Mechanisms of Long-Term Cardiac Ion Channel Regulation

Heart failure (HF) and atrial fibrillation (AF) represent two of the most prevalent heart diseases in the U.S. In moderate to severe HF and in AF numerous phenotypic changes occur including a reduction of voltage-gated calcium channel current density. Changes in Ca channel expression may have far-reaching effects on heart function due to their central role in excitation contraction coupling. Also, Ca channels have been implicated in excitation-transcription coupling. Thus alterations of Ca channel expression in the heart can also impact on disease progression. The principal voltage-gated Ca channel in the heart (CaV1.2) is extensively studied in the realm of its biophysical characteristics and second messenger modulation. However, very sparse information is available regarding longer term regulation of Ca channel expression. In this proposal we introduce a novel mechanistic hypothesis governing the functional expression of cardiac Ca channels in the sarcolemma of cardiac myocytes. In our preliminary data we show evidence for a ras-related monomeric G-protein interaction with the CaV1.2 accessory subunit CaVb2. We propose to test the global hypothesis that G-protein interaction with CaVb subunits reduces Ca channel current density by competing for CaV1.2 binding. Moreover, our preliminary data shows strong evidence that this is a dynamic process that is modulated by chronic PKA activity. There are four specific aims that guide our experimental design: Aim 1 will characterize the time course and sub-cellular localization of nascent G-protein and CaVb complexes. Aim 2 will characterize the intracellular second messenger modulation of G-protein-CaVb regulation of Ca channel current expression. Aim 3 will characterize the modulation of Ca channel current and G-protein modulation in native cells. Aim 4 will characterize the function of these G-proteins in heart function. These aims encompass approaches ranging from investigations of a purely molecular nature (protein-protein interactions), to in vitro model systems (heterologous expression of recombinant proteins), to native cell systems (primary isolates and cultures of heart cells), to the impact of a single class of G-proteins on tissue level heart function. This proposal may identify a novel class of therapeutic targets for alleviation of heart dysfunction in HF and AF.