Muscle Contraction
Background Information:
A skeletal muscle is a collection of muscle cells, or muscle fibers. Each muscle fiber is
composed of a cell membrane, cytosol, mitochondria, and a whole bunch of myofibrils (bundles
of contractile and elastic proteins that carry out the work of contraction). Each muscle cell also
contains an extensive sarcoplasmic reticulum for calcium storage, and release.
A Myofibril is comprised of several types of proteins: two contractile proteins, myosin and
actin, two regulatory proteins, troponin, and tropomyosin, and a few accessory proteins to boot.
The contraction of a muscle is a complex process, involving many ions, proteins, and certain
regulatory mechanisms. Myosin is motor protein that converts chemical bond energy from ATP
into mechanical energy of motion. Your textbook describes the molecular basis for contraction,
the power stroke, a six-step process very well (Please read it). Actin is associated with the
regulatory proteins that prevent the myosin heads fro m completing the power stroke.
Tropomyosin is a long polymer that wraps around the actin filament, directly blocking the
myosin head binding site, and inhibiting actin and myosin sliding. Ca++ inhibits troponin so
filament sliding can occur. Ca++ plus troponin, cause a tug on tropomyosin, pulling it deeper into
the groove between actin strands. This uncovers active sites, which react with cross bridges on
myosin strands causing the sliding process (power stroke). ATP breaks these reaction bonds and
forms new ones (power stroke), and is required for relaxation. You should be able to explain how
the following components work to gether to produce a controlled skeletal muscle contraction:
Actin, tropomyosin, troponin, myosin, myosin head (crossbridge), Ca++(salts) and ATP. The
control of contraction is summarized in Chapter 12 of your text for a full discussion.
Relaxation of the muscle occurs when the sarcoplasmic reticulum pumps Ca++ back into its
lumen using a Ca++ - ATPase. As the free cytosolic calcium decreases, Ca++ releases from
troponin, tropomyosin slides back to block the myosin binding site and the fiber relaxes.
The basic unit of contraction in an intact skeletal muscle is the motor unit. One motor unit is
comprised of one neuron and the muscle fibers it innervates. If the action potentials of a motor
unit continue to stimulate the muscle fiber repeatedly at short intervals (high frequency),
relaxation between contractions diminishes until a muscle fiber reaches a maximal level of
contraction called tetanus. Look at the figures describing summation of contractions leading to
maximum tension, and eventually fatigue.
The term fatigue describes a condition in which a muscle is no longer able to generate or
sustain the expected power output. Fatigue can be caused by a variety of factors, such as
depletion of neurotransmitter, ATP, oxygen, varying ion concentrations, and a build up of lactic
acid to name a few.
The purpose of today’s laboratory activities is to reinforce the concept that many chemicals
are required for skeletal muscle contraction to occur, and to gain a better understanding of the
process. In addition, you will have an opportunity to observe both skeletal muscle fatigue and
summation to tetanus.
Experimental Protocol:
Activity A
1.) In this activity, you will be using glycinerated skeletal muscle to witness contraction.
2.) Obtain a microscope, 3 slides, a ruler, 2 glass teasing needles.
3.) Carefully use the teasing needles to retrieve a fragment of muscle from the glycerine.
Take care to not touch the muscle fragment with your fingers.
4.) Using the glass teasing needles, tease the muscle fragment you obtained into separate
fibers. The goal is to get single muscle fibers (i.e., muscle cells), although this may be
difficult. Examine these fibers using the microscope, observing the striations of the
resting muscle.
5.) Place three single muscle fibers with a small amount of the glycerine on each of the 3
microscope slides. The glycerine simply ensures that the muscle fibers do not dry out.
6.) Label three of the slides with an A, B, or C.
7.) Arrange the fibers in a parallel fashion on the slide.
8.) Measure the length of each fiber by placing the slide on top of the ruler under the
microscope.
9.) Wick away the glycerine on the slides using a KimWipe. Rinse each of these fibers by
placing a drop or two of distilled water on the fibers.
10.) Wick away the water on slide A with another KimWipe.
