CA2-SENSITIVE AND VOLTAGE-SENSITIVE - DIFFERENT PATHWAYS FOR BASAL AND SWELLING-INDUCED CA2+ ENTRY( ENTRY IN ISOLATED RAT CORTICAL COLLECTING DUCT IS PH)

We studied effects of extracellular pH (pH(o)), voltage and Ca2+-chann el blockers on the basal and swelling-induced increase in intracellula r Ca2+ ([Ca2+](i)) activity of isolated rat cortical collecting duct ( CCD) segments. [Ca2+](i) and membrane voltages (V-m) were measured usi ng the fura-2 method and the slow-whole-cell patch-clamp technique, re spectively. Fura-2 fluorescence ratio (FR) at a pH(o) of 7.4 was 1.04 +/- 0.02 (n = 121), corresponding to a calculated [Ca2+](i) of 97 +/- 6 nM. Depolarization of V-m by changes in extracellular K+ reduced FR (n = 6), and hyperpolarization of V-m increased FR (n = 6). Lowering p H(o) to 6.0 decreased FR by 0.07 +/- 0.01 (n = 20), elevation of pH(o) to 8.0 induced an increase in FR by 0.14 +/- 0.01 (n = 67) exclusivel y via Ca2+ influx. Ca2+ influx showed an increased Mn2+ quench of:Fura -2 fluorescence with alkaline pH(o) and a decreased quench with acidic pH(o). The alkaline pHo-induced increase in FR was completely abolish ed by Ni2+ (5 mM, n = 6) and La3+ (10 mu m, n = 8) and partially by Gd 3+ (10 mu M, n = 7). Cell swelling induced by a decrease in extracellu lar osmolality by 130 mosm led to an increase in FR of 0.46 +/- 0.05 ( n = 20). The swelling-induced increase was reduced at a pH(o) of 6.0 t o 0.27 +/- 0.06 (n = 10) and enhanced at pH(o) of 8.0 to 0.57 +/- 0.10 (n = 8). Ni2+ (n = 6) and Gd3+ (n = 7) had no, and La3+ (n = 8) only a partial inhibitory effect on the swelling-induced increase in FR. Ni 2+ quenched fura-2 fluorescence during cell swelling. Verapamil dimini shed the basal and swelling-activated Ca2+ influx only at a concentrat ion greater than or equal to 1 mu M. Neither nifedipine (1 mu M, n = 7 ) nor nimodipine (1 mu M, n = 5) had any effect on the swelling-induce d increase in FR. In rat CCD [Ca2+](i) is sensitive to pH, and V,. Bas al and cell swelling-induced Ca2+ influx is apparently mediated via di fferent pathways. L-type Ca2+ channels are most likely not responsible for this Ca2+ permeability.