synaptic+transmission

LECTURE OBJECTIVES1. To understand the overall concept of synaptic transmission. 2. To understand the inputs into dendrites and the cell body.3. To analyze the differences between metabotropic and ionotropic

receptors.4. To understand the sequence of events in the presynaptic terminal

leading to neurotransmitter release.5. To list the methods of transmitter inactivation and discuss their

importance.6. To describe in the steps (synthesis, storage, release and inactivation

of acetylcholine) of neurotransmission at the neuromuscular junction.7. To analyze a disorder associated with the NMJ - Myasthenia Gravis.

Synaptic TransmissionRichard W. Keller, Jr., Ph.D.

Center for Neuropharmacology and NeuroscienceME-751 [email protected] 2-5253

Dendrites(inputs)

Cell Body(metaboliccenter)

Axon(conduction of

message)

Axon Terminals(output)

MAJOR FEATURES OF A NEURONThere are many different types of neurons but they almost all have similar major features.

Quick Review covered by Dr. Lindsley

Synaptictransmission

- AP – Dr. Shin

What does this really look like????

NeuromuscularJunction

Nerve – NerveConnections

See Figure 11-36 – Alberts, et al.http://education.vetmed.vt.edu/Curriculum/VM8054/Labs/Lab10/lab10.htm

Artist John Allison’s Rendering of the Cortex

No Exam Material!

Dendrites(inputs)

Cell Body(metaboliccenter)

Axon(conduction ofmessage)

Axon Terminals(output)

MAJOR FEATURES OF A NEURON

Chemical Signal Chemical SignalElectrical Signal

Why these changes in message??

Today’sFocus

Today’sFocus

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Major features common to almost all nerve cells.

1. Cell Bodya. Basic machinery of life. The cell body of the

neuron contains all the same organelles that arefound in all animals cells.

[nucleus, rough endoplasmic reticulum (ER),smooth ER, Golgi apparatus, mitochondria - lots!]

b. The cell body functions as the metabolic center,the site of protein synthesis & packaging, etc.

c. Diameter 5 - 60 um - small compared to most cells

d. Some inputs from other neurons - most via dendrites


The cell body =~0.1% of entire

cell volume!

Nucleus

Fig 1.3 – Neuroscience; 2nd Ed.Purves, et al 2001; Sinauer


THE CELL BODY

No Exam Material!

2. Dendritesa. Communication: neurons receive inputs (messages)

from adjacent cells to pass on to other cells. Manyof these inputs (afferent signals) are onprojections from the cell body called dendrites.

1. "antennas" of the neuron2. branched in a "tree-like" fashion

b. Several dendrites - number and extent of thebranching varies dramatically

1. average neuron receives about 10,000inputs in its dendritic tree

2. Purkinje cells - very large dendritic treefrequently receiving 150,000 inputs

INPUTS = NEUROTRANSMITTERS interacting with RECEPTORS (Ligand-gated Ion Channels!)

A typical Neuron hasabout10,000 inputs into the dendrites

Thousands of excitatory and thousands of

inhibitory inputs!

Inputs

ExciteInhibit

Receptors

A cell potential exists across all parts of a neuronincluding the cell body and dendrites.

The resting membrane potential is ~ -80 mVFrom last lecture - primarily due to a K leak!

In fact, all living cells have membrane potentials!

so.... What’s the big deal about potentials in neurons?

Significant Changes in the cell potential result from NTsinteracting with & opening LIGAND-GATED Ion-Channels!

Really 2 types of Neurotransmitter-Activated Receptors

Ligand-gatedIon channels

Metabotropicreceptors

2nd messengers

Fig 5.16 – Neuroscience; 5th Ed.Purves, et al 2012; Sinauer

1. 2.

neurotransmitter = ligand

Direct!Indirect!

FAST!SLOW!

Lots of Inhibitory and Excitatory Inputs

Some inputs function to "excite" the cell, while others tend to "inhibit"the cell.

At a cell potential level what does “excite” mean?

"inhibit"?

Excitation Inhibition Excitation & Inhibition

DepolarizingLess (-) more (+)-50mv 0 mv(+) current in

HyperpolarizingMore (-)-100mv

(+) current outor (-) current in

InitialResting pot.-80 mv

-80 mV -80 mV

Fig 12-16 Principles of Neural Science4th Ed. Kandel, Schwartz & Jessell

Reality is a combination of the

two effects.

Reality is a combination of Excitation & Inhibition

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What are the most common Inputs on these Dendrites??

Of the 100 billion nerve cells in the CNS:

50-60% OF ALL NEURONS IN THE CNS USE GLUTAMATE AS THEIR NEUROTRANSMITTER!

30-40% OF ALL NEURONS IN THE CNS USE GABA AS THEIR NEUROTRANSMITTER!

So significant number of inputs on the dendritesare likely via Glutamate and GABA.