11.) Flood the fibers with ATP. Immediately observe the fibers under the microscope.
12.) After 2 minutes, re- measure and record the length of the fibers.
13.) Wick away the water on slide B with another KimWipe.
14.) Flood the fibers on slide B with Salt solution (containing K+ and Mg++ ). Immediately
observe the fibers under the microscope.
15.) After 2 minutes, re- measure and record the length of the fibers.
16.) Wick away the water on slide C with another KimWipe.
17.) Flood the fibers with ATP + Salt solution. Immediately observe the fibers under the
microscope.
18.) After 2 minutes, re- measure a nd record the length of the fibers.
19.) Calculate the percent contraction for each of the nine fibers using the following formula:
% contraction = Initial length (mm) – Contracted length (mm)
100
Initial length (mm)
Activity B
1.) In this activity, you will directly observe the effects of muscle fatigue.
2.) Record the time as your laboratory partner stands facing you with his/her arms held
straight out, parallel with the floor, with palms facing upward toward the ceiling. Stop
recording when his/her arms begin to shake with fatigue. Determine the duration of time
it took until this fatigue was reached.
3.) Record the time as your laboratory partner repeats step 2, but this time, place a magazine
on his/her palms when you begin recording. Stop recording when your lab partner’s arms
begin to shake with fatigue, and determine the duration of time it took until this fatigue
was reached.
4.) Record the time as your lab partner repeats steps 2 and 3, but this time, place a text book
on his/her palms when you begin recording. Stop recording when his/her arms begin to
shake with fatigue. Determine the duration of time it took until this fatigue was reached.
5.) Weigh the magazine and the textbook.
6.) Based upon the results obtained, draw a hypothesis regarding the effect of ‘load’ (i.e.,
weight) on muscle fatigue.
Activity C
1.) In this activity, you will have the opportunity to measure skin-surface voltages associated
with skeletal muscle contraction through a procedure known as electromyography
(EMG). EMG will be used to measure the voltages associated with contractions of
increasing force.
2.) For this activity, it will be necessary to calibrate the BioPac for each individual.
3.) To calibrate the BioPac machine, attach three electrodes to your dominant forearm as
shown in the picture below. [If your lab partner timed YOU in Activity B, above, then it
will be necessary that your lab partner perform this Activity C].
4.) Open the BioPac program and select Lesson L01 (EMG-1).
5.) While sitting still, have your lab partner select “Calibrate”. This will begin the process
whereby the computer will automatically establish parameters to record the data properly.
6.) Follow the instructions given by the BioPac program. Clench your fist and relax as
instructed; the program will automatically stop the recording. The recording should look
similar to that in the picture below; if it does not, select “Redo Calibration” and repeat the
procedure.
7.) Have your lab partner select “Record ” and follow the instructions. Clench your fist
softly for 2 seconds, then relax for two seconds, then clench harder for two seconds,
then relax for two seconds, then clench even harder for two seconds, then relax for
two seconds, and finally clench with maximum strength for two seconds then relax.
When you have finished with all four ‘clench-relax’ cycles, have your lab partner
select “Suspend”.
8.) The result should be a series of four clenches of increasing intensity, as shown in the
figure below. If the data look very different, have your lab partner select “Redo” and
repeat step 7. If the data look similar, select “Stop”, followed by “Yes” (the latter in
response to the question, ‘Are you finished with both forearm recordings’), and
finally “Done”. Remove the electrodes from your forearm.
9.) Select “Analyze Current Data File ” and “OK” to analyze the results.
10.) Observe the Raw EMG recording and the computer calculated Integrated EMG. The raw
EMG is the actual voltage recordings (in mV) at each instant in time while the integrated
EMG reflects the absolute intensity of the voltage from baseline.
11.) To analyze the data, set up the first four pairs of the channel/measurement boxes at the
top of the screen by selecting the following channels and measurement types from the
drop down menus:
12.) Use the arrow cursor and click the ‘I-beam’ cursor box on the lower right side of the
screen to activate the ‘area selection’ function. Using the activated I-beam cursor,
highlight the first EMG cluster, representing the first clenching event.
