13/01/2009 13:55:00

← Ion Channels: Depolarizing

← Ionophores: non-gated channels
• Toxins:
o Monensin
 Source: Bacteria
 Mechanism of Action
 Macro – antibiotic
 Micro – Na+ ionophore
o Amphotericin
 Source: Strepococcus
 Mechanism of Action
 Macro – antifungal and antiprotozoal
 Micro – Na+/K+ ionophore
←
← ATPases (pumps) can change membrane polarization
Ex: Na+/K+ ATPase For every 3 Na+ pumped out, 2 K+ ions enter the cell;
causing a hyperpolarization.
←
← Pump toxins disrupt function
• Toxins:
o Digitalis
 Blocks Na+/K+ ATPase by inhibiting dephosphorylation
o Ouabain (wabain)
 Blocks Na+/K+ ATPase
 o Thrapsigargin
 Blocks Ca++ ATPase in ER

Depolarizing channels
• Voltage-gated
o Na+ channels
o Ca++ channels
←
← Gating Models
• *Regional conformational change – a selective portion of the protein structure
changes
• General conformational change – entire structure changes
• *Blocking particle
← * most likely to be the cause
←
• Three kinetic steps
 o Open  inactivation  closed
o The inactivation gate (msec) will be much slower than the activation
gate (nsec)
←
• *Voltage-clamp single-channel recording
o The voltage would rise during the voltage command step
o The current opens during the command step and it shuts off but the
voltage still remains depolarized.
←
• *Active, inactive and closed
o In the inactive state there is no current but the voltage still remains
depolarized.
←
← Channel structure conservation
• Na+ and K+ channels end up being made up of a single protein. There is an open
central space. There are inactive regions and active regions within the protein.
←
←
←
←
←
←
←
← Na+ Channels
← 4 subunits with 6 trans-membrane segments each
← Na channel types NaV1.x  don’t need to know
←
← Additional Na+ channel blockers
 • Toxin:
o Saxitoxin (STX)  red tide
o Tetrodotoxin (TTX)
o QX-314  works from the inside, used in recording electrode to
selectively turn off neurons individually
o **Cocaine
o **Lidocaine  widely used because of less cardiac and liver toxicity
← **Local anesthetics block Na+ channels

←
• Cardiostimulants: prevent inactivation of Na+ currents
o *Aconitine – monks hood
o *Veratridine – rhizomes of sabadilla lily
← * Both are considered neurotoxins but may be used for very low heart rates
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←
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←
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←
←
← Ca++ Channels
← Intracellular Ca++ Channels
• IP3 (neurons and muscles)
o IP3 is the agonist
o Heparin (blood thinning agent) and Caffeine are antagonists
• Ryanodine (skeletal muscle, cardiac muscle, neuronal)
o Ryanodine, Caffeine, and Heparin are all agonists
o Dantrolene is the antagonists

← Two major types of Ca++ Channels

• LVA (low-voltage activated)
o Rapidly inactivating
• HVA (high-voltage activated)
o Inactivating
o Noninactivating
← ++
Ca Channel subunit structure
• 4 subunits with 6 trans-membrane domains each
←
← Ca++ Homeostasis in Neurons
← Ca++ Influx
• VDCCs
• NRs (NMDA receptors)
• Na+/Ca++ exchanger
← Ca++ Stores
• CaBPs (Ca++ Bound Proteins)
• ER
• Mitochondria
← ++
Ca Release
• IP3Rs
• RyRs
• Ca++ dependent
• Ca++ release
← Ca++ Efflux
• ATP-ases
• Na+/Ca2+ exchanger
← **Ca++ imaging with fluorescent indicators Fura 2  most commonly used
←
← Voltage-Dependent Ca++ Channel (VDCC) types:
 ← L ← T ← N ← P ← R
← (tra /Q
nsient)
← Cell Type ← Cardia ← Car ← N ← N ← N
c diac euronal euronal euronal
← Muscl ← Neu Only Only Only

e ronal
← LVA (fast ← ← X ← ← ←
inactivating)
← HV ← Inactivat ← ← ← Sl ← S ← F
A ing owest low ast
← Noninact ← X ← ← ← ←
ivating
← Blo ← Dihydrop ← *Nifed ← ← ← ←
ckers yridine ipine
← **Nim
odipine
← Arylalkyl ← Verap ← ← ← ←
amine omil
← Benzothi ← Diltiaz ← Flun ← ← ←
azepine em arazine
← Mib
efradil
← Alkaloids ← Phlore ← ← ← ←
tin
← Conotoxi ← ← ← Ω- ← Ω- ←
CTX-GVIA CTX-MVIIC
n
← Ω- ← Ω-

