Summary
LAB
EXER
CISE:
Electrical Stimulation of
Skeletal Muscle
We use our skeletal muscles to move our bodies, maintain posture, stabilize joints, and
generate heat. We usually control skeletal muscle consciously; however, stretch
reflexes can control our skeletal muscle subconsciously. We must understand how
skeletal muscles respond to stimuli in order to understand how our nervous system and
our skeletal muscle functions as a unit. Today you will explore how your skeletal
muscles function by electrically stimulating muscles found in the arm.
Let’s review our skeletal muscle anatomy. Skeletal muscle is highly organized and
follows the following morphology: 1) Skeletal muscle is striated and is composed of
muscle fibers (cells), 2) These muscle fibers contain myofibrils, 3) muscle fibers are
grouped together to form muscle fascicles, 4) muscle fascicles are grouped together for
form muscle bundles, 5) the functional unit is the sarcomere, 6) myosin (thick) and
actin (thin) filaments are highly organized which results in striations, 7) the
neuromuscular junction is the site where a motor neuron communicates with a skeletal
muscle fiber.
Let’s review our skeletal muscle physiology.
Neuromuscular Junction Events:
The central nervous system (CNS) and skeletal muscle communicate to cause skeletal
muscles to contract. Motor neurons receive an action potential from the CNS. When
the action potential (AP) reaches the synaptic bulb, voltage gated calcium channels
open and calcium ions diffuse into the synaptic bulb to trigger exocytosis of the
neurotransmitter acetylcholine (ACh). ACh diffuses across the synaptic cleft and binds
to the ACh-gated channels on the motor end plate (MEP) of the sarcolemma (plasma
membrane of the muscle fiber). The ACh-gated channels open allowing sodium ions
and potassium ions to diffuse across the MEP. A rush of sodium ions crosses into the
muscle fiber and depolarizes the membrane resulting in an end-plate potential (EPP).
Excitation-Contraction Coupling:
If the EPP reaches threshold, then the action potential travels across the sarcolemma
and into the transverse tubules (t-tubules). The voltage-sensing receptors (L-type
calcium channels called dihydropyridine receptor, DHP) on the t-tubule change
conformation and open the calcium ion release channels (ryanodine receptors, RyR)
that are on the sarcoplasmic reticulum (SR; the smooth ER of the muscle cell). Calcium
ions diffuse into the cytosol of the muscle fiber and bind to troponin. Troponin changes
conformation and moves tropomyosin away from the myosin-binding site on actin.
Cross-Bridge Cycling:
Myosin and actin can now interact to form cross-bridges. The steps in the cross-bridge
cycle are as follows. 1) Myosin and actin are attached. The myosin head will bind an
ATP molecule on the ATP binding site. This will cause the myosin head to change
shape (configuration) and detach (dissociate) from actin. 2) ATP is hydrolyzed into ADP
and Pi. This will cause the myosin head to change shape and “cock” into position (the
angle between the head and tail increases when cocked). 3) The cocked head then
binds to the myosin binding site on actin and forms a “cross-bridge” (physical
connection between actin and myosin). 4) The Pi is released from the myosin head
causing it to change shape. The change in shape causes a “power stroke” which slides
the filaments past each other shortening the sarcomere. 5) The ADP is released from
the myosin head. These five steps will repeat until the signal for contraction is
removed.
Relaxation:
Cross-bridge cycling will continue until ACh is removed from the synaptic cleft by
acetylcholinesterase (AChE) or via diffusion. Once ACh is removed, ACh-gated
channels on the MEP close resulting in a loss of the AP on the muscle fiber. The SR
then closes its calcium ion channels and the calcium ion pumps return the calcium into
the SR. These events cause the skeletal muscle to relax and lessen the force that it
produces.
Force production is dictated by many factors and processes. The number and type of
motor units that are recruited affects how much force is produced. The resting position
of the muscle affects the force that can be produced. The frequency, duration, and
voltage of the stimulus from the stimulation machine will affect the outcome of the
contraction.
