I. General Characteristics

A. Control of ion permeability is but one way that transmitters influence function. There are a diversity of ways in which nervous systems control their own properties.

B. Receptors associated with ion channels thought to mediate fast depolarization or hyperpolarization via control over ion channel permeability.

C. Types

1. Nicotinic cholinergic: cation channel; located at neuromuscular junction and at cholinergic autonomic preganglionic synapses.
2. GABAA/Glycine receptors: anion channel; located in brain and spinal cord
3. Glutamate receptors: cation channels; located in CNS
4. 5-HT3 receptor: cation channel

D. Ligand-gated ion channels represent a gene-family possibly with a common ancestor; all are oligomeric receptors, single functional receptor composed of several polypeptides.

E. Basic structure of each peptide subunit: long extracellular N-terminus ligand binding site, 4 membrane spanning domains (one contributing to formation of the ion channel) short C- terminus extracellular end.

F. Receptors are widely distributed phylogenetically; great variety of subtypes.

II. Nicotinic Cholinergic Receptor

A. Mediates increased permeability to Na+ or Na+/K+ ions leading to depolarization in receptive cell.

B. Structure in skeletal muscle

1. Multiple peptide subunits: 4 different polypeptide chains arranged in a transmembrane pentamer__a(2), b,g,d.

2. Reconstruction of purified receptor into lipid bilayers has shown that this pentamer cation channel has all the properties required for its regulation.

3. alpha sites appear to be binding sites for agonists and competitive antagonists. Two sites must be occupied for activation.

4. Can influence receptor response at several sites:

a. Noncompetitive antagonists (NCB) interfere with ion flow without affecting ACh binding. Binding of NCBs increases when channel is open suggesting that these drugs seem to bind to interior of channel and plug it. NCBs appear to bind to sites on all subunits.

b. Multiple subunit structure allows for allosteric regulation of receptor response via conformational changes.

C. Structure of CNS receptor

1. Pentameric: a (2), 'non-alpha' (3); to date 7 a and 3 b cDNAs have been cloned and sequenced.

2. CNS receptor has different pharmacologic and functional properties from skeletal muscle receptor---probably arising from different combinations of subunits.

A. Mediates fast inhibition in the brain; gates a chloride channel, Effect on Em depends on relationship of VCl- to Em.

B. Structure

1. Pentamer

2. At least 15 possible subunits have been cloned-6a, 4b, 3g, 1d, 1e Expression of each subunit has defined distribution pattern The gamma subunit exist in 2 forms due to alternate splicing of mRNA

3. Each subunit has 4 membrane spanning domains.

4. Different composition of subunits could yield diverse receptors (theoretically, 15,000 possible receptors).

5. In actuality, the a1b2g2 subtype is most abundant: 43-50% of all GABAA receptors Next most abundant seem to be the a2b2/3g2 and a3bng2/3 combinations, ~18% each.

6. Sites on receptor for many compounds that modulate GABA function: Benzodiazepines, barbiturates, steroids, ethanol

C. Pharmacologic evidence for different types of GABAA receptors

1. Pharmacology of drug CL 218-872

a. Displacement of BZ binding demonstrated homogenity in cerebellum; heterogeneity elsewhere.

b. Receptor with high affinity for 872 seems to be a1b2g2 type

a. GABA-mediated Cl- uptake

b. Cl- facilitated BZD binding

c. Different subunit distribution: a1 in both structures, but a2 and a3 sparse in hypo

IV. Glutamate Receptors

A. Mediate fast excitatory transmission on CNS; at least 3 classes

1. AMPA receptor (A1): Found in majority of fast excitatory synapses; gates a Na+ channel, low Ca++ permeability

2. NMDA receptor: gates a voltage dependent, Ca++ channel; slower kinetics than AMPA receptor. To remove Mg++ block, membrane must be depolarized before ion channel opens; glycine is a co-agonist (but glycine site is not strychnine sensitive).

