Schizophrenia is a chronic psychotic disorder with heterogeneous (positive, negative and cognitive) symptoms1. Although the etiopathogenesis has not been completely elucidated due to the complexity of the molecular mechanisms involved, dysregulation of the dopaminergic, glutamatergic and serotonergic neurotransmitter systems has been identified, involving ion channels of NMDA, several G protein-coupled receptors (GPCRs), such as D2, D3, 5-HT2A and mGlu2, which represent main targets of typical and atypical antipsychotics used in the clinic of this disease2. The objective of this research is to evaluate D2 and NMDA receptor-modulated signaling in a previously developed in vitro phenotypic model of schizophrenia3.
Neuroblastoma cell line SH-SY5Y, differentiated by treatment with retinoic acid, GLP-1 and different supplements, was used to establish the in vitro model. To evaluate the functionality of dopaminergic, serotonergic and glutamatergic receptors, as well as to analyze the possible reciprocal modulation (crosstalk) among them, second messenger measurement assays were employed using functional cAMP assays (Cisbio, Ref: 62AM4PEC) and intracellular calcium mobilization assays (Molecular devices, Ref: R8190).
The study of GPCRs evidenced a dopamine response compatible with D2 receptor expression (EC50= 0.60 M), this response was inhibited after administration of its specific antagonist, haloperidol (0.1M) (EC50= 0.75 M; Kb= 0.409M). On the other hand, glutamate induced an increase in intracellular calcium levels (EC50= 5, 27M). When evaluating reciprocal modulation, it was found that NMDA receptor activation conditioned the effect of D2, increasing its potency (EC50= 0.090 M), likewise, D2 activation also modulated the NMDA response, reducing its EC50 to 0.27 M. Furthermore, it was observed that the dopaminergic response previously modulated by glutamate was inhibited after administration of the NMDA antagonist MK-801(1M) (EC50= 1.327 M), suggesting that the potentiating effect of glutamate on dopamine signaling depends on NMDA receptors, and that such synergy is lost upon blockade. In contrast, serotonin did not generate any response through 5-HT2A receptors in this model.
Accordingly, the presence of D2 and NMDA receptors was confirmed in our cellular model, validating it as a useful model for the study of mechanisms associated with schizophrenia. Likewise, a cross-modulation (crosstalk) between both receptors was evidenced, which could have significant implications for the development of new therapeutic strategies for this disease.
