Dihydropyridine-sensitive Ca(2+) channels in human glomerular mesangial cells.
In mesangial cells (MC), the response of intracellular Ca(2+) concentration ([Ca(2+)](i)) to a contractile agonist is biphasic with a large, transient increase in [Ca(2+)](i) followed by a smaller but sustained elevation as Ca(2+) flows into the cell from the extracellular fluid. It has been postulated that membrane depolarization precedes opening of Ca(2+) channels in the plasmalemmal membrane. However, a role for voltage-gated Ca(2+) channels (VGCC) in human MC has been controversial, and their existence has not been verified with single-channel analysis. We used fura 2 fluorescence and patch-clamp techniques to determine the properties of the Ca(2+) entry pathway responsible for the sustained response of [Ca(2+)](i) in human MC. We found that ANG II at 10 nM, 100 nM, and 1 microM increased [Ca(2+)](i) to sustained levels of 22%, 35%, and 49%, respectively, above baseline. The sustained response to 1 microM ANG II was attenuated by diltiazem and was reduced to a value less than baseline in the absence of external Ca(2+). None of the peak responses (due to release of intracellular stores of Ca(2+)) were affected by removal of external Ca(2+) or addition of diltiazem. Upon elevating the extracellular [K(+)] from 5 mM to 75 mM, [Ca(2+)](i) reached a sustained level of 48% greater than baseline. This effect of high K(+) was attenuated by either Ca(2+) removal or addition of diltiazem. In the presence of 75 or 140 mM K(+), the dihydropyridine agonist BAY K 8644 (1 microM and 10 microM) initiated sustained [Ca(2+)](i) responses averaging 18% and 25%, respectively, greater than baseline. With <10 nM Ca(2+) in the external solution, BAY K 8644 did not significantly affect [Ca(2+)](i). In separate patch-clamp experiments, barium-selective channels were found in cell-attached patches with 90 mM BaCl(2) and 10 microM BAY K 8644 in the pipette solution. The single-channel conductance was 11.2 pS, and the open probability increased steeply at membrane potentials between -30 mV and 0 mV. It is concluded that human glomerular MC contain dihydropyridine-sensitive Ca(2+) channels responsible for the voltage-regulated entry of Ca(2+) into the cell during an agonist-induced contraction.
Hall
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Department of Physiology and Biophysics, University of Nebraska Medical Center, Omaha, Nebraska 68198-4575, USA.
Carmines
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Sansom
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D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D000804:Angiotensin II / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000187:drug effects*; D002478:Cells, Cultured; D004110:Diltiazem / Q000494:pharmacology; D016257:Fura-2; D005920:Glomerular Mesangium / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000494:pharmacology
D020746:Calcium Channels, L-Type; D000804:Angiotensin II; D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D004110:Diltiazem; D011188:Potassium; D002118:Calcium; D016257:Fura-2