Neuromuscular Function: Neural Impulse and Neurotransmitter Release

Muscle Physiology

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Agenda

Nerve impulse IntroductionChannels and pumpsThe neural impulse

Neurotransmitter release

Nerve Impulse - Introduction

What is a nerve impulse? A transmitted electrical charge that stimulates or inhibits

a physiological event What type of event?

Stimulate/inhibit another neural impulse Stimulate a gland Increase/decrease heart rate Activate skeletal muscle

What is an action potential? Synonym for impulse

Nerve Impulse - Introduction

Basic progression of events:

1. Disruption of the cell membrane’s electrical state

2. Restoration of the cell membrane’s electrical state

Nerve Impulse - Introduction In order to disrupt or restore a cell

membrane’s electrical state channels and pumps are needed

Agenda

Nerve impulse IntroductionChannels and pumps

General propertiesRegulatory channels pumps and other

The neural impulse Neurotransmitter release

Channels and Pumps – General Properties Purpose of channels and pumps: Maintenance of the cell membrane’s resting

electrical state (resting membrane potential – RMP) Both channels and pumps

Disruption of the cell membrane’s RMP Primarily channels

Restoration of the cell membrane’s RMP: Primarily pumps

Sarcolemma as well

Channels and Pumps – General Properties How is the RMP maintained, disrupted,

restored? Channels and pumps move charged ions into

and out of the cell Channels: Ions flow along electrochemical gradient Pumps: Move ions against electrochemical gradient

Terminology: Anion: Negatively charged ion Cation: Positively charged ion

Channels and Pumps – General Properties Speed and direction of transfer: Channels:

Can move several million ions / second Use diffusion (no energy required) Channels are less frequent Channels stay open for short periods of time

Pumps: Can move several hundred ions / second Require energy There are many more pumps than channels Pumps work constantly

Channels and Pumps – General Properties Selectivity: Channels and

pumps only allow certain molecules to pass

Mechanisms: Size: Water shell

Remember hydrophobic interior of membrane

Affinity: Specific proteins within the channels/pumps

Agenda

Nerve impulse IntroductionChannels and pumps

General propertiesRegulatory channels, pumps and other

The neural impulse Neurotransmitter release

Regulatory Channels, Pumps, Other Sodium channels Sodium-potassium pumps Potassium channels Calcium channels and pumps Anion channels

Na+ Channels

Structure: Two subunits

Alpha: Larger Acts as actual channel

Beta: Purpose unclear

Na+ Channels

Function: Disruption of RMP Voltage-gated

Change in electrical state of plasmalella activates channel

Sodium passes with concentration gradient

MacIntosh et al. 2006, Fig 9.7

Na+ Channels

Distribution: Na+ channels are found:

Axon hillock and nodes of Ranvier Sarcolemma Synaptic cleft T-tubules

Highest Na+ channel densities are observed at: Synaptic clefts Transverse tubules Nodes of Ranvier

Regulatory Channels, Pumps, Other Sodium channels Sodium-potassium pumps Potassium channels Calcium channels and pumps Anion channels

Na-K+ Pumps

Structure: Two subunits:

Alpha: Larger Contains Na+, K+

and ATP binding sites

Beta: Function not clear

Na-K+ Pumps

Function:

1. Maintain RMP at rest

2. Restore RMP after disruption

Na-K+ Pumps

Maintenance of RMP: Each cycle of the Na-K+ pump results in:

Removal of 3 Na+ Retrieval of 2 K+

Net removal of 1 cation intracellular negativity

Restoration of RMP after disruption

Na-K+ Pumps

Na-K+ pumps require ATP Enzyme Na-K+ ATPase Two states:

E1E2

MacIntosh et al, 2006, Fig 7.8

1. E2 releases Pi and picks up ATP

2. Energy from ATP releases 2 K+ and changes to E1

3. E1 releases ADP and picks up 3 Na+ and Pi

4. Pi changes back to E2 and releases 3 Na+ and picks up 2 K+

Regulatory Channels, Pumps, Other Sodium channels Sodium-potassium pumps Potassium channels Calcium channels and pumps Anion channels

