Regulation of cytoplasmic pH of cultured bovine corneal endothelial cells in the absence and presence of bicarbonate

Summary Single sodium-channel currents were measured in neuroblastoma cells after inhibition of inactivation by chloramine-T (CHL-T), sea anemone toxin II (ATX-II) and scorpion toxin (SCT). The decaying phase of the averaged single-channel currents recorded with 90-msec pulses in cell-attached patches was clearly slower than that of the unmodified channels, suggesting inhibition of macroscopic inactivation. Each substance caused repetitive openings and a moderate increase in the channel open time. AtV _m =RP+20 mV andT=12°C, the mean channel open times were 1.4, 1.6 and 1.8 msec for CHL-T, ATX-II and SCT, respectively, as opposed to 1.07 msec for native channels. Open-time histograms could be best fitted by the sum of two exponentials. The time constants of the fits were similar for histograms constructed from single openings and from openings during bursts. This suggests that the population of channels is homogeneous and that in bursts the same open conformations of channels occur as in single openings. Mean burst durations for bursts consisting of more than one opening atV _m =RP+20 mV were 4.9, 5.8 and 6.1 msec for CHL-T, ATX-II and SCT, respectively. Burst open-time histograms constructed from two or three openings were fitted by the gamma function. The different time constants of the fits obtained for ATX-II and SCT suggested multiple open conformations of channels for openings of bursts. However, significantly different open-time histograms constructed from the first, second and third openings of bursts could not be obtained systematically. A positive correlation was found for the dwell time of the first and the second, as well as for the second and the third opening of bursts with each substance, but a negative one for the dwell time of an opening and the neighboring closing of bursts with ATX-II. The results suggest a model with multiple open and inactivated states. In this model the inactivated states are weakly absorbing.