The planned research program integrates two distinct areas of investigation: (1) The mechanisms by which ethanol interferes with excitation-contraction coupling and depresses the contractile function of the heart; (2) The mechanisms and functional consequences of ethanol interaction with the inositol lipid signaling system in the liver. These projects are linked by a common focus on signaling processes mediated by Ca2+. It is also likely that some of the targets of ethanol action are common to both tissues. The experimental work in the heart will focus on the use of advanced techniques for studying ethanol action, including measurements of the dynamics of cytosolic Ca2+ and sarcomere shortening using digital imaging fluorescence microscopy techniques, together with whole-cell voltage-clamp measurements of sarcolemmal ion currents. Hypotheses to be investigated for the mechanism of ethanol action include a reduction in Ca2+ sequestering properties of the sarcoplasmic reticulum, alterations in Ca2+ and/or Na+ channel activity in the sarcolemma and a shift in the relation between Ca2+ and contractile activity of the muscle proteins. In addition, the interactions between ethanol and catecholamines, and the deleterious effects of ethanol and cocaine in combination, will be investigated. These studies, carried out using isolated rat ventricular myocytes, will examine the effects of both acute ethanol and long term ethanol- feeding. Studies of ethanol action on Ca2+ signaling in liver will also utilize imaging of Ca2+ in single isolated hepatocytes. This work will investigate the effects of ethanol on the spatial and temporal organization of Ca2+ signaling at the subcellular level, determine how the direct acute effects of ethanol are modulated by protein kinases, and examine the mechanism by which ethanol modifies the responses to hormones that regulated hepatic metabolism through changes in cytosolic Ca2+. In addition, the mechanisms by which ethanol affects the inositol lipid signal transduction system will be investigated using an isolated turkey erythrocyte membrane preparation. This retains a functional, hormone-coupled inositol lipid-specific phospholipase C (PLC) that generates the second messengers inositol 1,4,5-triphosphate and diacylglycerol. The turkey erythrocyte membrane PLC is sensitive to both acute stimulation by ethanol alone, and to inhibition of hormone activation by ethanol pretreatment. This research program will contribute to the elucidation of the mechanisms by which ethanol perturbs the normal function of liver and heart tissue.
| Nicolas, J M; Renard-Rooney, D C; Thomas, A P (1998) Tonic regulation of excitation-contraction coupling by basal protein kinase C activity in isolated cardiac myocytes. J Mol Cell Cardiol 30:2591-604 |
