Anatomy

Critical to function

10m 10m

NMJ on the muscle fiber

Synaptic selectivity at developing NMJ

Synapse from a frog sartorius neuromuscular junction showing vesicles clustered in the active zone, some docked at the membrane (arrows). (from Heuser, 1977)

Synaptic Transmission

The Steps

• Precursor transport• NT synthesis• Storage• Release• Activation• Termination ~diffusion, degradation,

uptake, autoreceptors

Synaptic Transmission Model

PresynapticAxon Terminal

PostsynapticMembrane

Terminal Button

(1) Precursor Transport

_ _ _

NT

(2) Synthesis

enzymes/cofactors

(3) Storage

in vesicles

A quantum is the number of transmitters released from a single synaptic vesicle

Vesicles have a fairly uniform size and diameter ≈ 40- 50 nm

Individual vesicles contain 8000 - 10,000 phospholipid molecules and several proteins. The vesicle molecular weight is approx. 3-5 x 106

Proteins associated with synaptic vesicles(identified through sequencing and cloning of cDNA’s)

Membrane proteinsA. Synaptophysin (~ 36 kD)B. Synaptotagmin (~ 61 kD; the Ca2+ sensor)C. Snares (residents of either the vesicle [v-snare] or the target membrane [t-snare])

1. VAMP (also called synaptobrevin), a v-snare (~18 kD)2. Syntaxin, a t-snare that also associates with Ca2+

channels (~32 kD; technically not a vesicle protein)3. SNAP-25, a t-snare (~25 kD; also technically not a

vesicle protein)D. Electrogenic proton ATPase -creates emf that drives

neurotransmitter uptake against a concentration gradient

Proteins associated with synaptic vesicles(identified through sequencing and cloning of cDNA’s)

Membrane proteinsA. Synaptophysin (~ 36 kD)B. Synaptotagmin (~ 61 kD; the Ca2+ sensor)C. Snares (residents of either the vesicle [v-snare] or the target membrane [t-snare])

1. VAMP (also called synaptobrevin), a v-snare (~18 kD)2. Syntaxin, a t-snare that also associates with Ca2+

channels (~32 kD; technically not a vesicle protein)3. SNAP-25, a t-snare (~25 kD; also technically not a

vesicle protein)D. Electrogenic proton ATPase -creates emf that drives

neurotransmitter uptake against a concentration gradient

An alternative form of Ca2+-dependent vesicle fusion, termed fast tracking, or “kiss and run” predominates at low frequency stimulation.

Life cycle of a synaptic vesicle

Synapse

Terminal Button

Dendritic Spine

Synapse

Terminal Button

Dendritic Spine

(4) Release

Receptors

Synapse

Terminal Button

Dendritic Spine

AP

Ca2+

Exocytosis

From Kristin Harris Lectures.http://synapses.mcg.edu/lab/harris/lectures.htm

1X

4X

2X

Stimulation

1 mV

1X

2X

3X

4X

mini Mini histogram.

Evoked amplitudes.

Squire Fund. Neurosci.

From Kristin Harris Lectures.http://synapses.mcg.edu/lab/harris/lectures.htm

Electron micrographs of “omega figures” (fusing synaptic vesicles) after slam freezing a firing synapse provided clinching evidence for the vesicle hypothesis.

No firing

Firing

Heuser and Reese, 1981

“docked”

“fast”“slow”

A cholinergic synapseNerve fiber (axon)

Action potential

Choline

Na+, Cl-

Acetyl-CoA

Acetyl-Choline

Acetyl-Choline

Ca + +

Ca + +

A cholinergic synapse (2): Rapid transmitter inactivation by cholinesterase

Acetyl-Choline

Cholineesterase

Acetate

Acetyl-CoA

Choline

Action potential

Ca + +

(5) Activation

(1) Ionotropic ChannelsneurotransmitterNTChannel

Ionotropic Channels

NT

Pore

Ionotropic Channels

NT

Ionotropic Channels

NT

Acetylcholine Receptor

(or )

Miyazawa, A., Y. Fujiyoshi, and N. Unwin. 2003. Structure and gating mechanism of the acetylcholine receptor pore. Nature 423:949-955.

ACh

ACh

45

End Plate Potential (EPP)

Outside

Inside

Muscle membrane

Presynapticterminal M

uscl

e M

embr

ane

Volta

ge (m

V)Time (msec)

-90 mV

VK

VNa

0

Threshold

Presynaptic AP

EPP

The movement of Na+ and K+

depolarizes muscle membranepotential (EPP)

ACh Receptor Channels Voltage-gatedNa Channels Inward Rectifier

K Channels

• Normally, the average EPP amplitude = 60 mV -In frog, ~150 vesicles

• Safety factor for transmission is therefore high (greater than 1) - Frog example: VEPP VAPthreshold

= 60 mV │-90 mV*- [-50 mV] │

= 60 mV 40 mV = 1.5

(*muscle resting VM = -90 mV)

Normal EPPs invariably evoke muscle action potentials

(6) Termination

(6.1) Termination by... Diffusion

(6.2) Termination by...Enzymatic degradation

Acetylcholine Metabolism

AChacetylcholine

esterase (AChE)choline + acetate

• AChE is located in the synaptic cleft• Choline is taken back up into the presynaptic terminal – active process• Acetate diffuses away to be utilized in other metabolic roles

(6.3) Termination by... Reuptake

(6.4) Termination by... Autoreceptors

A

The Safety Factor !!!

• Number of Quanta

• The receptor density on the post synaptic membrane

• The activity of ACH esterase

• The folds of the PS membrabe

• The presence of active zones

Voltage-gated channels

Na+ channelopathies

Gene Channel Disease

Muscle SCN4A subunit of NaV1.4 Hyperkalaemic periodic paralysisHypokalaemic periodic paralysisParamyotonia congenitaPotassium-aggravated myotoniaMyotonia fluctuansMyotonia permanensetc

Neuronal SCN1A subunit of NaV1.1

(somatic)

Generalised Epilepsy with Febrile Seizures + (GEFS+), Severe myoclonic epilepsy of infancy (SMEI)

SCN2A subunit of NaV1.2

(axonal)

GEFS+

SCN1B 1 subunit

Ca2+ channel structure

2

1

Ca2+ channelopathies

Gene Channel Disease

Muscle CACNA1S subunit of CaV1.1 HypoK periodic paralysisMalignant hyperthermia

RYR1 Ryanodine receptor (sarcoplasmic channel)

Malignant hyperthermiaCentral core disease

Neuronal CACNA1A subunit of CaV2.1

(P/Q-type channel)

Familial hemiplegic migraineEpisodic ataxia type 2Spinocerebellar ataxia type 6Absence epilepsy?

CACNA1H subunit of CaV3.2

(T-type channel)

Absence epilepsy

Nicotinic receptor channelopathies

Gene Channel Disease

Muscle CHRNA1 1 subunit Congenital myasthenic syndrome

CHRNB1 1 subunit

CHRND subunit

CHRNE subunit

Neuronal CHRNA2 4 subunit AD nocturnal frontal lobe epilepsy

CHRNB4 2 subunit

Slow channel syndrome

Sine et al (1995)

Fast channel syndrome can be associated with congenital joint deformities (arthrogryposis multiplex)

Brownlow et al (2001)