THE DETERMINANTS OF CONTRACTION IN VOLTAGE-CLAMPED, INTERNALLY PERFUSED SINGLE MAMMALIAN HEART CELLS

Abstract

I have developed a new preparation to study excitation-contraction (E-C) coupling in the mammalian heart. I modified a whole cell voltage clamp technique to simultaneously measure contractile shortening and transmembrane currents in single, enzymatically isolated guinea pig heart cells internally perfused with a pCa = 7, ATP-containing solution. The microscopic appearance of the cells was observed and recorded with a television system, while contractile shortening was measured 1000 times/sec using a linear photodiode array. The contractile and electrical properties of this new preparation were characterized. The effects of channel blockers, altered ion concentrations, and altered electrical parameters on the inward Ca{dollar}\sp{lcub}2+{rcub}{dollar} current (I{dollar}\sb{lcub}\rm si{rcub}{dollar}) and on twitch shortening were examined. Thus, I was able to study the processes which determine the amplitude and time course of the twitch contraction in a cardiac muscle preparation with intact membranes.;Step depolarizations from the holding potential of {dollar}-{dollar}50 mV to 0 mV triggered I{dollar}\sb{lcub}\rm si{rcub}{dollar} and uniform synchronous shortening which resembled that seen in intact, nonperfused, electrically stimulated cells. The amplitude of I{dollar}\sb{lcub}\rm si{rcub}{dollar} was insufficient to directly activate the myofilaments given the constituents of the intracellular perfusate. Changes in the magnitude of I{dollar}\sb{lcub}\rm si{rcub}{dollar} (by electrical or pharmacological manipulations) were accompanied by immediate parallel changes in the extent and velocity of shortening. Changes in voltage per se did not correlate to immediate changes in shortening. Increases in the amplitude, duration, and rate of the depolarizing step did cause an increase in the extent and speed of twitch shortening over the course of several contractions, however. Increases in the duration of the depolarizing step prolonged contraction. Local electrical and ionic changes around the membranes of intact nonperfused cells demonstrated that the mechanisms which control contractile strength act at a subcellular level. Thus, (1) the transmembrane Ca{dollar}\sp{lcub}2+{rcub}{dollar} current triggers a large release of Ca{dollar}\sp{lcub}2+{rcub}{dollar} from the sarcoplasmic reticulum (SR), (2) the amplitude of I{dollar}\sb{lcub}\rm si{rcub}{dollar} at each point along the cell determines of the size of that local release, and (3) a voltage-dependent process independent of I{dollar}\sb{lcub}\rm si{rcub}{dollar} (probably an electrogenic Na/Ca exchange) also grades the size of contraction and affects its timecourse, presumably by modulating Ca{dollar}\sp {lcub}2+{rcub}{dollar} accumulation in the SR.