GBA mutations 22:1099–109, 2019. 12. E.M. Rocha et al., “Progressive decline of
and treating them with GZ/SAR402671, a 2. P.J. Hallett et al., “Glycosphingolipid levels and glucocerebrosidase in aging and Parkinson’s
drug designed to lower glucosylceramide, a glucocerebrosidase activity are altered in normal disease,” Ann Clin Transl Neurol, 2:433–38, 2015.
compound that accumulates as a result of aging of the mouse brain,” Neurobiol Aging, 13. E.M. Rocha et al., “Sustained systemic
67:189–200, 2018. glucocerebrosidase inhibition induces brain
lysosomal damage and can cause the aggre-
3. M.H. Polymeropoulos et al., “Mutation in the α-synuclein aggregation, microglia and
gation of α-synuclein. α-synuclein gene identified in families with complement C1q activation in mice,” Antioxid
By and large, however, the field seems Parkinson’s disease.” Science, 276:2045–47, 1997. Redox Signal, 23:550–64, 2015.
to be sticking with the idea of α-synuclein 4. H. Braak et al., “Staging of brain pathology 14. S.P. Sardi et al., “Glucosylceramide
as the underlying pathological driver of related to sporadic Parkinson’s disease,” Neurobiol synthase inhibition alleviates aberrations in
Aging, 24:197–211, 2003. synucleinopathy models,” PNAS, 114:2699–704,
Parkinson’s disease, Isacson says. Luk
5. H. Braak et al., “Idiopathic Parkinson’s disease: 2017.
concedes the field’s focus on the protein is possible routes by which vulnerable neuronal 15. O.R. Brekk et al., “Lipid-dependent deposition of
probably not going to shift any time soon, types may be subject to neuroinvasion by α-synuclein and Tau on neuronal Secretogranin
mainly because an overwhelming major- an unknown pathogen,” J Neural Transm, II-positive vesicular membranes with age,” Sci
ity of Parkinson’s patients have Lewy 110:517–36, 2003. Rep, 8:15207, 2018.
6. S. Holmqvist et al., “Direct evidence of Parkinson 16. E.B. Moloney, “The glycoprotein GPNMB is
bodies. “It’s very hard to find Parkin-
pathology spread from the gastrointestinal selectively elevated in the substantia nigra of
son’s cases that don’t have Lewy pathol- tract to the brain in rats,” Acta Neuropathol, Parkinson’s disease patients and increases after
ogy,” Luk says, and notes most scientists 128:805–20, 2014. lysosomal stress,” Neurobiol Dis, 120:1–11, 2018.
MALFUNCTIONING MITOCHONDRIA
While some researchers are studying lysosomal dysfunction as a poten- nergic neurons derived in culture from samples of Parkinson’s patients’
tial cause of Parkinson’s disease, others have been probing the connec- skin cells, Northwestern University Feinberg School of Medicine neu-
tions turned up by genetic studies, such as the link between mitochon- rogeneticist Dimitri Krainc and colleagues found that reactive oxygen
drial dysfunction and α-synuclein accumulation. For example, immature species in the cells damaged mitochondria and that oxidized dopamine
human neurons carrying knockout mutations in the PINK1 gene, which began to build up. This caused a drop in GBA enzyme activity, lyso-
encodes a protein involved in mitochondrial function and has been somal dysfunction, and eventually α-synuclein accumulation (Science,
linked to Parkinson’s, died sooner than cells without the mutation. 357:1255–61, 2017).
Recently, researchers found that the PINK1 protein is vital to stabiliz- “Lipid regulation, lipid function, and lysosomal function are tightly
ing another protein, MIC60, which is essential for mitochondria to gen- regulated normally,” says the University of Oxford’s Frances Platt, who
erate energy. Young fruit flies that didn’t produce PINK1, and therefore studies lysosomal storage diseases. “If you cause an imbalance . . . you
didn’t have healthy mitochondria, didn’t crawl well and died relatively end up causing collateral problems for other organelles, and ultimately
early in adulthood. But when researchers overexpressed the protein you trigger cell death pathways” and neurodegeneration.
MIC60 in the brains of flies lacking PINK1, the animals’ neuronal mito- Imaging living cells, Krainc’s group has found that lysosomes and
chondria started generating more energy—enough to prevent dopa- mitochondria come into direct contact in a cell (Nature, 554:382–86,
mine-producing nerve cells from dying (Mol Cell, 69:P744–56.E6, 2018). 2018), providing a mechanism by which damaged mitochondria might
The study suggests that mitochondrial problems might spark a cascade interact with and disrupt the function of lysosomal enzymes. The work,
of cellular issues that cause Parkinson’s disease. says Universidad del Desarrollo’s Andres Klein, suggests that problems
Other new research indicates that Parkinson’s could stem from a with the mitochondria and lysosomes may create a problematic loop
combination of lysosomal and mitochondrial problems. Using dopami- that lies at the heart of Parkinson’s disease.
BRAIN
FOG
Air pollution likely has detrimental
effects on human neurodevelopment
and cognitive function.
BY CATHERINE OFFORD
© ISTOCK.COM, DEBIBISHOP
W
hen Lilian Calderón- ogy of proteins such as amyloid-ß is not umenting the cognitive changes occur-
Garcidueñas discov- fully understood, Calderón-Garcidueñas ring in human populations exposed
ered abundant hall- argues that air pollution is the most to air pollution, and by looking inside
marks of Alzheimer’s likely culprit behind the development of human and animal brains to try to deci-
disease in a batch the abnormalities she saw in her post- pher the underlying mechanisms. “This
of human brain samples a few years mortem samples—plus many other det- is the beginnings of a whole new field,”
ago, she initially wasn’t sure what to rimental changes to the brains of Mexico says Caleb Finch, a gerontologist at the
make of it. The University of Montana City’s residents. University of Southern California (USC).
neuropathologist had been studying the Once controversial, the theory that “This is like tobacco research and cancer
brains as part of her research on envi- air pollution damages the brain is gain- 70 years ago.”
ronmental effects on neural developing traction in the research community.
ment, and this particular set of samples Although government officials in Mexico In the mind
came from autopsy examinations car- have worked to improve air quality since Yale University health economist Xi
ried out on people who had died sud- the 1990s, the last couple of years have Chen got interested in how poor air qual-
denly in Mexico City, where she used seen thick smog descend over buildings, ity can affect the brain years ago as he
to work as a researcher and physi- forcing periodic school and office clo- started thinking about pollution’s cost to
cian. Although Calderón-Garcidueñas sures to stop people from venturing into human society. To date, most researchers
had collected much of the tissue herself the toxic air. And it’s not just Mexico. As have focused on evaluating air pollution’s
while attending the autopsies in Mex- the rest of the world’s urban areas and effects on mortality or respiratory health.
ico, the light-microscope slides she was their associated congestion continue to But changes to cognition and social
analyzing had been prepared by her col- expand, most countries are witnessing behavior also have profound influences
leagues, so she was in the dark about increases in airborne contaminants, from on a person’s daily life, from financial
what patient each sample came from. noxious gases such as nitrogen oxides decision making to relationships, Chen
By the end of the project, she’d iden- and ozone to fine particulate matter such argues. “Society can lose even more with
tified accumulations of the Alzheim- as dust, soot, and nanospheres of met- [this] more indirect link.”
er’s disease–associated proteins amy- als that penetrate deep into the human A couple years ago, Chen teamed up
loid-ß and hyperphosphorylated tau in body. One 2018 report by the Boston- with colleagues in Beijing to assess such
almost all of the 203 brains she studied. based nonprofit Health Effects Institute effects in adults exposed to air pollution
“When I started opening envelopes to warned that up to 95 percent of the peo- across China. The researchers used the
see who [each sample] belonged to . . . ple on Earth were breathing unsafe air. China Family Panel Studies—a longitu-
I was devastated,” she says. The people These trends are matched by an dinal dataset that includes verbal and
whose brains she’d been studying were increasing incidence of respiratory and math test scores from more than 20,000
not only adults, but teens and even chil- cardiovascular problems—consequences adults across the country between 2010
dren. The youngest was 11 months old.1 of the inflammation and tissue damage and 2014—and estimated air pollution
“My first thought was, ‘What am I going provoked by multiple components of air exposure for each participant’s town on
to do with this? What am I going to tell pollution. At the World Health Organiza- the days of testing using national records
people?’” she says. “I was not expecting tion’s inaugural conference on air pollution of nitrogen dioxide, sulfur dioxide, and
such a devastating, extreme pathology.” last fall, health officials gathered to dis- particulate matter levels. They also
Despite her shock, Calderón- cuss data showing that dirty air is impli- assembled data on other variables such
Garcidueñas had a reason to be on cated in more than 7 million deaths per as daily weather and the participants’
the lookout for signs of a disease usu- year, with the organization’s director- educational background.
ally associated with the elderly in these general, Tedros Adhanom Ghebreyesus, The analysis revealed that the higher
samples. For the last three decades, she’d declaring the situation a “silent public a person’s estimated exposure to air pol-
been studying the health effects of Mex- health emergency.” Only in the last few lution, the greater the decline in his test
ico City’s notoriously polluted air—a years, however, have researchers begun scores over the study period.2 Less edu-
blight that earned the capital the dubi- raising the alarm about links between cated men over 45 years old were the
ous distinction of most polluted megacity humans’ exposure to air pollution and worst-affected group, and verbal scores
on the planet from the United Nations brain function. were hit harder than math scores. The
in 1992. During that time, she’s discov- Epidemiologists, psychologists, and large sample size and the multiple time
ered many links between exposure to air neuroscientists are now working to fill in points suggest it’s more than just a corre-
pollution and signs of neural damage in the gaps in knowledge of how air pollu- lation, says Chen. “We are more confident
animals and humans. Although her find- tion might contribute to these less visible to say that our finding is a causal effect of
ings are observational, and the pathol- effects on human health, both by doc- air pollution on cognitive functioning.”
The paper is just one of the latest to participants had experienced at least one after birth. The group recently reported
draw a line between chronic exposure psychotic episode between the ages of 12 findings from a study of more than 2,200
to air pollution and lowered cognitive and 18 years, and the experiences were children that found lower working mem-
abilities in adults—with several studies most common among teenagers who had ory scores in boys who had been exposed
extending the link to neurodegenerative grown up with the highest exposure to to greater particulate matter levels from
disease. In 2017, for example, research- nitrogen oxides and particulate matter.5 the womb until age seven.8
ers in Ontario reported that people liv- “You’re getting . . . greater odds of Of course, observational studies can’t
ing next to roads with heavy traffic were having those experiences among ado- definitively demonstrate that air pollu-
more likely to be diagnosed with demen- lescents who lived in the areas with the tion causes cognitive changes in humans,
tia. Although the researchers didn’t highest levels” of pollution, says Fisher. not least because it’s impossible to fully
include pollution in their analysis, they That’s not as high as the associations disentangle factors that closely correlate
highlighted it as a likely contributor.3 seen with, for example, childhood abuse, with air pollution, such as noise. “A lot
That same year, USC researchers which is more strongly linked to psy- of the associations we’re seeing are with
reported that elderly women of European chotic experiences, she says, “but it’s a traffic-related pollutants, particularly in
ancestry living in areas of the US with lot higher than you might think.” cities,” Fisher says. In some cases, “it’s
levels of particulate matter that exceeded Some labs have linked air pollu- possible that it’s the noise of the traffic,
the US Environmental Protection Agency tion and brain development even ear- not the actual air pollution it produces,
standard of 12 μg/m 3 were 81 percent lier in life—right back to the womb. that’s problematic.”
more likely to experience cognitive One 2014 study of 250 children born Several labs are trying to minimize
decline generally and 92 percent more to nonsmoking African American and such limitations by collecting real-time
likely to suffer dementia than if they had Dominican women in New York City pollution levels and other environmen-
been living with exposure below that found that greater prenatal exposure tal data, rather than estimating expo-
limit. 4 The increased risk in polluted
areas—a finding that mirrors the results
from cohorts in London and Taiwan—
was particularly high for people with This is the beginnings of a whole new
dementia- or Alzheimer’s-associated
genetic variants such as APOE ɛ4, hint- field. This is like tobacco research and
ing that air pollution might have more-
severe effects in those genetically predis-
cancer 70 years ago.
posed to neurodegeneration. —Caleb Finch, University of Southern California
Other researchers are amassing evi-
dence of cognitive and psychiatric dys-
function in pollution-exposed younger to polycyclic aromatic hydrocarbons sure from existing datasets. ISGlobal, for
people, whose developing brains may be (PAH), which are produced by burn- example, plans to recruit 1,200 pregnant
particularly vulnerable. In 2016, research- ing fossil fuels, correlated with a higher women for an in-depth look at the con-
ers found that Swedish children and ado- prevalence of behavioral and cogni- nections between prenatal exposure to
lescents were more likely to have been tive abnormalities, including symp- air pollution and childhood health. Par-
prescribed psychiatric medicine if they toms associated with attention-deficit/ ticipants will monitor their surround-
lived in areas with high nitrogen dioxide hyperactivity disorder (ADHD).6 ings with “backpacks, sensors at home,
concentrations, and research published a Environmental epidemiologist Jordi GPS, measures of physical activity, mea-
few months ago linked lifetime exposure Sunyer of the Barcelona Institute for sures of noise,” and so on, Sunyer says.
to air pollution with depression and anxi- Global Health (ISGlobal) recently col- To provide a stronger assessment of
ety in 12-year-olds living in Ohio. laborated on a project that aimed to air pollution’s harmful effects, research-
In a recent large-scale project, replicate the ADHD-pollution link. The ers are also taking a more interdisciplin-
research psychologist Helen Fisher of study, which included data from nearly ary approach, using magnetic resonance
King’s College London and colleagues 30,000 mother-child pairs across eight imaging (MRI) to look inside the brain
analyzed data, collected through the separate cohorts in Europe, failed to for neurological features that can com-
UK’s Environmental-Risk Longitudinal find a pattern of ADHD incidence across plement assessments of cognitive func-
Twin Study, on more than 2,000 chil- regions of varying air qualities. 7 But tion. In addition, they have begun to pair
dren born in England and Wales in 1994 Sunyer’s team has found other cognitive epidemiological data with experimental
and 1995. According to results published problems in children who were exposed studies in the lab to learn about the bio-
earlier this year, nearly a third of the to air pollution during gestation and logical pathways that may underlie links
1 0. 201 9 | T H E S C IE N T IST 3 9
PORTS OF ENTRY
TO THE BRAIN
Air pollution refers to a wide range of gases, liquids, and solids suspended
in the atmosphere. Known to have harmful effects on the respiratory
and cardiovascular systems, these contaminants are now implicated in
damage to the brain—an organ exposed to the air via multiple pathways. Sulfur
dioxide
Carbon
monoxide
Polyaromatic
hydrocarbons (PAH)
1 Particulate Ozone
matter
Nitrogen
dioxide
1 IN THROUGH THE NOSE