K+ Channels

Structure: Several types of K+ channels with varying structures

Some allow K+ to leave the cell Some allow K+ to enter the cell

Different stimuli activate different K+ channels Increased intracellular [Na+] Decreased intracellular [ATP] Disruption of RMP Increased intracellular [Ca2+] Sarcoplasmic reticulum activation

K+ Channels

Function: Restoration of the RMP following disruption

Fast K+ channels allow outflow of K+ Activated via membrane disruption

Restoration of RMP during fatigue Several types of K+ channels inflow of K+ Activated via increased intracellular [Na+], [ATP],

[Ca2+]

Regulatory Channels, Pumps, Other Sodium channels Sodium-potassium pumps Potassium channels Calcium channels and pumps Anion channels

Ca2+ Channels and Pumps Structure: Ca2+ channels:

Several types: Voltage-gated Ca2+ channels:

Embedded within the axolemma of neuron Dihydropyridine (DHP) channels

Embedded within the sarcolemma of muscle Ryanodine (RYR) channels

Embedded within the SR membrane of muscle

Ca2+ pumps: Two types:

Ca2+ surface membrane pumps (SMP): Larger

Sarcoplasmic reticulum pumps (SERCA or Ca2+ ATPase) Occupies ~90% of the SR membrane

Ca2+ Channels and Pumps Function: Ca2+ channels:

Link disruption of cell membrane of neuron/muscle fiber to a molecular event

Neuron: Ach release Muscle fiber: Cross-bridge formation

Ca2+ SMPs: Maintain low intracellular [Ca2+]

SERCA or Ca2+ SR pumps: Remove Ca2+ from the sarcoplasm back into the SR Requires ATP (Ca2+ ATPase)

Regulatory Channels, Pumps, Other Sodium channels Sodium-potassium pumps Potassium channels Calcium channels and pumps Anion channels

Anion Channels

Structure: Similar to Na+ channels Most common is Cl- channel

Function: Maintain the RMP by flowing out

Distribution: Cl- channel is most common of all channels High permeability of Cl-

Anion Channels

The myotonic goat Genetic mutation results in decreased

permeability to Cl- Result: Inability of muscle fiber to restore RMP

following initial disruption Myotonic goat video

Agenda

Nerve impulse IntroductionChannels and pumpsThe neural impulse

Neurotransmitter release

Neural Impulse

The resting membrane potential Basic progression of events:1. Cell body of the neuron must receive adequate

stimulus-All-or-nothing fashion

2. RMP is disrupted (depolarized)3. RMP is rapidly restored (repolarized)4. Propagation

Neural Impulse - RMP

What is the Resting Membrane Potential (RMP)?

The difference in charge between the inside and the outside of the cell

Typical value -70 mVThe inside of the cell has a charge of –70 mV

relative to the outside of the cell

Neural Impulse

How is the RMP maintained? Several mechanisms:

1. Fixed anion structures increase negativity within cell

2. Na-K+ pump:-High extracellular [Na+], high intracellular [K+]

3. Permeability of membrane

Permeability of Membrane to Na+ Concentration gradient: Na+ into cell Electrical gradient: Na+ into cell Electrochemical gradient: Strong inward Channels: Few Effect: Low permeability of Na+ into the

cell

Permeability of Membrane to Cl-

Concentration gradient: Strong into cell Electrical gradient: Strong out of cell Electro chemical gradient: Weak outward Channels: Moderate Effect: Moderate permeability of Cl- out of

the cell

Permeability of Membrane to K+

Concentration gradient: Strong out of cell Electrical gradient: Strong into cell Electrochemical gradient: Weak outward Channels: Many Effect: High permeability of K+ out of cell

As K+ leaks out, they collect along the outer membrane due to negativity inside the cell

Bottom Line

The resting membrane potential is that “membrane potential” when the forces driving the influx/efflux of all ions are at equilibrium (no net movement of ions)

If membrane were permeable to only K+:RMP ~ -90 mV

Addition of Na+ and removal of Cl-RMP ~ -70 mV

Extracellular fluid

Intracellular fluid

Various fixed anionic structures

[Na+]

[Na+]

[Cl-]

[Cl-]

Least permeable. More permeable.

