Dopamine recruits D1Areceptors to Na-K-ATPase-rich caveolar plasma membranes in rat renal proximal tubules

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Abstract
Activation of dopamine D1A receptors in renal proximal tubules causes inhibition of sodium transporters (Na-K-ATPase and Na/H exchanger), leading to a decrease in sodium reabsorption. In addition to being localized on the plasma membrane, D1A receptors are mainly present in intracellular compartments under basal conditions. We observed, using [3H]SCH-23390 binding and immunoblotting, that dopamine recruits D1A receptors to the plasma membrane in rat renal proximal tubules. Furthermore, radioligand binding and/or immunoblotting experiments using pharmacological modulators showed that dopamine-induced D1A receptor recruitment requires activation of cell surface D1-like receptors, activation of adenylyl cyclase, and intact endocytic vesicles with internal acidic pH. A key finding of this study was that these recruited D1A receptors were functional because they potentiated dopamine-induced [35S]GTPγS binding, cAMP accumulation, and Na-K-ATPase inhibition. Interestingly, dopamine increased immunoreactivity of D1A receptors specifically in caveolin-rich plasma membranes isolated by a sucrose density gradient. In support of this observation, coimmunoprecipitation studies showed that D1A receptors interacted with caveolin-2 in an agonist-dependent fashion. The caveolin-rich plasma membranes had a high content of the α1-subunit of Na-K-ATPase, which is a downstream target of D1A receptor signaling in proximal tubules. These results show that dopamine, via the D1-like receptor-adenylyl cyclase pathway, recruits D1A receptors to the plasma membrane. These newly recruited receptors couple to G proteins, increase cAMP, and participate in dopamine-mediated inhibition of Na-K-ATPase in proximal tubules. Moreover, dopamine-induced recruitment of D1A receptors to the caveolin-rich plasma membranes brings them in close proximity to targets such as Na-K-ATPase in proximal tubules of Sprague-Dawley rats.