THE INFLUENCE OF IONS ON THE CONTRACTION OF ISOLATED
EARTHWORM SMOOTH MUSCLE
BACKGROUND READING
Animal Physiology by Hill, Wyse & Anderson, 2004: pp. 484–486.
ANIMALS & EQUIPMENT
Living material
Large earthworms (Lumbricus terrestrius).
Equipment
Scissors; pins; forceps; thread; ring stands and clamps; force transducer; bioamplifier;
earthworm ringer; immersion bath; plastic waste beakers; known weights; Macintosh
computer; PowerLab virtual chart recorder; BNC cables. (Include a sketch of the
equipment set up in your lab book that is of sufficient quality that you could use it to set
up the same experiment at a later date)
STANDARD
A.
PROCEDURE
Preparation of the equipment
1. Study the controls on the Bioamplifier. Set the amplifier to Bridge Mode, 100X
amplification, and 50K filter.
2. Open the chart application and set up to record from Channel 1 only.
3. Set the controls to collect data at a slow speed ... about 2 samples/sec is a good place
to start.
4. Turn on the 50 Hz LP filter within the Input Amplifier.
*
B.
You are probably expecting us to say: "Now you must calibrate the force
transducer". Surprise!!! You don't have to! The forces that you measure today
will be examined RELATIVE to each other. For the actual force to be really
meaningful, you would need to determine the cross-section of the active muscle and
the angle at which the muscle is pulling. Both of these measurements are beyond
the scope of the lab. In other words, we're not letting you off easy, we're just not
making you do something that won't really add anything of value to your analysis.
Dissection of Earthworm (from Hoar and Hickman, 1975)
**THIS IS A DIFFICULT DISSECTION!
TAKE CARE BUT BE BOLD**
* IF AVAILABLE PLEASE LOOK AT DEMONSTRATION DISSECTION BEFORE
BEGINNING*
*KEEP THE PREPARATION MOIST WITH RINGERS*
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1. Place an earthworm in the dissecting tray and pin down the anterior end. Place
another pin just anterior to the clitellum (Figure 1). DO NOT PLACE PINS
DIRECTLY ON THE MIDLINE OF THE ANIMAL OR YOU MIGHT PUNCTURE THE ORGANS.
Anterior
Clitellum
Posterior
Figure 1. External view of Earthworm (from: http://biog-101104.bio.cornell.edu/BioG101_104/tutorials/animals/earthworm.html)
2. Make a mid-dorsal incision just anterior to the clitellum and proceed carefully
forward, cutting just through the skin and body wall. Cut through septa and pin
body wall open to reveal the internal organs. Identify the anatomical features in
Figure 2 below to get your bearings.
Figure 2. Illustration of the internal organs in an earthworm opened by a mid-dorsal
incision. (From: http://sps.k12.ar.us/massengale/earthworm_dissection.htm)
3. Loosen the esophagus, crop, gizzard and anterior portion of the intestine from the
underlying tissues by carefully cutting the septa.
4. Carefully pass a long piece of thread under the esophagus and tie off. Do the same
for the intestinal end, tying the intestine off very close to the end of the gizzard.
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5. Cut the intestine to free the anterior end and then cut the intestine to free the
posterior end.
6. Carefully mount the preparation in the muscle bath according to instructions given
in the lab.
You should see slow rhythmic contractions within a few minutes. If you do not, carefully
adjust the tension on the preparation by lowering the muscle bath SLIGHTLY. If you do
not see contractions within a few minutes, you'll have to do another dissection.
C.
Normal Activity and the Effects of Cation Concentration
1. After the preparation has recovered from its initial trauma, record normal
activity of the gut preparation for around 5 min. You'll want to note the magnitude
and frequency of the beat pattern.
2. Drain the normal ringer solution and replace with one containing either no CaCl2
or no KCl. Record for 5 min.
3. Rinse the preparation with at least 3 changes of normal ringer. Following the
washes, immerse the preparation in normal ringer and record the response for 5
min.
4. Drain the normal ringer solution and replace with one containing either 2X CaCl2
or 2X KCl. Record for 5 min. or until you see a clear response.
5. Rinse the preparation with at least 3 changes of normal ringer. Following the
washes, immerse the preparation in normal ringer and record the response for 5
min.
PRE-LAB
(Due at the start of lab)
1. List two ways in which the appearance of smooth muscle cells differ from skeletal
muscle cells. List three major sub-cellular differences between smooth muscle cells
and skeletal muscle cells.
2. Relative to skeletal muscle contractions, would you expect to use a faster, slower, or
the same chart recorder speed to measure smooth muscle contractions? Why?
3. In general, how does the regulation of smooth muscle contraction differ from that of
skeletal muscle?
4. In the experiment that you will do today, how will you induce the muscle to contract?
(Hint: this is sort-of a trick question)
POST-LAB
(Due next week in lab)
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1. Write a short paragraph that verbally describes your results.
2. Construct at least two figures and/or tables to back-up your statement(s) above. The
figures should clearly show your main result. The figures can be raw or processed
data, depending on what you believe best illustrates your main result(s).
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