This U.S.-Poland research project between Dr. Kathryn F. Lanoue of Pennsylvania State University Hershey Medical Center and Jerzy Duszinski of the Nencki Institute of Warsaw, Poland, is to identify and characterize the transporter which catalyzes hormone dependent influx of extracellular calcium into liver cells. The researchers will study fura-2 loaded hepatocytes in primary culture, using a fluorescence digital imaging system in conjunction with whole cell patch clamp microinjection techniques, to determine the current voltage relationships of the putative calcium channel, as well as the enthalpy of calcium influx and its Ph dependence. Preliminary experiments indicate the calcium transport into the cell is not dependent on hormone occupancy of calcium mobilizing receptors in the plasma membrane, or on IP3 levels in the cell but it is dependent on the extent of depletion of intracellular hormone releasable calcium stores. Calcium depletion of these stores stimulates calcium entry into the cell, in a graded manner. The researchers will test the hypothesis that a calcium sensitive enzyme in the endoplasmic reticulum elaborates and releases to the cytosol a metabolite which regulates plasma membrane calcium permeability. They will design experiments to demonstrate the existence of such a diffusible metabolite and to characterize it. The researchers plan to use an inhibitor of the calcium plasma membrane transporter as a means to monitor calcium transporter activity in hepatocyte plasma membrane vesicles loaded with fura-2. They will use the vesicular assay system to screen the influence of putative regulators and as a basis for later solubilization, reconstitution, and purification. %%% This project in biology fulfills the program objective of advancing scientific knowledge by enabling leading experts in the United States and Eastern Europe to combine complementary talents and pool resources in areas of strong mutual interest and competence.
