Techniques of skinned cardiac cells and of isolated cardiac fibers with disrupted sarcolemmas with reference to the effects of catecholamines and of caffeine

Abstract

Single cardiac cells with disrupted sarcolemmas were obtained by homogenization in low free [Ca2+] medium. Skinned cardiac cells were obtained by microdissection of remaining pieces of sarcolemmas. In both preparations the buffers bathing the cells were in contact with the structures controlling activation: myofilaments, sarcoplasmic reticulum, and mitochondria. Tension was recorded with a highly sensitive force transducer. The use of either high or low total [EGTA] permitted the study of the effect of free [Ca2+] on either the myofilaments or the internal stores. At physiological levels of myoplasmic free [Ca2+], the internal stores of Ca2+ were located mostly in the sarcoplasmic reticulum, whereas mitochondria were involved at higher free [Ca2+]. Single cardiac cells with simply disrupted sarcolemmas responded to variations of perfusion media in a manner similar to the skinned cardiac cells. However, the remaining pieces of sarcolemma represented a limit to the intracellular diffusion of externally applied media and might contain receptor sites for catecholamines. In multicellular fragments, evidence of surface membrane activity (resting and action potentials) were found during certain ionic modifications.