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REVIEW Volume 34, No.

2 Lab Animal February 2005

Using Obese Mouse Models in


Research: Special Considerations for
IACUC Members, Animal Care
Technicians, and Researchers
Deborah J. Good, PhD With >50 different rodent models of obesity available, Institutional Animal Care
and Use Committees (IACUCs) are seeing numerous protocols using these
models for basic research projects and drug testing. Although researchers still
The mouse is the animal most commonly
use rat and hamster models of obesity, the mouse has become the most preva-
used to study the underlying causes of and lent animal used to study this phenotype because of the availability of many
treatments for obesity. The author reviews transgenics, knockouts (KOs), and spontaneous mutants that have been created
many of the issues that should be consid- or isolated in the laboratory. We can categorize these various mutants according
to tissue physiology: white fat or adipocyte mutants, central nervous system
ered by all involved in research with mice
mutants, gut and enteric nervous system mutants, and a broad class of ‘others’,
expressing this phenotype, and describes including angiogenesis control mutants, immune function control mutants,
some procedures exclusive to obesity thermogenesis or brown fat mutants, and skeletal muscle control mutants (for
research. reviews see refs. 1–7). What these mutant animals have in common is the
propensity to gain weight on regular diets. Some show early-onset obesity, usu-
ally characterized by increased body weight gain before puberty; others show
maturity, or adult-onset obesity, characterized by increased body weight after
sexual maturity. This review examines issues that IACUC members should con-
sider during the review of protocols that use obese mice, and describes special-
ized experiments that reviewers will see researchers use in studying obese
rodents. IACUC members, as well as animal care technicians, veterinarians, and
researchers, may be interested in the animal welfare and care issues associated
with the obese phenotype.

Special Welfare Considerations for Using Obese Mice


In addition to standard concerns for reviewing animal protocols, research proto-
cols using obese mice must address the special circumstances involved with using
these models. These circumstances fit into several categories: husbandry, geno-
types, and total animal numbers; housing conditions and enrichment; food
intake; surgery and anesthesia considerations; and humane endpoints. The
researcher planning the project and the IACUC members reviewing the protocol
must take all of these into consideration.

Husbandry, Genotypes, and Total Animal Numbers


The vast majority of the obese mice used in research today are KO mice, which are
animals with a homozygous deletion of a particular gene. Because of this condi-
The author is an Associate Professor in the tion, most obese animal colonies require careful husbandry, along with meticu-
Department of Veterinary and Animal Sciences, and lous genotyping and record keeping. Compared with wild-type (WT) colonies,
IACUC Chair at the University of Massachusetts, the breeding requirements and limitations of many obese KO mice lead to an
Amherst, MA 01003. Please address increased number of unused animals.
correspondence to the author at In general, researchers use heterozygous (HET) animals as breeders, resulting
goodd@vasci.umass.edu. in a 1:2:1 ratio of WT, HET, and homozygous KO mice. Because most researchers

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February 2005 Lab Animal Volume 34, No. 2 REVIEW

