Histochemical characterization of adenosine triphosphatases in skeletal muscle fibers by selective extraction procedures.

Abstract

Selective extraction procedures have been applied to frozen sections of rat skeletal muscles to determine the relative contribution of myosin adenosine triphosphatase (ATPase) and of membrane-bound (mitochondrial and sarcotubular) ATPases to the histochemical ATPase reactions demonstrated in muscle fibers by the calcium precipitatation mehod (Padykula-Herman procedure) and by the lead precipitation medium (Wachstein-Meisel procedure). A detergent treatment was used to solubilize membrane-bound enzymes and hypertonic KCl solution to extract myosin. Pretreatment with the non-ionic detergent, Triton X-100, severely depressed the ATPase activity demonstrated by the lead method, as well as several membrane-bound dehydrogenase activities, but did not affect the intensity of the ATPase reaction demonstrated in the muscle fibers by the calcium method. After detergent treatment, however, the end product of the latter reaction showed a network distribution which contrasted with the characteristic myofibrillar localization observed in untreated sections. Hypertonic KCl pretreatment, when applied to undried sections, abolished the ATPase reaction demonstrated in the muscle fibers by the calcium method; the ATPase demonstrated by the lead method was slightly depressed and the dehydrogenase activities unmodified by the same treatment. The results indicate that the ATPase activity demonstrated histochemically in skeletal muscle fibers by the calcium method is due to myosin ATPase, and suggest that the intermyofibrillar localization of the end product which occurs under certain conditions is artifactual.