Signaling Pathways Leading to Phosphorylation of Akt and GSK-3β by Activation of Cloned Human and Rat Cerebral D2 and D3 Receptors

Although dopamine (DA) regulates the serine/threonine kinase Akt and its downstream substrate glycogen synthase kinase-3β (GSK-3β), the direct influence of dopaminergic receptors remains poorly characterized. Short-term incubation of Chinese hamster ovary (CHO)-expressed human (h)D2L and hD3 receptors with DA (maximal effect, 5–10 min) phosphorylated Akt (Thr308 and Ser473) and GSK-3β (Ser9), actions blocked by the selective D2 and D3 antagonists, 3-\[4-(4-chlorophenyl)-4-hydroxypiperidin-l-yl]methyl-1 H -indole (L741,626) and (3 aR ,9 bS )- N [4-(8-cyano-1,3 a ,4,9 b -tetrahydro-3 H -benzopyrano[3,4- c ]pyrrole-2-yl)-butyl\] (4-phenyl)benzamide (S33084), respectively. Similar findings were acquired with the specific D2/D3 receptor agonist quinelorane, which also enhanced (10 min after administration) levels of p-Akt and p-GSK-3β in rat nucleus accumbens, an action blocked by the D2/D3 receptor antagonist raclopride. Akt and GSK-3β phosphorylation mediated via CHO-expressed hD2L and hD3 receptors was prevented by pertussis toxin and by inhibitors of insulin-like growth factor-1 receptors as well as phosphatidylinositol 3-kinase and Src. Likewise, chelation of intracellular Ca2+ and interference with an “atypical” phorbol ester-insensitive protein kinase C (PKC) abolished recruitment of Akt and GSK-3β. Inactivation of PKCμ blocked Akt and GSK-3β phosphorylation at hD2L receptors. However, blockade of conventional PKC isoforms attenuated the actions of DA at hD3 receptors only. Furthermore, phospholipase C (PLC), calmodulin, and Akt inhibitors abolished DA-induced GSK-3β phosphorylation by hD3 receptors, whereas phosphorylation by hD2L receptors partially involved calmodulin, Akt, and extracellular signal-regulated kinase (ERK) 1/2. In conclusion, at both hD2L and hD3 receptors, DA elicited a Gi/o- and Ca2+/calmodulin-dependent phosphorylation of Akt and GSK-3β via transactivation of insulin-like growth factor 1 receptor. However, significant differences were seen regarding the involvement of PLC, calmodulin, and ERK1/2.