Expression of Dihydropyridine Binding Sites in Renal Epithelial Cells

Abstract

It has recently been shown that rabbit kidney epithelial cells (proximal tubule) possess two dihydropyridine (DHP)-sensitive calcium entry channels (1, 2). To evaluate the properties of the DHP binding sites, the binding of the DHP, [3H]PN200-110, was studied in epithelial cell membrane fractions (proximal tubule) of rabbit kidney cortex. High-affinity binding sites for the DHP were observed in both basolateral and apical membranes and in a membrane microsomal preparation from rabbit primary cultures of proximal tubule cells (cultured PT). In an extended analysis of the basolateral membrane preparation, two high-affinity binding sites were evident with binding dissociation constants, Kd, of 0.005 and 0.75 nM. The Kdvalues are similar to that observed for L-type calcium channel α1-subunits. Using a homology-based cloning strategy, a 388-base fragment of an α1-subunit was cloned from RNA isolated from rabbit cultured PT cells and freshly isolated proximal tubules and found to encode a protein identical to the cardiac form of the L-type α1-subunit (α1C-subunit). It is concluded that renal epithelial cells express high-affinity dihydropyridine receptors and that the receptors may be components of plasma membrane calcium channels, including L-type calcium channels, that control calcium entry in these cells.