Electrophysiological studies of primary cultures of XXXX distal tubule cells.

@entity268 @entity95 @entity6478 @entity3013 @entity3044
Primary monolayers grown from F1 band of a Percoll gradient centrifugation ("distal" monolayers) were studied, after confluency, 6-14 days after seeding. Transmission and scanning electron microscopy revealed that two cell types, resembling principal cells of the @entity95 cortical collecting tubule (CCT) and intercalated cells of either CCT or connecting tubule, constitute approximately 96% of the monolayer. About two-thirds of the intercalated cells fluoresced when treated with fluorescent peanut lectin. Indirect specific immunocytofluorescent staining revealed fluorescence in 96% of the cells, confirming that the monolayers were derived from CCT or connecting tubule cells. Exposure of monolayers to vasopressin or @entity3044 increases @entity268 ( @entity268 ) content in the cells and bathing medium, whereas parathyroid hormone was ineffective. Electrophysiological studies revealed a transepithelial voltage (VT) of -11 +/- 2 mV, and basolateral membrane voltage (Vb) of -77 +/- 5 mV (n = 20). The transepithelial electrical @entity3013 ( @entity3013 ) was 1,870 +/- 250 omega X cm2 (n = 13). In three out of six monolayers, @entity6478 (10(-5) M) applied to the apical side produced an increase in apical membrane voltage (Va) from -71 +/- 1 to -89 +/- 9 mV) and a decrease in VT (from -10 +/- 1 to -2 +/- 1 mV). The @entity3013 did not change during @entity6478 exposure. Exposure of the apical membrane to 140 mM K+-depolarized Va from -67 +/- 7 to -39 +/- 11 mV (P less than 0.002) and hyperpolarized VT from -7 +/- 2 to -15 +/- 3 mV (P less than 0.005). Exposure to high K+ from the basolateral side depolarized Vb from -76 +/- 11 to -43 +/- 10 mV (P less than 0.001) and depolarized VT from -9 +/- to 8 +/- 5 mV (P less than 0.001). This preparation is suitable to study basic aspects of epithelial transport by electrophysiological methods and other techniques. The findings are consistent with several of the known properties of cortical collecting tubules from @entity95 studied by the isolated perfused tubule technique.
