AGE-DEPENDENT CONTROL OF SLEEP HOMEOSTASIS BY LAYER 5 MPFC NEURONS

While subcortical networks are central to sleep regulation, growing evidence indicates that the cortex, notably pyramidal neurons in layer 5 (L5), actively contributes to sleep homeostasis. This role is most prominent in the medial prefrontal cortex (mPFC), a region highly sensitive to sleep deprivation (SD) and a major source of slow-wave activity (SWA). Ageing disrupts mPFC function and SWA generation, yet it remains unclear whether these deficits reflect specific alterations in L5 activity. Moreover, how cognitive load during wakefulness influences L5 homeostatic response and whether this is altered by ageing is unknown. To address these questions, we combined electroencephalography (EEG) telemetry with fibre photometry calcium imaging in young adult (4–5 months) and aged (>18 months) Rbp4-Cre male mice. L5 neuronal activity was recorded across sleep and wake during a 24-hour baseline period, and during 3 hours of SD induced by either gentle handling (low cognitive load) or novel object exposure (high cognitive load), followed by a recovery period to assess the homeostatic sleep rebound. Currently, we have obtained data from 3 young and 8 aged mice and finalised the MATLAB analysis pipeline. We hypothesise that L5 calcium dynamics track sleep pressure accumulation and dissipation, with higher cognitive load eliciting stronger L5 activation during SD. We further predict that these dynamics are impaired in aged mice, consistent with increased sleep fragmentation and a blunted homeostatic rebound. Results from this study will highlight the contribution of mPFC L5 pyramidal neurons to the homeostatic regulation of sleep and its dysfunction in ageing.