3 Na+

2 K+

[K+]

[K+]

Most permeable.

Neural Impulse

The resting membrane potential Basic progression of events:1. Cell body of the neuron must receive adequate

stimulus-All-or-nothing fashion

2. RMP is disrupted (depolarized)3. RMP is rapidly restored (repolarized)4. Propagation

Neural Impulse: Adequate Stimulus

Recall that the RMP is -70 mV The dendrites of a neuron will receive

multiple stimulus from multiple different neurons

Some of the neurons are excitatory and some are inhibitory

Neural Impulse: Adequate Stimulus

Excitatory neurons: Neurotransmitter: Acetylcholine Action: Activate sodium channels Na+ flows in which

increases the RMP (makes more positive) Inhibitory neurons:

Neurotransmitter: Gamma amino butyric acid (GABA) or glutatmate

Action: Open chloride channels Cl – flows in which decreases RMP

(more negative) Open potassium channels K+ flows out which decreases RMP

(more negative)

Neural Impulse: Adequate Stimulus

Excitatory neurons create EPSPs Excitatory postsynaptic potentials

Inhibitory neurons create IPSPs Inhibitory postsynaptic potentials

It is the sum of all EPSPs and IPSPs that determines the net stimulus

If the net stimulus exceeds ~15 mV, threshold is reached

http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/E/ExcitableCells.html

Neural Impulse: Adequate Stimulus

All-or-nothing principle: The strength of an impulse is an intrinsic

property of that neuron Stronger stimuli do not increase the

strength of the impulse

Neural Impulse

The resting membrane potential Basic progression of events:1. Cell body of the neuron must receive adequate

stimulus-All-or-nothing fashion

2. RMP is disrupted (depolarized)3. RMP is rapidly restored (repolarized)4. Propagation

Neural Impulse - Depolarization

Depolarization: RMP -70 mV +30 mV Stimulus exceeds threshold Voltage-gated Na+ channels open

M gate Na+ flows into cell increasing RMP

H gate Change in charge closes second gate Depolarization activates adjacent voltage-gated Na+

channel Process continues along axolemma

MacIntosh et al. 2006, Fig 9.7

Neural Impulse - Repolarization

Repolarization: RMP +30 mV -70 mV H gate shuts Voltage gated K+ channels open

K+ flows out of cell Na-K+ pump assists Voltage gated K+ channels stay close

Overshoot of K+ outflow = hyperpolarization

Neural Impulse - Hyperpolarization

Also known as the “refractory period” Two parts:

Absolute refractory period: Due to the inactivation of the h gate Time: ~ 2.2 – 4.6 ms

Relative refractory period: Due to overshoot of K+ ion outflow past RMP Greater stimulus needed to create another action

potential

Neural Impulse - Propagation Propagation: The pattern of depolarization followed

by rapid repolarization along a membrane Differences between neurons and muscle fibers

Speed of transmission Muscle fiber: 3-6 m/s Neuron: 40-65 m/s

Saltatory conduction nodes of Ranvier High channel density

End result Muscle fiber: Muscle contraction Neuron: Neurotransmitter release onto neuron, gland, muscle

etc.

http://human.physiol.arizona.edu/sched/cv/wright/16action.htm

http://www.accessexcellence.org/RC/VL/GG/action_Potent.html

http://www.accessexcellence.org/RC/VL/GG/action_Potent.html

Agenda

Neural impulse Neurotransmitter release

Structural considerations of the neuromuscular junction (NMJ)

