SYNTHESIS AND SECRETION OF ENDOTHELIN IN A CORTICAL COLLECTING DUCT CELL-LINE

Previous experiments have shown that epithelial cells in the renal med ulla produce endothelin-1 (ET-1) and possess ET(B) receptors. It has b een suggested that medullary ET-1 may affect water and sodium absorpti on along the collecting ducts in an autocrine fashion. To study possib le mechanisms responsible for the regulation of medullary ET-1 product ion, experiments were performed in M-1 cells and mIMCD-K2 cells, cell lines derived from cortical and inner medullary collecting ducts of SV 40 transgenic mice, grown to confluence on collagen-coated filter inse rts. Both cell lines were found to express ET-1 mRNA and to secrete ET almost exclusively into the basolateral medium as long as the transep ithelial resistance was high. Inhibition of transcription with actinom ycin D was followed by a decline in both ET mRNA [halftime (t(1/2)) = 30 min] and ET secretion (t(1/2) = similar to 90 min). The addition of arginine vasopressin (AVP, 10(-8) M; 2- or 4-h exposure) or incubatio n of M-1 cells in hypertonic media (+50 mM NaCl, 4- or 6-h exposure) d id not significantly alter ET secretion or ET-1 mRNA expression. In co ntrast, simultaneously increasing AVP (10(-8) M in the basolateral med ium) and tonicity (+50 mM NaCl) for 4 h increased ET secretion (from 2 8.9 +/- 3.9 to 41.8 +/- 3.8 pg . h(-1). mg protein(-1); P = 0.029, n = 10) and ET-1 mRNA (control = 2,138 cpm/mu l, log of 3.33 +/- 0.048, n = 4; AVP+NaCl = 3,548.1 cpm/mu l, log of 3.55 +/- 0.09; P = 0.045, n = 5). Exposure of M-1 cells to hypertonic media (+50 mM NaCl or 100 mM mannitol) for 24 h was associated with a marked reduction of ET secre tion (-83.9% with NaCl and -78.4% with mannitol; P < 0.001). This redu ction was attenuated, but not prevented, by the presence of AVP in the basolateral medium (-40%). ET-1 mRNA, in contrast, did not change wit h 24-h exposure to hypertonic media and increased when AVP was present . Results are compatible with the concept that generation of ET by col lecting duct cells may contribute in a complex and time-dependent fash ion tot he paracrine control of collecting duct cell function.