use the homozygous KO mice in the experiments, with WT mice as hanging cages (Fig. 1). One study using rats found no difference
controls, breeding results in a surfeit of HET mice. It is important in food consumption or body weight gain with either cage system,
to consider this, both for reporting animal numbers in the proto- although rats clearly preferred the solid-floor caging as measured
col as well as in justification for not meeting the ‘reduction’ crite- by the percentage time spent in the solid floor cage (88%) when
rion for the ‘3 Rs’ (reduction, refinement, and replacement)8. given access to both via a perspex tube20. With all caging systems,
The use of KO breeders to produce homozygous KO mice and there should be monitoring of the housing of obese mice with
WT breeders to produce the control mice would be one means by respect to density. The Guide for the Care and Use of Laboratory
which to eliminate the overproduction of HET mice. However, in Animals (Guide)21 provides guidelines, based on body weight and
many obese models, the KO animals are infertile or show reduced required floor space per animal, that the researcher can use to
fertility9–11; therefore, they cannot become reliable breeders. These determine the number of animals allowable in a cage. In general,
circumstances offer researchers no choice but to use the HET mice up to four mice with a combined weight of <125 g can live togeth-
as breeders, resulting in the production of twice as many animals er in a standard 11 × 7 × 5 in. mouse cage. Because some obese
as are needed for the proposed experiments. Charts and informa- mice can weigh as much as 60–70 g (or more than twice that of
tion in Guidelines for the Care and Use of Mammals in Neuroscience the average mouse), there should be frequent monitoring of obese
and Behavioral Research can be useful in determining the exact animals in group housing to assure that minimum space require-
numbers of animal needed when the male and female homozygous ments are met.
KOs have reduced fertility12. To address possible animal welfare concerns, researchers study-
There is evidence of a maternal environment effect on the ing diabetic animals should consider the bedding type, the amount
development of obesity. For example, in mice heterozygous for a of bedding in the cage, and the cage type. Diabetes, which can
mutation in the leptin receptor (Lepr (db/+)13–16, gestational dia- accompany the obese phenotype, results in increased urine pro-
betes might contribute to increased adult body weight in WT off- duction in both humans and rodent models of the disease22. The
spring of db/+ mothers, compared with WT offspring from WT diabetic phenotype may require more frequent cage changes or a
dams17. Other reports show that in mice heterozygous for a dele- more absorbent bedding to compensate for the increased urine
tion in a phospholipid transporter, Atp10c, inheritance of the production. Some IACUCs recommend for diabetic animals the
mutation from the dam resulted in increased body weight and use of wire-grid hanging cages, which prevent the animals from
serum leptin levels and an increased risk of type 2 diabetes, com- lying in the urine-soaked bedding that could be present in solid-
pared with WT or Atp10c heterozygotes that had inherited the floor caging. However, one study has found that diabetic rats
mutation paternally18. In mice heterozygous for a mutation in the exhibit distal tibial mononeuropathy earlier than nondiabetic rats
stimulatory G protein alpha subunit α (Gsα), those inheriting the when reared in wire-grid hanging caging23. There should therefore
mutation maternally are obese, whereas paternal inheritance be careful consideration of choice of caging with regard to animal
results in thinness19. Each of these examples represents cases in needs and experimental design.
which breeding schemes and therefore animal numbers would There are limited published data on the relationship in mice
have to be modified further to correctly study the genetic contri- between housing conditions and body weight. One study of
bution and phenotypic outcomes of the mutant alleles or loci;
moreover, the researcher during the preparation of the IACUC
protocol and the IACUC members during review would have to
take these characteristics into account.

Housing, Temperature, and Enrichment


Although most obese mice do not require specialized individual-
ly ventilated microisolator cages, the type of housing (solid floor-
ing or grid/hanging cages) and the housing conditions (group or
singly housed animals, cage shelf level, room temperature, and
environmental enrichment) can affect behavior, food intake, and
body weight gain in both normal-weight and obese-prone
rodents. Researchers should consider these conditions when
studying obese mice; animal caretakers and IACUC members
should also be cognizant of the need for controlling housing con-
ditions for these animals.
Most mice live in standard solid-floor caging with recycled
FIGURE 1. Wire-grid hanging cages. We include Nestlet pads and
paper or wood chip bedding. Some experiments, such as food plastic plumbing pipes to provide areas of solid flooring and
intake analysis (discussed later), require the use of wire-grid enrichment to the mice.

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REVIEW Volume 34, No. 2 Lab Animal February 2005