13.) The computer will automatically calculate the minimum value, the maximum value, the
‘peak to peak’ value (p-p; the difference between min and max), and the mean (average
value) for the selected area.
14.) Record each of these values in the table below. Round to the nearest 1/100 mV.
15.) Repeat the analysis for each of the remaining 3 clench events, and record the data in the
table.
MIN
MAX
P-P
MEAN
Clench 1
Clench 2
Clench 3
Clench 4
Activity D
1.) In this activity, skeletal muscle contraction to the point of fatigue will be visualized
using EMG on the BioPac. Gross muscular contraction depends upon the gradual and
coordinated activation of multiple muscle cells—at minimum, all of those cells in a
single motor unit. In most cases, multiple motor units are recruited simultaneously. The
force of contraction of an entire muscle increases with the involvement of a larger
number of motor units, as well as with an increase in the frequency of nerve impulses
delivered to these motor units. In this way, the muscle can produce graded contractions.
For example, the biceps muscle will activate a greater number of motor units when
lifting a 10kg object than when it lifts a 2kg object. In addition, when lifting the heavier
object, the motor neuron of each active motor unit will increase its frequency of action
potentials delivered to the cells within that unit. This eventually results in a phenomenon
called tetanus, which is a smooth, sustained muscle contraction. When all of the motor
units within a muscle are activated to a state of tetanus, maximum force for that muscle
is achieved.
2.) For this activity, it will be necessary to calibrate the BioPac for each individual.
3.) To calibrate the BioPac machine, attach three electrodes to your dominant forearm as
shown in the picture above for activity C.
4.) Open the BioPac program and select Lesson L02 (EMG-2). Enter a file name that you
will remember.
5.) With the hand dynamometer at rest on the table, select “Calibrate”, then select “OK”.
6.) When instructed, grasp the hand dynamometer with your do minant hand as close to the
crossbar as possible without actually touching the crossbar. The crossbar should be held
parallel to the floor, not at an angle. When this is so, select “OK”.
7.) Follow the instructions given by the BioPac program. When instructed to do so, wait
two seconds, then squeeze the hand dynamometer as hard as possible for two seconds,
then relax. Continue to follow instructions until calibration is complete. The computer
will automatically stop the calibration. The calibration should look similar to that shown
in the picture below. If it does not, select “Redo Calibration” and repeat the procedure.
8.) Looking at the force level from the calibration procedure (the top part of the graph),
estimate the maximum force that was generated (in kg).
9.) Divide this maximum force by four. In the next part of the activity, gradually increase
the force of contraction by these increments. For example, if the maximum force
generated was 20kg, in increment will be 5 kg. Begin by squeezing at 5kg, then 10kg,
then 15kg, then 20kg. Watch the tracing on the computer to reach the target force.
10.) When you are ready to proceed, have your lab partner select “Record” and follow the
instructions. Wait two seconds, then clench at the first force (e.g., 5kg), then relax for
two seconds, then clench at the second force increment (e.g., 10kg) for 2 seconds, then
relax for two seconds then clench at the third force increment (e.g., 15kg) for two
seconds then relax for two seconds, then finally clench at maximum force (e.g., 20kg)
for two seconds then relax. When this process is complete, have your lab partner select
“Suspend ” to stop the recording.
11.) The result should be a series of four clenches of increasing intensity, as shown in the
figure below. If the data look very different, have your lab partner select “Redo” and
repeat step 10. If the data look similar, proceed to step 12.
12.) Have your lab partner select “Resume” to continue recording. Clench the dynamometer
with maximum force for as long as possible. A triangular shaped “marker” will appear at
the top of the data, denoting the beginning of this recording segment. When the force
falls below 50% of maximum (e.g., 10kg for an individual with a maximum force of
20kg), your lab partner should select “Suspend ” to stop the recording. He/she should
inform you when this point is reached (you should not be looking at the computer), so
that you can relax.
13.) The recording obtained should look similar to that shown in the picture below. If it looks
very different, have your lab partner select “Redo ”, and repeat step 12. If the image
looks similar, have your lab partner select “Stop” and “Yes ”. Your data will be saved as
the filename you entered in step 4 above with an added extension of ‘1-L02’.
