Course Code BIOM1051

Course Title Introductory Cellular Physiology

Course Coordinator Dr Saleem Babri
Due Date 19/04/24
Assignment Title Skeletal Muscle Practical Report
Word Count 3685
Date Submitted 19/04/24

Extension applied for Yes / No X Revised Date _

Student Number Surname First Name

47601475 Cheah Qiao Ren

 SKELETAL MUSCLE

Case Study
As a physiotherapist at the Royal Brisbane and Women’s hospital, you are treating a patient with chronic
muscle atrophy, suffered as a result of being bedridden for 2 months following an operation. A vital aspect of
your role in the patient’s recovery is to improve their coordination and muscle strength and explain how this
can be achieved to the patient.

EXPERIMENT 1
Hypothesis 1
It is hypothesised that as the stimulus strength (V) applied to R. Marina gastrocnemius muscle increases, the peak
contractile force (mN) increases.

Prediction of Results for Experiment 1

2.5
Predicted peak of contractile force (mN)

1.5

0.5

0
0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80 0.90 1.00

Stimulus Strength (V)

Figure 1: Predicted effect of increased stimulus strength (V) on the peak contractile force (mN) in Rhinella marina
gastrocnemius muscle. Plots represent theoretical data if hypothesis is confirmed (red) and if the response is unaffected
(blue).

Results for Experiment 1

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 SKELETAL MUSCLE

2000.00
1800.00
1600.00
1400.00
1200.00
1000.00
Peak contractile force (mN)

800.00
600.00
400.00
200.00
0.00
0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80 0.90 1.00
Stimus Strength (V)

Figure 2:
The effect of increasing stimulus strength (V) on the peak contractile force (mN) in R. Marina gastrocnemius muscle. Graph

Comparative Analysis
Complete Table 1 using data from your own experiment and the experiment completed by two other groups. Provide an
informative Table title in the space at the top of the table.

Table 1:
The maximum contractile force generated (mN) and minimum stimulus required for near maximum contraction (V) in R. M

Minimum stimulus required for

Muscle Sample Maximum contractile force generated (mN)
near maximum contraction (V)
Your own 1877 0.2
Alternative 1 1859 0.1
Alternative 2 1793 0.1

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 SKELETAL MUSCLE

EXPERIMENT 2
Hypothesis 2
It is hypothesised that as the stimulus frequency (Hz) increases for the R. Marina gastrocnemius muscle, the peak
contractile force (mN) increases.

Materials and Methods 2

The sciatic nerve connected to the R marina gastrocnemius muscle was prepared through dissection from the R marina
frog leg. The dissected sciatic nerve was placed into the nerve bath, and the gastrocnemius muscle was attached to the
nerve bath at one end, as well as to the force transducer at the other end through string. Stimulating electrodes were
connected from the Powerlab to the nerve bath. A constant passive force in the range of 200-250mN was established
in the gastrocnemius muscle using the force transducer. A constant stimulus strength of 0.2V was generated in the
sciatic nerve through the stimulating electrodes, where 0.2V is the minimum stimulus required for near maximum
contraction in the gastrocnemius muscle. For each stimulus frequency, 3 replicates of 5 pulses were used, with the
same number of pulses used for each replicate. A total of 13 different stimulus frequencies were used, 1Hz, 2Hz, 3Hz,
4Hz, 5Hz, 6Hz, 7Hz, 8Hz, 9Hz, 10Hz, 15Hz, 20Hz and 25Hz. The resulting mean peak contractile force from 3
replicates of each stimulus frequency was recorded. Each peak contractile force was measured as the difference
between the last distinct peak of the contractile force trace and the baseline passive force right before that peak on
LabChart, for each reptition of each stimulus frequency. The gastrocnemius muscle was initially stimulated at a
stimulus frequency of 1Hz as the negative control. A plot of peak contractile force against stimulus frequency used
was generated for analysis.

Prediction of Results for Experiment 2

N.B. you will need to adjust the x axis range to display the values of your predicted independent variables. Double-click on the x axis to view the
formatting axis options, and enter the necessary maximum range value in the “Bounds” field of the Axis options.

1.2

0.8

0.6
Predicted Peak Contractile Force (mN)

0.4

0.2

0
1 6 11 16 21 26

Stimulus Frequency (Hz)

Figure 3:
Predicted effect of increased stimulus frequency (Hz) on the peak contractile force (mN) in R. Marina gastrocnemius muscle. P

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 SKELETAL MUSCLE

Results for Experiment 2

N.B. you will need to adjust the x axis range to display the values of your independent variable. Double-click on the x axis to view the formatting axis
options, and enter the necessary maximum range value in the “Bounds” field of the Axis options.

