Abstract

α_1a-Adrenergic receptors (ARs) couple to phosphoinositide hydrolysis, adenylyl cyclase, and mitogen-activated protein kinase (MAPK) pathways. However, the interaction among these signaling pathways in activating extracellular signal-regulated kinase 1/2 (ERK1/2) is not well understood. We investigated the coupling of α_1a-ARs to ERK1/2 in Chinese hamster ovary (CHO)-K1 cells stably transfected with mouse α_1a-ARs, as well as the interaction between ERK1/2 and norepinephrine-induced cAMP accumulation. α_1a-AR activation by norepinephrine increased the cytosolic Ca²⁺ concentration and phosphorylated ERK1/2 in a time- and concentration-dependent manner. ERK1/2 phosphorylation was blocked by the MAPK kinase 1/2 inhibitor 2′-amino-3′-methoxyflavone (PD 98059) and the α₁-AR antagonist prazosin. A transient elevation in intracellular Ca²⁺ was required for the phosphorylation of ERK1/2; however, activation of protein kinase C did not seem to be required for ERK1/2 phosphorylation. Norepinephrine also stimulated cAMP accumulation in transfected CHO-K1 cells in a concentration-dependent manner via α_1a-ARs, which was blocked by the Ca²⁺ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid. Norepinephrine-induced ERK1/2 phosphorylation was inhibited by the adenylyl cyclase activator forskolin and was enhanced by the adenylyl cyclase inhibitor 9-(tetrahydro-2-furanyl)-9H-purine-6-amine (SQ 22536) and the protein kinase A inhibitor 4-cyano-3-methylisoquinoline. In conclusion, in transfected CHO-K1 cells, α_1a-AR activation activates both phospholipase C and adenylyl cyclase-mediated signaling pathways. α_1a-AR-mediated ERK1/2 phosphorylation was dependent on a rise in intracellular Ca²⁺, and this pathway was reciprocally regulated by the concomitant activation of adenylyl cyclase, which inhibits ERK1/2 phosphorylation. Thus, α_1a-AR stimulation of cAMP production may play an important role in regulating ERK1/2 phosphorylation in cell lines and native tissues.