Characterization of acute inhibition of Na/H exchanger NHE-3 by dopamine in opossum kidney cells

Background. Dopamine (DA) is a principal natriuretic hormone that defends extracellular fluid volume from a Na load. Natriuresis is effected partly through inhibiting the proximal tubule Na/H exchanger NHE-3. Changes in NHE-3 phosphorylation is one mechanism by which NHE-3 activity is regulated. Methods. We used opossum kidney (OK) cells to characterize the differential and synergistic effects of DA receptor subtype-1 (DA₁) and -2 (DA₂) agonists and the effect of blockade of protein kinase A (PKA) or protein kinase C (PKC) on NHE-3 activity and phosphorylation. Results. DA and DA₁ agonists inhibited NHE-3 activity, and DA₁ antagonist blocked the effect of either DA or DA₁ agonist. DA₂ agonist alone had no effect, but DA₂ antagonist reduced the DA effect on NHE-3 activity. DA₁ and DA₂ agonists together were more potent than DA₁ alone. PKA inhibition eliminated the effect of DA₁ agonist and partially blocked the effect of DA on NHE-3 activity. PKC inhibition did not block the DA effect. DA₁ agonist and PKA activation phosphorylated NHE-3 on identical sites. Despite lack of effect on NHE-3 activity, DA₂ agonists increased NHE-3 phosphorylation. DA-induced NHE-3 phosphorylation was distinct from DA₁ and PKA but closely resembled DA₂. Conclusion. We postulate the following: (1) DA modifies NHE-3 phosphorylation by activating PKA through DA₁ and by other kinases/phosphatases via DA₂. (2) DA₁ is sufficient to inhibit NHE-3, while DA₂ is insufficient but plays a synergistic role by altering NHE-3 phosphorylation.