14.) Remove the electrodes from your dominant forearm.
15.) Place the electrodes on your non-dominant forearm in a pattern identical to that which
you used in step 3 above. Have your lab partner select “Forearm 2”.
16.) Follow the calibration instructions, which should be similar to those outlined in steps 6
& 7 above. Data obtained from the non-dominant forearm will be saved with the
filename you entered in step 4, but with the extension ‘2-L02’ added.
17.) Repeat steps 8-14 using the non-dominant arm to clench the hand dynamometer.
18.) Select “Review Saved Data” from the file menu, and select the file to be analyzed (from
your dominant forearm).
19.) Observe the Raw EMG recording and the computer calculated Integrated EMG. The raw
EMG is the actual voltage recordings (in mV) at each instant in time while the integrated
EMG reflects the absolute intensity of the voltage from baseline.
20.) To analyze the data, set up the first three pairs of the channel/measurement boxes at the
top of the screen by selecting the following channels and measurement types from the
drop down menus:
21.) Use the arrow cursor and click the ‘I-beam’ cursor box on the lower right side of the
screen to activate the ‘area selection’ function. Using the activated I-beam cursor,
highlight the ‘plateau phase’ of the first clench cluster. The ‘plateau phase’ should be the
relatively flat force in the cluster.
22.) The computer will automatically calculate the ‘peak to peak’ value (p-p; the difference
between min and max), and the mean (average value) for the selected area.
23.) Record each of these values in the table below. Round to the nearest 1/100 mV.
24.) Using the I-beam cursor, highlight the plateau phase of the three subsequent clench
clusters. Record each of these values in the table below.
Dominant Forearm Clench Increments
FORCE AT PLATEAU
MEAN (KG)
Clench 1
Clench 2
Clench 3
Clench 4
RAW EMG
P-P (MV)
INTEGRATED EMG
MEAN (MV)
25.) Scroll along the bottom of the data page to the segment that includes the recording of
muscle fatigue (it should begin after the “marker” that appears at the top of the data.
26.) Change the channel/measurement boxes so that the first two
selected appear as follows (the third should be set to ‘none’):
27.) ‘Value’ measures the highest value in the selected area (in kg) while ‘delta T’ measures
the time lapsed in the selected area (in seconds).
28.) Use the arrow cursor and click the ‘I-beam’ cursor box on the lower right side of the
screen to activate the ‘area selection’ function. Using the activated I-beam cursor, select
just the single point of maximum clench strength at the beginning of this data segment
as shown in the figure below. Record this maximum clench force in the table below.
29.) Calculate the ½- maximal clench force and record it in the table below.
30.) Using a tape measurer, measure the maximum circumference of the dominant forearm
(at its greatest diameter) and record it in the table below.
31.) Measure the amount of time that elapsed between the initial maximum force and the
point at which you fatigued to 50% of this level. To do this, use the activated I-beam
cursor to highlight the area from the point of 50% clench force back to the point of
maximal clench force as shown in the figure below. Record this in the table below.
Dominant Forearm Fatigue Measurement
MAXIMUM 50% OF MAXIMUM TIME TO FATIGUE FOREARM
CLENCH
CLENCH FORCE
(SEC)
CIRCUMFERENCE
FORCE (KG)
(CM)
32.) Select ‘Review Saved Data’ from the file menu and select the filename entered for the
non-dominant forearm (see step 16 above).
33.) Repeat steps 18-31 for your non-dominant forearm. Record your results in the tables
below.
Non-Dominant Forearm Clench Increments
FORCE AT PLATEAU RAW EMG
MEAN (KG)
P-P (MV)
Clench 1
Clench 2
Clench 3
Clench 4
INTEGRATED EMG
MEAN (MV)
Non-Dominant Forearm Fatigue Measurement
MAXIMUM 50% OF MAXIMUM TIME TO FATIGUE FOREARM
CLENCH
CLENCH FORCE
(SEC)
CIRCUMFERENCE
FORCE (KG)
(CM)