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Am J Physiol Renal Physiol 278: F97-F103, 2000;
0363-6127/00 $5.00
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Vol. 278, Issue 1, F97-F103, January 2000

Dihydropyridine-sensitive Ca2+ channels in human glomerular mesangial cells

David A. Hall, Pamela K. Carmines, and Steven C. Sansom

Department of Physiology and Biophysics, University of Nebraska Medical Center, Omaha, Nebraska 68198-4575

In mesangial cells (MC), the response of intracellular Ca2+ concentration ([Ca2+]i) to a contractile agonist is biphasic with a large, transient increase in [Ca2+]i followed by a smaller but sustained elevation as Ca2+ flows into the cell from the extracellular fluid. It has been postulated that membrane depolarization precedes opening of Ca2+ channels in the plasmalemmal membrane. However, a role for voltage-gated Ca2+ 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 Ca2+ entry pathway responsible for the sustained response of [Ca2+]i in human MC. We found that ANG II at 10 nM, 100 nM, and 1 µM increased [Ca2+]i to sustained levels of 22%, 35%, and 49%, respectively, above baseline. The sustained response to 1 µM ANG II was attenuated by diltiazem and was reduced to a value less than baseline in the absence of external Ca2+. None of the peak responses (due to release of intracellular stores of Ca2+) were affected by removal of external Ca2+ or addition of diltiazem. Upon elevating the extracellular [K+] from 5 mM to 75 mM, [Ca2+]i reached a sustained level of 48% greater than baseline. This effect of high K+ was attenuated by either Ca2+ removal or addition of diltiazem. In the presence of 75 or 140 mM K+, the dihydropyridine agonist BAY K 8644 (1 µM and 10 µM) initiated sustained [Ca2+]i responses averaging 18% and 25%, respectively, greater than baseline. With <10 nM Ca2+ in the external solution, BAY K 8644 did not significantly affect [Ca2+]i. In separate patch-clamp experiments, barium-selective channels were found in cell-attached patches with 90 mM BaCl2 and 10 µM 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 Ca2+ channels responsible for the voltage-regulated entry of Ca2+ into the cell during an agonist-induced contraction.

diltiazem; BAY K 8644; voltage-gated calcium channel; fura 2; patch clamp


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