GABA = Gamma-aminobutyric acid

Na+ Cl-

GlutamateExcitation

GABAInhibition

Glu Glu GABA GABA

InitialResting

pot.-80 mv

MorePositive;

depolarizing

MoreNegative;

hyperpolarizing

Glutamate = major excitatory NT GABA = major inhibitory NT

In CNS Most Excitatory & Inhibitory Inputs are Glu & GABA

xxx

K+

Does “xxx” produceexcitatory orinhibitoryeffects viathis receptor???

xxx

Self StudyYou think about

this!inside

outside

No Exam Material!

DendriteApply Glutamate (Glu) into area with Glu receptors

Adapted from Fig 2.3 – Neuroscience; 5th Ed. Purves, et al 2012; Sinauer

a “graded” potential !!

Activation of Receptors Results in GradedPotentials in the Dendrites & Cell Body

signal decays over distance…..

ExcitatoryInput

ligand-gated ion channels…..

axon

Axon Hillock

B. There are thousands of excitatory and inhibitory inputs on dendrites & the cell body1. “cellular decision” has to be made to fire or

not to fire an action potential2. potential is summed over area (spatial

summation) & time (temporal summation)

3. the “trigger zone” is located in the “initialsegment” of the axon; this is where the “cellular decision” is made. [Adjacent to the axon hillock region of the cell body.]

If the cell potential in the initial segment is

sufficiently positive to lead to the opening of

voltage-gated Na+

channels – the cell fires an action potential.

Initial segment - very high density of voltage-

gated Na+ channels

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Na+

Channel= AP

K+

Channel= recovery

Action Potential moves down the Axon!

Covered by Dr. Shin and in Membrane Transport

axon = neuron's output - the "wire" leading to where the message will be delivered

All or None?

No Exam Material!

Na+/K+ Pump

ENERGYREQUIRED!

Covered in Membrane TransportFrom Fig 4.11 – Neuroscience; 5th Ed. Purves, et al 2012; SinauerNo Exam Material!

TERMINAL FIELDCELL BODYand

DENDRITES

AXON

LIGAND-GATED ION CHANNELS(“Neurotransmitter-activated”)

METABOTROPIC RECEPTORS that modify ion channels

Excite - depolarizeInhibit - hyperpolarize

GRADED POTENTIALS!

AXON Terminalsor “boutons”

AXON HILLOCKthreshold - fire?spatial summationtemporal summation

VOLTAGE-GATED ION CHANNELSALL OR NONE ACTION POTENTIAL!INFO CODED IN FREQUENCY

Examine what happens here!

Presynaptic terminal:contains synaptic vesicles loaded with neurotransmitter molecules

Lots of mitochondria (synaptic activity consumes lots of energy) in the terminal.

Synaptic cleft: 20-30 nmCrisscrossed by thin filaments

Postsynaptic component:contains intrinsic membrane proteins (receptors) to which the released transmitter can bind. This can trigger a variety of postsynaptic events.

Synaptic Transmission“The big picture”

Chapter 5 - Box 5A –Neuroscience; 5th Ed. Purves, et al 2012; Sinauer

DENDRITE OR MUSCLE FIBER!!

Synthesis of Neurotransmitters

synthesis of Enzymes &

Vesicles

synthesis of Neuro-

transmitters

can be up to 2 meters in human!

Figure 5.5 – Neuroscience; 5th Ed. Purves, et al 2012; Sinauer

THE “NAME” OF A NEURON COMES FROM THE NEUROTRANSMITTER THAT IS RELEASED

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The Synthesis, Packaging and Secretion of Neurotransmitters

Storage Vesicles

BiosyntheticEnzymes


Synaptic Terminals / Synaptic Boutons -con’t• An action potential (AP) moves down the axon and

invades the terminal area. This AP opens voltage-dependent Na+ channels and then voltage-dependent Ca+2 channels leading to an influx of Ca+2.

• This influx of Ca+2 is the trigger for exocytosiswhich leads to the release of neurotransmitter.In this process, vesicles fuse with the plasma membrane of the terminal and dump their contents into the cleft of the synapse.

• Terminals conserve membrane by recycling it; afterrelease, vesicles are reformed and refilled.

EXOCYTOSIS - the simple version!


“DockedVesicle”

Plasmamembrane

A REALVESICLE!

Takamori, et al.Cell 127: 831-846

Fig 5.10 – Neuroscience; 2nd Ed.Purves, et al 2001; Sinauer

More of the components involved in exocytosis

… and then a miracle

happens!

No Exam Material!


Role of Ca+2 in Synaptic Transmission

1

2

Remove Ca+2 from Buffer!1

2

Control

Why this delay?

?

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“PROJECTION AREA”OR

“TERMINAL FIELD”Those with AXON terminals

There are 2 major types of release sites.

