CALCIUM-DEPENDENT INACTIVATION OF INWARDLY RECTIFYING K+ CHANNEL IN A TUMOR MAST-CELL LINE

Antigenic stimulation of rat basophilic leukemia (RBL-2H3) cells, a tumor mast cell line, is associated with an increase in intracellular free Ca2+ concentrations ([Ca2+]i) and membrane polarization. We recorded whole cell and single-channel currents through the inwardly rectifying K+ channel, a major resting conductance of cells, using the patch-clamp technique, and we examined interactions between channel activity and [Ca2+]i. With 10-mu-M Ca2+ in the pipette, the amplitude of whole cell currents gradually declined within 5 min to 48 +/- 13% of that immediately after rupture of the patch membrane, in the presence of 1 mM ATP which minimized intrinsic rundown. In inside-out patches, activity of the channel was reduced by increasing the concentration of Ca2+ in the internal medium, both in the presence and absence of 1 mM ATP, with no apparent change in single-channel conductance. Time-averaged mean current activity in inside-out patches in the presence of 5-mu-M Ca2+ was < 50% of that with Ca2+ of 100 nM or less. These results suggest that a rise in [Ca2+]i leads to a closure of the inwardly rectifying K+ channel. In some inside-out patches, inward currents characterized by burst composed of rapid transitions between open and closed states were observed (flickering currents). Single-channel properties of the flickering currents are similar to the inwardly rectifying K+ channel except for kinetics (single-channel conductance of 24.5 +/- 7.9 pS, inward rectification, and permeability to K+). In addition, time-averaged mean current activity with Ca2+ > 5-mu-M reduced to 42 +/- 22% of that with Ca2+ < 100 nM mainly due to a decrease in channel openings. Whether the flickering currents passed the same inwardly rectifying K+ channel is yet to be determined. Discussion was made on the possible contribution of Ca2+-dependent inactivation of the inwardly rectifying K+ channel to stimulus-induced membrane potential changes in RBL-2H3 cells.