Contaminants breathed through the nose can come into direct
contact with the olfactory bulb, a neural structure in the
vertebrate forebrain. Some research in humans and other
animals suggests that fine particulate matter smaller than 2.5
micrometers across can reach the olfactory cortex and other
brain regions via this route.
2 2 INHALED INTO THE LUNGS


Most gases can easily traverse the epithelium in the lungs to
make it into the bloodstream, and some studies in humans
and rats suggest that fine particulate matter can do the same.
Circulating contaminants may wear down the blood-brain
barrier and/or cross it to directly interact with neural tissue.
3 3 VIA THE GUT


Pollutants that make it to the gut in swallowed air may be
absorbed by the gut wall and into the bloodstream, where
they can travel to the brain. But researchers are currently
more interested in how pollution may influence the brain
via changes to the composition of the gut’s resident
microbial community. Recent research shows that high
exposure to air pollution is associated with altered gut
microbiomes in humans, while research in mice shows that
inhaled particulate matter can change the microbiome’s
LAURIE O’KEEFE
composition within weeks. Microbiome changes have

recently been linked to cognitive function, suggesting air
pollution could act on the brain via this indirect pathway.
EVIDENCE OF HARM
between air pollution and cognition. “We
have to combine studies in animals with in
vitro studies in cells, with epidemiological
Many of the studies on the effects of chronic exposure to air pollution are data in humans,” says Sunyer, who recently
observational, making researchers wary of saying air pollution causes reduced reviewed the research in this space. “It’s
cognitive function. Nevertheless, evidence from in vivo studies in humans the combination that has to fit.”
and nonhuman animals, combined with in vitro research and analyses of
postmortem brains, is beginning to lend support to the idea. Zooming in
By the early 2000s, Calderón-Garcidueñas
had come up with a way to test her theories
about air pollution’s effects on the brain.
PARTICULAR PRESENCE Dogs in Mexico City would be exposed to
Researchers have discovered the same air as people, she reasoned, so
magnetite particles that look similar to those proin 2003, using brains from 40 euthanized
duced by vehicle engines in the frontal cortex of mongrels—26 from Mexico City and 14
postmortem human brains. from the less polluted city of Tlaxcala to
act as controls—Calderón-Garcidueñas
and colleagues documented neuropatho-
logical differences between the animals. In
particular, the team identified signs of ele-
vated inflammation, DNA damage, deg-
WHITE MATTER radation of the blood-brain barrier, and
Some studies have found reduced volumes of Alzheimer’s-type pathology in the Mexico
white matter in people who have been exposed to City canines.9 She’s since replicated the
elevated levels of air pollution. findings using postmortem human brains
and additional dogs.
Calderón-Garcidueñas’s projects
were seminal examples of what is now
a rapidly expanding effort to come up
with mechanistic explanations linking
REDUCED DEVELOPMENT chronic air pollution exposure with cog-
MRI scans indicate that some regions of the brain, nitive changes in humans. Such work
such as the basal ganglia, develop more slowly is pointing to a key role for the related
than normal in children exposed to air pollution. processes of neuroinflammation and
oxidative stress—an overproduction of
reactive oxygen species that can cause
cellular damage—in response to con-
taminants reaching the brain via the
nose or lungs. (See illustration on oppo-
site page.) Research by Finch’s group, for
GETTING INFLAMED example, found that cultures of rodent
In vitro and in vivo research finds a link between glial cells responded to air pollution
components of air pollution and inflammation- sampled from a local freeway by acti-
related damage in brain cells. vating toll-like receptor 4, a transmem-
brane protein involved in inflammatory
signaling. And mice that inhaled the pol-
luted air for 10 weeks showed activation
of those same inflammatory pathways in
the hippocampus, a brain region known
AMYLOID PLAQUES for its roles in learning and memory.10
A number of research groups have reported a In subsequent work, Finch’s group
higher accumulation of Alzheimer’s-associated found that mouse cell lines expressing
proteins such as amyloid-ß in the brains of a mutated version of the gene coding
humans and animals exposed to air pollution.
1 0. 201 9 | T H E S C IE N T IST 41
for amyloid precursor protein, a peptide tions among children in Mexico City plaques in human brains contained par-
implicated in Alzheimer’s disease, pro- compared to matched controls living in ticles of magnetite, which, thanks to
duced greater levels of biomarkers for less polluted areas. their iron content and magnetic prop-
oxidative stress with increasing expo- A study carried out a couple of years erties, can promote the production of
sure to nanosize particulate matter. And ago by researchers at ISGlobal com- reactive oxygen species and contribute
last year, a team in China reported that pared different regions of the brain and to neurodegeneration. The study was
fine particulate matter triggered oxida- found that some areas might be partic- agnostic as to the origin of these parti-
tive stress in living rats’ neurons and led ularly susceptible. The team reported cles; they can be produced through iron
to the degradation of the myelin sheath that schoolchildren exposed to high metabolism in brain tissue. But when
surrounding cells’ axons. Exposed rats PAH levels showed particularly stunted Calderón-Garcidueñas and colleagues
showed deficits in learning and memory growth in the caudate nucleus, a region took a closer look at magnetite identi-
as well as impaired sensory functions deep within the brain that is linked to fied in post-mortem brains from people
compared with controls.11 behavioral disorders such as ADHD. 13 in Mexico City and in Manchester, UK,
How such findings might translate This region already generates relatively they found that the particles’ crystal
into humans chronically exposed to high levels of reactive oxygen species, structures showed signatures of having
air pollution is less clear, though some explains study coauthor Marion Morta- been generated at high temperatures—
MRI data are consistent with the idea mais, now at INSERM in France. “The just like magnetite particles found in
that dirty air promotes tissue degrada- hypothesis is that . . . the local mecha- traffic-related air pollution.14
tion. One USC study of more than 1,400 nism to fight the oxidative stress is com- Some researchers who spoke to The
women across the US, for example, pletely overwhelmed by another intru- Scientist stress that, despite the discovery
found that a person’s estimated expo- sion of oxidative stress provoked by the of biologically plausible mechanisms, it’s
sure to particulate matter was inversely exposure to PAH.” still too soon to conclusively draw a caus-
correlated with her total volume of white Meanwhile, Calderón-Garcidueñas ative link between air pollution and cog-
matter—the part of the brain made up of has suggested that certain components nitive impairment in humans. For start-
myelin-covered nerve fibers—a decrease of air pollution could play a direct role ers, it’s difficult to translate the chronic
in which is associated with dementia. 12 in Alzheimer’s pathology. In 2016, exposure that characterizes most people’s
MRI studies by Calderón-Garcidueñas University of Texas at San Antonio contact with air pollution into short-term
have also found white-matter reduc- researchers reported that amyloid-ß lab studies with mice or cells, says Chen.
© ISTOCK.COM, ESOLEX
References
What’s more, the fact that air pollutants, Meaningful change, however, will
1. L. Calderón-Garcidueñas et al., “Hallmarks of
and their concentrations, vary around the require more than upping air quality for a Alzheimer’s disease are evolving relentlessly in
world makes it challenging to pin down particular school or region, Chen says. It’s Metropolitan Mexico City infants, children and
which are most dangerous and why. long-term exposure that’s associated with young adults. APOE4 carriers have higher suicide
As researchers grapple with these harm, so the solution needs to be a global risk and higher odds of reaching NFT stage V at
≤ 40 years of age,” Environ Res, 164:475–87, 2018.
issues, findings from other disciplines reduction in air pollution, he argues—a
2. X. Zhang et al., “The impact of exposure to air
pollution on cognitive performance,” PNAS,
115:9193–97, 2018.
This year, the US government floated

3. H. Chen et al., “Living near major roads and the
incidence of dementia, Parkinson’s disease, and
multiple sclerosis: a population-based cohort
plans that would reduce monitoring of study,” Lancet, 389:P718–26, 2017.
4. M. Cacciottolo et al., “Particulate air pollutants,
air pollution and its health effects. APOE alleles and their contributions to
cognitive impairment in older women and to
amyloidogenesis in experimental models,” Transl
Psychiat, 7:e1022, 2017.
5. J.B. Newbury et al., “Association of air
hint that the big picture may be even challenge, “because we all know that air pollution exposure with psychotic experiences
more complicated. It appears likely, for pollution has no physical boundary.” He during adolescence,” JAMA Psychiat,
example, that air pollution can influence suggests that mitigating pollution requires 76:614–23, 2019.
the brain via indirect mechanisms. Last the same sort of mentality as tackling cli- 6. F.P. Perera et al., “Early-life exposure to polycyclic
aromatic hydrocarbons and ADHD behavior
year, a team in the Netherlands found mate change, with international agree-
problems,” PLOS ONE, 9:e111670, 2014.
that people exposed to air pollution ments that set global targets for outdoor 7. J. Forns et al., “Air pollution exposure during
for just two hours had a greater abun- air quality, much as they have for carbon pregnancy and symptoms of attention deficit and
dance of several circulating microRNAs, dioxide emissions, he says. “[Air] is a col- hyperactivity disorder in children in Europe,”
some of which may help drive pollution- lective good—everyone consumes it.” Epidemiology, 29:618–26, 2018.
8. I. Rivas et al., “Association between early life
related disease. And several studies now Fortunately, the more-established
exposure to air pollution and working memory
indicate that commensal bacteria living literature on respiratory and cardio- and attention,” Environ Health Perspect,
in the gut might influence neurodegen- vascular effects of air pollution has 127:57002, 2019.
eration, possibly via inflammation of already provided momentum to move 9. L. Calderón-Garcidueñas et al., “DNA damage in
the gut lining. Recent research in mice toward such international efforts. The nasal and brain tissues of canines exposed to air
pollutants is associated with evidence of chronic
shows that inhalation of particulate mat- WHO’s air pollution conference last year
brain inflammation and neurodegeneration,”
ter alters the microbiome’s composition resulted in the Geneva Action Agenda Toxicol Pathol, 31:524–38, 2003.
within weeks. to Combat Air Pollution, which lays out 10. N.C. Woodward et al., “Toll-like receptor 4 in glial
priorities ranging from better air-quality inflammatory responses to air pollution in vitro
Cleaning up monitoring to the provision of support and in vivo,” J Neuroinflamm, 14:84, 2017.
11. Q. Zhang et al., “PM2.5 impairs neurobehavior by
As researchers continue to probe the for cities trying to reduce urban pollu-
oxidative stress and myelin sheaths injury of brain
effects of air pollution, several groups tion. But countries’ adherence to such in the rat,” Environ Pollut, 242:994–1001, 2018.
are investigating whether the apparent goals is voluntary and difficult to guar- 12. J.C. Chen et al., “Ambient air pollution and
damage can be mitigated. For example, antee. Just this year, the US government neurotoxicity on brain structure: Evidence from
urban green spaces are recognized by floated plans that would reduce monitor- women’s health initiative memory study,” Ann
Neurol, 78:466–76, 2015.
the World Health Organization (WHO) ing of air pollution and its health effects
13. M. Mortamais et al., “Effect of exposure to
as contributors to mental health, partly and downgrade estimates of the harm polycyclic aromatic hydrocarbons on basal ganglia
because they may improve air quality. it causes. Mexico, meanwhile, has been and attention-deficit hyperactivity disorder
A study published this May found that struggling with allegations of corruption symptoms in primary school children,” Environ
silver birch, elder, and yew trees were as government measures to reduce emis- Int, 105:12–19, 2017.
14. B.A. Maher et al., “Magnetite pollution
particularly effective at soaking up fine sions are reportedly undermined by local
nanoparticles in the human brain,” PNAS,
particulate matter in a lab setting, and officials and car inspection centers. 113:10797–801, 2016.
research by ISGlobal suggests that green “I’m not going to give up,” Calderón- 15. P. Dadvand et al., “Green spaces and cognitive
spaces within and around schools could Garcidueñas says. “It’s a difficult sub- development in primary schoolchildren,” PNAS,
help boost cognitive development in chil- ject, but it’s very interesting. I tell my 112:7937–42, 2015.
16. P. Dadvand et al., “Lifelong residential exposure
dren15 and enhance attention,16 although husband that it’s probably going to be
to green space and attention: a population-based
the mechanisms behind these effects another twenty years to acknowledge prospective study,” Environ Health Perspect,
aren’t entirely clear. what we’re doing right now.” g 125:097016, 2017.
1 0. 201 9 | T H E S C IE N T IST 4 3
GOTCHA: A colored transmission