CTX-MVIIC Aga-IA

← Inorgani ← X ← X ← X ← X ← X
c ions
← En ← BAY-K- ← X ← ← ← ←
hancers 8644
← Maitotoxi ← X ← X ← X ← X ← X
n
← * Widely used for hypertension to regulate heart rate and cardiac output
← ** Brain specific

←
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←
←
← Voltage-Gated Channels: Hyperpolarizing
• Chloride (Cl-) Channels
• Potassium (K+) Channels
*(K+ channels can flow inward if the cell is above than the resting potential or
outward if the cell is below the resting potential voltage -65mV.
←
← Chloride (Cl+) Channel Toxins:
• Chlorotoxin
• Flufenamate (chlorea use)
• Tamoxifen (breast cancer treatment)
←
← K+ Channel toxin sources – organic and synthetic
←
← Four Major Classes of K+ Channels
• Voltage-Dependent K+ Channels (VDKC)
o IA (first observed)
o ID (delayed)
o IK
o IM (muscurine sensitive)
• Rectifier K+ Channels (making right; returning to right potential)
o IK,
o Anomalous Rectifier (activated by below resting potential)
 o IH – hyperpolarization activated
o Tandem Pore (forms two channels per molecule)
• Metabotropic-Gated K+ Channels
o IM (metabotropic regulated)
o ATP-sensitive (requires ATP)
o IAHP (After Hyperpolarization Potential)
• Ca2+-Dependent K+ Channels (CDKC)
o IC (calcium)
o IM
o IAHP
←
←
←
←
← K+ currents and action potential firing
• IA and ID are the first to be activated and try to keep the action potential back to
resting. As the voltage rises up, the delayed rectifier, IK, tries to bring the
resting potential back. Then Ca++ dependant K+ channels are activated . Then
the IC, IM, IAHP to lower the after hyperoplarization potential. Then the slowIAHP,
and ISAHP. These determine ability for a neuon to fire again. If we reduce the
capacity for the IC, IM, IAHP, & ISAHP it would be sweet.
←
← Voltage-Dependant K+ Channels (VDKC)
← ← IA ← ID ← IK ← IM
← Activ ← LVA ← LVA ← HVA ← LVA
ation
← Inacti ← FI ← MI ← SI ← NI
vating
← Grap ← ← ← ←
h
← Toxin ← 4-AP ← 4-AP ← 4-AP ← ACh
(1 µM) [50 µM] [100 µM]
← DTX ← DTX ← DTX
← TEA ← TEA

[100 µM] [25 µM]

← quinin
e
← capsa
sin
← Cs+
VDKC structure (Ia, Id, Ik): 4 separate subunits with 6 trans-membrane segments each

← Rectifier K+ Channels (RKC)

← ← Tan ← Anomol ← IH
dem us Rectifier
← Activation ← voltag ← Hyperpola ← Hyperpola
e rization rization
independant ← Activated ← Activated
(HA) (HA)
← Inactivatin ← SI ← SI ← SI
g
← Activators ← Haloth ← ← Retigabin
ane e
← Isoflur
ane
← Blockers ← ← ← Cs+
Anomalous: 4 separate subunits with 2 trans-membarne segments each
Tandem Pore: 4 trans-membrane segments each, dimerize for 2 pores VIA volatile
anesthetics

←
← Metabotropic-Gated K+ Channels (MGKC)
← ← IATP ← IAHP ← IM
← Activation ← ← LVA ← LVA
← Inactivatin ← NI ← MI ← NI
g
← Neurotran ← ATP ← NE, ← Musca
smitter DA, 5HT, Glu rinic Ach
sensitivity agonist
← Ca++ ← Insen ← High ← High
Sensitivity sitive
← Activators ← Ninoxi ← ← Retiga
dil bine
← Blockers ← ← ← Ach
← Linopi
rdine
←
Ca++-Dependent K+ Channels (CDKC)
← ← ISAHP ← *IAHP ← **IC ← IM
← Activation ← ← ← ← LVA
← Inactivatin ← NI ← NI ← SI ← NI
g
← Neurotran ← No ← No ← Yes ← Mu
smitter scarinic
sensitivity Ach
agonist
← Ca++ ← yes ← 200-600 ← > 1 µM ← Hig
Sensitivity nM h
← Activators ← ← ← Niflum ← Reti
ate gabine
← Blockers ← Neurotran ← apamin, ← Charyb ← Ach
smitters via dequalinium, dotoxin ← Lin
metabotropic neurotransmitters ← TEA
pathways opirdine
← * Uses SK-Channel Proteins
• CAM remains constitutively bound to SK channels, acting as their Ca2+ sensor

