A METHOD TO ESTIMATE MITOCHONDRIAL CA2+ UPTAKE IN INTACT CARDIAC MYOCYTES

In the present paper we describe a method to estimate mitochondrial Ca 2+ uptake during the declining phase of Ca2+ transients (cell relaxati on) in intact isolated myocardial cells. This method is based on inhib ition of sarcoplasmic reticulum (SR) Ca2+ accumulation by caffeine, bl ockade of Ca2+ transport via sarcolemmal Ca2+-ATPase by treatment with carboxyeosin and inhibition of sarcolemmal Na+/Ca2+ exchange by remov al of extracellular Na+ and Ca2+. Ca2+ transients were evoked in rabbi t ventricular myocytes by quick and sustained caffeine application (10 mM) after a 5-min period of electrical stimulation to load the SR wit h Ca2+. Mitochondrial Ca2+ transport was estimated using a model descr ibed by Sipido and Wier (Journal of Physiology (1991), 435:605-630), w hich was originally proposed to describe Ca2+ fluxes during excitation -contraction coupling in cardiac cells. Our results indicate that, in intact rabbit myocytes, the Ca2+ flux due to net mitochondrial Ca2+ up take may attain a value close to 1 mu M/sec.