Investigation On The Behavior And Related Neuro-Electrochemistry Of Potential Medications For The Treatment Of Substance Use Disorders.

National Institute On Drug Abuse

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Abstract

The dopamine (DA) transporter (DAT) is the main pharmacologic target of abused psychostimulants like cocaine but also of prescribed medications like modafinil (MOD), which has shown little, if any, propensity for abuse in clinical or preclinical studies. Recently, we have explored the neurochemical and behavioral actions of MOD to better characterize its atypical psychostimulant profile. We found that MOD had a lower potency and efficacy than cocaine in stimulating nucleus accumbens shell (NAS) and core (NAC) DA levels, but, at variance with abused psychostimulants there were no statistically significant regional differences between accumbens subregions. In drug-discrimination studies MOD showed cocaine-like subjective effects at lower doses and earlier onset times than expected based on its DA effects. Those results suggest that non-DA-dependent actions may be playing a role in its unique pharmacological profile. In order to discover a potential non-dopaminergic mechanism for these MOD actions, we compared its behavioral and neurochemical effects with those of methylphenidate, another clinically approved medication that inhibits the neuronal reuptake of DA. We showed that methylphenidate, but not MOD, maintained intravenous self-administration in Sprague-Dawley rats similarly to cocaine. Both MOD and methylphenidate pretreatments potentiated cocaine self-administration. Cocaine, at self-administered doses, stimulated mesolimbic DA levels. This effect was potentiated by methylphenidate, but not by MOD pretreatments, indicating DA-dependent actions for methylphenidate, but not MOD. MOD is known to facilitate electrotonic neuronal coupling by actions on gap junctions. Carbenoxolone, a gap junction inhibitor, antagonized MOD, but not methylphenidate potentiation of cocaine self-administration. Our results indicate that MOD shares mechanisms with cocaine and methylphenidate but has a unique pharmacological profile that includes facilitation of electrotonic coupling and lower abuse liability, which may be exploited in future therapeutic drug design for cocaine use disorder. Though MOD might prove useful as a treatment for specific addicted populations (e.g. heavy crack-cocaine users, or cocaine and methamphetamine addicts without alcohol abuse comorbidity), broader effective medications for psychostimulant use disorders are still an unmet medical need. To this end, several newly synthesized analogs of MOD, for example JJC8-091 and JJC8-088, have been tested in our preclinical models as potential medications for methamphetamine (METH) and cocaine use disorder. JJC8-091 and JJC8-088, were assessed alone and in combination with cocaine to elucidate neurochemical correlates to their divergent behavioral profiles. Despite sharing significant structural similarity, JJC8-088 was more cocaine-like, increasing extracellular DA concentrations in the NAS efficaciously and more potently than JJC8-091. In contrast, JJC8-091 was not self-administered and was effective in blocking cocaine-induced reinstatement to drug seeking. Electrophysiology experiments confirmed that JJC8-091 was more effective than JJC8-088 at inhibiting cocaine-mediated enhancement of DA neurotransmission. Further, when VTA DA neurons in DAT-cre mice were optically stimulated, JJC8-088 produced a significant leftward shift in the stimulation-response curve, similar to cocaine, while JJC8-091 shifted the curve downward, suggesting attenuation of DA-mediated brain reward. Computational models predicted that while JJC8-088 prefers or stabilizes an outward facing conformation of DAT, like cocaine, JJC8-091 stabilizes DAT towards a more occluded conformation. Recently, we have collaborated on a project related to the potential effects of cocaine on neural autophagy in vitro and in vivo. Autophagy inhibitors, i.e. hydroxychloroquine and SBI-0206965, reduce the locomotor stimulant effect of systemic administration of cocaine and its related stimulation of extracellular NAS DA levels in mice. Cocaine-induced autophagy degrades transporters for DA but not serotonin in the NAS. Autophagy inhibition impairs cocaine conditioned place preference in mice. These findings indicate that autophagic degradation of DAT modulates behavioral and neurochemical actions of cocaine. Our research collaborations also include an investigation on the role of the microRNA system on neuronal signaling in mice lacking the translin/trax microRNA-degrading enzyme (Tsn KO mice). We found an altered behavioral and neurochemical response to psychostimulants, and also robust adiposity in the context of normal body weight, suggesting that this phenotype might reflect elevated brain levels of the psychostimulants tested. Analysis of the effect of diet-induced increases in adiposity in wild type mice on brain amphetamine levels also demonstrated that brain amphetamine levels correlate with adiposity. Increased adiposity displayed by Tsn KO mice or by wild type mice fed a high-fat diet correlates with elevated brain amphetamine levels. As amphetamine and its analogues are widely used to treat attention deficit disorder, which is associated with obesity, further studies are warranted to assess the impact of adiposity on amphetamine levels in these patients. In a more recent collaboration, we investigated the role of Zinc in cocaine seeking and abuse vulnerability. Cocaine increases synaptic dopamine (DA) by blocking the DA transporter (DAT). This process regulates cocaine reinforcement, seeking, and abuse. Zn2+ is an essential element that binds to DAT, but the in vivo relevance of this is unknown. We found that cocaine use in humans correlated with Zn2+ content in the striatum, a brain region with high DAT density. In mice, cocaine increased synaptic Zn2+ release in the striatum. Striatal Zn2+ increased cocaines affinity and in vivo potency at the DAT and was required for cocaine-induced DAT upregulation. Finally, genetic or dietary Zn2+ manipulations modulated cocaine efficacy across different behavioral paradigms. These findings reveal new insights into cocaines pharmacological mechanism of action and establish synaptic Zn2+ as a critical environmentally-derived regulator of cocaine abuse vulnerability. Collectively, these data reveal the underlying molecular mechanism at DAT that may be leveraged to rationally optimize leads for the treatment of psychostimulant use disorders.