Advanced Neuronal Modulation with Semiconducting Graphitic Carbon Nitride: Insights from <i>In Vitro</i>, <i>In Vivo</i>, and <i>In Silico</i> Studies

Impaired neuronal functions and cell death within ailments such as neurodegenerative Parkinson's disease pose significant challenges due to their complex pathophysiology and limited treatment options. In this landscape, innovative materials with unique physicochemical properties that ameliorate the debilitated neuronal functions are critically required. Neuronal functions rely on the conduction of nerve impulses, a process that can be effectively targeted using advanced materials that exhibit conducive properties essential for modulating neural activity. For their semiconductor characteristics, combined with well-suited biocompatibility, graphitic carbon nitride (g-C₃N₄) nanosheets provide promising avenues for such neurotherapeutic applications. Our multidisciplinary study investigates the potential of g-C₃N₄ nanosheets in promoting neuronal differentiation and network formation across <i>in vitro</i> and <i>in vivo</i> systems. SH-SY5Y cells exposed to g-C₃N₄ demonstrated enhanced neuronal differentiation and neuritic outgrowth over a chronic 21-days period, accompanied by an increased intracellular Ca²⁺ influx, pivotal for dopamine biosynthesis, as evidenced by the upregulated expression of vesicular monoamine transporter 2 (VMAT2), aromatic l-amino acid decarboxylase (AADC), and tyrosine hydroxylase (TH) genes. Utilizing transgenic <i>Caenorhabditis elegans</i> model expressing human α-synuclein, we observed the neuroprotective potential of g-C₃N₄, as evidenced by reduced protein aggregation and improved dopaminergic functions. In the pursuit of exploring the mechanism of g-C₃N₄-induced neuronal stimulation, the semiconducting nature of g-C₃N₄ came forth, which was further validated using theoretical (<i>in silico</i>) models. These models demonstrated an increase in the chemical potential of the material upon the application of electrical biases. Studying Ca²⁺ channel inhibition, we also observed that phenotypic and molecular effects were the outcomes of the stimulation caused due to the presence of g-C₃N₄ nanosheets. Our findings, supported by experimental and <i>in silico</i> studies, suggest that g-C₃N₄ nanosheets can effectively modulate neuronal behavior through their semiconducting properties, offering promising avenues for therapeutic interventions in neurodegenerative diseases.