The effect of temperature on the pH stability of myosin ATPase as demonstrated histochemically.

Abstract

The histochemical reaction for actomyosin adenosine triphosphatase (ATPase) and its lability to pH variations characterize two major categories of fibre types in mixed mammalian skeletal muscles. These are designated as the “Type I” and “Type II” fibres. Myosin ATPase of Type I fibres has been shown to be relatively acid stable but alkali labile while that of Type II fibres is relatively alkali stable but acid labile. In this study we have investigated the effect of preincubation at varying pH and temperatures on the histochemical demonstration of myosin ATPase in animal and human mixed skeletal muscles. Sections fixed in cold fixative at either high (10.4) or low (4.35) pH showed identical staining patterns. When alternate serial sections were preincubated in either cold (4°C) acidic or cold alkaline solutions, only the Type II fibres were intensely stained. After alkaline preincubation at room temperature (19–21°C) varying degrees of ATPase inhibition were observed in Type II fibres while Type I fibres remained unstained. Acidic preincubation at room temperature (19–20°C) resulted in the enhancement of staining of the Type I fibres but variable inhibition of activity was found in the Type II fibres. This study demonstrates that during preincubation, pH is not the only critical factor and the specific temperature of the acidic or alkaline preincubation bath is a major contributing factor in the histochemical demonstration of myosin ATPase. It also demonstrates that a temperature dependent loss of myosin ATPase activity in the frozen sections of skeletal muscle following acidic or alkaline pretreatment is consistent with previous biochemical reports on purified actomyosin (Guth and Samaha, 1969). Moreover, the heterogeneity can be displayed within both Type II white and Type II red muscle fibres when examined for myosin ATPase after acid preincubation.