Low chloride stimulation of prostaglandin E-2 release and cyclooxygenase-2expression in a mouse macula densa cell line

Reducing luminal NaCl concentration in the macula densa region of the nephr on stimulates renin secretion, and this response is blocked by a specific i nhibitor of cyclooxygenase-2 (COX-2) (Traynor, T. R., Smart, A., Briggs, J. P., and Schnermann, J. (1999) Am. J. Physiol. Renal Physiol, 277, F706-710 ), To study whether low NaCl activates COX-2 activity or expression we clon ally derived a macula densa cell line (MMDD1 cells) from SV-40 transgenic m ice using fluorescence-activated cell sorting of renal tubular cells labele d with segment-specific fluorescent lectins, MMDD1 cells express COX-2, bNO S, NKCC2, and ROMK, but not Tamm-Horsfall protein, and showed rapid Rb-86() uptake that was inhibited by a reduction in NaCl concentration and by bum etanide or furosemide. Isosmotic exposure of MMDD1 cells to low NaCl (60 mM ) caused a prompt and time-dependent stimulation of prostaglandin E-2 (PGE( 2)) release that was prevented by the COX-2 specific inhibitor NS-398 (10 m uM). Reducing NaCl to 60 and 6 mM for 16 h increased COX-2 expression in a chloride-dependent fashion. Low NaCl phosphorylated p38 kinase within 30 mi n and ERK1/2 kinases within 15 min without changing total MAP kinase levels . Low NaCl-stimulated PGE(2) release and COX-2 expression was inhibited by SE 203580 and PD 98059 (10 muM), inhibitors of p38 and ERK kinase pathways. We conclude that low chloride stimulates PGE(2) release and COX-2 expressi on in MMDD1 cells through activation of MAP kinases.