Localization of Ca²⁺ at the Plasma Membrane of Bullfrog Myocardial Cells

Cellular distribution and localization of Ca2+ in bullfrog myocardial cells was investigated on the light and electron microscopical level using different preparation procedures. Fixation and dehydration in glutaraldehyde and ethanol solutions containing increased amounts of Ca2+ (5-25 mM) cause the formation of numerous electron-dense deposits at the external face of the sarcolemma. X-ray microanalysis demonstrates that the deposits are mainly composed of calcium. Similar results were obtained in cryopreparations by quick-freezing of fresh myocardial tissue. Granules with a diameter of 16.6 nm, which clearly exceed all other cell constituents in contrast, are found at the entire myocardial cell surface. Besides an increased amount of calcium the granules contain potassium and phosphorus. Fluorescence microscopy on single cells obtained from bullfrog atrium by treatment with digestive enzymes and stained with chlorotetracycline (CTC) as a fluorescent probe for membrane-bound Ca2+ clearly support the observations made on the fine structural level. A strong fluorescence signal depending on the CTC- and Ca2+-concentration of the incubation medium is observed at the sarcolemma and can be inhibited by addition of La3+. The results of the present investigation suggest that the surface coat of the sarcolemma of myocardial cells represents an important compartment for accumulation and release of external Ca2+. The external lamina which partly covers the surface of myocardial cells seems not to be involved in the Ca2+ exchange.