3400.00
3200.00
3000.00
2800.00
2600.00
2400.00
Peak Contractile Force (mN)

2200.00
2000.00
1800.00
1600.00
1400.00
0.00 5.00 10.00 15.00 20.00 25.00

Stimulus Frequency (Hz)

Figure 4:
The effect of increasing stimulus frequency (Hz) on the peak contractile force (mN) in R. marina gastrocnemius muscle. Gra

Description of Findings for Experiment 2

Describe the important trends demonstrated from your experiment, using data to illustrate and support these outcomes.
In general for Figure 4, as the stimulus frequency increased from 1Hz to 25 Hz, the mean peak contractile force in the
gastrocnemius muscle increased from 1880.00mN to 3233.67mN. Contrary to this trend, there was an initial decrease
in the mean peak contractile force in the gastrocnemius muscle with 1880.00mN, 1710.33mN, 1635.67mN and
1582.00mN for 1Hz, 2Hz, 3Hz and 4Hz respectively, with 1582.00mN at 4Hz being the overall minimum mean peak
contractile force. The mean peak contractile force also plateaued towards the end, with a noticeable plateau towards
the maximum mean peak contractile force occurring with 3198.67mN and 3233.67mN for 20Hz and 25Hz
respectively. The overall maximum mean peak contractile force was 3233.67mN at 25Hz.

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 SKELETAL MUSCLE

Discussion
Throughout your answers to the following questions, focus on being specific, clear and concise, whilst also being
biologically detailed. See the Guide to Report Criteria for more tips on meeting the criteria standards in this section
such as using Results as Evidence, and References.

1. Briefly describe whether the results of your first experiment confirm or disconfirm your hypothesis.
The results of the first experiment confirms the hypothesis, as in general in Figure 2, as the stimulus strength increased
from 0V to 1V, the mean peak contractile force increased from 0mN to 1848.00mN. This is despite a minor decrease
in the mean peak contractile force towards the end, from 1877.67mN to 1848.00mN for the corresponding stimulus
strengths of 0.5V to 1.00V.

2. Why does peak contractile force change with different stimulus strengths? Discuss the biological mechanisms
underpinning your outcomes in Experiment 1.
In Experiment 1, Figure 2, in general as the stimulus strength increased from 0V to 1V, the mean peak contractile
force increased from 0mN to 1848.00mN. A motor unit consists of the motor neuron originating from the spinal cord,
its axon to the muscle and the multiple muscle fibres it innervates through neuromuscular junctions (1). The
gastrocnemius muscle and sciatic nerve consists of many motor units. When a stimulus such as the depolarising
stimulus of the stimulating electrodes is applied, given the depolarising stimulus goes above a certain threshold, this
causes an action potential to fire in the sciatic nerve. An action potential is an all-or-nothing rapid depolarisation
followed by repolarisation of the neuron initiated by sodium ion channels within the plasma membrane, followed by
potassium ion channels respectively (2). The action potential propagates down the sciatic nerve, releasing
acetylcholine neurotransmitters at the neuromuscular junction to the muscle fibres of the motor unit (3). This action
potential travels along the t-tubules of the straited muscle fibre to the sarcoplasmic reticulum, where calcium ions are
released which change the conformation of troponin complexes and tropomyosin (4). This allows for the cyclic
interaction of myosin heads with actin filaments through a power stroke, where conformational changes in myosin
cause the sliding of myosin filaments relative to actin filaments, leading to compression of sarcomere length and
overall muscle contraction, generating a contractile force (5). As the stimulus strength increases, more of these motor
units are recruited as action potentials fire in them. Henneman’s Size Principle relates the input and output properties
of motoneurons and their muscle fibres to size, and is the basis for size-ordered activation or recruitment of motor
units during movement (6). There is a distribution of nerve cells in the sciatic nerve fibre, where nerve cells leading to
Type I slow twitch fibres are smaller, and thus require less of a depolarising stimuli for an action potential to fire in
the nerve cell, which would lead to the muscle fibre contracting in the form of muscle twitches (7).This is due to the
same depolarising stimuli leading to a greater degree of change in the sodium concentration gradient for smaller nerve
cells, and thus a greater frequency of firing action potentials. Type IIA twitch fibres have medium sized nerve cells,
requiring more depolarising stimuli for an action potential to fire and the muscle fibre to contract (7). Type IIB twitch
fibres have large sized nerve cells, requiring the greatest depolarising stimuli for an action potential to fire and the
muscle fibre to contract (7). This leads to a distribution where increasing the depolarising stimuli strength in the sciatic
nerve will increase the proportion of muscle fibres contracting due to having action potentials firing in them for the
gastrocnemius muscle. Hence, as the stimulus strength increases, the peak contractile force in the gastrocnemius
muscle increases, due to the summation of a greater proportion of muscle fibres producing contractile forces.
In Experiment 1, Figure 2, upon reaching the near maximum mean peak contractile force of 1877mN for a stimulus
strength of 0.2V, the mean peak contractile force plateaus from 1707.00mN to 1848.00mN for a stimulus strength of
0.25V to 1.00V respectively. This is due to most of the muscle fibres in the gastrocnemius muscle already being
recruited and producing contractile forces due to having action potentials fire in them. This causes the peak contractile
force that the gastrocnemius muscle can produce to approach a maximum.

