Elucidation of the purinergic P2X4‐dopamine receptor interaction in regulation of sensorimotor gating and underlying signaling molecules

Abstract

Purinergic P2X4 receptors (P2X4Rs) are ion channels gated by adenosine‐5′‐triphosphate in the brain. Recent investigations from our laboratory have reported a role for P2X4Rs in regulation of various behaviors including sensorimotor gating, emotional responses to acute stressors, motor behavior and drug reward. Amongst these behaviors, our laboratory was greatly interested in the role of P2X4Rs in regulation of sensorimotor gating considering that disruption of this function has major implications in wide spectrum of psychiatric disorders including schizophrenia, bipolar disorder and autism. Sensorimotor gating is the process by which irrelevant stimuli can be filtered out prior to generation of motor output in response to a sensory stimulus. This process allows an individual to efficiently navigate a stimulus‐laden environment with minimum distraction. Positive modulation of P2X4Rs by ivermectin (IVM) induced deficits in prepulse inhibition (PPI) of acoustic startle reflex, which is a reliable operational measure for sensorimotor gating. Additionally mice deficient in the p2rx4 gene [i.e. (P2X4R KO)] exhibited PPI deficits in comparison to the wildtype (WT) littermates. Interestingly, the PPI deficits in P2X4R KO mice were rescued by DA receptor antagonists indicating an interaction between P2X4Rs and DA receptors in PPI regulation. On the basis of findings from P2X4R KO mouse model, we hypothesized that modulation of DA receptors can regulate the effects of IVM on PPI. This was accomplished by testing the effects of DA receptor antagonists, SCH 23390 (for D1 receptors) (1mg/kg) and raclopride (for D2 receptors) (3 mg/kg) on IVM‐mediated PPI deficits. We also tested the effects of DA receptor agonists, SKF 82958 (for D1 receptors) (0.1mg/kg) and quinpirole (for D2 receptors) (0.1 mg/kg) on IVM‐mediated changes in behavior. To elucidate the mechanisms underlying the behavioral outcomes, we investigated the interaction between IVM and dopaminergic drugs on phosphorylation of signaling molecules linked to PPI including dopamine and cyclic‐AMP regulated phosphoprotein of 32kDa (DARPP‐32), Ca2+/calmodulin kinase II (CaMKII), neuronal nitric oxide synthase (nNOS) and secondary messengers such as guanosine‐3′,5′‐cyclic monophosphate(cGMP) in the ventral striatum . We found that SCH 23390 significantly attenuated the PPI disruptive effects of IVM to a much greater degree than raclopride. On the contrary, quinpirole, but not SKF 82958, potentiated IVM‐mediated PPI disruption. At the molecular level, SCH 23390 and raclopride significantly blocked IVM‐mediated changes in phosphorylation of all the signaling molecules in the ventral striatum. SKF 82958 significantly potentiated IVM’s effects on DARPP‐32 and CaMKIIα phosphorylation in the same brain region. Overall, these findings suggest that pharmacological activation of P2X4Rs can induce dysregulation of signaling cascades downstream of DA receptors which can result in PPI deficits. Finally, these findings represent P2X4Rs as a novel drug target for treatment of disorders with inherent DA dysfunction that can consequentially lead to sensorimotor gating perturbations.Support or Funding InformationNIAAA/NIH AA0224488 (D.L.D)IVM disrupted PPI function that was significantly reversed by SCH 23390 and raclopride (A). Neither of the DA receptor antagonists significantly impacted IVM‐mediated effects on startle magnitude (B). Values represent mean PPI % or startle amplitude ± SEM from 18–22 mice per treatment group. ***P <0.001 versus control group. #P<0.05, ##P<0.01 versus IVM‐treated mice. +++P<0.001 versus SCH 23390‐treated mice and $P<0.05 versus raclopride‐treated mice, Tukey’s multiple comparison test.Figure 1IVM modulated DARPP‐32 phosphorylation via P2X4R potentiation in a D1 receptor‐dependent manner in the ventral striatum. Inter‐dependence between P2X4Rs and D1 receptors in DARPP‐32 phosphorylation may be relevant to DA‐dependent behaviors such as sensorimotor gating (A). The ability of raclopride to block IVM‐mediated changes at behavioral and cellular level indicates a plausible interaction between P2X4Rs and D2 receptors in regulation of DA‐dependent behaviors and signaling pathways (B).Figure 2