Motor units & Recruitment:
Motor units are composed of somatic motor neuron(s) and all the muscle fibers they
innervate. Motor units range from small to large. The smaller motor units have a lower
threshold while the larger motor units have a higher threshold. The smaller motor units
tend to be composed of Type I (slow twitch) fibers, medium motor units tend to be
composed of Type IIa (fast-oxidative; fast fatigue resistant) fibers, and large motor units
tend to be composed of Type IIb (Type IIx; fast glycolytic; fast fatigable) fibers. while
the larger motor units have a higher threshold. During a physiological contraction, you
will recruit different motor units at different times to produce the force that you require;
this is termed asynchronous recruitment and it helps prevent fatigue.
Length Tension Curve:
A muscle fiber twitch is affected by the length of the muscle. Muscles at optimal length
produce optimal force because there is an optimal amount of cross bridges being
formed. Muscles that are shortened produce less than optimal force because the
myosin filaments run into the Z disks inhibiting them from sliding and produce less
tension. Muscles that are too long produce less than optimal force because there is not
enough overlap for optimal cross bridging resulting in less sliding and less force
production.
Twitch, Summation, Unfused Tetanus, Fused Tetanus:
Single twitches, summation, unfused tetanus (incomplete tetanus) and fused tetanus
(complete tetanus) are all different ways to produce tension.
A twitch is when a single action potential (voltage) causes contraction and full
relaxation of the muscle. During twitching we see full relaxation between stimuli. The
tension (force) a twitch creates can vary depending on the voltage of the action
potential. The duration of a twitch can vary depending on the duration of the action
potential.
Summation is when the frequency of the action potential is increased, and full relaxation
cannot be reached. During summation we reach submaximal force and we see partial
relaxation between stimuli. Therefore, each subsequent action potential creates a
larger tension.
If frequency increases even more, then summation may reach maximal tension. This is
when incomplete tetanus (unfused tetanus) has been reached. Incomplete tetanus
(unfused tetanus) is when there is partial relaxation at maximal force (shaky muscle
contraction).
If frequency increases even more, then incomplete tetanus (unfused tetanus) will
become complete tetanus (fused tetanus). Fused tetanus is when there is no
relaxation at maximal force (smooth muscle contraction).
Today you will experiment with a stimulation machine (Stim machine) on a volunteer to
study twitches, summation, incomplete tetanus and complete tetanus.
Goals
Physiology students will learn about the physiology of skeletal muscle by studying the
electrophysiology of skeletal muscles in the arm. Students will change variables and
see how it affects skeletal muscle contraction patterns.
● Review the anatomy and physiology of skeletal muscle.
● Understand the processes that occur at the neuromuscular junction,
during excitation-contraction coupling, cross-bridge cycling, and relaxation.
● Understand the concepts of: a twitch, summation, unfused tetanus
(incomplete tetanus), and fused tetanus (complete tetanus). Identify them as a
graphical representation.
o Understand the relationship between stimulus voltage and
tension produced.
o Understand the relationship between stimulus frequency and tension
produced.
o Understand the relationship between stimulus duration and
tension produced.
● Understand the concepts of: motor unit, recruitment, and the
length-tension curve.
Lab Exercises:
HINT: Do not participate in the simulator experiment if you have a
pacemaker or other electronic device.
HINT: Remove jewelry and watches before beginning the experiment.
HINT: Work in groups of 8.
Stimulator Experiment (Electrode Stimulation Experiment)
Equipment:
● Follow directions and ask
for guidance when
necessary.
● Designate One Volunteer that
you will use for the entire
protocol (communicate often
and ask for consent often)
● Alcohol swabs
● Electrodes
● Elastic bands
● Electrolyte gel (salt aids
in electrical stimulation)
● Stimulator with variable
stimulation settings:
○ Duration (msec) is the
length of time that you are
sending a stimulus.
○ Voltage (V) is the strength
of the stimulus that you
are sending.