electron micrograph of a neutro-
phil (blue) capturing Neisseria
gonorrhoeae bacteria (orange) in a
neutrophil extracellular trap.
NETs: Two-Faced
Players in Immunity
Webs of nuclear material expelled from neutrophils trap invading
pathogens, but these newly discovered structures also have ties to
autoimmunity and cancer.
BY BORKO AMULIC AND GABRIEL SOLLBERGER
I
n the early 2000s, Arturo Zychlin- copious amounts of DNA. In cultured passive lysis. This has motivated a new
sky at the Max Planck Institute for human neutrophils, the webs were line of research devoted to characteriz-
Infection Biology in Berlin found able to trap the bacteria that had trig- ing these unique structures, delineating
that mammalian immune cells called gered their formation, thereby limiting the mechanisms that prompt their for-
neutrophils use an enzyme called neu- infection, so Zychlinsky and colleagues mation, and identifying their relevance
trophil elastase (NE) to cleave bacterial dubbed them neutrophil extracellular in mammalian biology.
virulence factors.1 When Zychlinsky and traps, or NETs.2 As more and more researchers join
his colleagues delved deeper into this The fact that neutrophils used their the burgeoning field, the spectrum of
© SCIENCE SOURCE, SPL
defense mechanism, they realized that nuclear material to catch pathogens was pathogens known to induce NET release
when activated by bacteria, human neu- intriguing to immunologists and cell from neutrophils has expanded from a
trophils release NE in what, under the biologists alike. The work of the Zych- variety of bacteria to fungi and, most
microscope, looked like a fibrous struc- linsky lab suggested that the release of recently, to viruses. However, it has
ture. This structure turned out to be a NETs was an active process, and that also become clear that NETs can have
meshwork of NE, other proteins, and the material wasn’t simply released by negative consequences for the organ-
1 0. 201 9 | T H E S C IE N T IST 4 5
isms that produce them—by activating quickly infiltrate tissues if the neutro- necrosis of neutrophils. 4 Later studies
autoimmune pathways or encouraging phils detect a microbial threat. Belong- added complexity by demonstrating
tumor cells to metastasize, for example. ing to a class of white blood cells called that different inflammatory triggers
Today, it is widely accepted that granulocytes, they are characterized by induce various pathways that all lead
NETs have both a protective and a path- a cytoplasm packed with granules con- to the release of NETs, and that NET
ological impact on the host. In 2012, taining antimicrobial proteins. Neu- release doesn’t always result in lysis of
Mariana Kaplan, now of the National trophils can engulf pathogens and then the neutrophil. 5
Institutes of Health, and the Univer- fuse their granules with their phago- That said, most pathways of NET
sity of Tennessee’s Marko Radic termed somes, which contain the internalized formation do kill the immune cell, typ-
NETs a “double-edged sword of immu- microbes. Alternatively, the cells can ically as a result of the production of
nity” and suggested that healthy organ- fuse their granules with the plasma reactive oxygen species (ROS). Bacte-
isms must tightly control their release membrane to release antimicrobials to rial or fungal pathogens cause neutro-
to minimize negative consequences for attack extracellular parasites. phils to activate kinases that induce
the host.3 The details of NET regulation Research on neutrophils is compli- assembly of an enzyme complex called
and function are now a very active area cated by the fact that they are short- nicotinamide adenine dinucleotide
of research. lived cells. For instance, unlike some phosphate (NADPH) oxidase. NADPH
other human cell types, neutrophils oxidase then produces large amounts of
NET basics cannot be cultured for more than a few superoxide—a highly reactive oxygen
Neutrophils are essential for immune hours, and they are not amenable to compound that carries an extra elec-
defense and prevention of microbial gene editing. For this reason, we still tron—during a process called the neu-
overgrowth. They are very abundant— lack a detailed mechanistic picture of trophil oxidative burst. ROS resulting
around 100 billion are produced in a how exactly NETs are formed. Early from the oxidative burst trigger disin-
human’s bone marrow in a single day— reports confirmed the original hypothe- tegration of a multiprotein complex to
and they circulate in the bloodstream to sis that NETs do not result from passive release active NE, a primary component
NET FORMATION
When a neutrophil encounters a pathogen, it can respond in several ways: phagocytosis, degranulation, or by releasing neutrophil extracel-
lular traps (NETs). In NET release, shown here, the enzyme complex NADPH oxidase generates reactive oxide species (ROS), which in turn
TAMI TOLPA
initiate the disintegration of granules, releasing neutrophil elastase (NE). NE then migrates to the neutrophil’s nucleus, where it cleaves
proteins that package the cell’s DNA as chromosomes. The chromatin expands until it fills up the entire cell, which breaks open and
extrudes the NET into the extracellular space. There, the webs are thought to trap and kill the triggering pathogens.
Granules NAPDH
oxidase
Neutrophil
ROS
Nucleus
Granules
Pathogen NE
NE migrates to the nucleus and NET is released,

NET formation is triggered, and NE is assists the decondensation of Chromatin and the pathogen is
released into the cytoplasm. chromatin. expands. trapped.
of NETs, into the cytoplasm. (See illus- experimental deletion of the genes that
tration on page 45.) encode these NET-degrading enzymes
NE then migrates to the neutrophil’s in those microbes leads to reduced bac-
nucleus, where it cleaves histones and terial virulence.8
other proteins to decondense the chro- The idea that NETs form barriers at
matin. Eventually, the chromatin fills mucosal surfaces was recently strength-
up the entire cell until the cell lyses and ened by additional work from Kubes and
extrudes the NET into the extracellular Ajitha Thanabalasuriar, now at Med-
space, a process known as NETosis. We Immune, showing that NETs form an
recently identified an important role for exclusionary microbicidal “dead zone”
the pore-forming protein gasdermin D in infected corneas. This confines bac-
in both the nuclear expansion and the teria to the outer surface of the eye and
lysis processes, although the mecha- prevents them from entering the eye and
nisms aren’t yet clear. 6 In the extracel- spreading to the brain.9
lular space, the webs are thought to trap The role of NETs in infections is not
and kill the triggering pathogens. limited to trapping microbes, however.
The structures contain multiple anti-
NETs in immune defense microbial molecules. Histones, for exam-
NETs’ ability to trap microbial invad- ple, are major components of the chro-
ers has been demonstrated in vitro for matin in NETs, and these proteins have
a variety of pathogens. The structures important bactericidal and immuno-
appear to be particularly important in stimulatory functions. After discovering
defense against pathogenic fungi, sug- NETs, Zychlinsky proposed antimicrobial
Today, it is widely
gesting NETs may have evolved as a way defense as the alternative, less-studied
to trap large organisms that cannot be function of histones, whose more promi- accepted that
engulfed via phagocytosis. 7 But a lack nent and established function is to orga-
of specific genetic tools has made it dif- nize and package DNA in the nucleus. NETs have both a
ficult to nail down the exact contribu- NETs also contain several alarm-
protective and a
tion of NETs to antimicrobial defense in ins, molecules that activate the immune
vivo. Most of the molecules that have so system and help propagate the inflam- pathological impact
far been found to regulate NETs are also matory response. Release of alarmin-
involved in other immune responses, containing NETs alerts the rest of on the host.
so a “NET knockout” organism is still
beyond our reach.
In spite of these technical chal-
lenges, researchers are slowly elucidat-
ing the function of NETs in immunity.
In 2012, Paul Kubes of the University of
Calgary and colleagues depleted NETs
in mice with skin infections by inject-
ing recombinant DNase 1, an enzyme
that chops up the DNA backbone of
NETs after they are released. This led
to increased dissemination of Staph-
ylococcus aureus from the skin to the
bloodstream, demonstrating that NETs
participate in containing bacteria at
the site of entry. 5 This experimental
VOLKER BRINKMANN
strategy of NET destruction by nucle-

ases is also used naturally by patho-
genic bacteria as a way to escape from
NETs: Staphylococcus and Streptococ-
cus species both secrete DNases, and
1 0. 201 9 | T H E S C IE N T IST 47
NETS IN HEALTH
The exact contribution of NETs to antimicrobial defense has been difficult to nail down, but researchers are slowly elucidating their
roles in protecting the body from invaders and other threats, including runaway inflammation. Those roles include:
Pathogens, mitogens,
TRAPPING NETs
crystals, or other
triggers NETs immobilize Pathogens
microbes and prevent
their dissemination.
Pathogens
BARRIERS
NETs can form
dense, exclusionary
barriers in the
eye that prevent
microbes from
penetrating the body.
Eye stroma
Neutrophil
Dendritic
IMMUNE SIGNALING cell
NET components act as
alarm signals to activate
additional immune
cells and propagate the
inflammatory response.
Macrophages and
dendritic cells sense
various components of Proinflammatory
cytokines
the NETs, including DNA
and proteins, which
leads them to produce DNase
proinflammatory cleaves
NETs
mediators.
Macrophage
Cleaved
cytokines
COUNTERING
NET
INFLAMMATION
When present at high
density, NETs can
cleave proinflammatory
cytokines and help
resolve inflammation.
. . . AND DISEASE
NETs have a dark side that makes them dangerous when inappropriately deployed.
The structures have been implicated as contributors to a range of conditions. MALARIA
NET formation is triggered during
malaria, and then the structures are
CANCER cleaved into fragments by circulating
NET-associated proteins lead DNase1. These fragments lead to
to reawakening of dormant upregulation of cytoadhesion receptors
cancer cells and convert them DNase on the surface of endothelial cells lining
to proliferating metastatic cells. the blood vessels. Cells infected with
Plasmodium parasites bind to these
receptors, which helps them avoid the
immune response in the spleen and
causes damaging inflammation.
NETs
Cytoadhesion
receptors
Plasmodium-
Cancer cells infected blood
cell
THROMBOSIS
NET components
promote blood
Metastatic
coagulation and
cancer cells
obstruction of small
blood vessels.
Antibodies
against DNA ATHEROSCLEROSIS
Immunostimulatory NETs activate macrophages,
molecules inducing them to produce
proinflammatory cytokines.
B cells The histones associated
with NETs also damage
the smooth muscle of the
arterial walls.
Activated Proinflammatory
dendritic cell Macrophage cytokines
LUPUS
This autoimmune disease is characterized
by production of autoantibodies directed
against one’s own DNA. NETs are
thought to be a source of autoantigens,
as well as immunostimulatory molecules Smooth
TAMI TOLPA
that activate dendritic cells and fuel muscle cell

