6. Lectures 14, 15, 15.5

Question 1

What are electrical synapses?

Type of membrane protein, time of transmission, separation, etc

Answer 1

Direct electrical transmission used for communication between cells
Used connexon membrane protein
Instantaneous transmission
Tiny distance of separation (3nm)

During excitatory synaptic transmission at electrical synapse, voltage gated ion channels in presynaptic cell generate the current that depolarizes the postsynaptic cell
These channels depolarizes the presynaptic cell and generate sufficient ionic current to produce a change in potential in post synaptic cell

Whatever happens to cell on left, happens to cell on right

Slides 4-8 lecture 14

Question 2

What are chemical synapses?

Agonist, membrane proteins, separation

Answer 2

Indirect chemical modes of transmission cells use to communicate
Ionotropic (faster) and metabotropic (slower)
Uses receptor/channel as membrane protein
ACh is an agonist
Large distance of separation between two opposing membranes (30nm-50nm)
Have an affect on one cell, but ends a terminal where electrical turns to chemical
This conversion keeps effect from crossing cells

Slides 4-8 lecture 14

Question 3

What are gap junctions?

What is their structure?

Answer 3

Connect 2 cells with a large and unselective pore
Allow passage of ions and small molecules as big as 1kDa
Expressed virtually in all mammalian cells
They provide pathways for both chemical and electrical communication

Structure: 2 opposed hexameric connexons (one contributed by each cell)
Each connexon has 6 subunits surrounding a central pore (each subunit is a connexin)

Slides 9-11 lecture 14
Slides 13-14 lecture 14

Question 4

What is a homotropic channel?

What is a heterotropic channel?

Answer 4

Homotypic channel- opposition of 2 identical connexon hexamers

Heterotropic channel- opposition of dissimilar connexon hexamers

Homomeric- connexon formed by single type of connexin
Heteromeric- connexon firmed by mix of different connexins

Slide 12 lecture 14

Question 5

Is the current passing through the gap junctions ohmic?

Answer 5

Yes; it varies linearly with the transjunctional voltage (Vm membrane voltage difference between the 2 cells)

Question 6

How does gap junctions gating work?

Answer 6

Gap junction regulated by calcium (permeable to calcium)
Increases in [Ca]i can cause gap junctions to close

In absence of Ca, the pore is in an open configuration and the connexin subunits are tilted 7-8 degrees from an axis perpendicular to the plane of the membrane
After addition if Ca, the pore closes and the subunits move to a more parallel alignment

Slide 16 lecture 14

Question 7

What’s the difference between gap junctions and hemichannels?

(What they allow across)

Answer 7

Gap junctions allow indirect intracellular propagation if second messengers (Ca, IP3, cAMP), metabolites (glutamate, glucose), and nucleotides (ATP, ADP, RNA) between adjacent cells

Uncoupled “free” hemichannels facilitate 2 way transfer if molecules between cytosol and extracellular milieu

Slide 18 lecture 14

Question 8

What are connexin hemichannels?

Answer 8

Hemichannels involved in the release of intracellular molecules into extracellukar spaces, such as in pannexin channels, resulting in communication links with neighbouring cells via paracrine pathway

Way that many cells release neurotransmitters

Slide 19 lecture 14

Question 9

How can chemical synapses amplify signals?

Answer 9

No structural continuity between pre and post synaptic neurons
Synaptic cleft is the separation between the 2 cells
Chemical synaptic transmission depends on the diffusion if a neurotransmitter across the synaptic cleft
Presynaptic terminals contain 100-200 synaptic vesicles each is filled with several thousand molecules of neurotransmitter

Chemical transmission lacks the speed of electrical synapses, but amplified signals (one synaptic vesicle releases several thousand transmitters that can open thousands of ion channels on target cells

Slide 22 lecture 14

Question 10

What are ionotropic and metabotropic post synaptic receptors?

Answer 10

Chemical transmitter receptors
Neurotransmitter receptor molecules
Ionotropic- fast ligand gated ion channels
Metabotropic- G protein linked receptors (instead of activating channel they trigger G protein generate cascade of kinases that lead to regulation of a channel)

Slide 23 lecture 14 shows difference
Slide 3 lecture 15.5

2 things in common:
Both are membrane spanning proteins that recognize and bind the transmitter
Both carry out an effective function within the target cell: postsynaptic receptors gate ion channels either directly or indirectly

Question 11

What are miniature end-plate potentials (MEPPs)?

Answer 11

Electrophysiological “noise” when recording membrane potential
The kiss was tiny depolarizations of ~0.4mV at random intervals
They are blocked by curare (nicotinic AChR antagonist) and enhances in size by inhibitor of AChE

Since these occur in discrete multiples of a unitary amplitude, ACh release is quantized, with the quantum event corresponding to ACh release from one synaptic vesicle

Slide 4-5 lecture 15
Slide 15 lecture 15.5

Question 12

How can the quintal nature of transmitter release be expressed quantitatively?

Answer 12

By postulating that a nerve terminal contains a population of N quanta or vesicles, that each has a finite probability (P) of releasing
Thus the mean number (m) of quanta released after any single nerve impulse is:

m= N x P

Question 13

What is the relation between presynaptic Ca2+ concentration and neurotransmitter release?

Answer 13

Calcium causes delay

Transmitter release is steeply dependant on Ca concentration in the presynaptic terminal
The release of transmitter from a vesicle requires the binding of 5 Ca2+ ions to a calcium sensing synaptic vesicle protein

Vesicle has a system to detect calcium and fuse to membrane and release neurotransmitter

Neurotransmitter release can be blocked by preventing Ca2+ influx

Slides 7-9 lecture 15

Question 14

What catalyzes fusion of vesicles with membrane?

