During the present reporting period, modest laboratory work was conducted on this project. In addition, a backlog of laboratory findings on this project were brought forward to the publication stage during this reporting period. First, we reported that SB277011A does not alter Pavlovian conditioned approach behaviors in rats (sign-tracking versus goal-tracking). Second, we reported on a new and novel series of compounds with exceptionally high dopamine D3 receptor affinities and/or selectivities. In this series, compounds BAK4-54 and CAB2-015 are novel high affinity D3R antagonists/partial agonists with improved metabolic stability over our previous lead compound PG648. Both compounds have dual functional profiles as full D3R antagonists at low doses (IC50 = 8.0 nM or 7.4 nM for BAK4-54 or CAB2-015, respectively) and partial D3R agonists at higher doses (EC50 = 140 nM, 25% stimulation or 20 nM, 31% stimulation for BAK4-54 or CAB2-015, respectively). In radiolabeled binding experiments, both compounds bind to D3R with subnanomolar affinities whereas BAK4-54 is over 100-fold selective for hD3Rs over the highly homologous hD2R, and CAB2-015 is somewhat less D3R selective(45-fold). We first evaluated the reinforcing effects of oxycodone in a rat model of self-administration and then explored the potential utility of CAB2-015 and BAK4-54, for treatment of prescription opioid abuse and dependence. We found that rats acquired oxycodone self-administration rapidly within a range of unit doses that was similar to that for heroin, confirming that oxycodone has significant abuse potential. Strikingly, pretreatment with either CAB2-015 or BAK4-54 (0.4-10 mg/kg, i.p.) dose-dependently decreased oxycodone self-administration, and shifted the oxycodone dose-response curve downward. Repeated pretreatment with CAB2-015 or BAK4-54 (0.4-4 mg/kg) facilitated extinction and inhibited oxycodone-induced reinstatement of drug-seeking behavior. In addition, pretreatment with CAB2-015 or BAK4-54 (4-10 mg/kg) also dose-dependently decreased oxycodone-enhanced locomotor activity, but only CAB2-015 decreased oral sucrose self-administration. These data suggest that D3R antagonists may be suitable alternatives or adjunctives to opioid-based medications currently used clinically in treating opioid addiction and that the D3R-selective compounds CAB2-015 or BAK4-54 provide new lead molecules for development. Third, we explored whether dopamine D3 receptor alteration constitutes a vulnerability risk factor for opioid abuse. The neurobiological risk factors that convey vulnerability to opioid abuse are unknown. Recent studies suggest that the D3R is involved in opioid self-administration, but it remains unclear whether altered D3R availability is a risk factor for the development of opioid abuse and addiction. Therefore, we used dopamine D3 receptor-knockout (D3-KO) mice to investigate the role of this receptor in the different phases of opioid addiction. D3-KO mice learned to self-administer heroin faster and took more heroin than wild-type mice during acquisition and maintenance of self-administration. D3R-KO mice also displayed higher motivation to work to obtain heroin reward during self-administration under progressive-ratio reinforcement, as well as elevated heroin-seeking during extinction and reinstatement testing. In addition, deletion of the D3R induced higher baseline levels of extracellular dopamine in the nucleus accumbens (NAc), higher basal levels of locomotion, and reduced NAc dopamine and locomotor responses to lower doses of heroin. These findings suggest that the D3R is critically involved in regulatory processes that normally limit opioid intake via DA-related mechanisms. Deletion of the D3R augments opioid-taking and opioid-seeking behaviors. Therefore, low D3R availability in the brain may represent a risk factor for the development of opioid abuse and addiction. Finally, we developed a rodent model of post-traumatic stress disorder (PTSD), and studied whether selective D3R antagonism alters stress-induced behavior in this model. In both animals and humans, dopamine function appears to be dysregulated in brain areas involved in the conditioned fear response(s) that underlie PTSD. We therefore studied the effect of the selective DA D3 receptor antagonists YQA-14 (6.25, 12.5, or 25 mg/kg i.p.) and SB-277011A (6 mg/kg i.p.) on tone-induced fear (assessed by measuring freeze time) in a modified version of the single-prolonged stress (SPS) model of PTSD in adult male Sprague-Dawley rats. Rats pretreated with vehicle and then subjected to restraint stress, forced swim, and random foot shock - all in the presence of a distinctive tone - displayed significantly increased tone-induced contextual freeze time following re-exposure to the tone. Rats pretreated with a single i.p. injection of 6.25 or 12.5 mg/kg of YQA-14 or 6 mg/kg of SB-277011A showed significantly attenuated contextual freeze time in the presence of the tone when tested 14 days after exposure to the stressors. Overall, our results indicate that selectively antagonizing DA D3 receptors significantly decreases freezing time caused by an environment previously associated with stress. If our findings can be extrapolated to humans with PTSD, they suggest that DA D3 receptors may play a role in the pathophysiology of PTSD, and that selective D3R antagonists may have therapeutic utility for the clinical management of PTSD.
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