The involvement of PI3K/Akt/mTOR/GSK3β signaling pathways in the antidepressant-like effect of AZD6765

Highlights

• AZD6765 induced an antidepressant-like effect in mice submitted to TST.

• There is an involvement of PI3K/Akt/mTOR/GSK3β signaling in AZD6765 effects.

• AZD6765 decreased the latency to feed in the NFT, similarly to ketamine.

• The effects of AZD6765 in NFT were dependent on mTOR signaling.

• AZD6765 reversed the depressive-like behavior induced by CUS.

Abstract

AZD6765 (lanicemine) is a non-competitive NMDA receptor antagonist that induces a fast-acting antidepressant effect without presenting psychotomimetic effects. However, the mechanisms underlying its effects remain to be established. In this context, we demonstrated that a single administration of AZD6765 (1 mg/kg, i.p.) was able to induce an antidepressant-like effect in mice submitted to tail suspension test (TST), an effect reversed by LY294002 (a reversible PI3K inhibitor, 10 nmol/site, i.c.v.), wortmannin (an irreversible PI3K inhibitor, 0.1 μg/site, i.c.v.) and rapamycin (a selective mTOR inhibitor, 0.2 nmol/site, i.c.v.). In addition, the administration of sub-effective doses of AZD6765 (0.1 mg/kg, i.p.) in combination with lithium chloride (non-selective GSK-3β inhibitor, 10 mg/kg, p.o.) or AR-A014418 (selective GSK-3β inhibitor, (0.01 μg/site, i.c.v.) caused a synergistic antidepressant-like effect. These results suggest the involvement of PI3K/Akt/mTOR/GSK3β signaling in the AZD6765 antidepressant-like effect. In addition, western blotting analysis showed an increased immunocontent of synapsin in the prefrontal cortex and a tendency to an increased immunocontent of this protein in the hippocampus 30 min after AZD6765 administration, but no significant effect of AZD6765 was observed in P70S6K (Thr³⁸⁹) phosphorylation and GluA1 immunocontent. A single dose of AZD6765 (3 mg/kg, i.p.), similarly to ketamine (1 mg/kg, i.p.), decreased the latency to feed in the novelty suppressed feeding (NSF) test, a behavioral paradigm that evaluates depression/anxiety-related behavior. This effect was reversed by rapamycin administration, suggesting the activation of mTOR signaling in the effect of AZD in the NSF test. In addition, a single administration of AZD6765 (1 mg/kg, i.p.) or ketamine (1 mg/kg, i.p.) reversed the depressive-like behavior induced by chronic unpredictable stress (CUS). Altogether, the results provide evidence for the fast-acting antidepressant profile of AZD6765, by a mechanism likely dependent on PI3K/Akt/mTOR/GSK3β.

Introduction

NMDA receptors are a class of ionotropic glutamatergic receptors related to synaptic plasticity and cell survival (Glasgow et al., 2015). These receptors have been extensively studied in encephalic structures such as cerebral cortex and hippocampus and are highly implicated in psychiatric disorders such as major depressive disorder (MDD) (Hansen et al., 2014; Li et al., 2009). Glutamate excitotoxicity due to over-activation of these receptors has been implicated in the pathophysiology of these disorders (Fan and Raymond, 2007; Wang and Michaelis, 2010).

NMDA receptor antagonists have demonstrated beneficial effects for the treatment of MDD (Dang et al., 2014; Zarate et al., 2010). Preclinical studies have reported that these antagonists exhibit antidepressant-like effect in several animal models of depression (Belzung, 2014; Li et al., 2010). An NMDA receptor antagonist that has emerged as a promising therapeutic agent is ketamine, a dissociative anesthetic that has demonstrated fast and long-lasting antidepressant effects when acutely administered at subanesthetic doses in patients suffering from severe MDD symptoms (Berman et al., 2000; Zarate et al., 2006; Zarate et al., 2013). The antidepressant response elicited by ketamine is proposed to be mediated by glutamate increase with the consequent stimulation of BDNF receptors, which leads to the activation of phosphatidylinositol 3-kinase (PI3K)/Akt, inhibition of glycogen synthase kinase-3β (GSK3β) and stimulation of mammalian target of rapamycin (mTOR) (Monteggia et al., 2013; Wohleb et al., 2017). The activation of mTOR results in the synthesis of synaptic proteins such as synapsin, considered essential for the rapid antidepressant effect of ketamine (Li et al., 2010). However, the psychotomimetic effects of ketamine limit its clinical use (Zarate, 2020) raising the interest in other NMDA receptor antagonists with potential antidepressant effects.

AZD6765 (lanicemine), another non-selective, non-competitive NMDA receptor antagonist with lower trapping than ketamine, has demonstrated antidepressant properties in clinical studies without inducing psychotomimetic effects (Sanacora et al., 2014; Zarate et al., 2013). A single infusion of AZD6765 induced rapid but not sustained antidepressant effects compared to placebo (Sanacora et al., 2014). Of note, a single AZD6765 infusion was more effective than placebo when administered to 22 subjects without causing psychotic or dissociative effects (Zarate et al., 2013). However, a study by Sanacora et al. (2017) demonstrated no significant difference between lanicemine and placebo treatment on any outcome measures related to MDD. In preclinical studies, AZD6765 elicited antidepressant-like effect in rodents subjected to forced swim test and learned helplessness paradigm (Sanacora et al., 2014). Furthermore, the co-administration of hyperforin, a natural and biologically active compound extracted from Hypericum perforatum (Cervo et al., 2002) that attenuated symptoms of mild to moderate depression in several clinical trials, displayed long-lasting antidepressant-like effect when administered with AZD6765 in mice (Pochwat et al., 2018). Despite some promising antidepressant effects elicited by AZD6765, the mechanisms and signaling pathways related to its effects have not been investigated. Therefore, the purpose of the present study was to evaluate if the antidepressant effects elicited by AZD6765 are dependent on PI3K/Akt/mTOR and GSK-3β, targets implicated in fast antidepressant responses (Li et al., 2010; Pazini et al., 2016).