Distinct calcium channel isoforms mediate parathyroid hormone and chlorothiazide-stimulated calcium entry in transporting epithelial cells.
Some cells express multiple calcium channel isoforms that are likely to have distinct functions. The present study used molecular cloning and antisense techniques to identify calcium channel isoforms mediating calcium entry in mouse distal convoluted tubule (DCT) cells. The DCT is the major site of hormone- and diuretic-regulated calcium transport in the kidney. Cellular calcium absorption involves entry through apical membrane calcium channels that are sensitive to dihydropyridine-type calcium channel antagonists. Partial cDNA clones corresponding to one isoform of the calcium channel alpha1 pore-forming subunit, alpha1C, and one isoform of the calcium channel beta accessory subunit, beta3, were isolated by RT-PCR. Full-length transcripts were detected by Northern blot analysis in immortalized DCT cells. Antisense oligonucleotides complementary to the alpha1C sequence inhibited the rise of intracellular calcium ([Ca2+]i) induced by the thiazide diuretic, chlorothiazide (CTZ), but not that induced by parathyroid hormone (PTH). However, antisense oligonucleotides complementary to the beta3 sequence inhibited both CTZ- and PTH-induced rises of [Ca2+]i. beta3 antisense oligonucleotides also inhibited the membrane hyperpolarization induced by CTZ but not that triggered by PTH. Thus, members of the voltage-gated calcium channel family are expressed in DCT cells, where they are responsible for hormone- and drug-induced calcium uptake. The results suggest that DCT cells contain multiple calcium channels with distinct roles in the regulation of cellular calcium.
Barry
E L
EL
Department of Pharmacology and Toxicology, Dartmouth Medical School, Hanover, New Hampshire 03755-3835, USA.
Gesek
F A
FA
Yu
A S
AS
Lytton
J
J
Friedman
P A
PA
D000818:Animals; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000096:biosynthesis / Q000187:drug effects / Q000502:physiology*; D020746:Calcium Channels, L-Type; D002478:Cells, Cultured; D002740:Chlorothiazide / Q000494:pharmacology*; D017931:DNA Primers; D004847:Epithelial Cells / Q000187:drug effects / Q000502:physiology*; D007686:Kidney Tubules, Distal / Q000502:physiology*; D007700:Kinetics; D046911:Macromolecular Substances; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D010281:Parathyroid Hormone / Q000494:pharmacology*; D016133:Polymerase Chain Reaction
D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D017931:DNA Primers; D046911:Macromolecular Substances; D016376:Oligonucleotides, Antisense; D010281:Parathyroid Hormone; D002740:Chlorothiazide; D002118:Calcium