Transmitter receptors

And signal transduction

Nicotinic channels

•  Two molecules of acetylcholine bind. •  agonist from Nicotinia tabacum •  found in vertebrate •  all (sympathetic and parasympathetic)

autonomic ganglia (the first synapse, not the neuro-effector junction),

•  muscle and other places

cloning

•  Torpedo - electric ray, up to 75 V (not that much) but 20 Amps.

•  Lots of generator potentials added up •  (vs. Electrophorus - lots of spikes, used

to isolate the Na+ channel of the action potential).

•  (There are also fish with electric sense, not just those that stun prey.)

more

•  bound by alpha-bungarotoxin - from banded krait Bungarus multicinctus (snake), 74 amino acids binds receptor irreversibly

•  causes paralysis by blocking transmission, •  label receptor 125I alpha-bungarotoxin. •  5 subunits - 2 alpha, beta, gamma and delta •  in neurons, 3 alpha, 2 beta (and no alpha

bungarotoxin sensitivity)

more •  spans the membrane 4 times. •  M2 likely lines the pore. •  Nicotine is an agonist; but like an antagonist •  blocks transmission at autonomic ganglia by

depolarization blockade. •  antagonists (curare, a plant alkaloid from

Clondodendron tomentosum). •  muscular relaxatants used (like

succinylcholine). •  must prove that anesthesia is adequate.

Recent paper •  BAOrser, Lifting the fog around anesthesia, •  analgesia (absence of pain) is paramount •  Unconsciousness (hypnosis) •  Immobility •  Interestingly, amnesia (loss of memory) •  Dentist William Morton, ether in Boston, 1846 •  These days, drugs that keep GABA-A

channels open (benzodiazepines like Valium also affect these)

Additional points

•  Developmentally, diffuse receptors cluster. •  Acetylcholinesterase, is all over the place. •  molecules are very concentrated at n.m.j.

crest, 20,000 - 30,000 per square micron •  (about as tightly packed as possible in

contrast with punctate voltage gated sodium channels).

•  Probably water filled pore.

Finally •  All 4 subunits needed in expression •  10 to the 6th ACh molecules from one a.p.

into n.m.j. cleft. •  2.5 x 10 to the 5th channels transiently open. •  400 nA n.m.j. end plate current. •  1 ms open time. •  10,000 Na+'s flow through each channel in

this time. •  Channel conductances of 25 pS

Myasthenis gravis

•  Smaller miniature end plate potential.Great weakness Receptors low

•  autoimmunity to nicotinic receptors. •  (NS proteins generally separated from

immune surveillance by blood brain barrier.) •  Thomas Willis (1685, of circle of Willis fame.) •  that action potentials are increased by

neostigmine treatment. •  Involvement of thymus.

Comments

•  Glutamate (AMPA, NMDA, Kainate), GABA, glycine, setotonin, purine

•  Keeping in mind that we already discussed ACh (nicotinic) receptors (above), the notable ligands missing from the channel receptor list are epinephrine and dopamine.

•  There is a staggering diversity of different types.

NMDA

•  NMDA = N-methyl D-aspartate. •  blocked by AP5 (2-amino-5-

phosphonovalerate) •  central excitatory - like inputs to

hippocampus •  On the basis of the reversal potential, it

is inferred that the channel is nonselective cation channel.

NMDA

•  Na+ & Ca2+ Calcium influx - •  excitotoxicity in injury or stroke. •  Voltage, glutamate, calcium cause

"vicious cycle of glutamate release.” •  Ca2+ "second messenger" •  important neural processes, such as

"learning," are attributed to NMDA receptors

AMPA and Kainate

•  = alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate

•  Kainate (from red alga Digenea simplex) and Quisqualate (from seed of Quisqualis indica) are excitotoxic amino acids

•  reversal potential is at 0 mV so it is likely opening for K+ & Na+ channels

GABA

•  GABA-A channel is for Cl-. •  Different combinations of 5 different subunits •  makes for a lot of diversity. •  Diazepam (Valium) and chlordiazepoxide

(Librium) [tranquilizers] bind to alpha and delta subunits - enhance GABAergic transmission.

•  Barbiturates [hypnotics] like phenobarbital bind to gamma subunit.

GABA, glycine, 5HT

•  GABA-A receptors blocked by bicuculline from Dutchman's breeches and picrotoxin from Anamerta cocculus.

