The presynaptic region

Chemical synapse (general overview)

varicosity / en passant synapse terminal / bouton

Presynaptic structures

• similar presynaptic structures

• transport process: stabilisation of local structures and ongoing axonal transport happen simultaneously

Presynaptic structures

asymmetric / excitatory /

Gray I

symmetric / inhibitory/ Gray II

Timescale of synaptic transmission • AP and Ca2+ dependent synaptic transmission

• spontaneous „mini” release (mPSP): neuronal background activity, spontaneous Ca2+ waves

<100 msec!

- 2-3 h (exc) – 3 min (inh) intervals, general feature

• release mechanisms similar to Ca2+-dependent exocytosis (hormone, neuropeptide release, mast cell degranulation, acrosome reaction, etc)

Quantal release

vesicule fusion

fused vesicles

transmembrane proteins (voltage gated Ca2+ channels)

• 1 synaptic vesicule = 1 quantum

- ~ 1500-5000 NT/vesicule (fast chemical synapse)

- < 2 ms diffusion: ~1mM NT conc. at the postsynaptic site

• 1000-2000 AChR opening

• 25 pS conductance, 1,5 ms opening time -> ~35000 Na+ influx -> 20-30 mV EPSP -> AP

- 0,5 ms synaptic delay -> EPSP/IPSP

- GABA, Glu-erg synapses:

- NMJ:

• 5-10 SV/AP

• ~30 receptor activated /SV release

• 300 SV/AP, ~ 1000 AZ (active zone)

• <1 mV PSP

Ca2+ microdomains

• elevation of local Ca2+-concentration depends on the distance from the voltage-gated Ca2+ [CaV] channels: within 50 nm and 100 ms >100 mM [Ca2+] rise

• structural organization of the active zone and the „synaptic unit” is essential in functionality

• locally very fast increase or drop in [Ca2+]: cooperative Ca2+ binding is probable

• distance between the release sites and CaV channels is regulated upon activity (see scaffold proteins, CAZ)

Synaptic vesicule turnover [classic (small molecule) NTs]

(1) Na+-dependent uptake of transmitters / transmitter precursors

(9) Ca2+ microdomains within the AZ

(2) NT synthesis

(3) axonal transport of SV (along MTs)

(4) transvesicular H+ gradient (vesicular ATPase)

(5) concentrating NTs within the SV (H+/NT antiporter)

(6) synapsin I-dependent SV transport to the active zone (actin)

(8) presynaptic depolarisation

(7) SV docking in the active zone (AZ) along Ca2+ channels

(10) exocitosis, quantal release

(11) non-quantal NT-leakage – fusion

between the SV membrane and the

plasma membrane

(12) endocytosis of the vesicule-membrane

(dynamin, clathrin coat)

(13) fusion with the endosomal cisternae

(14) SV formation

+ postsynaptic receptors

+ NT-degrading enzymes within the

synaptic cleft

Synaptic vesicule turnover

Synaptic vesicule turnover

Structure of a synaptic vesicle (SV)

maintenance

of proton

gradient

proton – NT

antiporter; secondary

active transport

transport along

F-actin towards

the AZ

Ca2+ sensor

vesicular component

of the SNARE

complex

priming,

endocytosis,

local

interactions

with

effectors

transporter homologue; sets

the Ca2+ sensitivity of the

primed SV

synaptobrevin

recycling

exocytosis; molecular

chaperon?

Structure of a synaptic vesicle (SV)

• transporters, proton pumps: a low copy number (1-10/vesicule) is already sufficient – re-filling within max 20 sec

• > 80 transmembrane protein, very high diversity

Structure of a synaptic vesicle (SV)

Synaptic vesicle pool

• RRP (readily-releasable pool):

• recycling pool:

• already docked vesicles, immediate release

• a few %

• can be released upon medium-intensity stimulus

• reserve pool: • released only after prolonged

stimuli (?) • majority of SVs

• approx 100-150 SVs per synapse / active zone

CAZ: Cytoplasmic matrix

of the active zone

periactive zone

Synaptic vesicule turnover

Synaptic vesicule turnover

palmitoylation

docking

platform

Exocytosis of a SV

• similar to „normal” exocytosis but much faster:

- docking – priming – Ca2+ trigger – fusion is a fast process during NT and neuropeptide release

- excitation – secretion coupling < 60 msec !

neuronal SNARE (soluble N-ethylmaleimide-sensitive factor [NSF] attachment protein [SNAP] receptor) complex

- plasma membrane:

- vesicular membrane:

• syntaxin 1 • SNAP-25 (synaptosome

associated protein of 25 kDa)

• synaptobrevin/VAMP (Syb2)

trans-SNARE komplex

• this SNARE complex is required only for the fast, Ca-triggered vesicular fusion, but not for spontaneous release (redundant SNAREs)

• in vitro the SNARE complex initiates membrane fusion only within hours

- in vivo 1-3 SNARE complex is enough for the fast fusion

- stabilising / activating factors:

• Munc-18: SM (Sec1/Munc18-like) protein -> regulation of release

• synaptotagmin: Ca2+ sensor -> trigger

• complexin: „clamping”

-different types – with different kinetics!

- trans – cis SNARE: fusion

Exocytosis of a SV

• additional core active zone protein network (scaffold proteins, protein-protein interactive domains):

- RIM (Rab3-interacting molecule)

Exocytosis of a SV

• Rab3 binding: vesicle docking

• Mun13 binding: vesicle priming

• RIM-BP / CaV binding: distance from the Ca2+ channels

- RIM-BP (RIM binding protein)

- Liprins: multimerisation; trans-synaptic localisation via PTPRs

- ELKs: regulate readily-releasable pool size

- Munc 13: essential for vesicle priming (binding and regulation)

- piccolo, bassoon: SV „guidance” to the AZ; associated at the sides of the AZ -> presynaptic skeleton

Cytoskeleton of the active zone (CAZ)

Endocytosis of a SV

- „kiss-and-run”: local recycling following endocytosis

- endosomal recycling

• clathrin-mediated endocytosis (dynamin)

- „kiss-and-stay”: local filling without detachment

Active zone in humans and in rodents – evolutionary differences?

• in similar cortical synapses, AZ is larger with more docked vesicles in humans compared to rodents – 4x more functional release site

Active zone in humans and in rodents – evolutionary differences?

Dale’s principle: one neuron, one neurotransmitter???

• traditional classification of NTs:

small molecule NT (Glu, GABA, Gly, ACh, purin, monoamine)

neuropeptides (SST, NPY, SubstP, encephalin, etc)

• normally in separate vesicles, can be released at the same time but with different kinetics

excitatory inhibitory neuromodulatory cells

• new findings: frequent GABA co-release!

Gly-GABA dopamine-GABA Glu-GABA Glu-GABA?

dopamin-GABA

ACh-GABA

Dale’s principle: one neuron, one NT???

Essay questions (choose one) What does the phrase „quantal release” mean? Explain the main steps and their

timescale during synaptic transmission! / Mit jelent a kvantális release fogalma?

Ismertesse a kémiai szinapszisokra jellemző szinaptikus jelátvitel lépéseit, azok

időrendjét!

Explain the main steps during the turnover of synaptic vesicles! Describe the main

components and current hypotheses for the turnover! / Ismertesse a szinaptikus

vezikula-körforgás főbb lépéseit! Ismertesse a főbb molekuláris komponenseket és

a jelenleg elfogadott elmélet(ek)et az egyes lépésekre!

Describe the main membrane components of synaptic vesicles and explain their

physiological role and importance! / Ismertesse a szinaptikus vezikulák általános

felépítését és a legfontosabb membránfehérje komponenseket, azok funkcióját!

Describe the molecular events taking place within the acive zone of the

presynapse! Introduce the important components of the active zone and their

functions in neurotransmission! / Mi történik a preszinapszis aktív zónájában?

Ismertesse az itt azonosítható jellegzetes struktúrákat, komponenseket és ezek

szerepét is!

How are „Ca2+ microdomains” formed? How do they contribute to presynaptic

functions? / Hogy alakulnak ki a Ca2+ mikrodomének? Hogyan vesznek részt a

preszinapszis működésének szabályozásában?

Recommended literature