3. Kianate receptor (A2): gate fast desensitizing currents. Similar to A1 receptor.

B. Structure

1. AMPA receptors

a. Receptor function can be reconstituted by expressing one or co-expressing any 2 of 4 subunits: Glu RA-Glu RD (or Glu R1-GluR4).

b. The receptor can exist in two forms due to alternate splicing of mRNAs.

2. NMDA receptors

a. Two subunit types: NR1 in combination with any one of NR2a-d

b. NR1 exists in different splice forms

c. Expression of NR1 alone yields a receptor with glycine requirement and Mg++ sensitivity, but whole cell currents are smaller than when in combination with other subunit.

C. Glutamate induced neurotoxicity

1. NMDA receptor appears critical 2. Appears due to excessive Ca++ buildup in cell, can draw water into cell and lead to cell lysis. Ca++ also activates enzymes that may cause damage to mitochondria, lipases, proteases 3. Ca++ may elicit excess glutamate release, possibly via Ca++ activation of nitric oxide

D. Interaction between excitatory and inhibitory receptors controls overall level of neural excitation.

V. Desensitization

A. Ligand-gated ion channels open in response to binding of transmitter but can also close for varying periods with agonist still bound. Does this desenstization contribute to the functioning of these fast synapses?

B. Markov model shows that a bound receptor can either be open or desensitized. In this model, a bound receptor is 10x more likely to open or unbind than to desensitize. Whether desensitization would occur depends on rate of rise of agonist concentration;

C. The nicotinic ACh receptor: slow desensitization and fast recovery

1. Application of ACh to muscle does result in a depression of subsequent ACh responses lasting from seconds to min. But at the NMJ, the miniature endplate current (mEPC) only lasts a few ms because ACh unbinds rapidly from the receptor and free ACh is quickly cleared from the synaptic cleft by ACh-esterase.

2. Modest desens. of the EPC probably only occurs when clearance of ACh from the cleft is slowed by esterase inhibitors-and only at high freq. of stimulation.

3. Nicotinic receptors then are similar to theoretical model in that channels are more likely to unbind than to desensitize after a short pulse of agonist. Nicotine tolerance in smokers might involve receptor desens. caused by low levels of circulating nicotine.

D. AMPA receptors: Fast desensitization and slow recovery

1. Profound desens. of AMPA receptors occurs on the time scale of glutamate-mediated mEPSC. Receptor does not fit Markov model; when glutamate binds, receptor is as l likely to desensitize as open.

2. Current decay after a brief saturating pulse of glutamate mimics shape of mEPSC and desens during a long pulse is only a little slower.

3. Desensitization of AMPA receptor ensures brief mEPSC even if the transmitter timecourse in the synaptic cleft is prolonged. Desens. limits frequency at which AMPA receptors can produce full amplitude responses to glutamate.

4. Channel requires 10s of ms to recover to unbound (activatable) state--very slow

5. Modulation of desensitizing mechanisms may influence receptor function

E. NMDA receptor: insight into regulation of desenstiization

1. Duration of EPSC mediated by glutamate is much longer than the lifetime of the transmitter in the cleft.

2. Mechanisms via which NMDA receptors desensitize include decrease in affinity for glycine co-agonist and Ca++ mediated events.

3. Desens. seems to relate to activation of receptors; a test EPSC is smaller after a conditioning train of stimuli when receptors are activated during the train but not in presence of NMDA antagonist AP-5. If Ca++ related kinases are blocked, then receptor activation has no effect on size of test EPSC. Thus, Ca++ entering cell through NMDA receptor triggers enzymatic events that inhibit subsequent receptor activity.

4. Desent. of NMDA receptors might be a mechanism to regulate postsynaptic Ca++ levels.

F. GABAA receptors: Fast desensitization and fast recovery

1. Desens. usually viewed as a mechanism to prevent undesirable consequences of receptor excitation.

2. Desen. of receptors may prolong rather than curtail synaptic events.

3. After a single pulse, Cl- channels oscillate between open and long desensitized states before unbinding. Rates into and out of each state relatively fast. Visits to the exposure to GABA.

4. Model describes impact of different types of desentization on cell response