Ethanol Effects on SR Ca²+ Release in Cardiac Myocytes

DESCRIPTION (provided by applicant): One of the most serious secondary effects of alcohol abuse is the development of specific alcohol-induced cardiomyopathies which often lead to congestive heart failure. Previous findings suggest that one of the mechanisms underlying reduced cardiac function by ethanol (EtOH) may involve the direct suppression of cardiac excitation-contraction (E-C) coupling. Interestingly, this direct effect does not appear to involve alterations in the trigger for cardiac contraction (via L-type Ca 2+ current) raising the possibility that the primary negative inotropic effects of EtOH might reside in an altered response to that trigger. The goal of this project is to investigate how EtOH affects Ca2+ release from internal stores in the sarcoplasmic reticulum (SR) during the cardiac cycle. This will be accomplished through measurement of Ca 2+ sparks, which are thought to represent the fundamental Ca 2+ release units responsible for contraction. The Specific Aims of this project are: 1) to determine if the cardiodepressant effects of EtOH occur as the result of a direct suppression of SR Ca 2+ release; and 2) to investigate whether or not these changes in SR Ca 2+ release contribute to the depression in cardiac function that occurs in a chronic model of alcohol-induced cardiomyopathy. Ca 2+ sparks will be measured in rat ventricular myocytes with the Ca2+-sensitive fluorescent indicator fluo-4 using laser scanning confocal microscopy in combination with whole cell voltage clamp techniques. Some experiments will be performed in saponin-permeabilized myocytes exposed to different cytosolic Ca2+ concentrations in order to assess Ca 2+ sensitivity of Ca 2+ sparks activated purely by Ca 2+ in the absence of functional sarcolemmal Ca 2+ current. These methodological approaches will allow the investigation of how EtOH affects SR Ca 2+ release that occurs both as a result of its normal trigger (L-type Ca 2+ channels) and in response to the final common p