Objective Define the role of Nrf2 in regulating cellular cobalamin content. Background Vitamin B12 (cobalamin) serves as a cofactor for methionine synthase, which catalyzes regeneration of methionine from homocysteine. Cellular cobalamin processing requires the availability of cytoplasmic glutathione and nicotinamide adenine dinucleotide phosphate (NADPH). Neuregulin-1 is a neurotrophic factor that stimulates phosphatidylinositol 3 kinase (PI3K) signaling. PI3K activation downstream of neuregulin-1 has been demonstrated to promote cobalamin processing to active species, by enhancing cysteine uptake and subsequent glutathione production. PI3K signaling and lithium can upregulate Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) activity by inhibiting glycogen synthase kinase 3 beta. Whether Nrf2 promotes cobalamin processing by modulating redox conditions through its role as a transcription factor is unclear. Methods SH-SY5Y cells grown in AMEM with 10% FBS were treated with 10 mM lithium chloride (LiCl). Cobalamin and thiol analysis were done via HPLC. SH-SY5Y cells grown in AMEM with 1% FBS were pre-treated with PI3K inhibitors pictilisib (pict) (100 nM) or wortmannin (wort) (100 nM), prior to 1h neuregulin-1 (1 nM) exposure. RT-qPCR was performed to measure gene expression of enzymes involved in redox-related cobalamin processing. Results 1 or 4h LiCl exposure increased glutathione. Cyanocobalamin (CNCbl) was decreased at both time points. Adenosylcobalamin (AdoCbl) was increased at 1h, but decreased after 4h. Neuregulin-1 treatment following pre-treatment with pict or wort modulated expression of Methylmalonic aciduria and homocystinuria type C protein (MMACHC) and Methionine synthase reductase (MTRR). MMACHC was increased by neuregulin-1 alone and decreased by neuregulin-1 with wort. MTRR was increased by wort alone or combination with neuregulin-1. Conclusion Our research strengthens the role of PI3K in regulating cobalamin processing and suggests Nrf2 may positively regulate cobalamin processing. Future research may directly implicate Nrf2.