Beta-hydroxybutyric acid alleviates anxiety- and depression-like behaviors in mice by impeding complement 3-mediated microglial engulfment

Depression, as a common mental disorder, seriously endangers the health of the population, especially for adolescents. With the increasing prevalence of depression in adolescents and the limited use of antidepressant medications in adolescents, non-pharmacological treatments for adolescent depression also need to be explored more often. Beta-hydroxybutyric acid (BHB), the main active ingredient of the ketogenic diet, helps to protect neurological health, but whether it has a mitigating effect on depression in adolescents remains unclear. In this study, we investigated the mitigating effect of BHB on depressive-like behavior in adolescence and its underlying mechanisms. Mice were subjected to chronic unpredictable mild stress (CUMS) for modeling depression, and BHB (200 mg/kd/d) were administrated to mice for 10 days after CUMS. First, behavioral experiments, including the open-field test (OFT), elevated plus maze test (EPM), tail suspension test (TST), and sucrose preference test (SPT), revealed that BHB supplementation significantly restored anxiety- and depression-like phenotypes in mice. Further investigation using fiber photometry technology demonstrated that BHB restored the excitatory neuronal response in the medial prefrontal cortex (mPFC) during the TST, as evidenced by increased peak latency and amplitude of neuronal activity. Electrophysiological and morphological analyses further showed that BHB rescued CUMS-induced impairments in synaptic transmission, as indicated by increased frequency of miniature excitatory postsynaptic currents (mEPSC), and restored dendritic spine density in the mPFC. Synaptic structures and functions could be regulated by microglial engulfment. Mechanistically, BHB reduced the expression of Iba1 and restored microglial branching in CUMS mice, which suggested that BHB suppressed the pro-inflammatory polarization of microglia. Importantly, BHB decreased the expression of complement 3 (C3) and CD18 (a subunit of C3 receptors), suggesting that BHB may inhibit C3-mediated microglial engulfment of neuronal synapses. These findings demonstrate that BHB mitigates anxiety- and depression-like behaviors by impeding C3-mediated microglial engulfment in the mPFC. Collectively, BHB may represent a promising therapeutic approach for alleviating complement-related anxiety and depression, shedding light on novel strategies for neurodevelopmental and neuropsychiatric disorders.