inflammation. damage
nal via the alarmins in NETs. This is but it seems that neutrophils remain
illustrated nicely in the case of Pseu- viable, phagocytic, and are even able
domonas aeruginosa, an opportunis- to “push” the NET forward as they
tic pathogen that is able to colonize migrate. These findings demonstrate
the lungs of cystic fibrosis patients. P. that neutrophils do not necessarily die
aeruginosa only triggers NETs when after they form NETs.
it expresses virulence factors such as
flagella or the toxin pyocyanin. In the Downsides of NETs
absence of these virulence factors, neu- Unfortunately, as with other neutro-
trophils mostly respond to these bacte- phil responses, NETs have a dark side
ria by attempting to engulf them. that makes them dangerous when
inappropriately deployed. This is the
Triggers of NET formation case with malaria, which is caused by
It’s not just pathogens that can initiate Plasmodium parasites that invade and
NET formation. A variety of host mol- proliferate in red blood cells. Infected
ecules can also activate neutrophils to cells adhere to the walls of blood ves-
extrude their innards into the extra- sels throughout the body, leading to
cellular space. One example is crystals pathological changes such as obstruc-
of cholesterol, a form of the lipid that tion of capillaries and inflammation.
causes inflammation. And in 2017, we Our group made the surprising dis-
and our colleagues found that mito- covery that NETs cause blood vessels
genic signaling, which induces cell divi- to become sticky by expressing cytoad-
sion in some immune cells, also triggers hesive proteins, which facilitate bind-
various cell cycle–related phenomena in ing by Plasmodium-infected cells. In
neutrophils, but with the consequence the absence of NETs, infected mice tol-
The process of cells of NET formation rather than division.11 erated the presence of parasites and
Notably, NETs can form indepen- showed no signs of disease. 13
releasing their DNA dently of the canonical pathway involv- Similarly harmful effects of NETs
as a protective ing ROS-producing NADPH oxidase, have been described in noninfectious
stimulated by pathogens or compounds diseases such as lupus, an autoim-
mechanism might be such as ionophores, agents that lead mune condition. In these instances,
to ion fluxes through the cell mem- NET release is dysregulated, trigger-
ancient. brane.11 It is unclear whether NADPH ing chronic pathological inflammation,
oxidase–independent pathways rely with NETs potentially acting as a source
on ROS produced by other means or of autoantigens and immunostimula-
the immune system to the presence whether these are truly ROS-indepen- tory molecules. (See “Commensal Men-
of microbes or foreign substances. dent processes. Interestingly, iono- ace,” The Scientist, June 2019.)
Twenty-five years ago, immunologist phore-driven activation of neutrophils As large, fibrous extracellular struc-
Polly Matzinger of the National Insti- triggers strong calcium fluxes, which tures, NETs are also excellent scaf-
tute of Allergy and Infectious Diseases activate the enzyme peptidyl arginine folds for thrombosis, the formation of
proposed the “danger theory” of immu- deiminase 4 (PAD4). PAD4 converts blood clots that are the main cause of
nity, postulating that the immune sys- the amino acid arginine to citrulline heart attack and stroke. Denisa Wag-
tem is concerned with limiting virulent and acts on various substrates, includ- ner’s group at Harvard Medical School
bacteria and tolerating nontoxic ones.10 ing histones. Citrullination of histones has shown that NETs are found in both
NET release may serve as one mecha- is thus a good biomarker for NETs in arterial and deep vein thrombosis and
nism of flagging intolerable bacteria. the extracellular space. that their degradation by DNase 1 is
Instead of engulfing the microbes and Perhaps the most surprising mech- protective in animal models of the dis-
initiating the immunologically silent anism of NET formation involves DNA ease. 14 Before being linked to NETs,
apoptotic pathway, the NET-producing release by living neutrophils, a process histone-packaged bits of DNA called
form of cell death utilizes proinflam- termed vital NETosis by Kubes and nucleosomes were known to have pro-
matory pathways. Bacteria that are colleagues.12 It is unclear which mol- coagulant properties.
toxic enough to lead to NETosis will ecules mediate DNA release in this NETs can also be subverted by
send a powerful inflammatory sig- case—or how the release even occurs— malignant cells to facilitate metasta-
sis. The link between inflammation Borko Amulic is an assistant professor

and cancer progression has long been at the University of Bristol in the UK
established, but it was a seminal study and a Medical Research Council Fel-
by the laboratory of Mikala Egeblad at low. Gabriel Sollberger is a postdoctoral
Cold Spring Harbor Laboratory that researcher at the Max Planck Institute
demonstrated NETs can reawaken dor- for Infection Biology in Berlin.
mant tumor cells. 15 Working in mouse
models of breast and prostate cancer,
the Egeblad lab showed that bacterial
References
antigen–triggered NETs remodel the 1. Y. Weinrauch et al., “Neutrophil elastase
microenvironment to encourage acti- targets virulence factors of enterobacteria,”
vation and proliferation of tumor cells. Nature, 417:91–94, 2002.
Importantly, researchers were able to 2. V. Brinkmann et al., “Neutrophil extracellular
traps kill bacteria,” Science, 303:1532–35, 2004.
block metastasis by either interfering
3. M.J. Kaplan, M. Radic, “Neutrophil extracellular
with NET production or by blocking traps: Double-edged swords of innate immunity,” J
downstream signaling effects. Inhibi- Immunol, 189:2689–95, 2012.
tion of NETs is thus a potential strategy 4. T.A. Fuchs et al., “Novel cell death program
for developing novel cancer therapies, leads to neutrophil extracellular traps,” J Cell
Biol, 176:231–41, 2007.
and is likely to be the focus of intense
5. B.G. Yipp et al., “Infection-induced NETosis
research in coming years. is a dynamic process involving neutrophil
multitasking in vivo,” Nat Med, 18:1386–93, 2012.
Open questions 6. G. Sollberger et al., “Gasdermin D plays a vital
Despite very active research, there are role in the generation of neutrophil extracellular
traps,” Sci Immunol, 3:eaar6689, 2018.
still a number of unknowns concerning
7. N. Branzk et al., “Neutrophils sense microbe
NETs. Mechanistically, we do not fully size and selectively release neutrophil
understand how many pathways exist extracellular traps in response to large
to induce neutrophils to make NETs, pathogens,” Nat Immunol, 15:1017–25, 2014.
or how these pathways might interact. 8. J.T. Buchanan et al., “DNase expression allows
the pathogen group A Streptococcus to escape
And the pathological effects of NETs
killing in neutrophil extracellular traps,” Curr
are only beginning to be detailed, while Biol, 16:396–400, 2006.
their potential healing properties are 9. A. Thanabalasuriar et al., “Neutrophil
currently understudied. extracellular traps confine Pseudomonas
Another important question is aeruginosa ocular biofilms and restrict brain
invasion,” Cell Host Microbe, 25:526–36.e4, 2019.
NIAMS SYSTEMIC AUTOIMMUNITY BRANCH, MARIANA J. KAPLAN, M.D., CHIEF
what determines whether a neutrophil

10. P. Matzinger, “Tolerance, Danger, and the
undergoes lytic or non-lytic NET forma- Extended Family,” Ann Rev Immunol, 12:991-
tion. The concept of neutrophils releas- 1045, 1994.
ing NETs while remaining motile and 11. B. Amulic et al., “Cell-cycle proteins control
capable of engulfing bacteria is intrigu- production of neutrophil extracellular traps,”
Dev Cell, 43:449–62.e5, 2017.
ing, but more studies are required to
12. B.G. Yipp, P. Kubes, “NETosis: How vital is
elucidate when and where this occurs. it?” Blood, 122:2784–94, 2013.
Finally, an interesting and unsolved 13. S.L. Knackstedt, “Neutrophil extracellular traps
question is how evolutionarily conserved drive inflammatory pathogenesis in malaria,”
the process of NET formation is. Plant root PhD diss., Humboldt University, 2018.
14. T.A. Fuchs et al., “Extracellular DNA traps promote
cells release chromatin to defend against
thrombosis,“ PNAS, 107:15880–85, 2010.
pathogens,16 and a recent study described 15. J. Albrengues et al., “Neutrophil extracellular
NET-like structures released from phago- traps produced during inflammation awaken
cytes of invertebrates, such as crabs, mus- dormant cancer cells in mice,” Science,
sels, and anemones.17 The process of cells 361:eaao4227, 2018.
16. M.C. Hawes et al., “Extracellular DNA: The tip
releasing their DNA as a protective mech-
of root defenses?” Plant Sci, 180:741–45, 2011.
anism might therefore be ancient. Further 17. C.T. Robb et al., “Invertebrate extracellular
studies will hopefully identify the origin of phagocyte traps show that chromatin is an
NETs, and thereby shed light on this fasci- ancient defence weapon,” Nat Comm, 5:4627,
nating and important process. g 2014.
1 0. 201 9 | T H E S C IE N T IST 51
EDITOR’S CHOICE PAPERS
The Literature
NEUROSCIENCE
Stem Cell Stimulation for Stroke Recovery

THE PAPER
S.P. Yu et al., “Optochemogenetic stimula-
tion of transplanted iPS-NPCs enhances
neuronal repair and functional recovery
after ischemic stroke,” J Neurosci, 39:6571–
94, 2019.
GLuc
Cell transplantation therapy offers a prom- VChR1
ising route to recovery after stroke, but the channel
grafted cells face a harsh environment, with
elevated levels of free radicals and proin- CTZ
Neurotransmitters
flammatory cytokines, compromised blood
supply, and degraded neural connectivity, Sodium ions
says Shan Ping Yu, a neurology researcher
at Emory University School of Medicine. He
and his colleagues aimed to build a new tool
INNER GLOW: A new technique, optochemogenetics, activates light-sensitive channels inside the brain with a
to help stem cells integrate with host neural drug-like compound administered through the nose. Once the compound, CTZ, has entered the brain, it interfaces
circuitry after implantation. with the protein Gaussia luciferase (GLuc) on modified ion channels—in this case a channelrhodopsin protein,
Scientists have long known that stimu- VChR1. GLuc emits light, causing the VChR1 to open and allow sodium ions to flow inside the neuron, thus
lating transplanted neural stem cells encour- stimulating the cell.
ages them to differentiate into neurons
and connect with nearby host cells. Many fusion protein can be hooked up to either heightened sensation in their paws and
researchers turn to optogenetics to excite excitatory or inhibitory channels in the neu- whiskers, among other parameters. “On
grafted stem cells, but because light travels rons to either stimulate or tamp down the some of the tasks, they can even recover to
poorly through dense tissue, the technique cells’ function. Yu and his colleagues dubbed the normal [pre-stroke] level, so that’s a very
requires researchers to stick a laser into the new technique “optochemogenetics.” exciting result,” says Yu.
their subjects’ brains. So Yu and his coau- The team treated a portion of the iPSC- Promoting cell survival remains a cen-
thors turned instead to a type of enzyme that injected mice with daily CTZ, while another tral challenge in stem cell–based stroke
grants fireflies and jellyfish their glow: lucif- group received light stimulation. Com- treatment, says Marcel Daadi, a neuro-
erase. “These proteins carry their own light, pared with cells grafted into mice that did scientist and medical researcher at the Texas
so they do not need a light source,” says Yu. not receive any stimulation, the cells in both Biomedical Research Institute who was not
The researchers injected neural pro- groups of treated animals formed more con- involved in the study. He adds that being
genitor cells that had been derived from nections with neighboring neurons. Cells able to stimulate stem cells through nasal
induced pluripotent stem cells (iPSCs) into stimulated by CTZ or light also grew more- administration of a drug was “a clever way
the brains of mouse models of stroke. The heavily-insulated axons, had increased rates to ensure wide distribution of the drug, and
cells were genetically engineered to express a of signal transmission between neurons, to be able to stimulate pretty much the total-
fusion protein called luminopsin 3 (LMO3), and better repaired damaged connections ity of the grafted cells.”
crafted from the bioluminescent enzyme between the thalamus and cortex than cells If the approach proves viable in peo-
Gaussia luciferase and the light-sensitive that received no stimulation. ple, it could find applications beyond stroke
© KELLY FINAN
protein VChR1. LMO3 activates in response A suite of behavioral tests revealed that treatment, Daadi says—such as in helping to
to either physical light or a molecule called treated mice had also regained lost senso- rebuild neural circuits damaged by Parkin-
CTZ, which can be delivered noninvasively rimotor function within a few weeks, show- son’s disease or traumatic brain injury.
through the nose into the brain tissue. The ing significantly improved dexterity and —Nicoletta Lanese
ASSOCIATIONS IN THE BRAIN: Researchers correlate the age of one’s NEURAL DAMAGE: A brain slice from a patient with Alzheimer’s disease
father at birth with abnormalities in the brain previously linked to autism. reveals protein clumps (dark spots) characteristic of the disease.
DISEASE & MEDICINE NEUROSCIENCE
Paternal Age and Autism Alzheimer’s in Sleep