3. Many biological factors can affect the function of a muscle, some of which you have learnt about. Discuss the
biological reasons for the differences observed in maximal contractile force and minimum stimulus strength required to
generate that contractile force, between muscles in the comparative analysis.

The frog may have had a genetic abnormality in misregulation of the FRG1 gene, leading to Facioscapulohumeral

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 SKELETAL MUSCLE

muscular dystrophy, which causes progressive muscle atrophy (8). Muscle atrophy refers to a removal of contractile
proteins such as actin and myosin filaments, leading to a shrinkage of muscle fibres (9). Muscle physiological cross-
sectional area is defined as the volume of a muscle divided by its fibre length, and is proportional to the maximum
strength or contractile force of the muscle (10). This decrease in muscle volume from muscular atrophy in the frog leg
thus leads to a decrease in maximum strength or contractile force of the muscle. Hence, in Experiment 1, Table 1,
there would be a decrease in maximum contractile force generated from the gastrocnemius muscle from 1877mN in
our group, to 1859mN in alternative group 1 and 1793mN in alternative group 2, due to progressively stronger forms
of Facioscapulohumeral muscular dystrophy and muscle atrophy in the R. Marina frog leg.
The age of the frog can affect the relative muscle fibre compositions in the frog. For instance, there tends to be an
increased percentage of type I slow twitch fibres among the female population as they age (11). Assuming this trend
applies to the frog sample from alternative group 1 and 2, and assuming those frog samples came from an older female
frog, that means there would be a higher composition of type I slow twitch fibres in their gastrocnemius muscle. Type
I slow twitch fibres are smaller, and thus require less of a depolarising stimuli for an action potential to fire in the
nerve cell, which would lead to the muscle fibre contracting in the form of muscle twitches (12). Thus, a greater
proportion of type I slow twitch fibres leads to less depolarising stimuli being needed on average to recruit all of the
gastrocnemius muscle fibres, due to the distribution of individual depolarising stimulus required to recruit the muscle
fibres skewing towards the smaller type I slow twitch fibres with lower depolarising stimulus required. Hence, the
minimum stimulus required for near maximum contraction decreases from 0.2V in our group to 0.1V in both
alternative group 1 and 2.

4. Briefly describe whether the results of your second experiment confirm or disconfirm your hypothesis.
The results of the second experiment confirms the hypothesis, as in Experiment 2 Figure 4, in general as the stimulus
frequency increased from 1Hz to 25Hz, the mean peak contractile force increased from 1880.00mN to 3233.67mN.
This is despite an initial slight decrease in the mean peak contractile force of 1880.00mN, 1710.33mN, 1635.67mN
and 1582.00mN for the corresponding stimulus frequencies of 1Hz, 2Hz, 3Hz and 4Hz.

5. Discuss the biological mechanisms underpinning these outcomes.

In Experiment 2 Figure 4, in general as the stimulus frequency increased from 1Hz to 25Hz, the mean peak contractile
force increased from 1880.00mN to 3233.67mN. As the stimulus frequency increases, the time interval decreases
between repeated depolarising stimuli leading to repeated all-or-nothing action potentials in the sciatic nerve (2). This
refiring of the action potential occurs as long as the inter-pulse interval between the two action potentials is longer
than the absolute refractory period of the motor axon (13), within which any stimuli will not cause the nerve to fire an
action potential due to inactivation of sodium ion voltage-gated channels (14). With smaller time intervals in between
action potentials in the sciatic nerve, this leads to increased temporal summation of action potentials as the action
potentials increasingly superpose. This leads to a higher summated compound action potential being generated in the
muscle fibre through transmission of the sciatic nerve action potential in the neuromuscular junction. Hence, there is a
greater extent of summation of twitches, where a greater proportion of muscle fibres are activated by the same
stimulus. These muscle twitches end up merging into higher force contractions with a higher peak contractile force
(15). Hence, as the stimulus frequency increased in Experiment 2, the mean peak contractile force increased.

