Regulation of serum‐ and glucocorticoid‐inducible kinase phosphorylation and catalytic activity in ventral tegmental area by chronic drugs of abuse

Abstract

Changes in the activity and output of dopamine neurons in the ventral tegmental area (VTA) by chronic drugs of abuse contribute to addiction‐related behaviors. Our lab has previously demonstrated that serum‐ and glucocorticoid‐inducible kinase 1 (SGK1) phosphorylation and catalytic activity are increased by chronic administration of cocaine and morphine. Further, using viral‐mediated overexpression studies, we have observed that decreasing VTA SGK1 catalytic activity decreases drug reward and intake, and that altering VTA SGK1 phosphorylation at serine 78 (S78) is sufficient to modulate cocaine reward and morphine intake, as measured by conditioned place preference and a two‐bottle choice task, respectively. However, little is known about regulation of SGK1 S78 phosphorylation and its role in kinase activity. Therefore, the aim of my study is to identify the kinases responsible for SGK1 S78 phosphorylation by chronic drugs of abuse, since this may serve as a novel node for therapeutic intervention. Using an in silico approach, I identified promising candidate kinases, namely: glycogen synthase kinase 3 beta (GSK3β), cyclin‐dependent kinase 5 (cdk5), extracellular signal‐regulated kinase 5 (ERK5), and ERK1/2. I utilized the Neuro2A mouse neuroblastoma cell line and established an induction protocol that robustly increases both SGK1 S78 phosphorylation and catalytic activity, as measured by phosphorylation of N‐myc downstream regulated gene (NDGR) protein, the only known SGK1‐specific substrate. I then evaluated the role of candidate kinases using specific pharmacological inhibitors: roscovitine (cdk5), GSK3β inhibitor XXVII (GSK3β kinase), U0126 inhibitor (ERK1/2) and BIX02189 (ERK5). I found that inhibition of any of upstream candidate kinases can dose‐dependently decrease SGK1 S78 phosphorylation without affecting the SGK1 catalytic activity, while SGK1‐specific inhibition (GSK650394) downregulates both S78 phosphorylation and SGK1 catalytic activity. These data suggest that our in silico candidates are valid and S78 phosphorylation is not necessary for kinase activity, but that changes in catalytic activity may influence S78 phosphorylation. We have now developed a series of mutant constructs to directly test the relationship between S78 phosphorylation and catalytic activity. Additionally, given our data that VTA overexpression of an SGK1 mutant that lacks S78 phosphorylation decreases morphine and cocaine reward, I will next test whether intra‐VTA infusion of pharmacological inhibitors is sufficient to prevent the cocaine‐ and morphine‐induced increase in SGK1 S78 phosphorylation and associated behavioral changes. Together, my work seeks to characterize the regulation of SGK1 S78 phosphorylation in drug‐related behaviors to identify a potential novel pharmacological target for improved treatment of addiction.Support or Funding InformationPhRMA Foundation Research Starter GrantR01 DA039895, NIDAThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.