Nrf2 Regulates Basal Glutathione Production in Astrocytes

Astrocytes produce and export glutathione (GSH), an important thiol antioxidant essential for protecting neural cells from oxidative stress and maintaining optimal brain health. While it has been established that oxidative stress increases GSH production in astrocytes, with Nrf2 acting as a critical transcription factor regulating key components of the GSH synthetic pathway, the role of Nrf2 in controlling constitutive GSH synthetic and release mechanisms remains incompletely investigated. Our data show that naïve primary mouse astrocytes cultured from the cerebral cortices of Nrf2 knockout (Nrf2^−/−) pups have significantly less intracellular and extracellular GSH levels when compared to astrocytes cultured from Nrf2 wild-type (Nrf2^+/+) pups. Key components of the GSH synthetic pathway, including xCT (the substrate-specific light chain of the substrate-importing transporter, system x_c⁻), glutamate-cysteine ligase [catalytic (GCLc) and modifying (GCLm) subunits], were affected. To wit: qRT-PCR analysis demonstrates that naïve Nrf2^−/− astrocytes have significantly lower basal mRNA levels of xCT and both GCL subunits compared to naïve Nrf2^+/+ astrocytes. No change in mRNA levels of glutathione synthetase (GS) or the GSH exporting transporter, Mrp1, was found. Western blot analysis reveals reduced protein levels of both subunits of GCL, while (seleno)cystine uptake into Nrf2^−/− astrocytes was reduced compared to Nrf2^+/+ astrocytes, confirming decreased system x_c⁻ activity. These findings suggest that Nrf2 regulates the basal production of GSH in astrocytes through constitutive transcriptional regulation of GCL and xCT.