The Signal Transducer for the Dopamine-1 Regulated Sodium Transport in Renal Cortical Brush Border Membrane Vesicles

Abstract

We have reported the presence of dopamine-1 (DA-1) and dopamine-2 (DA-2) receptors in renal brush border and basolateral membranes. DA-1 agonists stimulate adenylate cyclase (AC) and phospholipase C (PLC) activity in both membranes. Moreover, the ability of a DA-1 agonist (fenoldopam) to stimulate PLC activity is independent of AC activity. A DA-2 agonist (LY171555) by itself was without effect and did not enhance the ability of the DA-1 agonist to stimulate PLC activity. The DA-1 but not DA-2 agonists inhibit Na⁺/H⁺ exchange activity in brush border membrane vesicles (BBMV) and Na⁺/K⁺-ATPase activity in basolateral membranes. However, cAMP inhibits, while protein kinase C (presumably via PLC activity) stimulates, Na⁺/H+ exchange activity. We therefore determined the effect of DA-1 agonists on Na⁺/H⁺ exchange activity when PLC or AC activity was blocked using neomycin or dideoxyadenosine, respectively. The drugs were incubated with minced renal cortex prior to preparation of BBMV by differential centrifugation and MnCl₂ precipitation. Enrichment of BBMV was not affected by drug treatment. The Na⁺/H+ exchange activity was assessed by measuring amiloride (1 mmol/L) sensitive ²²Na⁺ uptake in BBMV (pH_i = 5.5, pH_o = 7.5, Na_i⁺ = O, Na_o⁺ = 1 mmol/L). Neomycin inhibited DA and DA-1-stimulated PLC activity in BBMV in a concentration dependent manner (10^-6 to 10^-4 mol/L). Neomycin (10^-4 mol/L) completely blocked the ability of DA and DA-1 agonist to stimulate PLC activity but had no consistent effect on DA-1 inhibited Na⁺/H+ exchange activity. Dideoxyadenosine inhibited DA and DA-1 stimulated AC activity without affecting DA-1 stimulated PLC activity. Dideoxyadenosine (10^-4 mol/L) completely inhibited DA-1 stimulated AC activity and also blocked the inhibitory effect of DA-1 agonist on amiloride sensitive ²²Na⁺ uptake. Kinetic analysis of ²²Na⁺ uptake revealed that the DA-1 agonist (5 × 10~⁵ mol/L) decreased the maximum velocity (V_max) without affecting the affinity (K_m). Dideoxyadenosine blocked the DA-1 effect on V_max and had no effect on K_m. In additional studies, the isoquinoline sulfonamide derivative, H4, which inhibits protein kinase A activity also blunted the inhibitory effect of DA-1 agonist on ²²Na⁺ uptake. These studies suggest that the inhibitory effect of DA-l agonist on Na⁺/H+ exchange activity in BBMV is mediated via the stimulation of AC but not PLC activity. Am J Hypertens 1990;3:47S-50S