nonobese diabetes-prone mice found no difference in the onset of ences in body weight gain described earlier for group-housed
diabetes if mice were housed singly, or with up to seven other age- compared with singly housed animals are due at least in part to
and sex-matched mice24, whereas another study suggests that ‘huddling’, which reduces thermoregulatory energy needs. A study
singly housed mice show less variation in body weight and body fat that examined the effect of cage enrichment, including parame-
content than mice housed five per cage25. One study by Leiter ters such as group housing, nesting materials, nest boxes, gnawing
shows that group-housed male mice subjected to stress induced by sticks, and PVC tubes found that the enriched animals (i.e., in
repetitive handling developed glucose intolerance that could be cages containing nesting material or nesting material plus one or
ameliorated by adrenalectomy at 3 weeks of age26. Notably, studies more of the extras) gained more weight than mice that were sim-
by Van Loo and colleagues suggest that male mice prefer group ply group housed, even though they showed reduced food intake,
housing over single housing, especially in regard to sleeping sites, suggesting that the enrichment materials may also reduce energy
but that neither housing condition altered body weight or food needs of the mice33. Thus, researchers working with obese rodents
intake27. Conversely, group-housed hamsters show a >30-g should monitor housing environment, as well as room tempera-
increase in body weight and a 6% increase in body fat compared ture, in their colony maintenance and experimental conditions.
with individually housed hamsters28.
Shelf level on the cage rack may also affect body weight, with a Food Intake
higher shelf level on the cage rack corresponding to a lower inci- With few exceptions, mice with targeted or spontaneous mutations
dence of diabetes and the least amount of weight gain28,29. Thus, in genes that result in obesity show hyperphagia (i.e., increased
the literature suggests that researchers should design their housing appetite)7. In addition, a diet high in fat can result in obesity in nor-
for obese mice with consideration for conditions that will affect mal mice5. Animal care facilities purchase rodent diets that can have
their own testing results, and based on consistency in housing a crude fat content anywhere between 4.5% and 11% (e.g., Purina
parameters between obese and normal control mice, keeping in 5001, Purina Prolab RMH 2000, Purina 5015, Purina LabDiet, St.
mind the density limitation specified by the Guide21. Louis, MO). In animals with which this author works, moving from
It is clear that conditions that affect the rate of body heat loss, the National Institutes of Health (NIH) to the University of
such as room temperature and group housing, can affect weight Massachusetts resulted in a slight reduction in body weight, espe-
gain in rodents. The Guide suggests housing mice at a room tem- cially in our older Nhlh2 KO mice. We traced this weight loss to a
perature of 18°–26°C (64°–79°F), with minimal temperature fluc- change from Purina 5015, containing a minimum of 9% fat, which
tuations21. Many obese animals show reduced core body tempera- was fed to the mice at NIH, to Purina 5001, containing a minimum
tures, and some show an inability to respond effectively to cold of 4.5% fat, which is currently their diet at the University of
exposure by increasing their own heat production5. One study Massachusetts. Additional discussions with representatives from
examined the ability of the ob/ob mouse, which has a spontaneous several rodent diet suppliers have led our laboratory to investigate
mutation in the adipocyte hormone leptin, to respond to different the possibility that differences between these rodent diets and,
environmental temperatures. At room temperatures between 30°C more importantly, between individual lots of the same rodent diet,
and 10°C, the body temperature of ob/ob mice is consistently which may vary in the maximum amount of fat, could lead to
reduced by up to 2.5°C compared with normal mice, most proba- experimental differences in the body weight gain in our obese KO
bly due to reduced energy expended on thermogenesis30. When mice. Researchers should be aware of the type of rodent food used
room temperature is reduced to 4°C, ob/ob mice cannot adapt and by their animal facility and ensure that the animals’ diet most close-
die of hypothermia, whereas normal mice maintain their body ly fits the particular study needs.
temperature through cold-induced thermogenesis30.
Carrying a spontaneous mutation in the leptin receptor, db/db Surgery and Anesthesia of Obese Mice
mice show a similar response to a room temperature of 4°C, and To this author’s knowledge, there has been no comprehensive or
their body temperature is 1°–2°C lower than normal mice when comparative study of the effect of different anesthesia regimens on
maintained at 23°C (ref. 31). At 23°C, db/db mice deposit 75% obese mice, although many veterinary medicine textbooks address
more fat than normal mice, because of reduced thermoregulatory dosage calculations that compensate for obesity and the comitial
thermogenesis32. Furthermore, at temperatures considered to be decrease in water compartmentalization. In general, increased body
thermoneutral (33°C), db/db mice still deposited 25% more body weight, in particular the excess fat accumulation that occurs in
fat than normal mice, most probably because the mutant mouse obese mammals, alters drug kinetics and may cause cardiopul-
was unable to show a compensatory reduction in food intake in monary complications during surgery under general anesthesia. In
response to the reduced energy requirements32. Thus, fluctuations addition, as a result of the function of some genes, KO mice could
in room temperature during experimental testing of obese rodents show both obesity and changes in sensitivity to anesthesia.
could have slight, but meaningful effects on weight gain, fat depo- Examples of this sensitivity are the KO mice for the brain NPY
sition, and core body temperature. receptors, Y1 and Y2. The Y1-receptor KO mouse displays both
Moreover, the prevailing opinion is that some of the differ- obesity and reduced sensitivity to pentobarbital and avertin34–36,

