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EXPERIMENT 7
Neuromuscular Physiology and Muscle Mechanics
(revised Jul/12)
LEARNING GOALS:
Be able to:
Explain the steps involved in inducing a single muscle contraction, naming the
structures and types of signaling involved.
Set up a frog neuromuscular preparation and explain the function of the
instrumentation used.
Properly use the PowerLab stimulator and Chart functions for collecting,
displaying, and analyzing data.
Demonstrate recordings of twitch, summation, treppe, tetany, and fatigue and
know the difference between what is observed in the laboratory and what occurs in
the body.
Analyze records to measure latency, threshold, recruitment, twitch duration,
summation, tetany, fatigue, and the effects of various pharmacological agents.
Develop and test hypotheses (with a scientific rationale) to determine what a
specific unknown drug is (from among several possibilities), based on its effects on
nerve, muscle and neuromuscular junction.
Before coming to class: This is a complicated set of experiments, so come well
prepared with a flow chart that simplifies all the instructions you will carry out.
Think of all the sites or steps, from the activation of the motor neuron to muscle
fiber contraction that may be affected by a given drug, and their effects on muscle
contraction. How could you test where the site of action may be for a particular
pharmacological agent? In this lab, you will test an unknown (to you) compound to
determine its site of action.
The possible drugs to be used are listed below (they will be randomly labeled A, B,
C and D):
Eserine (= physostigmine), an acetylcholine esterase inhibitor
Curare, an antagonist of the nicotinic acetylcholine receptor
Tetraethylammonium, a blocker of voltage-gated potassium channels
You will be exploring the nature of neuromuscular transmission. The isolated frog
neuromuscular preparation the sciatic nerve and the gastrocnemius muscle will
allow you to investigate basic neuromuscular physiology. Although under normal
conditions a nerve AP originates within the central nervous system (CNS), in the
laboratory you will mimic CNS stimulation by electrically stimulating the sciatic
nerve, thus producing typical action potentials on motor neurons within the sciatic
nerve, which will in turn generate APs that trigger contraction of the innervated
muscle fibers. Be sure you understand the consequences of this difference
(between what you do in lab and what occurs in vivo). Note that the electrical
stimulation will be applied to the nerve, which is a bundle of sensory and motor
neuron axons. Those closest to the site of stimulus will be depolarized the most,
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Stimulating electrodes
Computer and PowerLab interface, Chart
software
Frog Ringers
Force transducer
Calibration weights
Unknown drugs handle these with care and wash your hands
after handling!
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Stimulus
Figure 7-1. Graded response of whole muscle due to recruitment of motor units.
Strong stimuli lead to recruitment of fibers (A). As the stimulating voltage is
diminished, fewer fibers are stimulated until a threshold stimulus is achieved (B).
Stimuli with lower stimulus strength than the threshold stimulus fail to elicit a
contraction (C).
Fatigue
Tension
Complete
Tetany
Treppe (Staircase
effect)
Summation begins
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Figure 7-5. Steps for dissecting the frog leg. Step one will have been completed
for you.
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glass probe
sciatic nerve
Stimulating hook
electrodes under
sciatic nerve
Gastrocnemius muscle
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ON
NEURON, MUSCLE,
AND THE
NEUROMUSCULAR
JUNCTION:
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Threshold measurements.
Minimum Stimulus Voltage for Twitch
Minimum Stimulus Voltage for Maximum
Contraction
Measurements for Time Course of a Twitch and the Latency Period
A. Your Data Pad should also have a value for Time and Mean for both
channels (stimulator and muscle). In the tracings from the recruitment
experiment, find the stimulus that produced a muscle contractile response
that was approximately 50% of the maximal response. Insert a COMMENT
somewhere at the beginning of this trace to mark it. Highlight both the
stimulus and entire response by holding the SHIFT key so both traces are
highlighted. Use the Zoom Window to expand both traces.
B. Use the overlay function in the top right corner of the Zoom Box. Refer to
Fig. 7-3 depicting parts of a twitch response to obtain data for the Time
Course of Twitch Table in Report 7. Annotate the data pad to indicate which
values are in which row. SAVE the DATA PAD as a textfile with an
appropriate filename for the data.
Take the following time measurements by double clicking on the following points.
(Check your data pad to make sure the measurements are added).
1. Time at the beginning of the stimulus.
2. Time at the beginning of the twitch response.
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(time in sec)
C. To obtain data for fatigue, determine the stimulus frequency that elicited fatigue
and the time from the start of stimulation to when fatigue was first noted.
1. Use the SAVE AS command under FILE to save the DATA PAD as a textfile.
Enter the name of the file and its folder above.
2. IN PREPARATION FOR PRINTING, delete irrelevant traces, compress, and
highlight the relevant traces. PRINT your best trace showing the different
phases of tetany.
D. Effects of the unknown drug will be descriptive and no data need to be analyzed
but you will need to append the Chart traces that pertain to this experiment. As
before, PRINT only selected traces, deleting irrelevant traces and compressing the
traces to be printed.
Experiment Report 7
Neuromuscular Physiology and Muscle Mechanics
WORK DONE BEFORE THE EXPERIMENT:
List your working hypothesis or hypotheses, your predictions about the
experimental results, and the rationale for your hypotheses and predictions.
Hypotheses:
Predicted results and rational:
DISCUSSION QUESTIONS:
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