Vitamin C Modulates the PI3K/AKT Pathway via Glutamate and Nitric Oxide in Developing Avian Retina Cells in Culture

<b>Background:</b> In addition to its known antioxidant function, the reduced form of vitamin C, ascorbate, also acts as a neuromodulator in the nervous system. Previous work showed a reciprocal interaction of ascorbate with glutamate in chicken embryo retinal cultures. Ascorbate modulates extracellular glutamate levels by inhibiting excitatory amino acid transporter 3 and promoting the activation of NMDA receptors and the consequent activation of intracellular signaling pathways involved in transcription and survival. <b>Objective:</b> In the present work, we investigated the regulation of AKT phosphorylation by ascorbate in chicken embryo retina cultures. <b>Methodology:</b> Cultures of chicken embryo retina cells were tested using Western blot, immunocytochemistry, fluorescent probe transfection, and cellular imaging techniques. <b>Results:</b> Our results show that ascorbate induces a concentration and time-dependent increase in AKT phosphorylation via the accumulation of extracellular glutamate, the activation of glutamate receptors, and the activation of the PI3K pathway. Ascorbate produces an increase in intracellular calcium accumulation and, accordingly, AKT phosphorylation by ascorbate is blocked by the calcium chelator BAPTA-AM. Moreover, AKT phosphorylation is also blocked by the nitric oxide synthase inhibitor 7-nitroindazole, indicating that it is mediated by calcium and nitric oxide-dependent mechanisms. <b>Conclusions:</b> We demonstrate that ascorbate modulates the PI3K/AKT pathway in retinal cultures through the activation of glutamate receptors and NO production in a calcium-dependent manner. Given that previous research has shown that glutamate induces ascorbate release in retinal cultures, our findings emphasize the significance of the reciprocal interactions between ascorbate and glutamate in retinal development. These findings provide further evidence supporting the role of ascorbate as a neuromodulator in retinal development.