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February 2005 Lab Animal Volume 34, No. 2 REVIEW

whereas deletion of the Y2 receptor results in increased body weight Specialized Procedures for Studying Body
in females and increased sensitivity to pentobarbital35,37. Weight Regulation in Rodents
When selecting the anesthesia agent to use with obese rodents, IACUC reviewers of protocols from researchers studying body
the surgeon should take into account the agent’s fat solubility. For weight regulation will certainly encounter proposed experiments
highly lipophilic drugs, including barbiturates (e.g., pentobarbital) that are somewhat unique to the obesity research field. In particu-
and cyclohexamines (e.g., ketamine), animals with large amounts lar, food intake and food deprivation studies are common, along
of body fat show delayed recoveries, and sometimes death, due not with specialized diets designed to promote either weight gain or
only to the uptake and storage of these drugs in the fat, but also to loss in the animals. Body temperature and the effect of different
severely depressed respiration from slower metabolism38. The temperatures to simulate thermoneutrality or cold-induced ther-
inhalant drug isoflurane, which is insoluble in both blood and mogenesis are probable considerations. Finally, researchers often
fat38, achieves rapid and easily reversible anesthesia with no death propose measurements of energy expenditure through calorimetry
or breathing difficulties in our Nhlh2 obese mice, which contain a and physical activity. IACUC reviewers should be aware of special
targeted deletion of the neuronal transcription factor. Inhaled considerations for these types of experiments and ensure that
anesthesia requires the use of a mask or nose cone for the rodent, researchers follow guidelines in their design and implementation
a vaporizer for the anesthesia, and a regulator to deliver a supply of that comply with applicable guidelines and regulations.
oxygen during anesthesia. Workers should have protection from
the inhalant anesthesia, which enters the atmosphere during the Food Intake, Food Deprivation, and Specialized Diets
procedure. They most commonly do surgery in a fume hood or When a new KO or mutant mouse is determined to have a pheno-
with a scavenger unit (e.g., NP 60-0979 F/AIR Filer canister, type of increased body weight, studies of food intake are some of
Harvard Apparatus, Inc., Holliston, MA) to remove the vapors the first to be undertaken. Next on the list are food deprivation and
from the surgery suite. IACUC members should make certain that refeeding studies to analyze changes in gene expression, body
the choice of anesthesia is appropriate for the animal, especially weight, and other physiological and behavioral changes in the
ensuring that the researcher understands the potential complica- mutant animal, followed by analysis of the response of the mutant
tions of anesthesia in obese rodents. If the scientist is choosing mice to specialized diets with high-fat, low-fat, or pharmacological
inhaled anesthesia, then the IACUC should ensure that the sur- additives.
geons follow the requirements to protect themselves. The analysis of food intake usually involves placement of mice
in specialized cages so that daily food intake can be measured, tak-
Humane Endpoints ing into account both ingested material and spillage. The most
Researchers should establish humane endpoints for all studies economical and technologically simple method for doing this
involving animal use, but for obese and diabetic animals, some analysis involves the use of a hanging metal cage with a wire-grid
guidelines may require implementation to prevent distress or dete- bottom. Animals live individually in these cages, with food spillage
rioration of the animals due to the nature of their disease39. For a collected daily on paper mats beneath the cage. Workers weigh
diabetic animal, if diabetes is ongoing, end-organ effects will occur, both the remaining food in the bin and the spillage to determine
including diabetic retinopathy, diabetic nephropathy, and uncon- the daily intake for each animal. Vendors also sell cages with elec-
trollable hyperglycemia40. For an obese animal without diabetes, tronic and computerized weighing systems, although most still use
severe weight gain can result in hypertension, renal problems, and a grid flooring in their design to allow the measurement of
even respiratory depression41,42. The IACUC should be certain that spillage. The primary problem with each of these experimental
the researcher has established a humane endpoint with regard to designs concerns the guidelines for animal cage flooring published
the phenotype of the disease model. In particular, there should be in the 1996 revised version of the Guide12. Although these guide-
no possibility of the animals developing morbidities associated lines do not prohibit wire-grid flooring for rodents, they recom-
with obesity or diabetes, if it is not necessary to the research ques- mend solid flooring on the basis of research showing that rodents
tion being asked. Because it is not possible for the animal care staff prefer solid flooring and may develop foot lesions on wire-grid
or researcher to have a generalized list of clinical signs that will flooring20. In our laboratory, we use wire-grid flooring for food
cover all types of experimental paradigms, one paper has suggested intake studies, but we include plastic plumbing pipes 4 in. in diam-
the use of an individualized score sheet system that would allow eter and ∼5 in. long, as well as Nestlet pads (Ancare Corp.,
caretakers or researchers to assess the level of pain or distress in Bellmore, NY) to provide some areas of solid flooring and enrich-
their animals39. The sheet would be available in the animal room ment to the mice (Fig. 1)43.
and would list all possible clinical signs, scoring details, and the Following the results of food intake studies, many researchers
steps to take if one or more of the clinical signs are present, includ- continue with analysis of food deprivation and refeeding. They
ing scientific steps such as isolation of particular tissues for analy- normally use these experiments to analyze the role of the gene of
sis. In this way, there can be assessment of the welfare of the animals interest (i.e., the knocked out or mutant gene) on physiological
without compromising the scientific goals of the study. and behavioral parameters. The Guide21 specifies that food can be