•  Glycine receptor blocked by strychnine, alkaloid from seeds of Strychnos nux-vomica - causes seizures.

•  5HT3 is also an ion channel - maybe the molecule only spans the membrane 3 times.

Metabotropic

•  1971 Nobel Earl W. Sutherland, Jr. (US) •  "mechanisms of actions of hormones," •  father of signal transduction. •  cAMP as a second messenger •  adrenergic effects on metabolism in

liver (which mobilizes glucose from glycogen).

Autonomic: Ach, NE

•  nicotinic (ionotropic) ganglia, •  muscarinic (metabotropic") neuro-effector •  Adrenergic only in neuro-effector. •  alpha excitatory, arteriole, agonist

Neosynephrine (phenylephrine). •  Beta inhibitory (excitatory heart) •  beta blocker propranolol for hypertension. •  several alphas & betas. •  adrenergic is metabotropic, not ionotropic

Lefkowitz

•  Cloning of the gene and cDNA for mammalian beta-adrenergic receptor and homology with rhodopsin, Nature 321, 75, 1986

G protein coupled receptors

By hydrophobicity, they all cross membrane in 7 alpha-helices.

•  Rhodopsin was the prototype, •  followed closely by the beta adrenergic

receptor. •  Then many neurotransmitter and

hormone receptors were found.

Super(duper)family

•  1990's, olfactory Axel & Buch 2004 Nobel •  In summary, there is an enormous diversity! •  (>1000 in mammals). •  N terminus outside cell, glycosylation •  C-inside (heptahelical) -phosphorylation, •  2nd and 3rd loops and C terminus for

interaction with a subunit of G protein

Many for transmitters

•  mGluR1-8, •  NE alpha 6 types, beta 1 , 2, & 3, •  D1-D5, •  GABA-B(1&2), •  5-HT-12 types •  Purines 12 types •  And muscarinic (1-5)

Muscarinic

•  (postganglionic parasympathetic) - •  muscarine - from poisonous red mushroom

(Amanita muscaria) stimulates, •  atropine (from deadly nightshade) blocks

(belladonna = beautiful lady). •  Acetylcholine from Vagus (X) slows heart, •  acetylcholinesterase inhibitor such as

insecticide malation or nerve gas) would stop •  atropine, block receptor, would save your life.

notes

•  “G” -alpha subunit binds GTP •  AGGilman and M Rodbell, Nobel 1994 •  heterotrimer alpha, beta and gamma •  alpha and beta are about the same size,

gamma is smaller •  alpha and gamma linked to membrane by

fatty acid •  alpha subunit affects effector and is GTPase

notes

•  cascades with different second messenger (signalling) systems (effectors)

•  beta adrenergic, Gs (stimulatory), adenylyl cyclase, cAMP, protein kinase (PKA) phosphorylation

•  For glutamate, Gq (q is designation here) activates Phospholipase C (PLC) or PLA2.

•  For dopamine, a Gi (inhibitory) inhibits the cAMP cascade

Phosphoinositide cascade

•  phosphatidylinositol-4,5-bisphosphate PIP2 •  DAG (diacyl glycerol) IP3 (inositol

trisphosphate •  IP3 causes release of Ca2+ from

nonmitochondrial stores, IP3 receptor. •  Ca2+ intracellular messenger. •  binds calcium binding proteins like calmodulin •  DAG activates PKC (protein kinase C) •  [kinase is an enzyme that phosphorylates].

Lipids are interesting

•  In 1970, lipids seemed boring, hold proteins. •  1980's, turn over, and to be signaling •  NorpA (no receptor potential) Drosophila •  have rhodopsin but lack phospholipase C. •  I did not isolate the mutant or make this

discovery, but I did work on norpA. on lipids and fatty acids in Drosophila with reference to phototransduction.

also

•  ATP -> adenylyl cyclase -> cAMP -> phosphodiesterase -> 5'AMP.

•  Caffeine and theophylline inhibit PDE (phosphodiesterase for cAMP),

•  thus potentiating the "upper" action of norepinephrine by making its second messenger longer lasting.

Recent paper

•  LTD=Long term depression, •  model for learning. •  Conditioned eyelid response •  paradigm for learning. •  Transgenic with Purkinje-specific promotor •  used to express a molecule to inhibit PKC. •  These mice lack LTD for this learning.