Role of cytochrome P-450 arachidonate metabolites in endothelin signaling in rat proximal tubule

We examined the rat proximal tubule (PT) response to endothelin-1 (ET-1) in terms of 20-hydroxyeicosatetraenoic acid (HETE) dependency. Arachidonic acid (AA) (1 muM) decreased ouabain-sensitive (86)Rb uptake from 2.1 +/-0.1 to 0.3 +/-0.08 ng Rb.10 mug protein(-1).2min(-1) (P<0.05); 20-HETE (1 M) had similar effects. Dibromododecenoic acid (DBDD) (2 muM), an inhibitor of omega -hydroxylase, abolished the inhibitory action of AA on (86)Rb uptake whereas the PT response to 20-HETE was unaffected. ET-1 at 0.1, 1, 10, and 100 nM reduced (86)Rb uptake from 2.8 +/-0.3 in control PTs to 2.4 +/-0.2, 1.7 +/-0.1, 0.67 +/-0.08, and 0.1 +/-0.03 ng Rb.10 mug protein(-1).2 min(-1), respectively. DBDD (2 mM) abolished the inhibitory effect of ET-1 on (86)Rb uptake as did BMS182874 (1 muM), an ET(A)-selective receptor antagonist. ET-1 (100 nM) significantly increased PT 20-HETE release by similar to 50%, an effect prevented by DBDD. N(omega)-nitro-L-arginine-methyl ester (L-NAME), given for 4 days to inhibit nitric oxide synthase (NOS), increased arterial pressure from 92 +/- 12 to 140 +/-8 mmHg and increased endogenous release of 20-HETE from isolated PTs (measured by gas chromatography/mass spectrometry). In L-NAME-treated PTs, but not in control PTs, 0.1 mM AA inhibited ouabain-sensitive (86)Rb uptake by >40%; the response to AA was attenuated by DBDD. We conclude that, in the PTs, 1) 20-HETE is a second messenger for ET-1 and 2) conversion of AA to 20-HETE is augmented when NOS is inhibited.