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REVIEW Volume 34, No. 2 Lab Animal February 2005

withheld for research purposes but that the animal should be organ function represents the majority of energy usage by the
monitored for distress. Mice can go without food for up to 48 h body, adaptive thermogenesis, which is induced by cold exposure
(refs. 44–46), but effects on gene expression can occur at time or diet, accounts for ∼25% of the required daily energy expendi-
points ≤24 h (refs. 47,48), so design of the study should allow for ture and is usually defective in obese mutant mouse models50.
the minimum required time needed for an effect. In addition, There are two methods for measuring core body temperature
mice should have, and generally do have, unrestricted access to in mice. First, surgically implanted transmitters (e.g., Mini Mitter
water during these deprivation periods. Fasting experiments often Co., Inc., Bend, OR) can transmit core body temperatures contin-
take place in cages with wire-grid bottoms, because fasting- uously and throughout the lifetime of the mouse. The obvious
refeeding paradigms are common endpoints. Thus, the same con- concern is that the animal must undergo a survival surgery for the
cerns mentioned for grid flooring in food intake studies should implant, but because these devices are placed under the skin and
receive consideration in the fasting-refeeding experiments that no body cavity opening is required, surgical complications are
use these cage types. In addition, several reports have suggested usually minimal. The second method for body temperature mon-
that the ambient temperature of the housing conditions influence itoring is manual, using rectal probes. In our laboratory, we have
the effects of food restriction themselves49. In particular, reports used a tissue-implantable thermocouple microprobe (Physitemp
indicate that standard housing temperatures of 23°C represent IT-18, Physitemp Instruments, Inc., Clifton, NJ) as a mouse rectal
mild cold stress for mice and that this temperature actually results probe attached to a Physitemp Thermalert TH-5 digital ther-
in elevated food intake. Thus, colder room temperatures in addi- mometer (Fig. 2)43. To avoid irritation to the mice, we clean the
tion to food deprivation could represent an added stress on the probe in 70% ethanol between animals, and we coat it with min-
animal. In addition, as discussed earlier, group housing can affect eral oil before insertion. We measure temperatures at a depth of
the energy requirements of animals because of their tendency to 2 cm, for ∼15 s, or until a stable reading is obtained. We have
huddle to conserve energy. Whether the researcher uses single or marked our probe with a dot of white correction fluid at the 2-cm
group housing will most probably affect the severity of the fasting depth to ensure the consistency of our reading and animal safety.
regimen. These issues should receive consideration for all food Whereas the advantage of this method over the implanted devices
deprivation studies. is that animals do not need to undergo surgery for temperature
Diet-induced obesity is one of the most common methods for monitoring, the disadvantage is that rectal measurements can be
studying the role of diet in the development of obesity 5. stressful to the animal and can result in injury if a probe is insert-
Specialized diets, formulated in the amount of fat, calories, or ed into the rectum beyond 3 cm. Although there are only anecdo-
other components, are available from a number of suppliers (e.g., tal reports on the injuries that can occur with rectal probes, there
Research Diets, Inc., New Brunswick, NJ; Harlan Teklad, Madison, are many studies on the stress effects of rectal temperature mea-
WI; and Purina LabDiets). Research specifications can also dictate surements on mice and rats. In mice, repeated measurements
the formulation of diets, with the addition of specific types of car- using a rectal probe can result in hyperthermia of >1°C (ref. 51).
bohydrate or fat sources or drug to modulate food intake. For In addition, stress-induced hyperthermia can occur when group-
IACUC protocols that request the use of a nonstandard diet for housed mice are measured sequentially, with the core temperature
body weight studies, reviewers should request that the expected of the last mouse increased by nearly 2°C (ref. 52). Use of this
effects of this diet be adequately explained and justified. Some of method for body temperature reading does therefore have the
the more palatable diets, such as the rodent cafeteria diet, com- potential to confound results. Thus, IACUC members reviewing
posed of cookies, breads, peanuts, cheese, and other ‘delectables’ protocols in which body temperature measurements will be made
for mice, can cause animals to overeat or become hyperphagic, in should make sure the researcher is aware of the disadvantages to
addition to having a higher overall fat content than a standard each method and has implemented safeguards to protect the wel-
rodent diet5. These diets can lead mice to become extremely fare of the animal, either from stress or anxiety induced by the
obese, sometimes doubling their total body weight over the nor- manual method for body temperature measurement, or from pos-
mal 10- to 30-week period of feeding. Because this situation can sible surgical complications with the implanted devices.
lead to cage densities that are higher than those allowed, the Indirect calorimetry is the measurement of the O2 consumed
IACUC reviewer should carefully consider the time frame for spe- and the CO2 produced by an animal. Researchers then use the
cialized diets, as well as the total weight gain expected over that measurements of these variables to calculate the metabolic rate or
time period. energy usage of the animal. Several suppliers provide individual
units for measuring respiration in mice (e.g., Oxymax cages,
Body Temperature Measurements and Experimentally Columbus Instruments, Columbus, OH). In these units, mice usu-
Induced Energy Expenditure ally have ad lib access to food and water, and have plenty of room
Most obese mice show reduced energy expenditure, measured by for exercise. Thus, there are no animal welfare issues associated
body temperature, respiration rate, or physical activity (discussed with use of these cages for metabolic measurements. If the animals
later)6,50. Although obligatory energy expenditure for body and are held in calorimetry chambers for >24 h in an area outside of