THE PAPER THE PAPER
W. Yassin et al., “Paternal age contribution to brain white J.R. Winer et al., “Sleep as a potential biomarker of tau and
matter aberrations in autism spectrum disorder,” Psychiatry Clin β-amyloid burden in the human brain,” J Neurosci, 39:6315–
Neurosci, doi:10.1111/pcn.12909, 2019. 24, 2019.
In the past few years, a number of high-profile studies have Everybody sleeps, but not all sleep is created equal. Good sleep
linked parental age at birth, and paternal age in particular, with appears to keep the brain healthy, clearing toxins from the organ.
a child’s autism risk. Walid Yassin, a neuropsychiatric researcher But bad sleep might be a sign of neurodegenerative disease, with
at the University of Tokyo, wanted to know if having older past studies linking sleep disturbances to the progression of
parents correlated with characteristics of the brain that have Alzheimer’s disease.
been linked to autism. Interested in whether brain waves correlated to accumulation
When Yassin and his colleagues examined the brain scans of of tau and amyloid-ß, two proteins considered hallmarks of the
39 adult males with high-functioning autism spectrum disorder disease, University of California, Berkeley, graduate student
(ASD) and of 37 typically developing males, they found that Joseph Winer and colleagues took positron emission tomography
paternal age correlated with characteristics of the white matter measures of the proteins in 31 cognitively healthy individuals in
in regions of the brain responsible for social interactions in their 70s, then gathered electroencephalogram (EEG) readings of
analyses of all 76 individuals. Specifically, in the men with older brain activity patterns while the participants slept.
fathers, these areas had higher radial diffusivity, a measure of Analyzing the data, the team found that the degree of
water diffusing toward the axonal membrane instead of along synchronization of two types of brain waves prominent during
the axon, suggesting damage to nerve cells’ myelin sheaths, sleep—bursts of oscillations called sleep spindles and a slow
says Yassin. “And such difference in radial diffusivity has oscillation or slow wave—correlated to levels of tau protein
been previously reported in ASD.” in the medial temporal lobe, a region of the brain important
Magdalena Janecka, an epidemiologist who specializes for memory. More tau buildup was associated with less
in autism at the Icahn School of Medicine at Mount Sinai in synchronization of the brain waves. A drop in the amplitude of
New York, applauds the study’s focus on the brain. “We have slow wave was also predictive of higher amounts of amyloid-ß
a lot of epidemiological associations . . . but what [underlies in the cortex of the brain. “What was interesting to us was that
© ISTOCK.COM, WENHT; WIKIMEDIA, PATHO
them] is still very much underexplored,” she says. “The you could separate the EEG signals,” Winer says, suggesting that
authors did a great job at exploring the mechanism that the signals and their synchronization could potentially serve as
could connect the two.” biomarkers of Alzheimer’s disease that may be apparent before
But Janecka adds that the results can’t distinguish whether more-severe symptoms such as memory loss emerge.
the link between age and autism is due to an accumulation of The results are intriguing, says Brendan Lucey, a neuroscientist
mutations in the sperm of older men, or if men who choose at Washington University in St. Louis who was not involved
to have children later in life are enriched for certain traits in the study. But the team’s claim that the accumulation of
associated with autism. “Is the effect we’re observing due to tau or amyloid-ß generates distinct EEG signals may be a
age or is it due to some underlying propensity of men who delay bit premature, he says. “We need a lot more replication and
fatherhood?” she asks. larger samples to see if the differences are true.”
—Jef Akst —Ashley Yeager
1 0. 201 9 | T H E S C IE N T IST 53
PROFILE
The Cerebellum’s Secrets

Neuroscientist Kamran Khodakhah has discovered that the brain region does more than coordinate
movement; it also plays a role in reward circuits and in addiction.
BY ANNA AZVOLINSKY
N
early 30 years ago, Kamran Khodakhah, now a neuro- Purkinje neurons, he found, used the high concentration
scientist at Albert Einstein College of Medicine, signed of InsP3 plus calcium for rapid calcium signaling, and they
up for a TV repair course that met several times a week released tenfold more calcium, on a much faster timescale
at night at a local community college in London. While many than non-neuronal cells did.
of the other students were attending with the obvious goal of The work piqued Khodakhah’s interest in the cerebellum,
repairing TVs and other appliances, Khodakhah had a different which he would go on to study for the rest of his career.
aim. He reasoned that if he could understand how a television
worked, he could design new tools to study the rat brain slices EAVESDROPPING ON NEURONS
he had collected. Khodakhah was born in Tehran, Iran, in 1967, the oldest of
Khodakhah was working as a PhD student in the lab of three siblings. His mother, Mahin, was an English teacher,
neuroscientist David Ogden at the National Institute for Medical and his father, Akbar, who held a doctorate in pharmacy, had
Research, trying to determine whether a particular signaling served as the chief executive officer of several Iran-based
pathway—the inositol trisphosphate (InsP3)/calcium signaling biopharmaceutical companies. Khodakhah recalls a happy
pathway—could be activated in nerve cells called Purkinje neurons. childhood: as a kid, he loved solving puzzles and making lit-
They are found in the cerebellum and have a high density of tle electronic gadgets. He also says he enjoyed reading liter-
InsP3 receptors. By taking the TV repair class, Khodakhah ature and philosophy. After the Iranian Revolution precipi-
wanted to learn to build an electronic circuit to enhance his tated the ouster of the ruling monarchy in 1979, it became
camera images in order to better visualize the Purkinje cells more difficult to obtain Western books, but not impossible,
within slices of the cerebellum and to study the InsP3/calcium Khodakhah recalls. There was a stretch of road in front of
ion signaling pathway. Tehran University that was lined with bookstores that would
He used the new imaging setup, combined with existing stack Western books behind the ones on display. “I really
lab tools such as flash photolysis, to introduce inert precursor enjoyed seeking out the books that were difficult to come by,”
molecules of InsP3—called caged InsP3—into Purkinje neurons he says. “I found it really exhilarating, and that helped form
in cerebellar slices prepared from rat brains. When stimulated my personality and character.”
with light, a caged InsP3 molecule is rapidly converted into an At age 16, he went to Bryanston School, a boarding school
active form that binds to InsP3 receptors. Khodakhah then used in Dorset, England. Then, in 1985, he enrolled at King’s Col-
a fluorescent calcium indicator and recorded the calcium channel lege at the University of London to study pharmacology and
activity to see if the binding of InsP3 receptors caused release of became intrigued by the puzzles of biology. He began to do
calcium from internal stores. At the time, researchers knew that research in the late Alan Prince’s electrophysiology labora-
in liver and other non-neuronal cells, InsP3 molecules act as tory, learning to measure neurotransmitter release from rat
messengers, stimulating the release of calcium ions, which then brain slices. During his senior year, Khodakhah met Ogden
activates internal cellular pathways. Whether something similar and was introduced to methods to measure single ionic chan-
happened in Purkinje neurons wasn’t clear, but if it did, the process nel activity. “You could hear the gurgling sound when the
might reveal something about how the cerebellum coordinates channels opened and current was flowing,” he recalls. “Seeing
movement, Khodakhah thought. and hearing a live protein perform its function—that was the
After a year of trying, the study seemed to be a failure—the moment I fell in love with physiology,” he recalls.
young doctoral candidate couldn’t show that InsP3 stimulates “How the brain works was and still is a complete mystery,”
intracellular release of calcium ions in Purkinje neurons. Then, Khodakhah says, “and I was drawn to trying to solve this ulti-
at 1 a.m. one night in the lab, Khodakhah decided to do something mate puzzle of how consciousness comes from this organ.”
extreme. He added 10 times the dose of caged InsP3 he had
JASON TORRES
been previously using, reasoning that the cerebellum slice itself NEW TOOLS, NEW FOCUS
might be absorbing too much of the light needed to activate After joining Ogden’s lab, which was in the process of mov-
the InsP3 molecules. “I did the experiment, and the result was ing to the National Institute for Medical Research, now the
beautiful,” he says. Francis Crick Institute, Khodakhah started working on the
InsP3-calcium signaling in Purkinje neurons and, when he

had finished his PhD, moved to the US in 1992 to do a postdoc
in Clay Armstrong’s biophysics laboratory at the University of
Pennsylvania. There, Khodakhah continued to study Purkinje
neurons, this time as a model of synaptic plasticity—the way
that neurons modulate their signaling strength to learn new
things, such as how to perform appropriate muscle movements.
“Synaptic plasticity is akin to learning to throw a ball at the
right strength, so you don’t throw it too far or not far enough,”
Khodakhah explains.
Armstrong told Khodakhah that he needed to build his
own equipment—including his own amplifier, to increase the
power of the measured electrical signal in the neurons. “The
experience was invaluable because it taught me to be nimble
KAMRAN KHODAKHAH in creating the tools I need,” Khodakhah says. “In my own lab,
Professor, Departments of Neuroscience, Psychiatry and Behavioral if we want to push technology forward, we sit and build our
Sciences, and Neurology, Albert Einstein College of Medicine own circuits, or create a new computer program rather than
Chair, Department of Neuroscience, Albert Einstein College of Medicine waiting around for someone else to do it for us.”
Vice Chair, Department of Psychiatry and Behavioral Sciences, Albert
Einstein College of Medicine
Florence and Irving Rubinstein Chair in Neuroscience, Albert
Einstein College of Medicine How the brain works was and still is
Inaugurating Chair, Cerebellum, Gordon Research Conference (2011) a complete mystery.
Director, Grass Fellowship Program, Marine Biological
Laboratory (2015–2018)
Greatest Hits In Armstrong’s lab, Khodakhah demonstrated that calcium

• Demonstrated that inositol trisphosphate (InsP3) binds to InsP3 release in rat Purkinje neurons is sufficient to tamp down
receptors in cerebellar Purkinje neurons to rapidly release calcium the strength of the cells’ synaptic inputs. This dialing down
ions and modulate the neurons’ excitability. of the input is called long-term synaptic depression and is a
• Found that InsP3-evoked calcium signaling in cerebellar Purkinje neuronal mechanism by which the cerebellum learns to
neurons leads to long-term depression of synaptic inputs, a form of perform motor tasks. Khodakhah’s work also showed that
plasticity that is thought to contribute to motor learning. InsP3 molecules are needed to execute the dialing down of
• Provided evidence that the precision of intrinsic pacemaking the signal.
activity of Purkinje cells is crucial for their function in motor Khodakhah spent five years in Armstrong’s lab before accepting
coordination, and that factors that affect this precision lead to an assistant professorship at the University of Colorado Health
various forms of ataxia. Sciences Center in Denver. A year later, the National Ataxia
• Showed that a number of hereditary dystonias—movement Foundation awarded him a $15,000 grant—the first grant
disorders in which a person’s muscles contract uncontrollably— of his career and one that he says changed his professional
may be caused, primarily, by dysfunction of the cerebellum, rather trajectory. Unlike the basic science grants that he had previously
than malfunction of the basal ganglia. applied for, this award forced Khodakhah to link the dysfunction
• Showed that aside from coordinating movement, the cerebellum of Purkinje cells to motor conditions such as impaired coordination,
also contributes to controlling the brain’s dopaminergic signaling or ataxia. “I realized that those who donated the money were
and may play a role in mental disorders and addictive behaviors. putting their hope in me to help patients,” he says.
1 0. 201 9 | T H E S C IE N T IST 5 5
PROFILE
This also made him realize that neuroscientists did not have provided a number of new targets for pharmacological
precisely know how the cerebellum controlled movement and surgical management of dystonias,” Khodakhah says.
and how its dysfunction led to impaired coordination.
Prior to receiving the grant, Khodakhah recalls struggling BROADENING BRAIN CONNECTIONS
to explain the significance of his research, but that lack of Digging deeper into the literature, Khodakhah found that
understanding of how brain activity is tied to motor output there are striking correlations between perturbations of
made it abundantly clear. the cerebellum and certain brain disorders. For example,
In 2001, Khodakhah moved to the department of neuroscience people with damage to the cerebellum have an increased
at the Albert Einstein College of Medicine in New York, where risk of developing autism, and a smaller cerebellum has been
he continued to study Purkinje cells. These neurons, he explains, associated with a diagnosis of schizophrenia. Additionally,
have a regular pacemaker-like firing, which is crucial to motor brain scans of individuals recovering from addiction who were
coordination. In 2002, Khodakhah, with postdoc Mary Womack, presented with cues related to their addiction showed higher
found that calcium ion signaling is crucial for the intrinsic blood flow in the cerebellum compared to scans of the brains
baseline firing and function of these neurons. of healthy individuals.
Research has also pointed to the cerebellum’s possible
role in release of dopamine, a neurotransmitter involved in
the brain’s reward pathways. Based on the literature, two
trainees in Khodakhah’s lab, Ilaria Carta and Christopher
In my own lab, if we want to push
Chen, decided to examine whether the cerebellum is indeed
technology forward, we sit and build our interacting with the ventral tegmental area (VTA)—a part of
own circuits, or create a new computer the midbrain brain involved in reward circuitry—and plays a
program rather than waiting around for role in rewards and social behaviors. The team inserted genes
someone else to do it for us. that produce light-sensitive proteins into select cerebellar
neurons in vitro and in mice and stimulated those neurons
with light. When the cerebellar neurons that extended to the
VTA were optogenetically stimulated, the VTA neurons fired
more often.
If Purkinje cell pacemaking is crucial for motor control, Chen and Carta then assessed whether these connections
Khodakhah reasoned, then there are probably genetic between the cerebellum and VTA influenced the mice’s
mutations that affect the pacemaking activity that are linked behavior, stimulating the cerebellar neurons while the
to ataxia disorders. The lab found one such mutation in the animals were free to move around a brightly lit chamber.
literature, in the gene that encodes the calcium channel in Mice generally avoid such bright spaces, but after repeated
Purkinje neurons. The mutation, previous work showed, causes stimulation while in the chamber, the mice began to prefer
episodic ataxia type-2 (EA2). In 2006, Khodakhah’s lab showed it over a dark chamber when given the option. The finding
that mice with a mutation in the calcium channel have Purkinje supports the cerebellum-VTA connection’s role in reward
neurons that produce an irregular firing pattern. Introducing seeking and addictive behavior, Khodakhah and colleagues
a drug called 1-ethyl-2-benzimidazolinone (1-EBIO), which reported in Science this January.
targets calcium-dependent potassium channels, directly to “I think these results are going to reintroduce the
the cerebellum reestablished the rhythmic pacemaker-like cerebellum as a major brain region that contributes to
firing, which improved the motor movements of the mice. The cognitive and emotional processes,” Khodakhah says.
researchers then found that a FDA–approved analog called His team already has more findings that support the
chlorzoxazone was also effective in reducing ataxia in mice cerebellum’s contribution to addictive behavior, and in
when delivered orally. Several pharmaceutical companies are particular to the solidifying of a neurally stimulating
now planning to test such drugs in clinical trials to potentially behavior such as drug use. Such memory-making may
correct EA2 and other forms of ataxia in humans. render some individuals more susceptible to addictions.
While investigating factors that control Purkinje neuron “One of the biggest problems is that those who are addicts
pacemaking, Khodakhah realized that cerebellar dysfunction [or former addicts] can be weaned from their addiction
can also contribute to dystonia, a different form of motor but if there is a new stress, the person is very susceptible
dysfunction. Dystonia was previously thought to arise as a to relapse,” Khodakhah says. “We think the reason is that
consequence of dysfunctional basal ganglia—brain regions there is a signature of the memory within the cerebellum. .
deep within the cerebral hemispheres. However, the lab found . . If we understand that better we might be able to provide
that cerebellar dysfunction affects basal ganglia activity via a pharmacological or other therapeutic interventions to help
synaptic pathway routed through the thalamus. “Our findings these individuals.” g
SCIENTIST TO WATCH
Johannes Kohl: Circuit Breaker