Basic progression of events

NMJ Structure

The NMJ includes:The distal neuron

Synaptic knobs/terminal endings/axon terminals Synaptic vesicles

The muscle fiber Motor end plate Primary and secondary synaptic clefts Acetylcholine receptors Sarcolemma

NMJ Structure – Distal Neuron

The distal neuron gradually loses its myelin as it approaches the muscle fiber

The neurons branch excessively and end in “boutons”Synaptic knobsTerminal endingsAxon terminals

NMJ Structure – Synaptic Knobs Lay in a semi-circle manner Do not make direct contact with muscle

fiber Function: Release neurotransmitter

AcetylcholineNEED FIGURE 3.1, MacIntosh

NMJ Structure – Synaptic Vesicles Small spheres located within

synaptic knobs Contain the neurotransmitter

Ach Formed when axolemma

becomes invaginated and “pinches off”

NMJ Structure

The NMJ includes:The distal neuron

Synaptic knobs/terminal endings/axon terminals Synaptic vesicles

The muscle fiber Motor end plate Primary and secondary synaptic clefts Acetylcholine receptors Sarcolemma

NMJ Structure – Muscle Fiber Motor end plate: The area of the muscle

fiber that makes “near” contact with the synaptic knob

NMJ Structure – Muscle Fiber Primary synaptic cleft: A small gap that separates the

membranes of the synaptic knobs and the muscle fiber ~70 nm

Secondary synaptic cleft: Regular repeated invaginations of the sarcolemma underneat the primary synaptic cleft Add Fig 3.1, MacIntosh

Acetylcholine receptors Embedded within plasmalella in junctional folds ~10,000/micrometer2 (2 binding sites/receptor) 5 subunits (2 bind Ach)

NMJ Structure – Muscle Fiber

Sarcolemma:Basement membrane lays over both synaptic

clefts Insulation

Contains acetylcholinesterase (AchE) Hydrolyzes Ach and stops synaptic transmission

Agenda

Neural impulse Neurotransmitter release

Structural considerations of the neuromuscular junction (NMJ)

Basic progression of events

Neurotransmitter Release

Basic Progression of Events: Action potential reaches synaptic knob Neurotransmitter release from synaptic vesicles Motor end plate depolarization Acetylcholinesterase hydrolyzes Ach Hydrolyzed Ach is resynthesized Resynthesized Ach is taken up by synaptic vesicles

Action Potential Reaches Synaptic Knobs Depolarization of synaptic knob activates

voltage-gated Ca2+ channels Ca2+ rushes into synaptic knob Role of Ca2+:

Assist with fusion of synaptic vesicles Assist with release of Ach from vesicles

Ca2+ mediated release of Ach is the rate limiting step (~0.2 ms)

Ach Release from Vesicles

Prior to Ca2+ influx, vesicles are “docked” Ca2+ assists with fusion with axolemma Ca2+ assists with Ach release from

vesicles via exocytosis New vesicles are created via endocytosis

Prevents build-up of tissue at synaptic knob

Ca2+

Marieb & Mallett, 2005, Fig 12.8

Motor End Plate Depolarization

2 Ach bind receptors opening center pore of receptorNa+ flows into cellK+ flows out less rapidly

Voltage-gated Na+ channels activated around the motor end plate

Sarcolemma depolarizes and action potentials propagate

Ach

Na+ K+

-Decreased negativity inside

-Increased RMP

-Motor end plate depolarized

AchE Hydrolyzes Ach

Upon receptor activation, Ach molecules dissociate

Ach molecules fall into secondary synaptic cleft

AchE hydrolyze acetate + choline molecules

Ach

AchE

A + Ch

Acetate Choline

AchE

Ach

Ach is Resynthesized

Choline molecules are absorbed into the synaptic knob

Choline acetyltransferase resynthesizes AchAcetyl CoA + choline acetylcholine

Acetyl CoA from mitochondria

New Ach Vesicles

Acetylcholine transporter assists with uptake of resynthesized Ach into vesicles

Filled vesicles dock near the axolemma