title: Regulation of PI3K/Akt signaling by N-desmethylclozapine through activation of δ-opioid receptor.
abstract: We have previously reported that N-desmethylclozapine (NDMC), a major clozapine metabolite, acts as a δ-opioid receptor agonist. Here, we show that in different cellular systems NDMC regulates protein kinase B/Akt (Akt) signaling through the activation of δ-opioid receptors. In Chinese hamster ovary cells transfected with the human δ-opioid receptor (CHO/DOR), NDMC induced a time- and concentration-dependent phosphorylation of Akt at Thr308 and glycogen synthase kinase-3β (GSK-3β) at Ser9 and these effects were fully blocked by the δ-opioid receptor antagonist naltrindole. NDMC-induced Akt and GSK-3β phosphorylations were completely prevented by pertussis toxin, the Src tyrosine kinase inhibitor PP2 and the selective insulin-like growth factor-I (IGF-I) receptor tyrosine kinase inhibitor tyrphostin AG 1024. NDMC stimulated IGF-I receptor β subunit tyrosine phosphorylation and this effect was prevented by either naltrindole or PP2. Blockade of phosphatidylinositol 3-kinase (PI3K) α, but not PI3Kγ, suppressed NDMC-induced Akt and GSK-3β phosphorylation, whereas inhibition of Akt curtailed the stimulation of GSK-3β phosphorylation. In rat nucleus accumbens, NDMC induced Akt and GSK-3β phosphorylation either in vitro or in vivo and these effects were prevented by naltrindole. NDMC also regulated Akt and GSK-3β phosphorylation through δ-opioid receptors in NG108-15 cells. In these cells NDMC counteracted oxidative stress-induced apoptosis and the effect was lost following PI3K inhibition. These data demonstrate that in different cell systems NDMC can stimulate Akt signaling by activating Gi/Go-coupled δ-opioid receptors, which, at least in CHO/DOR cells, regulate PI3Kα through Src-dependent transactivation of the IGF-I receptor, and indicate that through this mechanism NDMC can exert neuroprotective effects.
authors.affiliation: Section of Biochemical Pharmacology, Department of Neuroscience, University of Cagliari, Monserrato, Cagliari, Italy.
authors.forename: Maria C
authors.initials: MC
authors.lastname: Olianas
authors.forename: Simona
authors.initials: S
authors.lastname: Dedoni
authors.forename: Pierluigi
authors.initials: P
authors.lastname: Onali
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D016466:CHO Cells; D003024:Clozapine / Q000031:analogs & derivatives* / Q000494:pharmacology; D006224:Cricetinae; D003412:Cricetulus; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006822:Hybrid Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D008297:Male; D051379:Mice; D009714:Nucleus Accumbens / Q000187:drug effects / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D051381:Rats; D017465:Receptors, Opioid, delta / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*
chemical_list: D017465:Receptors, Opioid, delta; C058272:norclozapine; D019869:Phosphatidylinositol 3-Kinases; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; C000605508:Gsk3b protein, rat; D051057:Proto-Oncogene Proteins c-akt; D038362:Glycogen Synthase Kinase 3; D003024:Clozapine