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February 2005 Lab Animal Volume 34, No. 2 REVIEW

FIGURE 2. Core body temperature measurement in an obese FIGURE 3. Running-wheel cages. The mouse running on the
mouse. We use a mouse rectal probe inserted to a depth of 2 cm. wheel trips a circuit each time it makes one full revolution.
A small amount of correction fluid indicates the depth mark to the Computer software calculates the number of revolutions in 20-
researcher; this is indicated in the figure as the white mark on the min time bins.
probe stem nearest to the mouse’s anus.

the normal animal room, IACUC reviewers should inspect the decrease in body temperature in some mouse models that show
holding area or room, as required by PHS Policy for Satellite Areas. normal response to cold with ad lib food access (e.g., DGAT1
Once basal measurements of either body temperature or meta- mutant mice54). Accordingly, IACUCs and researchers using this
bolic rate are made, researchers studying body weight regulation method to analyze body weight and gene function in mutant mice
often test the ability of their obesity models to respond to diet- or should provide adequate monitoring of animals during cold expo-
cold-induced thermogenesis. Normal animals sense augmented sure to ensure that death from severe hypothermia is prevented
food or caloric intake, and the body and brain respond with a com- and is not an unexpected endpoint in their studies. In addition,
pensatory increase in thermogenesis. For studies of diet-induced IACUC review of the protocol should include inspection of the
thermogenesis, mice usually receive a high-fat diet and have their cold room housing any rodents for >24 h.
body weight changes monitored weekly. Many obese mouse models
do not respond with a compensatory increase in thermogenesis and Physical Activity Measurements
show massive weight gain on these diets. As discussed earlier, The amount of physical activity done by an animal can influence
researchers and IACUCs should monitor the use of high-fat diets to the amount of energy used by that animal. Mild food restriction or
ensure that maximum cage densities are not exceeded. cold exposure induces in some obese mouse mutants (e.g., ob/ob
For cold-induced thermogenesis, researchers measure core mice55) a torpor or a period of physical inactivity accompanied by
body temperatures of animals in response to differing lengths of reduced core body temperature. Many obese mice, including the
time at thermonegative temperatures, usually 4°C. Normal mice ob/ob, melonocortin-4 receptor KOs, and those that we use in our
can adapt to this temperature by increasing thermogenesis and laboratory, the Nhlh2 KO mice, show reduced daily physical activ-
maintaining relatively normal core body temperatures, but many ity when given access to a running wheel or when monitored using
mutant obese mice fail to respond and show severe declines in beam breaks with open-field testing equipment38,56,57.
body temperature in relatively short time frames. For example, Measurement of daily activity with either computerized running
mice lacking all three β-adrenergic receptors fail to increase ther- wheels (Fig. 3) or open-field testing equipment does not pose any
mogenesis after 10–20 h of exposure to 6°C, showing core body animal welfare issue, and, in fact, animals with running-wheel
temperature reductions of 4°–10°C (ref. 53). Some of the animals cages receive an added enrichment benefit.
in the study appeared to have been taken out at earlier time points, An extension of physical activity measurements with which
with body temperatures of <27°C after only several hours of expo- there could be some animal welfare concerns is the use of forced
sure54. Fasting animals in conjunction with cold exposure to max- exercise either to measure gene expression changes in response to
imize changes in energy balance can also result in a pronounced physical activity or to measure the ability of increased physical

