Abstract
Using the single-channel patch-clamp technique, we identified a Ca2+ channel in the apical membranes rabbit cultured proximal tubule cells. The channel is permeable to both Ca2+ and Ba2+ but not to monovalent cations. In on-cell patches, the channel opened infrequently and had a conductance of 4.6 +/- 0.9 pS (n = 5) with 105 mM CaCl2 in the pipette. Although addition of forskolin (12.5 microM) Cl2 is without effect, addition of phorbol 12-myristate 13-acetate (PMA, 1 microM) activated the channel. At 0 mV pipette voltage (resting state) in the on-cell patches, PMA increased open probability (P0) from 0 to 6.9 +/- 2.3% (n = 5) within 1-3 min of PMA application. Likewise, stretching the membrane patch (-10 to -30 mmHg) activated this channel (P0 increased to 5.3 +/- 2.1%, n = 3, at 0 mV applied pipette potential), with results consistent with a mechanosensitive channel. The channel displayed only modest voltage sensitivity, with mild activation on membrane hyperpolarization but with inactivation on strong depolarization. The addition of the L-type Ca2+ channel blocker (antagonist), nifedipine (10 microM), completely blocked this channel in both on-cell and inside-out patches, whereas the agonist, BAY K 8644 (5 microM) was without...Continue Reading
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