Those with AXON “boutons”

AcetylcholineGlutamateGABA

DopamineNorepinephrineSerotonin

10,000terminals

500,000boutons

Associated with these two different fields are

1. AxonTerminals axon

axon

Extracellular levels – IMP!

Volume Transmission

2. Axon Boutons

ClassicalNeurotransmission

Synaptic cleft levels - IMP!

2 types of synaptic contacts that release NTs.

Catecholamines & Serotonin

GlutamateGABA

(ACh - NMJ)

• Release must be terminated and the synapse must be cleared of transmitter so that the next message can produce its effect.

• The mechanisms of NT removal used are:

1. Uptake or Reuptakea. Reuptake in the cells that released the NTb. Uptake into another cell

2. Enzymatic degradation

3. Simple diffusion away

Removal of NeurotransmittersResetting the system for the next message!

Which drug blocks the removal of ACh at the NMJ and is a threat to life?Nerve Gas – loss of the ability to breath – can’t move diaphragm!!

DA

DADADA

DA

AP

DA-term.ppt

vesicular uptakeVMAT

DAT DAT

Other Cells

DA

COMTmetabolism

80%

Post SynapticTarget Cell

20%

1

2

1

2DA Uptake 1DA Uptake 2

DAT = Dopamine Transporter (Reuptake)

DAT MAOmetabolism

60%

40%

DA recp DA recpDA recp

DopamineNorepinephrineEpinephrineSerotonin

ReUptakeinto Boutons

Transporters (Trans) are protein carriers that move molecules through the plasma membrane

ECF

1 of the 500,000 boutons

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Uptake intoAstrocytes

GlutamateGABA

Fig 7.2 – Fundamental Neuroscience;2nd Ed. Squire, et al 2003; Academic Press

Major removal Site! Glu & GABA

Transporters

(Astrocyte)

Nerve Terminal

1. Uptake

2. Enzymatic degradation1. In the ECF2. In other cells3. In the releasing terminal / bouton


Removal of Neurotransmitters

Thanvi & Lo - Postgrad Med J 2004;80:690-700

Enzymatic Degradation

AChCholine+ acetate

NeuroMuscularJunction

AChHistamine

Acetylcholinesterase

1. Uptake

2. Enzymatic degradation


Removal of Neurotransmitters

All NTs to some extent!

Fig 7.2 – Fundamental Neuroscience;

2nd Ed. Squire, et al 2003; Academic Press

Diffusion

Recovery from Depolarization of Release(Recovery of the Terminal)


Receptor Binding in Post synaptic cell

Metabotrophic Receptor - Second

Messengers

Neurotransmitter Release

Ligand-gated Ion channelopening or closing

Postsynaptic cells – membrane potential change - excited or inhibited

Events from Neurotransmitter Release to Postsynaptic Excitation or Inhibition

Does binding to a receptor play a role

in NT removal?

NtNt Nt

Nt

NtNtNt

Nt

Nt2

Locations ofReceptors

Nt2

PostsynapticPresynaptic

autoreceptors(Feedback – enough NT! )

Presynapticheteroreceptors

Nt3

Nt3

Ion channel

Metabotropic receptor

Not much evidence for thesehetero -receptors at NMJ.

There are presynaptic auto-receptors at NMJ.

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Nt = neurotransmitter

The Combined Effect of Neurotransmitters and their Receptors

Many different kinds of chemical messages

• Fast vs slow – Some neurotransmitters (NTs) work veryquickly and are rapidly removed (ACh at NMJ) vsothers that are more neuromodulators (DA in brain)

• Receptors also add variability. The same NT can beExcitatory at one receptor and Inhibitory at another.

• There are receptors with different affinities for the same NT; so low levels of NT can stimulate one set of receptors and high levels can stimulate additional receptors.

Same NT Different Responses at Different Acetylcholine Receptors

Excitatory

Inhibitory

see animation of inhibition @ http://www.blackwellpublishing.com/matthews/neurotrans.html

Nicotiniccholinergicion channelreceptor

Muscariniccholinergicmetabotropicreceptor

Many Different Receptors Respond to Different levels of

Neurotransmitter

Ion Channel Receptors

Metabotropic ReceptorsIon Channel vs

Metabotropic

Excitatory vsInhibitory

Some respond to low NTlevels others respondto high NT levels

Fig 6.3 & 6.4 – Neuroscience; 5th Ed.Purves, et al 2012; Sinauer Do Not Memorize

The Neuromuscular Junction (NMJ)Specialized chemical synapse between a motor neuron and striated muscle fibers. The NT used is acetylcholine(ACh)

Synthesis: from acetyl CoA and choline by choline-acetyl transferase (CAT) in the terminal

Storage & release: Uses mechanisms described previously

Receptors: NMJ uses nicotinic acetylcholine receptors. These are ligand-gated Na+ (some K+) ion channels that will generate a depolarization when bound to ACh. This depolarization is called the endplate potential (EPP).