Answer 14

SNARE proteins catalyze fusion of vesicles with membrane
Tight interaction between VAMP and SNAP-25 proteins

Slide 10-11 lecture 15

First complex keeps vesicle close to membrane but doesn’t fuse it

Question 15

What are synapsins?

Answer 15

Peripheral vesicle protein
Regulates availability of vesicles from the reserve pool

Slide 10 lecture 15

Question 16

What is the SNARE complex cycle?

Answer 16

Slide 12 lecture 15

1. Synaptobrevin (VAMP) interacts with 2 plasma membrane target proteins (syntaxin and SNAP-25)
2. The 3 proteins form complex bringing vesicle and presynaptic membranes close (4 α helicies, one each from VAMP and syntaxin, 2 from SNAP-25)
   Munc18 bunds to SNARE complex
3. Calcium influx triggers rapid fusion of vesicle to membrane (SNARE complex now resides in plasma membrane
4. 2 proteins (NSF and SNAP) bind to SNARE complex and cause it to dissociate (ATP dependant reaction)

Question 17

What is the structure of synaptotagmin?

Answer 17

Synaptotagmin is an integral membrane protein of synaptic vesicles

Short N terminal tail
Single hydrophobic domain that spans the vesicle membrane
Long cytoplasmic tail that contains 2 C2 domains (C2A and C2B)
C terminus

Slide 13 lecture 15

Question 18

What are the 4 different active zone morphologies of exocytosis?

Answer 18

Dense projections- neuromuscular junction

T-bar structure- neuromuscular junction

Ribbon synapse- vertebrate photoreceptor cell ribbon synapse

Central synapse- vertebrate

Slide 14-15 lecture 15

Question 19

How is capacitance used as indicator of exo/endocytosis?

Answer 19

Slide 16 lecture 15

Exocytosis can release so much membrane capacitance goes up

Question 20

What is amperometry?

Answer 20

Presynaptic detection of transmitter release by carbon fiber electrode
Carbon fiber is an electrochemical detector of serotonin

At [Ca]o of 5mM, the current is large and composed of many small spikes
At 1mM, individual spikes of serotonin release are shown

Small and large spikes

Slide 17 lecture 15

Question 21

Do all synapses contain ionotropic and metabotropic receptors?

Answer 21

Yes!

Question 22

What is the nAChR structure in muscle?

Answer 22

Slide 4-6 lecture 15.5

Not highly selective, just need positive charge (cation)
Turns itself and twists to pull the gate open to open the pore

2 α, 1 β, 1 γ, and 1 δ
Need 2 alpha subunits because the combo of alpha subunit with other subunits generates binding sites
Receptor opened with 2 molecules of neurotransmitter so need 2 alpha subunits
α subunits contain receptor sites for acetylcholine

Question 23

What are homopentamers in nAChR composition?

What are heteropentamers?

Answer 23

Homopentamers
Functional with just alpha subunits 
α7- central nervous system
α8- only expresses cheek
α9- inner ear

Very fast (like voltage gated sodium channels)

Heteropentamers
α3β4- PNS
α4β2- CNS

All these receptors are activated by nicotine- when they open they allow formation if EPSP and cause excitation
A smoker will have trouble walking

Slide 7 lecture 15.5

Question 24

Where is nAChR location?

Answer 24

In the CNS- most are α4β2 locates in presynaptic neuron- involved in neurotransmitter release (increase it)
α4 has highest affinity in CNS and brain where addictive centres are

Muscle and PNS- only receptor- α3β4
Nicotinic receptors are post synaptic and involved in driving synapse transmission

Slide 8 lecture 15.5

Question 25

What’s the difference in function of muscle nAChR and neuronal nAChR?

Answer 25

Slide 9 lecture 15.5

Muscle receptors- alpha beta gamma delta subunits
Ohmic receptor more voltage applied the more current you get
Reverse potential at 0
Pore of muscle channel is 3x bigger than neuronal

Neuronal receptors- looks similar to Kir channel
Inward rectifier
Polyamines mop up negative charges (have a lot of positive charges)
Polyamines block neuronal receptors not muscle

Question 26

How can synaptic potentials at neuronal synapses be either excitatory or inhibitory?

Answer 26

Synaptic potentials are subthreshold

One can begin to elucidate the nature of the currents underlying these synaptic potentials by determining their reversal potentials

Action potential of presynaptic excitatory cell A leads to ESPS (excitatory post synaptic potential) of cell C
Action potential of presynaptic inhibitory cell B leads to IPSP (inhibitory post synaptic potential) of cell C

Slide 10 lecture 15.5

Question 27

What is the result of ACh binding to the post synaptic membrane?

Answer 27

It causes an EPSP (excitatory post synaptic potential) that decays with distance from the site of action
EPSP from ACh are graded potentials

Slide 11 lecture 15.5

Question 28

What ions pass through the AChR channels during generation of EPP?

Answer 28

ACh binding to the post synaptic membrane causes opening if ion channels and reversal potential of this channel can be determined using voltage clamp of post synaptic cell

nAChRs are non-selective cation channels (permeable time Na+, K+, Ca2+)

Question 29

How do the subunits of nAChRs affect their function?

Answer 29

Oocytes with injected mRNA encoding either α, β, γ, δ or α, β, ε, δ

Channel with ε subunit has unitary conductance of 59pS
Channel with γ subunit has conductance if 40pS

Different functional properties of nicotinic AChRs with different subunit compositions reflect their specialized roles in synaptic transmission

Fetus (γ, open for longer time) and adults (ε, currents larger and open more frequently) have different subunits

Question 30

What is α-bungarotoxin (α-BTX)?

Answer 30

Snake venom, neurotoxin

Binds competitively and irreversibly to nAChRs at the neuromuscular junction, causing paralysis, respiratory failure, and death

Antagonist