• Apamin (bee venom) binding to SK channel

**BK Channel Proteins

← Glutamate receptors
← Ionotropic neurotransmission cell neighboring another cell that fires can also
have excitatory post synaptic potentials. Glutamate is the neurotransmitter that brings
about EPSPs (Glu for short). Inhibitory (IPSPs) are GABA modulated. EPSP depolarize and
IPSP hyperpolarize. They are not separate from each other but sum. The sum of the two
identical units of stimulation from both EPSP and IPSP is still an inhibition not an excitation!
You need a larger amount of excitation than inhibition (they are not linear function).
←
Excitatory amino acids
← - Glu
← - AMPA **the most abundant receptor type in the nervous system
← - KA
← - NMDA
← Glu Receptors: Ligand gated Rs (non-NMDA Receptors)
• AMPA
o R1-R4
o Sodium (Na+) channels
o Typically contain at least one GluR2, at least one other subtype, thus act
as Na+ channels (hill coefficient = 2)
• KARs
o Glur5- glur7
o KA-1 & KA-2

AMPA KA
Agonist Endogenous Glu Glu
Exogenous AMPA KA
Antagonist (competitive) NBQX CNZX
Desensitize Removal cyclothiazide concavilin A (conA)
 *GluRs subunits form tetramers

GluR Biology: unique GluR2 subunits. Amino terminals on exterior and carboxyl group in
the cytosol. There are 4 membrane spanning and a subunit Q/R site where you can either
have a Q = Gln or R = Asp. That area determines the specificity for Na+ if not then they will
also allow Ca2+ ions through also. The sensitivity will increase the amount of Na+ influx
under the right conditions.
Iontropic effects of Glu
← AMPA receptors and NMDA receptors. An EPSP (voltage) and EPSC
(current (mA)) create an action potential.
←
← Glu EPSPs are typically mediated by multiple receptor types.
←
← Glu: multicomponent EPSCs See notes
←
← Sulfur-containing EAA transmitters
← Homocysteic acid – produced an anoxious state drownding and
taurine does not cross the blood brain barrier but in diseased states then it
stimulates the inhibitory function.
←
←
← NMDA Receptors
← Ionotropic  NMDA, AMPA, Kainate (functional classes)
←
← NMDA gene families Ex: (NR1, NR2a-d, NR3a &b)
← Unique protein structure of NRs *** NR= 2 NR1 + 2 NR2 ** (dual-
heterodimer = tetramer) The subunits NR2 determine the flow and glutamate
binds to the NR2s. The NR1 have binding sites for glycene (Gly). The receptors
have a co-agonist… it requires all sites activated (2 gly and 2 glu) **unique
←
← **Typical Structure NR = 2 NR1 + 2 NR2 subunits
← NR1 are found everywhere in the brain
← NR2a – d are regionally distributed
← Ex: RN2a are found in hypocampus & Nr2c – cerebellum

←
← NMDA Receptor Sites
← Site: ← Ligand ← A ← Antag ← Mechanism
gonist onist
← NR1 ← Gly ← X ← ←
← ← Ser ← X ← ←
← ← DCS ← X ← ←
(partial)
← ← kynurenate ← ← X ←
(Kyn)
← NR2 ← Glu ← X ← ←
← ← NMDA ← X ← ←
(exo)
← ← APV (not x ← ← X ←
BBB) (exo)
← ← CPP (x BBB) ← ← X ←
(exo)
← PCP ← PCP ← ← X ← blocks open
channel
← MK-801 ← ← X ← blocks open
channel
← Angeldustin ← ← X ← blocks open
channel
← Ketamine ← ← X ← blocks open
channel
← Dextrometh ← ← X ← blocks open
orphan channel
← Memantine ← ← X ← blocks open
channel
← Mg++ ← Mg++ ← ← X ← blocks open
← channel

← Memantine ← ← X ← blocks open

channel
← EtOH ← EtOH ← ← X ← blocks open
channel
← Polyami If ← ← X ← decrease Glu
ne e binding
n
p
r
o
d
il
← Cadaverine ← ← X ← decrease Glu
binding
← Spermine ← X ← ← increase Glu
binding
← Heavy ← Zn++ ← X ← ← prolongs open
Metal channel
H H ← X ← ← prolongs open
P channel
O
-
4
← **Uses/abuses of NMDAR ligands
• Therapeutic
o Dissociative anesthesia
o Cough suppression
o Learning and /or memory enhancement
o Stroke/drowning/ischemia neuroprotection
• Non-therapeutic
o Dissociative hallucinogen
←
← **“Use-dependent” Mg++ block  When magnesium isn’t present there is a
continuous opening but with normal [Mg++] it will “flicker”
←
← Regulation of NRs by phosphorylation
• NMDAR is attached to PSD-95 connected to other proteins and mGluR is bound to
the homer, which can cause phosphorylation of the NMDA receptor and enhances
the current flow. Review Hammond…
← Excitotoxicity via NRs
← NMDA induced apoptosis via Ca++ influx… APV, MK-801, Kyn block apoptosis
← NR roles in plasticity
← Antagonists block learning, and agonists generate seizures
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← Inhibitory amino acid receptors – GlyRs, GABAARs, GABACRs
← *Activation of the receptors cause and influx of Cl- ions.

