and/or aims): Intracellular Ca2+ plays a role in regulating many of the hormonally mediated functions of intestinal epithelial cells (enterocytes), including secretion and Ca2+ uptake. In many type of non-excitable cells, a dominant mechanism for mobilizing Ca2+ from intracellular compartments involves phosphoinositide hydrolysis to produce inositol 1,4,5-triphosphate (InsP3) which then opens intracellular Ca2+ channels. There is evidence that enterocytes also utilize this phosphoinositide pathway. However, direct evidence for InsP3-gated Ca2+ channels has been lacking. In preliminary experiments from the applicants laboratory, saturable InsP3 binding sites have been detected in enterocyte microsomal membranes, suggesting the presence of InsP3 receptors. The long term objective of this proposal is to characterize these putative InsP3 receptors in enterocytes. Specifically, the aims of the planned research are: 1). to characterize these receptors by their InsP3 binding properties with radioligand binding studies, and 2). to characterize their single-channel gating properties by voltage-clamping reconstituted channels in planar lipid bilayers. The studies in this proposal aim to show how InsP3 and other modulatory agents control the opening of this channel. Therefore, these studies will help to elucidate how drugs and second messengers might be able to alter cytosolic Ca2+ levels by interacting with this channel. Such information could be useful for understanding and treating disorders stemming from the disruption of Ca2+-dependent pathways.
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