○ Frequency (Hz) is the
number of times you are
sending a stimulus in one
second (On the Stim, Hz is
labeled as events per
second or pulse per
second)
Protocol:
1. Pick an arm on your volunteer. Choose one muscle group from the list below to
study for this experiment and locate the motor points where electrodes should
be placed:
i. Isolated finger flexion. This video is a great resource.
ii.
Lumbrical grip. This video is a great resource.
iii. Curious? In your free time checkout this playlist to explore
other electrode placements for muscle groups.
2. Place a drop of electrolyte gel on each motor point and rub it on the area of the
skin where you will place the electrodes. Remove excess gel by wiping it off
with a paper towel.
3. Apply a thin layer of electrolyte gel on each electrode and spread it
evenly. Remove excess gel by wiping it off with a paper towel.
4. Place the electrodes on the motor points and secure them with the elastic bands.
5. Initial Twitch Experiment:
a. Set the Stimulator Machine to:
i. Voltage = 20 V
ii.
Duration of the stimulus = 2 msec
iii.
Frequency = single pulse
b. Look at the fingers and deliver a stimulus by pressing “Single” stimulus.
c. Determine if a twitch was seen.
i. If so, then this is your threshold setting. Move on to “Step 5-f”.
ii.
If not, then continue to the next step.
d. Change the Voltage to 30 V and deliver a stimulus by pressing “Single”
stimulus.
e. Determine if a twitch was seen.
i. If so, then congratulations you found the proper motor points.
Move on to “Step 5-f”. Do not move the electrodes for the rest of
this experiment.
ii.
If not, adjust the electrode placement and repeat this
experiment until you see a twitch. You may need to increase the
voltage on folks with larger muscles. Ask for support if needed.
f. At which voltage did you see a twitch? 70
V
6. Twitch Threshold Experiment:
a. Begin by using the settings that you determined in the Initial
Twitch Experiment.
i. Set the Stimulator Machine to:
1. Voltage = 70
V (taken from Step 5-f)
2. Duration of the stimulus = 2 msec
3. Frequency = single pulse (1 Hz; 1 pulse per second)
b. For each test, lower the voltage in a stepwise manner until you
determine the threshold. Look at the fingers and deliver a stimulus
by pressing “Single” stimulus.
c. Determine the lowest voltage needed to stimulate a twitch.
i. What is the lowest voltage needed for you to see a twitch?
70
V
ii.
When stimulating your volunteer using the Twitch Threshold
settings, you are recruiting the SMALL
[small /
medium / large] motor units.
7. Twitch Voltage Experiment:
a. Begin by using the settings that you determined in the Twitch
Threshold Experiment.
i. Set the Stimulator Machine to:
1. Voltage = 75
V (taken from Step 5-f)
2. Duration of the stimulus = 2 msec
3. Frequency = single pulse (1 Hz)
b. Increase the voltage by 5 V and deliver a stimulus by pressing “Single”
stimulus.
c. Repeat the previous step
i. You should not surpass 30 V until you chat with your professor
to get approval.
ii.
If your volunteer does not want to proceed to a higher voltage,
then you should move on to the duration experiment.
d. Determine how the twitch changes as voltage changes.
i. Increasing voltage will result in a(n) INCREASE
[decrease / increase / no change] in twitch force production.
ii. Decreasing voltage will result in a(n) DECREASE
[decrease / increase / no change] in twitch force production.
iii. Increasing voltage will result in a(n) INCREASE
[decrease / increase / no change] in twitch duration.
iv. Decreasing voltage will result in a(n) DECREASE
[decrease / increase / no change] in twitch duration.
v.
Increasing voltage will result in recruiting LARGER
[smaller / larger] motor units.
vi. Decreasing voltage will result in recruiting SMALLER
[smaller / larger] motor units.
vii.
