Kinetics and regulation of a Ca -activated Cl conductance in mouse renal inner medullary collecting duct cells

Boese, S. H., O. Aziz, N. L. Simmons, and M. A. Gray. Kinetics and regulation of a Ca -activated Cl conductance in mouse renal inner medullary collecting duct cells. Am J Physiol Renal Physiol 286: F682–F692, 2004. First published December 16, 2003; 10.1152/ ajprenal.00123.2003.—Using the whole cell patch-clamp technique, a Ca -activated Cl conductance (CaCC) was transiently activated by extracellular ATP (100 M) in primary cultures of mouse inner medullary collecting duct (IMCD) cells and in the mouse IMCD-K2 cell line. ATP also transiently increased intracellular Ca concentration ([Ca ]i) from 100 nM to peak values of 750 nM in mIMCD-K2 cells, with a time course similar to the ATP-induced activation and decay of the CaCC. Removal of extracellular Ca had no major effect on the peak Cl conductance or the increase in [Ca ]i induced by ATP, suggesting that Ca released from intracellular stores directly activates the CaCC. In mIMCD-K2 cells, a rectifying timeand voltage-dependent current was observed when [Ca ]i was fixed via the patch pipette to between 100 and 500 nM. Maximal activation occurred at 1 M [Ca ]i, with currents losing any kinetics and displaying a linear current-voltage relationship. From Ca -dose-response curves, an EC50 value of 650 nM at 80 mV was obtained, suggesting that under physiological conditions the CaCC would be near fully activated by mucosal nucleotides. Noise analysis of whole cell currents in mIMCD-K2 cells suggests a singlechannel conductance of 6–8 pS and a density of 5,000 channels/cell. In conclusion, the CaCC in mouse IMCD cells is a low-conductance, nucleotide-sensitive Cl channel, whose activity is tightly coupled to changes in [Ca ]i over the normal physiological range.