CHLORIDE AND FLUID SECRETION BY CULTURED HUMAN POLYCYSTIC KIDNEY-CELLS

Epithelial cells cultured from the renal cysts of patients with autoso mal dominant polycystic kidney disease (ADPKD) secrete fluid via a pro cess stimulated by adenosine 3',5'-cyclic monophosphate (cAMP). We hav e investigated the hypothesis that fluid secretion by these cells is d ependent on cAMP-mediated chloride secretion. Individual cultured ADPK D cells were suspended within a polymerized collagen matrix and stimul ated to form cysts. Individual cultured cysts were placed in a chamber on the stage of an inverted microscope equipped with epifluorescent a nd video analysis attachments. The rate of fluid secretion. cell volum e and changes in intracellular Cl- were measured. In the absence of se cretagogues, fluid was absorbed from the cyst cavity (-2.36 +/- 0.64 n l/min/cm(2) inner surface area). S-Bromoadenosine 3',5'-cyclic monopho sphate (8-Br-cAMP) plus 3-isobutyl-1-methlyxanthine (IBMX) induced a r apid reversal in the net movement of fluid to secretion (6.79 +/- 1.28 nl/min/cm(2)). Bumetanide reversibly reduced fluid secretion to 0.95 +/- 0.60 nl/min/cm(2). Cell volume rapidly decreased by 7.5 +/- 0.9% w ith the initiation of secretion and bumetanide caused an additional lo ss (4.2 +/- 1.0%). Furosemide had a similar effect on forskolin-induce d fluid secretion. Cellular chloride concentration was monitored with the use of the indicator, 6-methoxy-N-ethylquinolinium chloride (MEQ). Removal of Cl- from the bath reduced intracellular [Cl-] (MEQ fluores cence increased by 11.4 +/- 2.3%). In cysts pretreated with furosemide to prevent Cl- entry, the application of forskolin caused a decrease in Cl- concentration (MEQ fluorescence increased by 9.3 +/- 2.6%). Usi ng monolayers of cultured ADPKD cells, grown on permeant supports, we compared the changes in short circuit current (I-SC) induced by forsko lin in the presence and absence of external Cl-. Forskolin increased I -SC (from 8.9 +/- 2.7 to 10.6 +/- 2.7 mu A/cm(2)) in the presence of C l-, but did nor significantly affect I-SC in its absence. These data i ndicate that cultured ADPKD cells can direct fluid transport in either the absorptive or the secretory direction, and that cAMP stimulates s ecretion and this secretion is accompanied by a net loss of cell solut e. Inhibition of secretion by bumetanide or furosemide caused an addit ional loss of cell solute, including Cl-. The ionic transepithelial cu rrent induced by forskolin is dependent on the presence of Cl-. These data support the thesis that chloride secretion drives fluid secretion by cultured ADPKD cells.