I propose to study the mechanisms by which cardiac cells regulate gap-junction function (permeability). Gap junctions represent the pathways of intercellular communication in cardiac tissue and as such are integrally involved in the conduction properties of the heart. Inspite of this critical role, the factors which modulate the permeability of cardiac cell gap junctions and the mechanisms used by the cardiac cell to maintain 'normal' junction permeability remain to be elucidated. Two methods will be used to assess the function of the intercellular conduction pathway. The first method involves a test of the permeability of the junction to intracellularly injected fluorescent dyes of varying molecular weight. This technique is a very sensitive method for detecting small changes in gap junction permeability. The second method involves measurement of conduction time (or delay) from a stimulating electrode to the recording electrode. Alteration of junction permeability and conduction time will be correlated with alterations of electrical and contractile activity and ionic fluxes. This comprehensive approach allows distinction between alteration of conduction time due to alteration of junction permeability versus alteration of other properties which influence conduction. In particular, I will: 1) examine the role of the Ca++ binding and sequestering organelles in the protection of the intercellular conduction pathway from elevated intracellular Ca++ levels, and 2) determine whether these roles, established under control conditions, are altered under the abnormal conditions of increased cellular Ca++ load, anoxia or ischemia. The ultimate goal is to determine whether one of the steps leading to irreversible tissue damage following anoxic or ischemic insults is compromised junction function. Cultured neonatal rat myocardial cells and the isolated arterially perfused interventricular septum of the rabbit will be used for these studies. The data gained from these studies will be used to formulate new strategies for protection of conduction parameters during and following anoxic and ischemic interventions. The proposed research represents a unique approach to the questions of how the heart maintains normal conduction parameters, and of how those parameters are modified during myocardial disease.