← **GlyR activated IPSCs has a hill coefficient of 2

←
← Structure Pentameric receptors (five sets of subunits make up their structure)
• GABA-A – contain alpha and beta subunits (and gama)
o GABAAR mediated Cl- currents are ligand- and voltage-dependent
• GABA-C – contains rho subunits
• Regional heterogeneity of GABAAR alpha subunits
←
← Gly Receptor Anchoring to the cytoskeleton, “Clustering”
• They are attached to gepherin to form the clusters and attracted to microtubules
and neurofilaments and other gephrin. If there is a knockout of the gepherin the
GlyRs are not clustered and inhibition is affected.
←
←
← Gly Receptor ligands (strychnine sensitive)
← Agonists (competitive)
• Gly
• Taurine
← Partial agonists (competitive)
• B-aminobutyrate (BABA)
← Antagonists (competitive)
• Strychnine (muscle spasms and breathing issues)
← Antagonists (noncompetitive)
• Picrotoxin
• Picrotoxinin
← **Nipecotate suppresses but prolongs IPSPs
←
Uses/abuses of GABAAR ligands
• Therapeutic
o Sedatives
o Hypnotics (sleeping pills)
o Anxiolytics
o Seizure prevention, reduction (AEDs) Anti-Epileptic Drugs
• Recreational
o Date rape
o Sedative-hypnotics
o Anxiolytics
← Related issues:
• Development of tolerance
• Development of dependence/addiction

GABAA Receptor Ligands

 Agoni Antagonist *Alloster
st ic
Modulato
r
Lig Competitive Noncom
an petitive
d GABA bicuculli picrotoxin Barbiturate
ne s**
muscim GABAzin picrotoxini BDZs
ol e n
gavoza -cyphat neurosteroi
dol ds
isoguva flucybene EtOH
cine
propybicy
phat
← *Allosteric modulators effect GABAAR IPSPs
← **Benzodiazepines (BDZs) enhance the affinity of GABARs for GABA thus enhance Cl-

conductance of channel. More is likely to bind and stay bound.

← Barbiturates Ligands - enhance Cl- conductance of channel

• Agonists (competitive)
o Phenovarbital (luminal)
o Pentobarbital (nembutal)
←
Benzodiazepines (BDZs) Ligands
• Agonist (competitive) – both classes of omega receptors
o Chlordiazepoxide
o Diazepam (valium)
o Carbamazephine
• Partial Agonist
o Bretazenil
• Antagonists (competitive)
o Flumazenil
• Agonists (inverse)
o Carboline
←
← Ω1 Receptors are associated with hypnotic affects
• Zolpidem (ambien) – non-benzodiazepine hypnotic (imidazopyridine)
• Eszopiclone (lunesta) – non-benzodiazepine hypnotic (cyclopyrrone)
← Ω 2 Receptors are associated with anxylitic
←
← ** Diazepam  liver conversion to nordazepam  oxazepam and two products after
breakdown are still BDZs
←
← Neruosteroid Ligands
• Agonist (competitive)
o Pregnenolone – decrease anxiety and increase GABA affinity
• Antagonists (competitive)
o Allopregnanolone – increases anxiety – reduce GABA affinity
←
← GABACR are unique (and rare) –
 • Found in the retina horizontal cells. They have long durations and high current.
• Pentameric but unique subunit expression : Rho
• Mostly insensitive to most GABAa and GABAb antagonists
• Insensitive to modulators (no sensitivity to
BDX/Barbiturates/neurosteriods/ethanol)
• Very slow channel gating (activation inactivation occurs in secs not ms
• Little or no desensitization
• Cl- channel blocked by picrotoxin/picrotoxinin
←
← GABAC Receptor Ligands
• Agonists (competitive)
o GABA
• Partial Agonists
o CACA (cis-4-aminocrotonate)
• Antagonists (competitive)
o TPMPA
• Antagonists (non-competitive)
o Picrotoxinpicrotoxinin