Purpose

Energy, in the form of ATP (Adenosine triphosphate), is necessary for muscle contraction. In living muscle, the ATP is generated "on the spot" for use by individual fibers. In this experiment, you will learn several things:

1. That ATP as a chemical can be applied to preserved fibers, and contraction will result;
2. That the ATP is effective only in the presence of certain salts.
3. That the mechanism for investigating these relationships utilizes the Scientific Method.

Rabbit psoas muscle preserved in 50% glycerol can be frozen and stored almost indefinitely (Could you use this for "deep sleep"?) ATP solutions are refrigerated but should be as fresh as possible when used (Why?).

Procedure

1. Remove form the test tube the stick to which the bundle of muscle has been tied. Pour the glycerol into a glass dish. Cut the muscle bundle into pieces about 2 cm. In length. Drop these into the glycerol in the dish. One piece is sufficient for each individual or team.

2. Tease the segment of muscle into very thin groups of fibers (muscle cells). Try to get strands no thicker than a fine human hair; single fibers, if obtained, will demonstrate the greatest contraction. Do not use strands of muscle exceeding 0.2 mm in thickness.

3. Transfer three or more of the thinnest strands to a minimal amount of glycerol on a slide, with no cover slip. Position the strands straight and parallel to each other. (There must be enough glycerol to prevent drying out; however, the less you use, the easier the fibers are to measure.) Place this slide under a dissecting microscope and measure the length of the fibers with a millimeter ruler held under the slide. Record these lengths.

NOTE: If you work in groups of three, each member should do either 4, 5, or 6 and the three pool their results.

4. Flood the fibers with several drops of the solution containing ATP plus potassium (K+) and magnesium (Mn++)(divalent cation) ions. Observe the reaction of the fibers. After 30 seconds or more, measure the fibers again. Record.

5. Repeat the experiment using new fibers and the solution of ATP alone. Record.

6. Repeat the experiment with new fibers and the solution of salts alone. Record

Name ____________________________
Partner 1 ________________________ Partner 2________________________________

Briefly describe and draw the appearance of the fibers before treatment.

How do they look after contraction? Pay special attention to the spacing of the striations and the smoothness of the sides. Describe and draw.

Conclusions

1. Was it necessary to try ATP alone and salts alone?

2. What is the technical term for the three setups used?

3. What conclusions may be drawn from your results?

Discussion

Compare your data with those of other students. Assuming everyone got positive results with ATP + salts, what additional factors seem to affect the amount of contraction?

Hint: The speed and extent of the reaction are influenced by the amount of glycerol on the slide, the concentration of active ATP, and the width of the muscle strand (Why?). Under favorable conditions, a myofiber can be expected to contract to almost 50% of its starting length within 10 seconds. (Is there a concomitant change in width?)

This page last updated: 3/4/01