In Experiment 2 Figure 4, there was an initial slight decrease in the mean peak contractile force of 1880.00mN,
1710.33mN, 1635.67mN and 1582.00mN for the corresponding stimulus frequencies of 1Hz, 2Hz, 3Hz and 4Hz. This
may be due to the effects of temporal summation being negligible, as the time interval between action potentials and
muscle twitches is still too long to allow for significant summation of action potentials leading to significant
summation of muscle twitches. Hence, the 5 individual contractile force peaks in each replicate of a given stimulus
frequency only depend on the individual compound action potentials which caused them. This explains why the mean
peak contractile force would not increase in this range of stimulus frequencies. The decrease in mean peak contractile
force may be due to other external factors in the environment, such as small fluctuations in the positioning of the
sciatic nerve or the drying up of the sciatic nerve as its exposed to the external environment, which may lead to nerve
damage and loss of the capacity of the sciatic nerve to transfer signals, leading to lower action potentials and resulting
muscle twitches.

In Experiment 2 Figure 4, there was a plateau in the mean peak contractile force of 3198.67mN and 3233.67mN for
the respective stimulus frequencies of 20Hz to 25Hz towards the maximum peak contractile force of 3233.67mN and

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 SKELETAL MUSCLE

the maximum stimulus frequency used of 25Hz. As the stimulus frequency increases, the time interval between action
potentials from depolarising stimuli decreases. The refiring of the action potential may not occur towards very high
frequencies such as 20Hz and 25Hz, as the inter-pulse interval between the two action potentials is not longer than the
absolute refractory period of the motor axon (13), where any stimuli within this time interval will not cause the nerve
to fire an action potential due to inactivation of sodium ion voltage-gated channels (14). Hence, there is a minimum
time interval for temporal summation of action potentials leading to temporal summation of muscle twitches in the
innervated muscle fibres to be achieved (15), within which there will be no second action potential firing. This leads to
a maximum possible extent of temporal summation of muscle twitches and maximum possible peak contractile force,
within which a greater extent of second action potentials will not fire as they are within the absolute refractory period
of the first action potential, and a plateau in the mean peak contractile force in Experiment 2.

6. In the case study, you are working on muscle atrophy. Discuss:

a) what happens in atrophy and how this affects muscle activity/performance, and
b) how muscle composition and mass can increase upon recovery from atrophy.
(A)
Muscle atrophy refers to a removal of contractile proteins such as actin and myosin filaments, leading to a shrinkage
of muscle fibres (9). This can be due to disuse of muscles or neurogenic conditions (16). Muscle physiological cross-
sectional area is defined as the volume of a muscle divided by its fibre length, and is proportional to the maximum
strength or contractile force of the muscle (17). This decrease in muscle volume from muscular atrophy thus leads to a
decrease in maximum strength or contractile force of the muscle, decreasing muscle performance. Symptoms of
atrophy can also include numbness, weakness and tingling in limbs (16). As the loss of muscle predominantly occurs
in type IIB fast twitch fibres first than other fibres, higher output fast twitch fibres with higher contractile force peaks
are the first to start shrinking (18). This leads to an overall decrease in the peak contractile force of the entire
distribution of muscle fibres in the muscle group.

(B)
Upon recovery from atrophy, with increased resistance training and protein rich diets, this leads to greater extent of
protein synthesis which exceeds protein breakdown in muscle cells, especially synthesis of contractile components
such as actin and myosin protein filaments (19). This leads to an overall increase in muscle mass and volume. Muscle
physiological cross-sectional area is defined as the volume of a muscle divided by its fibre length, and is proportional
to the maximum strength or contractile force of the muscle (17). As the volume of the muscle increases and muscle
fibres get longer with more protein synthesis, there is an increase in physiological cross-sectional area and maximum
strength or contractile force of the muscle.

7. The patient you are treating does not have a strong science background; explain to them how stimulation of skeletal
muscle can result in different strength contractions using language which they can understand.
A lack stimulation of muscles, where you don’t use your muscles often to do small exercises and you don’t regenerate
them often with meat diets rich in protein, can cause the important parts of your muscles that help you contract and
move your muscles and body to shrink. Your muscles get smaller in size, each fibre has less surface area, and that
causes them to be less able to contract or move, and they get weaker. On the other hand, with more stimulation of your
muscles, by working out with small exercises and regenerating them with meat diets rich in protein, your muscle cells
start to build themselves up again with more protein. This increases your muscle mass and volume. As each muscle
fibre has more surface area, the muscle gets stronger and more able to contract or move.

References
List any references you have used in your answers, in the panel below.
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Medicine; 2001
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2023
3. Sam C., Bordoni B. Physiology, Acetylcholine. United States: National Library of Medicine; 2023

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2015
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tendon. United States: J Hand Surg Br.; 1991
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