Histochemistry of creatine phosphokinase

Abstract

In a preliminary experiment, brief formaldehyde fixation of rat myocardium permitted preservation of creatine phosphokinase (CPK) activity and fine structure. Electrophoresis of the homogenate of the fresh and fixed myocardium demonstrated mitochondrial and three non-mitochondrial CPK isoenzymes. An osmiophilic tetrazolium salt and ferricyanide were used for the electron microscopic demonstration of CPK. With tetrazolium, the reaction products were localized near the inner mitochondrial membrane in the intermembranous space of mitochondria, but the reaction was noted on the matrix side of the membranes with ferricyanide. Reaction was also present in the sarcoplasmic reticulum (SR) of the skeletal muscle. Both heart and skeletal muscle showed diffuse background cytoplasmic reaction. The heat-sensitive brain-type isoenzyme of CPK was present in the extramitochondrial sarcoplasm of the heart muscle. CPK activity was demonstrated in isolated heart mitochondria and skeletal muscle SR. Myocardial degeneration was produced in the rat myocardium by injection of isoproterenol. Two to 4 h after the injection there was an extensive histochemical reduction of CPK activity in the myocardium. Twelve to 24 h after injection, most of the areas with initially reduced CPK reaction recovered the enzyme activity and only the subendocardial layer underwent necrosis without recovery of the CPK activity. The maximal rate of increase in serum CPK activity coincided with the most extensive depletion of the myocardium CPK. Mitochondrial isoenzyme did contribute to elevation of serum CPK and appeared to remain in the injured myocardial cells.