If you stimulate your volunteer with a relatively small voltage,
then you are recruiting the
SMALL
[small / medium
/ large] motor units.
viii. If you stimulate your volunteer with a relatively medium
voltage, then you are recruiting the
SMALL AND MEDIUM
[medium / small
& medium / medium & large] motor units.
ix. If you stimulate your volunteer with a relatively large voltage, then
you are recruiting the
SMALL MEDIUM AND LARGE
[large / medium
& large / small, medium, & large] motor units.
x.
xi.
xii.
If you stimulate your volunteer with a relatively small voltage,
then you are recruiting the
SLOW OXIDATIVE
[slow / fast
oxidative / fast glycolytic] motor units.
If you stimulate your volunteer with a relatively medium
voltage, then you are recruiting the
SLOW AND FAST OXIDATIVE
[fast oxidative /
slow & fast oxidative / fast oxidative & fast glycolytic] motor units.
If you stimulate your volunteer with a relatively large voltage, then
you are recruiting the
SLOW AND FAST OXIDATIVE, AND FAST GLYCOLYTIC [fast
glycolytic / fast oxidative & fast glycolytic / slow, fast oxidative, &
fast glycolytic] motor units.
8. Twitch Duration Experiment:
a. Begin by using the settings that you determined in the Twitch
Threshold Experiment.
i. Set the Stimulator Machine to:
1. Voltage = 70
V (taken from Step 5-f)
2. Duration of the stimulus = 2 msec
3. Frequency = single pulse (1 Hz)
b. Lower the duration to 1 msec and deliver a stimulus by pressing
“Single” stimulus.
c. Increase the duration to 5 msec and deliver a stimulus by pressing
“Single” stimulus.
d. Determine how the twitch changes as duration changes.
i. Increasing duration will result in a(n) INCREASE
[decrease / increase / no change] in twitch force production.
ii. Decreasing duration will result in a(n) DECREASING
[decrease / increase / no change] in twitch force production.
iii. Increasing duration will result in a(n) DECREASE
[decrease / increase / no change] in twitch duration.
iv. Decreasing duration will result in a(n) INCREASE
[decrease / increase / no change] in twitch duration.
9. Frequency Experiment (summation, incomplete tetanus, complete tetanus):
a. Begin by using the settings that you determined in the Twitch
Threshold Experiment.
i. Set the Stimulator Machine to:
1. Voltage = 70
V (taken from Step 5-f)
2. Duration of the stimulus = 2 msec
3. Frequency = single pulse (1 Hz)
b. Increase the frequency in a stepwise manner until summation occurs.
Look at the fingers and deliver a stimulus by pressing “Continuous”
stimulation.
i. You saw summation at 6.5
Hz.
ii.
Describe how the fingers contract:
The middle figure underwent rapid twitching while the other fingers all began
to contract slower
c. Continue to increase the frequency in a stepwise manner until
incomplete (unfused) tetanus occurs. Look at the fingers and deliver a
stimulus by pressing “Continuous” stimulation.
i. You saw incomplete (unfused) tetanus at 11
Hz.
ii. Does incomplete (unfused) tetanus reach maximal force? NO
[yes / no]
iii.
Describe how the fingers contract:
they blend into almost one singular contraction, happening very quickly with
little to no relaxation period.
d. Continue to increase the frequency in a stepwise manner until complete
(fused) tetanus occurs. Look at the fingers and deliver a stimulus by
pressing “Continuous” stimulation.
i. You saw complete (fused) tetanus at 13
Hz.
ii. Does complete (fused) tetanus reach maximal force? yes [yes/no]
iii. Describe how the fingers contract:
They completely folded inward, causing complete flexion, and
my wrist began to flex forward as the contraction reached its
peak.
10. Clean-Up:
a. Turn off the stimulator machine and unhook your volunteer. Set all
the controls to their lowest setting.
b. Use a paper towel to clean off the electrodes. Dispose in regular trash.
c. Use an alcohol swab to clean the electrodes. Dispose in regular trash.
d. Clean your lab bench.
e. Wash your hands. Wash your arm if needed.