Receptors and transduction 1Receptors and transduction 1

References:Chapter 11 – Neuron by Levitan & Kaczmarek ORChapter 6 – Neuroscience by Purves et al

1. K. Tsuzuki and T. Ozawa. Glutamate receptors www.els.net2. Madden, D. Structure and function of glutamate receptor ion

channels (2002). Nature Revs Neurosci. 3, 91.

Dr. MV HejmadiDr. MV Hejmadi

SynapseSynapse: the point where electrical conduction is converted to chemical conduction (mostly)

The Chemical The Chemical SynapseSynapse1) AP invades

2) Ca++ influx 3) Depolarization

release of NT 4) NT diffusion

5) Ligand binding6) Depolarization7) transmitter recycling8) vesicular membrane recycling

Action potential

Calcium channels

Calcium ions

L-glutamate vesicles

L-glutamate release

AMPAR

Na+

NMDAR

Mg2+

Synaptic cleft

Presynaptic Terminal

Postsynaptic spine

Excitatory synaptic transmission by L-glutamate

GlutamateAcetylcholine etc

GABA

Specificity of responsesSpecificity of responses

Following activation by a NT, how does a neuron know what the response should be?

Depending on the type of NT (excitatory or inhibitory), key receptors transduce this signal and dictate the nature of the response

GluGlu

DADA

AChACh

Substance P

GABA

Output

• i.e. glutamatergic, GABAergic, i.e. glutamatergic, GABAergic, cholinergic etccholinergic etc

Each neuron uses one NT to send a signal Each neuron uses one NT to send a signal (usually)(usually)

Synaptic inputs to a GABA-ergic Synaptic inputs to a GABA-ergic medium spiny neuronemedium spiny neurone

Each NT can activate diverse Each NT can activate diverse receptorsreceptors

e.g. acetylcholine can activate 2 classes of receptors, nicotinic or muscarinic

Nicotinic AChRNicotinic AChR• Mimicked by nicotine

(agonist)

• Only in certain tissues (skeletal muscle, parts of the CNS and PNS)

Muscarinic AChRMuscarinic AChR• Mimicked by

muscarine (agonist)

• Cardiac muscle, cholinergic synapses of the CNS

How do you measure specificity? How do you measure specificity? Pharmacological toolsPharmacological tools

Nicotinic AChRNicotinic AChR

blocked by antagonists

-bungarotoxin

curare

Muscarinic AChRMuscarinic AChR

Blocked by antagonists like

atropine

Bungarus multicinctus

Henbane plant

Receptors categorised based on their Receptors categorised based on their transduction mechanismstransduction mechanisms

A)A) Ionotropic receptorsIonotropic receptorsIonotropic receptors are directly coupled to ion channels and are activated by neurotransmitters which open the channel pore to allow movement of specific ions, creating postsynaptic potentials

B) Metabotropic receptorsB) Metabotropic receptorsThese receptors are not directly coupled to their ion channels and transduce the signal via guanyl nucleotide-binding proteins (G-proteins) that activate intracellular second messenger pathways

FAST DIRECT

SLOW INDIRECT PSPs

Fig 5.22; Purves et al

FAST DIRECT PSPs

SLOW INDIRECT PSPs

Ionotropic receptorsIonotropic receptors

Fig 11.8; Levitan and Kaczmarek

Ionotropic receptor typesIonotropic receptor types

pentameric

glutamateglutamate

tetrameric

Assembly of either similar subunits (homomeric) or different subunits (heteromeric)

Heterogeneity of nAChRin mammalian tissues

BrainBrainaddictionanalgesia

anxietyattentioncognition

AutonomicAutonomicGangliaGanglia

cardiovasculargastro-intestinal

MuscleMusclecontraction

Ionotropic receptors - Generic structureExample:

Nicotinic acetylcholine receptor (pentameric)

SubunitsEach subunit has four hydrophobic regions

(TMI – IV)

glutamateglutamate

Ionotropic receptors-Generic Ionotropic receptors-Generic structurestructure

Unlike other iR, iGluR have a re-entrant loop inTMII domain

extracellular N-terminus and intracellular C-terminus

Long loop between TMIII and IV forms part of the binding domain with the C-terminal half of the N-terminus

The modular nature of iGluR subunits

Extracellular NTD followed by S1 - sequence between NTD and TMIS2 - sequence between M3 and M4TMIII and TMIV domains with an intervening re-entrant P loop,. C terminus intracellular

S1 and S2 half-domains form the ligand-binding domain.

