UTILIZATION OF BOUND CALCIUM IN THE ACETYLCHOLINE CONTRACTURE OF FROG SKELETAL MUSCLE

¹ Department of Pharmacology and Therapeutics, University of Manitoba Faculty of Medicine, Winnipeg, Manitoba, Canada

It was shown previously that keeping a frog toe muscle in a calcium-free solution for a few minutes rapidly and completely eliminated the potassium-induced contracture without reducing the potassium-induced depolarization. These experiments were performed using solutions containing choline in place of sodium ions. In similar experiments it was observed that a "residual response" to potassium remained after the muscle had been kept in a calcium-free Ringer solution for more than 1 hour. This "residual response" to potassium was blocked by d-tubocurarine, 10^-4 g/ml and it was not produced in a muscle denervated 14 days or more prior to testing, showing that the "residual response" was produced by acetylcholine released from intramuscular nerve endings by potassium. Both acetylcholine and carbachol could induce a contracture of the toe muscle in the absence of extracellular calcium ions.

"Residual responses" to potassium, acetylcholine contractures and carbachol contractures could be obtained when using solutions containing lithium, but not sucrose or choline in place of sodium ions. Neither acetylcholine nor carbachol contractures could be produced in muscles completely depolarized by potassium. These results indicate that acetylcholine and carbachol must depolarize the muscle membrane in order to produce a contracture.

With prolonged exposure of the muscles to calcium-free solutions (1 to 3 hours) "residual responses" to potassium, carbachol and acetylcholine contractures were eventually eliminated. Placing the muscles in solutions with calcium ions restored the sensitivity to acetylcholine and carbachol and after a relatively brief exposure to calcium ions (30 minutes or less) all three types of responses could be obtained again with the muscles in calcium-free solutions. These results show that acetylcholine and carbachol induce contractures in calcium-free solutions by releasing calcium from a binding site in or on the muscle fibers. This binding site also could be depleted by treating a muscle in a calcium-free solution with caffeine or by repeated testing with potassium in a calcium-free solution containing cobalt ions.

In conclusion, acetylcholine and carbachol induce contractures of frog toe muscles by depolarizing the membrane and thereby promoting the entrance of calcium ions into the cells. These drugs can also release calcium ions from a binding site in or on the muscle fibers and in this way induce contractures in the absence of extracellular calcium ions.