Group leader, Francis Crick Institute, Age: 36
IBY NICOLETTA LANESE
A
fter pipetting countless solutions into peripheral neurons react identically to the By the time he earned his degree in 2013, Kohl
tiny tubes as a biochemistry under- pheromone. It is a circuit “switch” the flies have was ready to study an animal model other than
graduate at the University of Bayreuth deep inside their brains that leads to sex-specific flies, he says. “Flies are a fantastic model system,
in Germany, Johannes Kohl needed a break responses, Kohl explains. The transcription but I was looking for a more intriguing behavior.”
from science. So, upon completing his two-year factor fruitless, long implicated in Drosophila He chose to do a postdoc with Catherine Dulac,
pre-diploma, similar to an associate’s degree, sexual behavior, controls the position of specific a molecular biologist at Harvard University and
he embarked on a yearlong trip with his girl- dendrites and thus wires the circuit in a sex- Howard Hughes Medical Institute. Dulac’s lab
friend to South America. While he traipsed specific manner. Eliminating the fruitless gene studies mice, and the team had recently dis-
through the continent’s many jungles and cities, in males leads them to react to cVA as females covered that the neuropeptide galanin plays a
the 22-year-old Bavaria native began to itch for would, whereas expressing the gene in females crucial role in the animals’ parenting behav-
a new academic challenge. masculinizes their behavior, Kohl found (Cell, ior. The group’s research had also revealed that
“I actually can trace it back to this one day 155:1610–23, 2013). galanin is heavily expressed in a region of the
where we were basically stranded in this city hypothalamus called the medial preoptic area
in the Peruvian jungle,” he says. “Internet cafes (MPOA), and Kohl wanted to explore how this
[were] still a thing, and we were sort of bored and tiny population of neurons orchestrates parenting
didn’t know what to do. So I just started browsing and other complex behaviors.
Wikipedia aimlessly.” Kohl kept landing on neuro- “Things like feeding, fleeing, mating, and
science pages and quickly became absorbed parenting—in nature, there’s very little room
in the subject—so much so that later that year for error in these behaviors,” he says. Tracing
in 2006, he applied and was accepted to the the galanin-expressing MPOA neurons showed
Leibniz Institute for Neurobiology in Magdeburg, that they receive signals from 20 different brain
Germany, to earn his bachelor’s degree. areas and are grouped into specific neuronal
At Leibniz, Kohl began interrogating the subsets. Each subset funnels its outputs to a
forces that sculpt the neural circuits underlying different brain region. By modulating the activity
behaviors in fruit flies, as well as what physio- of the specific groups of neurons, Kohl
logical and environmental factors drive their learned that each one seems to be tuned
function. “At first, the circuit seemed mind- to specific parenting behaviors, such as
blowingly complex, and it would seem quite grooming or interacting with pups (Nature,
hopeless to find any functional principles in 556:326–31, 2018).
this tangled mess,” he says. But after he gradu- The research helped Kohl win the Peter
ated and joined Gregory Jefferis’s Drosophila and Patricia Gruber International Research
group at the MRC Laboratory of Molecular Award from the Society for Neuroscience—
Biology in Cambridge, UK, Kohl found that the a $25,000 prize, which he used to help
fruit fly “was a perfect system to investigate the launch his own lab at the Francis Crick Insti-
© CHRISTOPHER BEAUCHAMP PHOTOGRAPHY
relationship between genetics—how genes wire tute in London in January 2019.

up circuits—and how these circuits control an Kohl is now investigating how transient
animal’s behavior.” physiological states such as hunger, stress, or
As a PhD student in Jefferis’s lab, Kohl studied tiredness affect the neural circuits underlying
why male and female D. melanogaster display parenting, feeding, and aggressive behaviors.
opposite reactions to the male sex pheromone There’s no doubt that Kohl will tackle his new
11-cis-vaccenyl acetate (cVA). To females, cVA work with the same impressive gusto that he
is an aphrodisiac, while males respond to the applied to his postdoc, Dulac says. “In the lab,
scent with aggression. Combining single-cell we were making jokes that there were everyone’s
electrophysiology in the brains of immobilized standards, and then there were Jonny’s stan-
fruit flies, high-resolution confocal microscopy, dards,” she explains. “Jonny had breakthroughs
and genetic analysis, Kohl found that both sexes’ all the time.” g
1 0. 201 9 | T H E S C IE N T IST 57
LAB TOOLS
Peering into the Brain

Researchers are developing a variety of approaches for clearing
neural tissue to get a better view of the brain’s circuitry.
BY ANDY TAY
T
he mammalian brain consists
of billions of neurons wired
together in various circuits, each
one involved in specific physiological
functions. To better understand how
these different neurons and circuits are
associated with mental activities and
diseases, researchers are reconstruct-
ing detailed, three-dimensional maps
of neural networks.
However, 3-D imaging of the mam-
malian brain is challenging. Light
scatters as it travels through layers of
tissue, dispersed by a variety of mole-
cules such as water, lipids, and proteins.
This reduces image resolution. Since then, tissue clearing methods FILAMENTOUS: In a whole mouse embryo
cleared with RTF and labeled with antibody tar-
One way to improve resolution is have continued to improve as researchers
geting neurofilament, axons innervating the
to reduce the scattering. Researchers have developed chemical mixtures that bet- brain, whisker pad, and forelimb become visible
achieve this by first removing water and ter preserve tissue architecture and protein in greater detail. (Scale bars = 1,000 μm in left
lipids from tissue. Next, chemicals are structures. The latest techniques are also two images, 200 μm in right four images.)
introduced that have a refractive index— compatible with fluorescent labels. When
a measure of how much the molecules they are integrated with advanced tissue
bend light that passes through them—in processing methods such as automated cell APPROACH: Zhu and her team devel-
the range of that of proteins. Establish- counting, researchers can identify neurons oped a tissue clearing method called
ing near-homogenous refractive indices and even intracellular components with RTF (Rapid clearing method based on
in the molecules that populate the tissue ever-improving precision. Triethanolamine and Formamide) that
environment allows light rays to converge Here, The Scientist highlights some is compatible with lipid-based dyes (Sci
to improve image resolution. This is the of the recent innovations that combine Rep, 8:1964, 2018).
working principle of most tissue clearing emerging tissue clearing and tissue pro- RTF is a solution containing water,
methods, which have been used success- cessing techniques for high-resolution formamide, and a high refractive index
fully for decades on hard tissues like bone. imaging in the brain. chemical known as triethanolamine.
Researchers have performed brain In a serendipitous accident, another
tissue clearing with limited success, as LIPID-COMPATIBLE CLEARING lab found that formamide, a chemical
the chemicals available were too harsh commonly used in buffers for molec-
on delicate neural tissues. In 2013, Karl INVESTIGATOR: Dan Zhu, biomedical ular biology applications, could clear
Deisseroth and his team at Stanford engineer, Huazhong University of Sci- tissues. Some tests suggest that for-
University revolutionized the approach ence and Technology in China mamide most likely partially denatures
with a hydrogel-based technique called proteins, lowers their refractive indi-
CLARITY. This technique enabled PROBLEM: Many tissue clearing methods ces and brings them closer to those of
researchers to label neurons in mouse rely on organic solvents or detergents, water and lipids. This helps to create a
neural tissue with fluorescent mark- which degrade lipid cell membranes, as environment with greater uniformity in
ers and then to image an entire mouse well as lipid-based dyes that are typically refractive indices.
DAN ZHU
brain without sectioning it, while pre- used to stain neurons and intracellular The team showed that after clearing a
serving the fluorescence signals. organelles. mouse brain with RTF, the fluorescence
10. 201 9 | T H E S C IE N T IST 59

LAB TOOLS
signals of endogenous proteins and a TIPS: To optimize tissue transparency, hard tissues and decolorization of col-
lipid-based dye called Dil, which is widely Zhu recommends adjusting the clearing ored tissues such as liver, which looks red
used to stain axons, were not quenched. time outlined in their protocol. Although because it contains high levels of iron.
RTF is not commercially available, the For those working with small body parts,
DOWNSIDES: RTF is unable to clear reagents are cheap and can be bought off the tissues can be passively immersed in
adult mouse brain tissue that is more the shelf. The setup for clearing is also chemical reagents. Alternatively, clear-
than one millimeter thick. There is simpler than most other tissue processing solutions can be circulated through-
also an undesirable shrinkage of about ing techniques, Zhu says. out the body using a perfusion pump for
2 percent to 5 percent in cleared tis- whole body clearing.
sues, which can reduce the accuracy of CLEARING BOTH HARD AND The researchers used PEGASOS to
imaging tissue architecture and neural SOFT TISSUES trace single axons between two differ-
networks. ent segments of the spine of rats and
INVESTIGATOR: Hu Zhao, dentist, to visualize neurons within the spi-
TIME FRAME: Unlike other techniques, Texas A&M University nal cord without surgically separat-
which take about a week for clearing ing them from the vertebrae. They
soft brain tissues, RTF clears tissues Problem: Existing clearing solutions also visualized the 3-D distribution
after a day of chemical incubation. The are not optimized to clear both soft of nerves within the bone marrow
process is also reversible: cleared tis- tissues, such as brain and spinal cord, and found that nerves are spatially
sues can be restored to their original and calcified hard tissues such as bone enriched in certain bone regions.
state by soaking in saline. This is useful at the same time. This makes it chal-
for long-term storage because cleared lenging to visualize intact connections DOWNSIDES: After PEGASOS pro-
tissues are less stable. between the central and peripheral ner- cessing, tissue shrinks 30 percent to
vous systems, because peripheral nerves 40 percent, which may distort tissue
WHAT’S NEXT? Immunostaining with in the spinal cord are connected to the architecture.
antibodies allows researchers to visual- vertebrae, and vertebrae are too opaque
ize specific cells and proteins, but tissue for imaging. TIME FRAME: For immersed soft/hard
clearing typically slows down antibody tissues, clearing takes about 7 to 12 days.
penetration into tissues. Zhu is working APPROACH: Zhao and team developed a Whole-body clearing takes longer because
on a way to accelerate antibody pene- procedure, which they named PEGASOS clearing solutions must be circulated for
tration into brain tissues cleared with (the polyethylene glycol associated sol- 10 days, bringing the entire clearing pro-
RFT so as to further reduce tissue pro- vent system), to clear both soft and cess to three weeks for a young rat. The
cessing time. hard tissues at the same time (Cell Rese, duration for the circulation process must
28:803–18, 2018). The process is similar be doubled for adult rats, which have
to other clearing protocols except it uses more tissues. This lengthens the entire
VASCULAR SPOTLIGHT: Researchers used
PEGASOS tissue clearing and light-sheet micro-
polyethylene glycol (PEG) for enhanced clearing process to about a month.
scopy to visualize the mouse whole brain vascu- fluorescence preservation and contains
lature at high resolution. two additional steps: decalcification of WHAT’S NEXT? Tissue shrinkage may
not be uniform for all types of tissues.
Zhao plans to develop an improved
chemical formula of PEGASOS to over-
come this challenge. The team also
plans to reduce the autofluorescence—
natural emissions from the body’s pro-
teins that can create noise in the data—
that is associated with tissue shrinkage.
TIPS: Tissue clearing is just the first step,

says Zhao; the subsequent imaging and
data processing are more challenging. He
suggests new users start with thin tissue
slices and not an entire organ to famil-
HU ZHAO
iarize themselves with the entire work-

flow. The technique uses off-the-shelf
reagents; Zhao estimates that clearing

an entire mouse brain costs about $30.
ENLARGE FOR A BETTER VIEW
INVESTIGATOR: Hiroki Ueda, biologist,

University of Tokyo
PROBLEM: Most existing brain clearing

techniques do not leave tissue entirely
transparent, making it difficult to construct
brain maps with single-cell resolution.
APPROACH: After screening close to

