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The calcium release process of the sarcoplasmic reticulum of mammalian skeletal muscle will be examined in studies involving 1) muscle fibers, 2) isolated triads (junctions of t-tubules and sarcoplasmic reticulum) and 3) isolated sarcoplasmic reticulum (SR). The mechanism(s) of Ca++ release that contribute toward physiological contractile activation during excitation-contraction coupling will be identified by comparing the sensitivity of various ways of releasing calcium from SR blockade by different pharmacological agents. The pharmacology of muscle excitation-contraction coupling will be examined in electrophysiological and optical studies involving mammalian skeletal muscle fibers. The contractile threshold for different durations of depolarization under voltage clamp conditions will be determined in the presence and absence of different blockers of various forms of Ca++ release from isolated SR. Those substances affecting the contractile properties of the fibers under these conditions will be further scrutinized in experiments involving direct measurement of myoplasmic Ca++ transients with arsenazo III or antipyrylazo III. Isolated triads will be utilized to reproduce calcium release from the SR portion of the isolated structure in response to depolarization of the associated transverse tubule membrane. These experiments will assess the pharmacological sensitivity of the physiological SR calcium release process, which will then be compared to the sensitivities of various forms of calcium release elicited directly from isolated SR. SR isolated from both rabbit skeletal and cardiac muscle will be induced to release Ca++ by calcium-induced, "depolarization"-induced, alkalinization-induced, various forms of drug-induced, and spontaneous release of calcium. The sensitivity of each of these kinds of release to different pharmacological agents will be tested. Comparison to the sensitivity of the physiological release in muscle fibers, above, should assess the physiological relevance of any of these forms of calcium release and may demonstrate alternative means of opening the presumed SR Ca++ channels involved. Once alternative means of opening the physiologically relevant channels are known, patch clamp/single channel recording techniques will be applied to isolated SR to assess sensitivity to SR membrane potential and other electrical properties.

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