Renal sympathetic nerves play a central role in the regulation of tubular Na⁺ reabsorption. Norepinephrine (NE) and neuropeptide Y (NPY) are colocalized in renal sympathetic nerve endings. The purpose of this study is to examine the integrated effects of these neurotransmitters on the regulation of Na⁺-K⁺-ATPase, the enzyme responsible for active Na⁺ reabsorption in renal tubular cells. Studies were performed on proximal tubular segments, which express adrenergic - and -receptors, as well as NPY-Y₂ receptors. It was found that - and -adrenergic agonists had opposing effects on Na⁺-K⁺-ATPase activity. -Adrenergic agonists induced a dose-dependent inhibition of the Na⁺-K⁺-ATPase activity, whereas -adrenergic agonists stimulated the enzyme. NPY abolished -agonist-induced deactivation of Na⁺-K⁺-ATPase and enhanced -agonist-induced activation of Na⁺-K⁺-ATPase. The -adrenergic agonist appeared to inhibit Na⁺-K⁺-ATPase activity via a cAMP pathway. NPY antagonized -agonist-induced accumulation of cAMP. In our preparation, NE alone had no net effect but stimulated the Na⁺-K⁺-ATPase activity in the presence of -adrenergic antagonists, as well as in the presence of NPY. The results indicate that, in renal tissue, NPY determines the net effect of its colocalized transmitter, NE, by its ability to attenuate the - and enhance the -adrenergic effect.