This grant will investigate the mechanism of chronic alcohol- induced dilated cardiomyopathy (ADCM), by longitudinally assessing the structural and functional responses of the left ventricle (LV) and isolated cardiomyocytes to chronic alcohol intake. In addition, the role of the renin-angiotensin system (RAS) and the alterations in its intracellular signaling in producing ADCM will be determined. We will test the following specific Hypotheses: [H1] Chronic alcohol intake produces direct depressions in cardiomyocyte contraction, relaxation, [Ca2+]i transient and sarcolemmal Ca2+ channel activity (ICa,L), and thus causes [H2] a progressive impairment of LV systolic, diastolic function, and leads to congestive heart failure, and [H3] the most important factor in promoting the transition to ADCM is the alcohol-induced, sustained activation of circulating and cardiac RAS. This results in alterations in the AT1 receptor-coupled, protein kinase C and/or inhibitory G protein (Gi)-mediated responses of the LV and myocytes to angiotensin II (ANG II). Thus, [H4] blocking the RAS with chronic ACE inhibition or ANG II AT1 receptor blocker will blunt or prevent the chronic alcohol- induced functional and structural changes. The studies will be conducted in three control and four experimental groups studied over eight months: 1) uninstrumented alcohol-fed (22 percent alcohol once per day, providing 33 percent of total daily caloric intake); 2) instrumented alcohol-fed; 3) instrumented alcohol- ramipril (ACE inhibitor, 0.1 mg/kg/day)-fed; 4) instrumented alcohol-irbesartan (ANG II AT1 blocker, 5 mg/kg/day)-fed. The control groups will consist of age-matched dogs under identical conditions except dogs in the control groups will not receive alcohol. Serial changes of cardiac systolic and diastolic function and RAS activation will be quantitated in these conscious dogs, chronically instrumented to measure LV pressures and volume. Cell contractile function, [Ca2+]i transient and ICa,L will be serially measured in freshly isolated cardiomyocytes from the LV obtained by biopsy from the same animals. Myocytes will be studied under conditions with superfusion of: 1) calcium, 2) isoproterenol, 3) pimobendan, 4) alcohol, and 5) ANG II. In addition, the studies of myocytes response to ANG II will be repeated after preincubation with an AT1 receptor blocker, a protein kinase C inhibitor, or a Gi protein inhibitor, before, during and after the development of ADCM. These studies will be the first detailed longitudinal studies of cardiac and myocyte structure, function, [Ca2+]i transient, ICa,L during chronic alcohol intake. In addition, these studies will provide unique information on the role of the RAS and the mechanism of ADCM. These studies are necessary to extend our knowledge about the mechanism in the development of ADCM and help target therapy for ADCM.
