In vivo chemogenetic generation of intraneuronal H₂O₂

Recombinant D-amino acid oxidase (DAAO) is extensively employed for controlled generation of intracellular H₂O₂. This chemogenetic tool offers unique opportunities for dissecting the contribution of oxidative stress in the occurrence of aging-related degenerative disorders. However, the employment of this tool to induce oxidative stress in the brain in vivo has not yet been tested. Here, we combine viral vector-mediated expression of DAAO and HyPer7, a genetically encoded fluorescent biosensor for H₂O₂, in hippocampal neurons with real-time in vivo fiber photometry to examine chemogenetic production of H₂O₂ upon delivery of D-norvaline via intraperitoneal injections or drinking water. Observed changes in the HyPer7 ratiometric signal in response to D-norvaline delivery through both routes confirmed controlled generation of intraneuronal H₂O₂in vivo. Raman microspectroscopy revealed a decrease of electron transport chain loading with electrons upon in vivo chemogenetic generation of intraneuronal H₂O₂. In sum, our findings enable us to suggest that the DAAO-based chemogenetic tool combined with delivery of D-norvaline via either intraperitoneal injections or drinking water is suitable for induction of chronic oxidative stress in the brain in vivo.