The structure of this domain is shown as a ribbon diagram. It consists of two lobes (lobe I, blue; lobe II, red), separated by a ligand-binding cleft.

Lobe 1- structural unit attached to the amino-terminal domainLobe 2 - structural unit that links to M1 and M3

Madden, D. Structure and function of glutamate receptor ion channels (2002). Nature Revs Neurosci. 3, 91.

Modular structure of iGluRsModular structure of iGluRs

Emerging structural explanations of ionotropic glutamate receptor

function ROBERT L. McFEETERS and

ROBERT E. OSWALDThe FASEB Journal. 2004;18:428-438

transmembrane topology, the two extracellular domains, and the carboxyl-terminal domain. Flip/flop refers to an alternatively spliced segment that plays a role in desensitization. S1 refers to the sequence between the amino-terminal domain and M1; S2 refers to the sequence between M3 and M4. Lobe 1 is the structural unit attached to the amino-terminal domain, and Lobe 2 is the structural unit that links to M1 and M3. Note that segments of S1 and S2 are found in both Lobes 1 and 2.

Monomers associate most strongly through interactions between their amino-terminal domains (NTDs) (star in middle figure). Dimers undergo a secondary dimerization, mediated by interactions in the S2 and/or transmembrane domains (stars in right-hand figure). The crystallographically observed S1S2 dimer probably corresponds to this secondary dimerization interaction.

The 'dimer-of-dimers' model of iGluR assembly

NMDA receptor(N-methyl- D-apartate)

AMPA receptor(-amino 3hydroxy 4methyl 5propionic acid)

Kainate receptors(kainic acid)

EpileptogenesisSynaptic plasticity?

Flip/flop region-spliced variants (exon 14 –flop and Exon 15 for flip)

Modulates LTPResults in calcium entry, changes in secondary messages altered protein synthesis

Calcium as a signal

The NMDA receptor is blocked by a Mg++ ion at resting potentialThe NMDA receptor is blocked by a Mg++ ion at resting potential

Glutamate directly gates NMDA and AMPAR. NMDAR controls movement of Ca 2+ Na+ and K+

binding sites for glycine, zinc, phencyclidine (angel dust) and Mg2+.

Under resting conditions Mg2+ blocks the site and keeps the receptors silent.

Upon depolarisation the Mg is released and there is movement of Ca into the cell.

NMDA receptorsNMDA receptorsGLYCINE GLUTAMATE

MgMK801

NR1-1a/b-4a/b

NR2ANR2BNR2CNR2D

NR3A

Ca Na

calmodulin

PSD proteins

CamKIIP P

NR1 NR2

2+ +

2+

OUT

IN

P = multiple sites for phosphorylation

Zn2+ POLYAMINE

NR1

NR1

NR2NR2

?

So what are the most important NT in the mammalian brain?

• glutamate and GABA are the most abundant which mediate synaptic transmission in the CNS via ionotropic receptors (LGICs)

• In general,

• GABA - inhibitory whereas

• glutamate - excitatory

Roles of glutamate in the CNSRoles of glutamate in the CNS

• Synaptic transmission (EPSPs)• Long term depression• Long term potentiation

– Dendritic sproutin– Synaptic modification– Control of gene expression

Excess of glutamate release can cause neuronal Excess of glutamate release can cause neuronal death (excitotoxicity)death (excitotoxicity)

e.g. during stroke, epilepsy, Parkinson’s diseasee.g. during stroke, epilepsy, Parkinson’s disease

Effects of glutamate-induced excitotoxicity

Nature Reviews Neuroscience 4; 399-414 (2003

GABA receptorsGABA - major inhibitory NT in the mammalian CNS GABA receptors are pentameric in structure3 classes GABAA and GABAC receptors are ionotropic, GABAB receptors are metabotropic

Cl-

GABA

Cl-Cl-

GABA

GABAA receptor binding sites

Benzodiazepine – allosteric agonist - tranquilisers / anticonvulsantsBarbiturates – prolong action of GABA - anaesthetics / hypnotics

So why have multiple iRSo why have multiple iR??• The post-synaptic response to stimulation can be

modulated in the short term (i.e. for hundreds of milliseconds) or for the long term (hours, days or even weeks!!).

• Synaptic strength is increased or decreased by altering the level of post-synaptic depolarisation. This is achieved through changing how well receptors respond to stimulation,– by altering the length of time they are active, – the number of receptors physically present or – by altering the amount of L-glutamate that is released

into the synaptic cleft