Inactivation: ACh is inactivated enzymatically. The synaptic cleft in NMJ contains acetylcholinesterase (ACE) which breaks ACh into acetyl CoA and choline. Choline is brought into the terminal via a transporter for reuse in synthesis.

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The Neuromuscular Junction (NMJ):

http://education.vetmed.vt.edu/Curriculum/VM8054/Labs/Lab10/lab10.htmhttp://demo.classontheweb.com

NMJ

Saladin – Anatomy and Physiology:The Unity of Form and Function 1996

McGraw-Hill

CNS Synapse

2,000-6,000 um2 2-10 um2

Surface Area of a Presynaptic TerminalThe Neuromuscular Junction (NMJ) A lot of what we

know was first worked out at the

NMJ!!

Not toScale!

Freeze Fracture EM

Heuser, et al. 1979J.Cell.Biol 81 275John Heuser = http://www.heuserlab.wustl.edu/v2.0/index.shtml

Unstimulated

Stimulated

NMJ Freeze

FractureSlam the tissue against a copper plate frozen in liquid nitrogen and then break it apart.

Do Not Memorize

Recordings in the junction reveal “graded” local potential changes (EPP) before an Action Potential is produced.

Stimulated ACh release ~150 MEPPs (vesicles) = EPP

An EPP involves ligand-gated ion channels – if the membrane potential change is sufficient (exceeds the voltage-gated Na+

channel threshold) then an AP that involves voltage-gated ion channels occurs in the muscle fiber.

The discovery of Miniature Endplate Potentials“mini’s” or MEPPs

Bernard Katz1950’s

spontaneous 0.5mV changes in potential(single vesicle leak)

evoked change with stimulation (AP-stimulated release of ~150 vesicles of ACh)

Nerve stimulation

Membrane potential of muscle fiber

EPP

Skeletal Muscle Action Potential• in the muscle, away from the neuromuscular junction

the AP is again all-or-nothing (no EPP!)

When the depolarization produced by the EPP is sufficient to activate voltage-gated Na+ channels, an action potential is generated in the muscle fiber.

This action potential results in opening of voltage-gated Ca+2 channelslocated both in the cell membrane and in endoplasmic reticulum membranes of the muscle fiber. The consequence is a massive entry of Ca+2 into the cytoplasm, both from outside the cell and from intracellular storage sites, which results in contraction of the muscle fiber.

EPP is playing a role similar to which region in

the neuron?

Curare (a cholinergic nicotinic antagonist) binds to nicotinic cholinergic receptors but produces no effects. If it is added to the bath, it blocks the binding and effects of ACh at these receptors – reducing the EPP below the voltage-gated Na+ channel thresholdthus preventing the AP.

Curare can limit the size of the EPP and thus block the initiation of the Action Potential.

60/68French physiologist Claude Bernard mid 1800s

Myasthenia Gravis• MG is an autoimmune disease that disrupts transmission at the

NMJ.• The body forms antibodies to nicotinic ACh receptors

- destroys some receptors and blocks others• There are less functional receptors and thus weak muscles.

Y Y

Receptorsblocked byantibodies!

Myasthenia Gravis cont…

• MG typically affects the cranial muscles first– weak neck muscles cause the head to fall forward or

backward– drooping eyelids (ptosis)– inability to move the eyes side to side– double vision (diplopia)– difficulty speaking (dysarthria)– difficulty swallowing (dysphagia)

Weakening of theMuscles!!!

Myasthenia Gravis cont…• One of the most treatable neuromuscular disorders!• Administer acetylcholinesterase blockers

such as neostigmine or pyridostigmine(+ immunosuppressive drugs)

This reduces the ACh destruction by AChE & increases the accumulation of ACh at the neuromuscular junction. Hence there is more ACh to interact the remaining receptors.

Before Neostigmine

After Neostigmine

Walker MB (1934). Treatment of myasthenia

gravis with physostigmine. Lancet 1:1200-1201.

Sample Questions:

1. Neurotransmitters (ligands) activate ion channels in the dendrites resulting in membrane potential changes that are described as:

a. all or noneb. negativec. gradedd. positivee. integrated

2. A rabbit shot with a curare-tipped arrow would quickly lose its ability to run away, because at the neuromuscular junction

a. the breakdown of acetylcholine would be inhibited.b. the acetylcholine receptors would be blocked.c. acetylcholine could no longer be released from the presynaptic terminals.d. the voltage-dependent sodium channels would be blocked.e. synthesis of acetylcholine from choline would be inhibited.

The End!c b

synaptic+transmission

Documents

cell body functions

cell body of theneuron

cell bodya

cell potential result

animals cells

nerve cells

purkinje cells

thousands of inhibitory