1,700 chemicals, Ueda and his colleagues
developed a method, named CUBIC-X,
for expanding brain tissue after clear-
ing it (Nat Neurosci, 21:625–37, 2018).
They immersed the tissue in hydro-
philic (water-loving) reagents identified
through the chemical screen, enlarging
the volume of the brain approximately brain atlas with single-cell annota- BRAIN LANDSCAPE: Expansion microscopy
tenfold—the optimal volume expansion, tion. Users can compare their data to and tissue clearing using CUBIC-X provides
a whole-brain view in a mouse engineered to
as tissues can become fragile after that. the atlas to assess the reproducibility
express a fluorescent marker in neurons.
Using their protocol, Ueda and his col- of brain clearing. CUBIC-X is commer-
leagues were able to image entire 1 cm– cially available ($50 for 25 mL, which
thick mouse brain without losing any is enough for one mouse brain; Tokyo because small changes in a protein’s
cells. And because the expanded tis- Chemical Industry). 3-D shape can cause it to lose its fluo-
sue is 99 percent water, the refractive rescence and binding abilities.
index stays mostly the same throughout SHIELD PROTECTORS The team screened six types of
the brain’s layers. This causes tissues to epoxides and used computational sim-
become transparent and allows nearly INVESTIGATOR: Kwanghun Chung, ulations to identify the one that best
100 percent light transmission for chemical engineer, Massachusetts Insti- preserved the fluorescence of green
single-cell resolution imaging. tute of Technology fluorescent protein (GFP). They then
infused the chosen epoxide into the
DOWNSIDES: The chemicals used to PROBLEM: During tissue clearing, brain tissues of mice genetically mod-
expand tissues tend to quench fluores- enzymes and chemicals are used to ify to express GFP in neurons. There,
cence signals for labeling neurons and digest tissues, remove lipids, and some- the epoxy reacted with GFP to form
subcellular components such as RNA. times dehydrate the sample. In addition flexible chemical bonds. These bonds
to quenching fluorescent signals from protected GFP from denaturation dur-
TIMEFRAME: Tissue clearing with labeled molecular, these procedures ing tissue clearing, and allowed GFP to
CUBIC-X takes about 16 days for a inevitably degrade proteins and RNA, refold back into its natural conforma-
whole mouse brain. destroying key molecular information. tion after tissue clearing.
Chung and his colleagues showed
WHAT’S NEXT? In the future, Ueda APPROACH: Chung and his colleagues that their technique, which they named
hopes to apply CUBIC-X to other turned to a material called polyepox- SHIELD (Stabilization to Harsh condi-
organs, including heart and lungs. He ide to solve this problem. Commonly tions via Intramolecular Epoxide Link-
also plans to use the technology to clear known as epoxy resin, polyepoxide is ages to prevent Degradation), main-
the brains of larger mammals such as widely used in the aerospace indus- tained fluorescent protein signals. It
nonhuman primates and humans. try as glue to provide structural sup- also protected the ability of proteins to
HIROKI UEDA
port to weaker materials such as wood. bind to antibodies and RNA transcripts
TIPS: The researchers created the On a molecular level, it helps maintain to RNA probes after tissue clearing (Nat
CUBIC-Atlas, an open access mouse protein structure. This is important Biotechnol, 37:73–83, 2019). The team
10. 201 9 | T H E S C IE N T IST 61

LAB TOOLS
NEURAL HIGHWAYS: Researchers used

SHIELD clearing and 3-D multimodal imaging
to visualize neural circuitry in the mouse brain.
(Red and green fluorescence proteins label
neural fibers and synapses, respectively. Their
overlap yields yellow.)
combined SHIELD with tissue clearing

with CLARITY to image 2 mm–thick
segments of human brain and mouse
brain with single-cell resolution.
TIMEFRAME: SHIELD tissue treat-

ment takes about four days for a whole
mouse brain. Subsequent tissue clear-
ing requires an additional four to six
days, for a total of two weeks.
WHAT’S NEXT? The team plans to
KWANGHUN CHUNG
apply SHIELD to larger organs and
even tumors. They are also developing TIPS: Due to epoxide processing, longer for a whole mouse brain. SHIELD
an improved system to facilitate anti- SHIELD-treated tissues are stiffer and is available through Life Canvas Tech-
body penetration through tissues for have a slower antibody penetration rate, nologies. A 500 mL kit ($1,000) is suffi-
faster labeling. so antibody labeling will take two days cient for clearing about 16 mouse brains.
RESERVE YOUR SPOT BY OCTOBER 25 FOR SAVINGS UP TO $400!
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BIO BUSINESS
Breaking the Antidepressant Drought

FDA approvals of the first major new classes of antidepressant therapies in
decades open up new possibilities for the development of psychiatric drugs.
BY BIANCA NOGRADY
A
s droughts go, the one plaguing the tive metabolite of the sex hormone pro- with similar results. Known by the
antidepressant drug development gesterone, which plays key roles in the generic name brexanolone, the drug sped
landscape for the past few decades female reproductive system. through Phase 2 and Phase 3 trials before
has been noteworthy. Since the advent of Progesterone and allopregnanolone being approved by the US Food and Drug
serotonin and norepinephrine reuptake levels peak during the third trimester of Administration (FDA) on March 19.
inhibitors in the 1980s and 1990s, there pregnancy, then crash immediately after Now marketed by Sage Therapeutics
has been a dearth of new pharmacological delivery. Preclinical data suggested the as Zulresso, the therapy is the vanguard of
therapies for mood disorders, says psychi- drop in allopregnanolone could be a trigger a new wave of antidepressants. Although
atrist Samantha Meltzer-Brody, director of for postpartum depression in some women. the path to market hasn’t been straightfor-
the University of North Carolina’s Perina- The company-funded trial involved admin- ward for all drug candidates, these treat-
tal Psychiatry Program. “The same medica- istering SAGE-547 to a handful of patients ments are known for being fast-acting
tions largely that were there when I went to with postpartum depression as an intrave- and effective, and have fewer side effects
medical school a long time ago were still the nous infusion over 48 hours. than previous therapies. These improve-
© ISTOCK.COM, MARTIN BARRAUD
ones we’ve been using.” The response in the first patient ments are reflected in the price tag: the
Given this state of affairs, Meltzer- treated with SAGE-547 was dramatic. first of these new antidepressants to
Brody says she had the “most modest” of From being withdrawn and depressed reach the market—Zulresso and Janssen
expectations a few years ago when she got with no appetite before treatment, she Pharmaceuticals’ Spravato (esketamine),
involved in the first clinical trial testing began smiling, talking, eating, and inter- approved just two weeks earlier for major
a new drug, SAGE-547, for postpartum acting, Meltzer-Brody says. “After that depressive disorder—cost up to tens of thou-
depression. Developed by Massachusetts- first patient, we thought either that’s one sands of dollars for a course of treatment.
based Sage Therapeutics, SAGE-547 is a heck of a placebo or maybe there’s a sig- But what really sets these new depres-
solution of allopregnanolone, a neuroac- nal.” Three more patients were treated, sion-treating drugs apart is the “circuit-
1 0. 201 9 | T H E S C IE N T IST 63
driven” approach to their development, you actually come out with some options researches ketamine but wasn’t involved
says Sage Therapeutics’ chief research that are really good” as potential targets in the 2006 study.
officer Jim Doherty. A research focus on for antidepressant drugs. Since that research was published,
basic neuroscience has expanded under- Brexanolone, for example, is the prod- interest has surged in developing new
standing of how different neural circuits uct of research on how to modulate the func- ketamine-based therapies for depression,
are involved in brain function—and how tion of the brain’s gamma-aminobutyric and esketamine is the first ketamine-derived
to target those circuits therapeutically. acid type A (GABAA) receptors, which product on the market. It’s the s-enantiomer
“The purpose of the brain is as a commu- normally interact with allopregnanolone of ketamine—one of two mirror-image mol-
nication network,” Doherty says. Instead and other neuroactive hormones. The ecules that together make up ketamine—and
of thinking only in terms of candidate drug drug began life as an epilepsy therapy, but is administered in a nasal spray formulation.
molecules and receptors, “we try to think Sage soon realized its potential for treat- The drug was approved by the FDA last March
as much as we can at that level [of the ing postpartum depression. as an add-on therapy for treatment-resistant
whole communication network] to under- Esketamine, meanwhile, is one of major depression, but not without some
stand what are going to be the circuit-level another new class of antidepressants, controversy. “The FDA gave Janssen quite a
consequences of our molecules.” based on a drug that has been in clinical bit of flexibility,” says Todd Gould, a neuro-
use for half a century. The general anes- pharmacologist at the University of Maryland
Based on biology thetic and painkiller ketamine is one of School of Medicine. “They only met their pri-
For a long time, the only treatments avail- the World Health Organization’s essential mary outcome in one of three acute studies.”
able for depression were two classes of medicines because of its safety and effi- A typical course of esketamine
antidepressants known as tricyclics and cacy in both adults and children. A couple involves four weeks of twice-weekly
monoamine oxidase inhibitors (MAOIs), of decades ago, with growing awareness treatments, followed by maintenance
both of which were discovered in the of the role that the neurotransmitter doses once every one or two weeks in
1950s. Three decades passed before a glutamate and its interactions with the patients who respond, continuing for up to
new class emerged—the SSRIs, with the N-methyl-D-aspartate (NMDA) recep- nine months based on clinical judgement.
first drug Prozac (fluoxetine) launched tor play in mood disorders, researchers The choice of nasal delivery was deliber-
on the market by pharma giant Eli Lilly began to investigate whether ketamine, ate, says Ella Daly, therapeutic area lead
in 1988. (See timeline on page 65.) Still which blocks the NMDA receptor, might for mood in US Scientific Affairs at Jans-
the most widely prescribed antidepres- also be effective in treating depression. sen. “Unlike the intravenous formulation,
sants in the world, SSRIs are thought The first clinical study of ketamine the intranasal route is noninvasive, [and]
to influence mood by increasing levels for depression, published in 2000, found we felt it would facilitate outpatient access
of the neurotransmitter serotonin in the significant and rapid improvement in and administration,” Daly says.
brain’s synapses. But their exact mecha-
nism of action is unknown. They’re also
ineffective for many people, and even After that first patient, we thought either that’s one heck of a
when they help, can require weeks or
placebo or maybe there’s a signal.
even months to alleviate patients’ symp-
—Samantha Meltzer-Brody, University of North Carolina
toms. Researchers began to ask whether
approaches to antidepressant develop-
ment based on more-recent neurosci-
ence might prove more successful. depression symptoms in seven individu- However, because esketamine, like
“The exciting thing for a clinician- als with major depression. A second ran- ketamine, can have cognitive, dissociative,
researcher like me is to be able to see domized, placebo-controlled, double- and even psychedelic side effects, the nasal
that the field is broadening in the under- blind crossover study in 2006 confirmed spray must be administered in a super-
standing of what’s creating depression,” the benefits, and showed that they could vised medical setting, and the patient
says Jayashri Kulkarni, psychiatrist and be delivered within just two hours of an has to remain at the clinic for at least
director of the Monash Alfred Psychia- intravenous infusion, based on patient two hours after administration. “Gener-
try Research Centre in Melbourne, Aus- questionnaires. “You don’t have a suicidal ally [side effects attenuate], though, with
tralia, who is involved in a clinical trial patient sitting around for weeks or months repeated dosing, so we see that reducing
of esketamine funded by Janssen. “The trying to see if the next medication is actu- and being less significant,” Daly says.
move in the last ten years has been to look ally going to work,” says psychiatrist and Neither esketamine nor brexanolone
at causes of depression in terms of brain neuroscientist Ronald Duman, director of are cheap. The list price for Spravato is
chemistry as well as brain circuitry or the Abraham Ribicoff Research Facilities $590–$885 per treatment session, or
brain physiology, and when you do that, at Yale University School of Medicine who up to more than $30,000 for a full nine
months of treatment at maximum dos- A HISTORY OF ANTIDEPRESSANTS

