Multi-Regulatory Effects of Agmatine on Akt/GSK-3β/β-Catenin Signaling Pathway in SH-SY5Y cells: A Potential Explanation for its Conflicting Roles in Schizophrenia

Abstract

Objectives: Schizophrenia is a common psychiatric disease with its complex neurobiology. The current knowledge about the neurobiology of schizophrenia is based on dopaminergic and glutamatergic dysregulation of the central nervous system. The facts that all current antipsychotic drugs act on dopaminergic D2 receptor antagonism and glutamatergic N-Methyl D-Aspartate (NMDA) receptor antagonists reveal schizophrenia-like behavioral and structural alteration in rodents and humans are the major evidences of these hypotheses. However, they are not sufficient to explain the disease neurobiology and lead novel therapeutics. In recent years, it has been shown that the functions of certain intracellular pathways are disrupted in schizophrenia, especially Akt/glycogen synthase kinase 3beta (GSK-3β)/β-catenin signaling pathway. Certain studies showed that GSK-3β inhibition accompanied to the effects of antipsychotic drugs and specific inhibition of the GSK-3β might be a valuable approach for the novel treatments for schizophrenia. Agmatine is an endogenous amine that interacts with many receptors and enzymes including adrenergic, glutamatergic, cholinergic, serotonergic and imidazoline receptors and nitric oxide synthase enzyme. The limited number of studies showed conflicting results about the role of agmatine on schizophrenia and there was no study to evaluate the intracellular molecular mechanisms of agmatine`s schizophrenia-related effects. Herein, we aimed to investigate the effects of agmatine on MK-801 induced neurotoxicity and the disruption of the Akt/GSK-3β/β-catenin signaling pathway in SH-SY5Y human neuroblastoma cells. Methods: SH-SY5Y cells were grouped as the medium, agmatine (50μM), agmatine (100μM), olanzapine (10μM), MK-801 (100μM), MK-801+Agmatine (50μM), MK-801+Agmatine (100μM) and MK-801+Olanzapine (10μM). Real-time cell analysis (RTCA), real-time polymerase chain reaction (Rt-PCR) and western blot (WB) were used for monitoring the cell viabilities, quantifying gene and protein expressions, respectively. Cell viabilities monitored for the 24 hours and comparative viabilities (%) were calculated at the 6th, 12th and 24th hours of treatments. Akt, GSK-3β and β-Catenin gene expressions phosphorylated (p) and non-phosphorylated protein expressions were investigated at the 24th hour of treatments. Results: In RTCA, MK-801 administration decreased cell viabilities at the 6th (p<0.01) and 24th (p<0.001) hours compared to medium in cells. Agmatine (50μM) and agmatine (100μM) pre-treatments increased the cell viabilities at the 6th (p<0.05), 12th (p<0.01), and 24th (p<0.05) hours while olanzapine at 24th (p<0.05) compared to MK-801. In Rt-PCR, MK-801 administration increased GSK-3B decreased CTNNB1 gene expressions (p<0.001) compared to the medium. Agmatine (50μM) and olanzapine reversed (p<0.001) the effects of MK-801 for both gene expressions while agmatine (100μM) reversed (p<0.001) only GSK-3B. In WB, MK-801 administration decreased the p-GSK-3β/GSK-3β ratio compared to medium while only agmatine (50μM) increased the ratio compared to MK-801 administration. Conclusion: Herein, it has been shown that agmatine had a neuroprotective effect on MK-801 induced cell toxicity in SH-SY5Y cells. Our results have suggested a low dose (50μM) agmatine shows antipsychotic like effects on Akt/GSK-3β/β-catenin signaling pathway in SH-SY5Y cells bot, not high dose (100μM). We have suggested that these doses dependent regulatory effects of agmatine might be a possible explanation for its conflicting role in schizophrenia.