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REVIEW Volume 34, No. 2 Lab Animal February 2005

2. Arch, J.R. Lessons in obesity from transgenic animals. J. Endocrinol.


activity to reduce body weight in obese rodents. In these experi- Invest. 25(10), 867–875 (2002).
ments, which are becoming more commonplace for obesity 3. Brockmann, G.A. & Bevova, M.R. Using mouse models to dissect the
genetics of obesity. Trends Genet. 18(7), 367–376 (2002).
researchers, mice exercise on miniature treadmills either at a set
4. Mauvais-Jarvis, F., Kulkarni, R.N. & Kahn, C.R. Knockout models are
speed or with increasing speed or increasing inclines for a set peri- useful tools to dissect the pathophysiology and genetics of insulin resis-
od of time. Scientists in the field of skeletal and cardiac muscle tance. Clin. Endocrinol. (Oxf.) 57(1), 1–9 (2002).
5. Tschop, M. & Heiman, M.L. Rodent obesity models: an overview. Exp.
research have used such procedures for quite some time, with Clin. Endocrinol. Diabetes 109(6), 307–319 (2001).
treadmill speeds of up to 20 m/min and times ranging from 15 min 6. Robinson, S.W., Dinulescu, D.M. & Cone, R.D. Genetic models of obesi-
ty and energy balance in the mouse. Annu. Rev. Genet. 34, 687–745
to two 3-h bouts of exercise per day58,59. Laboratories studying body (2000).
weight often measured O2 consumption along with exercise, with 7. Beck, B. KO’s and organisation of peptidergic feeding behavior mecha-
moderate exercise protocols consisting of 12 m/min speed and up nisms. Neurosci. Biobehav. Rev. 25(2), 143–158 (2001).
8. Russell, W.M.S. & Burch, R.L. The Principles of Humane Experimental
to 1 h of running (e.g., Bouthegourd et al.60), or intense bouts of up Technique (Methuen, London, 1959).
to 18 m/min until exhaustion, usually between 17 and 30 min (e.g., 9. Magni, P., Motta, M. & Martini, L. Leptin: a possible link between food
intake, energy expenditure, and reproductive function. Regul. Pept.
Faldt et al.61). The Animal Welfare Act (AWA) mandates exercise for 92(1–3), 51–56 (2000).
laboratory dogs and stipulates that this exercise must be voluntary 10. Good, D.J., Porter, F.D., Mahon, K.A., Parlow, A.F., Westphal, H. &
and cannot be in the form of forced exercise on a treadmill, except Kirsch, I.R. Hypogonadism and obesity in mice with a targeted deletion
of the Nhlh2 gene. Nat. Genet. 15(4), 397–401 (1997).
when exemptions are given for scientific research62,63. Because lab- 11. Johnson, S.A., Marin-Bivens, C.L., Miele, M., Coyle, C.A., Fissore, R. &
oratory mice are not covered by AWA, no such mandates exist. Good, D.J. The Nhlh2 transcription factor is required for female sexual
behavior and reproductive longevity. Horm. Behav. 46(4), 420–427 (2004).
Under Public Health Service guidelines, especially in the area of 12. Institute for Laboratory Animal Research, National Research Council.
alternatives and refinement of procedures, there should be assur- Guidelines for the Care and Use of Mammals in Neuroscience and
Behavioral Research (National Academies Press, Washington, DC,
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