WHOLE-CELL SODIUM CONDUCTANCE OF PRINCIPAL CELLS FRESHLY ISOLATED FROM RAT CORTICAL COLLECTING DUCT

Cortical collecting duct fragments were manually dissected from 6-wk-o ld Sprague-Dawley rats. The fragments were enzymatically digested (col lagenase A) into single cells, washed, and resuspended in serum-free R PMI 1640. Individual cells were examined electrophysiologically using the whole cell patch-clamp technique. Two morphologically distinct cel l types were present in the cell suspension. Small round cells that ha d a capacitance of 7 pF and larger oval cells with a capacitance of 29 pF were consistently observed. Whole cell electrophysiological examin ation revealed that the small round cells had virtually no plasma memb rane ionic conductance, whereas both inward and outward currents were observed in the larger oval-type cells. Also, superfusion of 250 pM ar ginine vasopressin specifically increased the inward conductance of on ly the larger cells. The effect could be completely inhibited by 2 mu M amiloride or 100 mu mol of the Rp diastereomer of 8-(4-chlorophenylt hio)-adenosine 3',5'-cyclic monophosphate (a specific adenosine 3',5'- cyclic monophosphate inhibitor). These findings are consistent with th e hypothesis that the larger cells are principal cells and the smaller cells are intercalated cells and directly demonstrate that an amilori de-sensitive whole cell conductance is readily observable in freshly i solated cortical collecting duct cells. Thus the whole cell configurat ion of the patch-clamp technique appears to be well suited for assessi ng cellular mechanisms that regulate the ionic conductances of cortica l collecting duct cells.