Properties of

Skeletal Muscle
Laboratory 2

Except where noted, Images and content are sourced from Sherwood textbook

Experimental Overview

Objective: study skeletal

muscle structure and
function including:

Setup / Procedures:

Isolate gastrocnemius and

sciatic nerve from doublepithed frogs

Test stimulus intensity

relationship

Spatial and Temporal

Summation

Test stimulus frequency

relationship

Steps at the neuromuscular

junction

Test pharmacological
antagonism

Test direct stimulation

Motor units
Excitation Contraction
Coupling

Skeletal Muscle Anatomy

Muscle comprises the largest group of tissues in the

body, accounting for approximately half the bodys
weight.

Skeletal Muscle is made up of muscle fibers (individual

muscle cells)

Myofibrils Composed of thick (myosin)

and thin (actin) filaments

Each thick filament is surrounded

by 6 thin filaments

Each thin filament is surrounded

by 3 thick filaments

The Sarcomere is the functional

unit of the skeletal muscle
A

bundle of thick & thin

filaments comprising the area
between two z-lines

The Motor Unit

Motor neurons branch to innervate

many individual muscle fibers

A motor unit consists of the motor

neuron and all of the muscle fibers it
innervates

Small motor units allow for very fine

muscle control with small force (eyes)

Large motor units are incapable of fine

control but are able to create larger
amounts of tension (leg muscles)

A muscle often has different sized

motor units which can control force of
contraction through recruitment

Recruitment The Henneman Size

Principle

Physiologically, motor units are

activated from smallest to
largest

This allows the body to easily

use the smallest necessary
force to accomplish the task

More force can be created as

needed for difficult tasks

Small motor units also have

higher fatigue resistance

Motor Nerve Structure

A nerve (as an organ, not an individual

cell) such as the sciatic nerve we are
studying contains many axons bundled in
groups called fascicles

Motor neuron axons vary in diameter based

on how many muscle fibers they innervate

Normally the spinal cord can activate motor

neurons in its preferred order (Henneman
Size)

Artificial stimulation recruits in the

opposite order (largest to smallest) because
large diameter axons have a lower
threshold to stimulation
Resistance

R=L/A

ExcitationContraction
Coupling:
Summary

1: The Neuromuscular Junction

NMJ: Area where the motor neuron

synapses on the muscle fiber

Depolarization at the axon terminal

causes opening of voltage-gated
Calcium channels

Calcium entry and binding to

vesicles promotes fusion and release
of Acetylcholine (ACh) into the
synaptic cleft

ACh binds to nicotinic ACh Receptors

(nAChR) on the muscle fiber, causing
them to open

ACh reuptake channels refill axon

terminal and end stimulation

2: The Motor End Plate

MEP: muscle fiber side of the NMJ

Acetylcholine binding to nAChR

opens its associated Na+ channel

Na+ entry causes a graded

depolarization (Excitatory Post
Synaptic Potential EPSP, or
Inhibitory IPSP)

If enough depolarization occurs to

reach threshold, nearby voltagegated Na+/K+ channels open,
triggering an action potential that
propagates along the muscle fiber

3: The T-Tubules

The T-Tubules or transverse tubules

are invaginations of the plasma
membrane

They function to bring action

potentials into the interior of
skeletal muscle fibers

This spreads action potentials across

a greater surface area of the cells.

4: The Sarcoplasmic Reticulum

Action potentials travel into the

T-Tubules alongside the
Sarcoplasmic Reticulum

At the boundary, paired

dihydropyridine (DHPR) and
Ryanodine (RyR) receptors sit
on the T-tubule and SR

Depolarization at the T-Tubule

opens the voltage gated DHPR,
which causes opening of the
mechanically paired RyR

Opening of RyR allows Ca2+

efflux out of the SR

5: Actin-Myosin Crossbridging

The troponintropomyosin complex

inhibits cross-bridge
formation

Ca2+ binds to troponin.

Ca-Troponin Complex
pulls tropomyosin off
the myosin binding site,
allowing
A cycle of cross-bridge
ATP
formation
hydrolysis ratchets

the myosin head along

actin to contract the
muscle fiber

The contraction
cycle is ended when
intracellular Ca2+ is
depleted and Ca2+
is no longer bound
to troponin

Laboratory Measurements

A twitch (small muscle contraction)

can be caused by a single action
potential or by direct stimulation

Increasing Voltage will cause

increased motor unit recruitment
and higher tension (Spatial
Summation)

At low Frequency tension will

appear as single unfused twitches

Higher Frequency will increase

tension as twitches begin to merge
(Temporal Summation)

Max tension will see complete

fusion (Tetanic Contraction)

AP: 1-2ms, twitch: 100ms

Part 1: Equipment Set-Up

Team A

Set-up computer and calibrate force transducer

Help Team B when done

Team B

Isolate gastrocnemius from frog and attach to transducer

Isolate sciatic nerve and connect to electrode

Notes:

Gloves are recommended when working on the frog. Do not touch

the computer with gloves or froggy hands!!!

Wash hands after touching the frog, esp. if you do not wear gloves!!!

Be very careful with the sciatic nerve do not over stretch it!!

Remember to F9 and make notes about voltages used!

Part 2a: Stimulus Intensity-Response

Summary:

Find Threshold and Vmax

Acquire approx. 10 data points ranging from Threshold to Vmax

Important Notes:

Maintain passive tension (20g) (Why is this important? Length-Tension

relationship!!)

Keep nerve/muscle moist with Ringers, keep rest of frog covered &
moist (Cutaneous respiration!!)

Make sure electrode isnt shorting (touching other tissue, too wet, etc)

Dont spend excessive time on this section

Expected Results?

Part 2b: Stimulus Frequency-Response

Summary

Use Vmax from Part 2a (how does amplitude continue to rise

if we are already at Vmax?)

Acquire a series of points changing freq from 0.5 to 25 pps

Important notes

As previous, maintain passive tension & moisture, check for

possible short in electrode

If there is a sudden dropout of tension in the higher

frequencies check that the muscle is still securely tied to
the transducer

Expected results?

Part 2c: Pharmacological Antagonism

(Tubocurare)

Summary

Use Vmax from Part 2a

Record the entire trial continuously

Acquire pre-TC baseline, inject TC, and record approx. 10 min

Important Notes

As previous, maintain passive tension & moisture, and check that

electrode is not shorting out on tissue

Note start and stop of injection (why?)

Expected Results?

Part 2d: Direct Stimulation

Summary

Evenly space electrodes about 1/3 from top & bottom of

muscle & poke firmly through muscle.

Begin at Vmax from Part 2a

Record entire trial without stopping

Find Direct Threshold, then record at 10x Direct Threshold

Important Notes

As previous, maintain passive tension and moisture

Expected Results?