age, while a one-time, 60-hour intrave- Researchers have been working for decades on new ways to treat depression, but the US
nous infusion of Zulresso costs around market is still dominated by drugs that were developed in the late 1980s and early 1990s.
$34,000. But their success has caught
the attention of the pharmaceutical
industry, which had been moving away Iproniazid, the first of the monoamine oxidase inhibitors
from psychiatric drug development due (MAOIs), is developed, after doctors realize that isoniazid,
to challenges in translating animal find- a tuberculosis drug with a similar structure, has an
ings into humans, says Duman, who has 1952 unexpected euphoric effect on patients. The drug inhibits the
received fees and grant support from monoamine oxidase enzyme, which interacts with several
Johnson & Johnson, the parent com- neurotransmitters in the brain, including serotonin.
pany of Janssen. “There’s a very renewed
interest now because of ketamine and
Spravato,” he says. “This is going to help Imipramine, the first tricyclic antidepressant, is introduced for
bring big pharma back to the table.” medical use. Derived from antihistamine compounds, this drug
1957 class blocks the reuptake of serotonin and norepinephrine into
Pursuing derivatives presynaptic neurons, thereby increasing extracellular levels of
Esketamine’s mirror twin, the r-enantiomer the neurotransmitters in the brain.
of ketamine, is also being investigated as a
potential therapeutic molecule. “The pre-
clinical data from our lab and other labs Fluoxetine, better known as Prozac, makes its debut on the
indicates that the r-ketamine is the more market as the first selective serotonin reuptake inhibitor
potent antidepressant, [but] that remains (SSRI)—still the class of antidepressants most commonly
to be tested in humans,” says Gould. While 1988 prescribed today. By reducing the reuptake of serotonin,
the s-enantiomer is a more potent antago- the drug increases the extracellular concentration of the
nist of the NMDA receptor, Gould says that neurotransmitter.
doesn’t necessarily translate to stronger
antidepressant effects.
Gould and others are also interested in Bupropion, a type of antidepressant that doesn’t fit into
the metabolites that result from ketamine’s existing drug classes, is approved as a treatment for major
breakdown in the body, after research in 1989 depressive disorder. It increases dopamine and norepinephrine
animals found that ketamine’s metabo- levels in the brain by inhibiting the neurotransmitters’
lites were not only necessary for its antide- reuptake.
pressant effects but could, by themselves,
induce ketamine-like responses. One of
those metabolites, known as (2R,6R)- Venlafaxine, the first of the serotonin-norepinephrine reuptake
hydroxynorketamine and patented by inhibitors (SNRIs), hits the market. Like the SSRIs, these drugs
Gould and others, is about to start Phase 1 1993 inhibit the reuptake of serotonin, but they additionally do the
clinical trials funded by the National Insti- same for norepinephrine.
tutes of Health.
At dosages that relieve depression-
like symptoms in animals, the compound Vortioxetine, another atypical antidepressant, is approved. In
“does not block the NMDA receptor, it addition to inhibiting the reuptake of serotonin, vortioxetine
does not produce the side effects of ket- acts as an agonist and antagonist of different serotonin
amine, and it does not appear to have the 2013 receptors, with the net effect of increasing extracellular
potential for addiction or abuse,” says amounts of serotonin and modulating the release of other,
clinical pharmacologist Carlos Zarate, downstream neurotransmitters.
chief of the section on the Neurobiology
and Treatment of Mood Disorders at the
National Institute of Mental Health who, Brexanolone and esketamine, the first of a new wave of
with colleagues, also has patents for ket- 2019 drugs born of research into the underlying brain circuitry of
amine and its metabolites for the treat- depression, are approved and put on the market.
ment of mood disorders.
Zarate believes that ketamine-based pharmacology of Zulresso but to have an in the elderly population in depression
drugs have great potential, especially if oral once-a-day pharmacokinetic profile,” are more challenging because response
the abuse potential and dissociative side Doherty says. The company also has other rates typically are lower,” Daly says. That
effects are reduced. “What we do know drugs in early development that target the study also used a lower starting dose, she
is that ketamine, at least in our research, same NMDA receptor system as ketamine. notes, adding that an older population
seems to have more-broad therapeutic It hasn’t all been smooth sailing. may need a longer duration of treatment
effects, called pan-therapeutic effects,” Pharmaceutical company Allergan had to show benefit.
Zarate says. “It seems to work very well in a high-profile failure of three Phase 3 Despite the setbacks, there is gen-
anxiety, [post-traumatic stress disorder] placebo-controlled clinical trials of its eral agreement that the antidepressant
symptoms, anhedonia or lack of pleasure, NMDA receptor–targeting drug rapas- landscape is undergoing a profound
suicidal thinking, and in fact sometimes tinel, which did not meet the primary change for the better. “It’s going to be
even in people who have failed electro- endpoint of preventing relapses of major the new norm, in that next-generation
convulsive therapy.” depression. And both the brexanolone treatments will be required to have a
The drug development pipeline for and esketamine Phase 3 trials detected rapid onset of action unless they’re spe-
treatments that, like brexanolone, target high placebo response rates, a common cial or unique in some other therapeu-
the GABA receptor system may also be feature of late-stage trials in depres- tic property,” Zarate says. “Imagine, for
opening up. Sage Therapeutics is starting sion that can make it difficult to dem- every episode of depression you inter-
Phase 3 trials of another GABAA receptor onstrate that a treatment is achieving a vene [in] very early, you could signifi-
modulator, called SAGE-217, for adults clinical benefit. cantly reduce the amount of time our
with major depressive disorder. A recent In the case of esketamine, one of its patients spend in depression, [are] not
placebo-controlled Phase 2 study showed Phase 3 trials, carried out in patients able to function, have poor quality of
that the compound achieved significant aged 65 years and older with treatment- life, and are at risk of suicide.” g
improvements in depressive symptoms, resistant depression, failed to show sta-
without any major safety signals. “That tistically significant efficacy compared Bianca Nogrady is a freelance science
molecule was designed to have the same to placebo. “It’s fair to say that studies writer based in Sydney, Australia.
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READING FRAMES
Warm-Blooded, Warm-Hearted
How evolution geared the mammalian brain toward kindness.
I
BY PATRICIA CHURCHLAND
T
hree myths about morality remain highly advantageous, endothermy is calorie-
alluring: only humans act on moral demanding: gram for gram, warm-blooded
emotions, moral precepts are divine animals must consume 10 times as many
in origin, and learning to behave morally calories as their cold-blooded cousins. To
goes against our thoroughly selfish nature. meet this intense metabolic demand, evolu-
Converging data from many sciences, tion stumbled upon Big Learning as a way
including ethology, anthropology, genetics, to upgrade foraging, molding the brain’s
and neuroscience, have challenged all three cortex as a tool to yield Big Intelligence.
of these myths. First, self-sacrifice, given Unique in its six-layer architecture, the
the pressing needs of close kin or conspe- cerebral cortex exists in all mammals, and
cifics to whom they are attached, has been only in mammals, though birds do have an
documented in many mammalian species— analogous brain structure. But our amazing
wolves, marmosets, dolphins, and even cortex comes with a cost, and that cost is
rodents. Birds display it too. In sharp con- profound immaturity of the newborn. WW Norton & Company, June 2019
trast, reptiles show no hint of this impulse. Learning involves sprouting neuronal
Second, until very recently, hominins structures to embody what is learned. If Big
lived in small groups with robust social Learning is to yield Big Intelligence, neo- us the flexibility and intelligence to pre-
practices fostering well-being and survival natal brains must have plenty of potential dict, plan, and problem solve. Variability in
in a wide range of ecologies. The idea of a for their cortical neurons to grow and forge genes as well as in the environment means
divine lawgiver likely played no part in their new network connections. To accommo- that human moral behavior has deep com-
moral practices for some two million years, date the extreme vulnerability of newborn monalities but also strong differences across
emerging only with the advent of agriculture mammals, the wiring in mammalian brains cultures, ideologies, and religions. Norma-
and larger communities where not everyone was modified to ensure that mothers—and tive differences notwithstanding, the chief
knew everyone else. The divine lawgiver sometimes fathers, siblings, and other rela- social skills we acquire in development are
idea is still absent from some large-scale reli- tives—cared for vulnerable offspring. Evo- typically transportable to other families and
gions, such as Confucianism and Buddhism. lution’s trick was to expand the wiring for even other cultures, with some adjusting
Third, it is part of our genetic heritage to care from “me” to “me and mine.” Ancient here and there.
care for kith and kin. Although self-sacrifice neural networks for pain and pleasure were Are there any practical implications of
is common in termites and bees, the altruistic modified with the upshot that we mam- this account? Possibly. One is that appre-
behavior of mammals and birds is vastly more mals feel pleasure when those to whom we ciation of the common biological under-
flexible, variable, and farsighted. Attachment are attached are safe and fed and close by, pinnings of sociality and morality may
to others, mediated by powerful brain hor- and pain and anxiety when they are threat- soften the hard edge of moral arrogance.
mones, is the biological platform for morality. ened or faraway. Perhaps it will make a little room for
As I write in my new book, Conscience: Mammalian brains are thus soft-wired humility when we are otherwise tempted
“Between them, the circuitry supporting for love and affection. Our genes see to it to assume that only our group has exclu-
sociality and self-care and the circuitry for that our brains have neural networks to sive insight into what is right and wrong.
internalizing social norms create what we form powerful attachments, which lead to Additionally, it may fuel skepticism
call conscience. In this sense, your con- social behavior such as cuddling, sharing toward those who advertise themselves as
science is a brain construct, whereby your food, and defending against attack. Our moral authorities with special knowledge
instincts for caring, for self and others, are enduring sense of self-care impels us to seek denied the rest of us. g
channeled into specific behaviors through balance between selfishness and selflessness.
development, imitation, and learning.” Our big cortex learns how to flourish in Patricia Churchland is professor emerita of
The ultimate driver of the evolution- both the social and the physical world, act- philosophy at the University of California,
ary changes that eventually produced ing as a kind of buffer to the tight genetic San Diego, and the recipient of a MacArthur
mammalian and avian brains was endo- control typical of social insects. The cerebral Fellowship. Read an excerpt of Conscience
thermy—being warm-blooded. Although cortex, especially large in humans, gives at the-scientist.com.
1 0. 201 9 | T H E S C IE N T IST 67
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Wine Therapy, Middle Ages

BY KERRY GRENS
I
n 1395, the Hôpital Civil de Strasbourg,
at the time an almost-300-year-old hos-
pital, dug a wine cellar specifically to
serve its patients—and not just with meals.
From high cholesterol to herpes, doctors
of the Middle Ages were prescribing wine
for “pretty much any disease,” says Azélina
Jaboulet-Vercherre, a wine historian at
Ferrandi Paris, a culinary and hotel man-
agement institute. For instance, she notes
that melancholia, as depression was called
back then, was treated with thin white
wine—never red. “You smell it first. It has
to smell good,” says Jaboulet-Vercherre.
“And if your case is not too severe you are
allowed a little amount to drink.”
European physicians of the Middle
Ages were by no means the first to use
wine medicinally. A millennium earlier,
the Greeks and Romans were using wine
in concoctions to treat various ailments
and as a wound cleaner. In the 11th cen-
tury, the Persian scientist Ibn Sina (called SUPPLY ROOM: The facility now known as the Cave Historique des Hospices de Strasbourg was
Avicenna in the West) wrote in his influ- installed at the Hôpital Civil de Strasbourg in France in 1395 to store wine. Doctors of the Middle
ential textbook Canon of Medicine that Ages used wine to treat various illnesses, support patients’ well-being, clean wounds, and sterilize
surgical equipment. The wine cellar at the Hôpital Civil was in use for hundreds of years, until physi-
wine “conserveth the body, expelleth dis- cians turned away from the medical use of wine and the barrels began deteriorating from neglect.
ease from the joints, purifieth the frame In 1996, the hospital revived the use of the barrels, and local winemakers continue to age wine in
of corrupt humours, engendereth cheer- them—for recreational rather than therapeutic consumption.
fulness . . . ,” according to a translation
in Philip Norrie’s chapter, “The history Jaboulet-Vercherre says a number of sometimes contaminated. For the same
of wine as a medicine,” in the 2003 book hospitals in Europe had their own wine reason, patients often got wine to drink.
Wine: A Scientific Exploration. Ibn Sina’s cellars. While physicians often offered “If you drank the water you were going
list of wine’s nearly endless applications wine as a tonic to patients, its principal use to die, and if you drank the wine you
goes on, from aiding digestion to staving was for the treatment of wounds, says Nor- wouldn’t,” says Norrie.
off gray hairs. rie, a family physician and wine historian Wine has since fallen out of favor as
Continuing this long tradition, wine in Sydney. Clinicians soaked sponges or medicine. Doctors are no longer pre-
was in such regular rotation as a pana- cloth in wine and applied it to the wound. scribing wine on the grounds that it
cea during the Middle Ages in Europe “The most important thing in the Middle “enforceth the liver,” as Ibn Sina advo-
that the Hôpital Civil would accept vine- Ages was infection,” he tells The Scientist. cated, and there is quite a bit of debate
yard plots in lieu of gold for patients’ “You were going to die of some infectious about its possible benefits versus harms.
payments. “To stock all the wine that was disease, and wine was a good antiseptic.” But Norrie is an evangelist of wine’s
produced, they had to build a cellar,” says Norrie says it wasn’t the alcohol in therapeutic advantages. He sells “the
WIKIMEDIA, IAN COATES
Thibaut Baldinger, the cellar’s manager, the wine that killed off pathogens, but world’s first resveratrol enhanced wine”
via email. The “wine therapy” given to the grapes’ polyphenols, and red wine and invokes a concept credited to the 16th
patients for wellness could be up to two was preferred over white because it con- century physician-scientist Philippus
liters a day, but Baldinger points out that tains more of these compounds. Hospi- Aureolus Theophrastus Bombastus von
it was at a much lower alcohol level than tals would also use wine to clean their Hohenheim (a.k.a. Paracelsus): it’s the
what modern drinkers imbibe. surgical instruments, as the water was dose that makes the poison. g
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