Activation of two types of Ca2+‐permeable nonselective cation channel by endothelin‐1 in A7r5 cells

In A7r5 cells loaded with the Ca2+ indicator fura-2, we examined the effect of a Ca2+ channel blocker SK&F 96365 on increases in intracellular free Ca2+ concentrations (Ca2+i) and Mn2+ quenching of fura-2 fluorescence by endothelin-1 (ET-1). Whole-cell patch-clamp was also performed. 2. Higher concentrations (> or = 10 nM) of ET-1 (higher ET-1) evoked a transient peak and a subsequent sustained elevation in Ca2+i: removal of extracellular Ca2+ abolished only the latter. A blocker of L-type voltage-operated Ca2+ channel (VOC) nifedipine at 1 microM reduced the sustained phase to about 50%, which was partially sensitive to SK&F 96365 (30 microM). 3. Lower ET-1 (< or = 1 nM) evoked only a sustained elevation in Ca2+i which depends on extracellular Ca2+. The elevation was partly sensitive to nifedipine but not SK&F 96365. 4. In the presence of 1 microM nifedipine, higher ET-1 increased the rate of Mn2+ quenching but lower ET-1 had little effect. 5. In whole-cell recordings, both lower and higher ET-1 induced inward currents at a holding potential of -60 mV with linear I-V relationships and reversal potentials close to 0 mV. The current at lower ET-1 was resistant to SK&F 96365 but was abolished by replacement of Ca2+ in the bath solution with Mn2+. The current at higher ET-1 was abolished by the replacement plus SK&F 96365. 6. In a bath solution containing only Ca2+ as a movable cation, ET-1 evoked currents: the current at lower ET-1 was sensitive to Mn2+, whereas that at higher ET-1 was partly sensitive to SK&F 96365. 7. These results indicate that in addition to VOC, ET-1 activates two types of Ca2+-permeable nonselective cation channel depending on its concentrations which differ in terms of sensitivity to SK&F 96365 and permeability to Mn2+.