Caloric Constraint During Refeeding Optimizes the Neuroprotective Efficacy of Alternate-Day Fasting

Abstract Intermittent fasting (IF) confers neuroprotective effects in models of neurodegeneration, yet whether compensatory caloric intake during refeeding limits these benefits remains unclear. Here we combine longitudinal in vivo two-photon imaging with transcriptomic profiling in a TDP-43 proteinopathy mouse model to resolve the temporal dynamics of alternate-day fasting (ADF). Neuronal degeneration and microglial engagement oscillated with fasting–feeding cycles, decreasing during fasting but increasing during refeeding. Transcriptomic analyses revealed that refeeding triggered a rapid activation of metabolic and biosynthetic programs alongside inflammatory signaling, while suppressing fasting-induced neuroprotective pathways, indicating acute sensitivity of the neuroimmune axis to caloric transitions. Constraining caloric intake during the refeeding phase through a calorie-restricted ADF (crADF) regimen eliminated these oscillatory neuronal and microglial responses. Our findings identify the refeeding phase as a critical determinant of fasting efficacy and show that caloric precision during this window stabilizes neuroimmune homeostasis, thereby enhancing the therapeutic potential of intermittent fasting.