Hypothesis

During sleep, autophagic flux in adrenal zona reticularis cells acts as a selective editor that removes damaged mitochondria and misfolded steroidogenic enzymes, thereby preserving DHEA synthesis capacity. Chronic sleep disruption blunts this nocturnal autophagy, shifting the zona reticularis toward a catabolic state marked by accumulated oxidative damage and accelerated DHEA loss.

Mechanistic Basis

The glymphatic autopsy metaphor extends to peripheral endocrine organs: sleep provides a stereotyped low‑glucocorticoid window that permits AMPK activation and ULK1‑dependent autophagy initiation. In aging, flattened cortisol rhythms elevate nighttime glucocorticoid receptor (GR) signaling, which suppresses AMPK via increased REDD1 expression and mTORC1 reactivation. Consequently, autophagosome formation (LC3‑II conversion) falls, p62 accumulates, and damaged CYP17A1 and StAR proteins are not cleared. The resulting enzymatic deficit reduces pregnenolone conversion to DHEA, reinforcing the observed 1–2 % yearly decline.

Testable Predictions

1. Mice subjected to chronic fragmented sleep will show reduced LC3‑II/I ratios and elevated p62 specifically in zona reticularis (but not zona fasciculata) compared with rested controls.
2. This autophagy deficit will correlate with lower adrenal DHEA output and blunted corticosterone awakening response (CAR analogue).
3. Pharmacological enhancement of autophagy (e.g., spermidine or low‑dose rapamycin administered during the rest phase) will restore LC3‑II flux, rescue DHEA synthesis, and normalize the CAR analogue without altering overall corticosterone levels.
4. Genetic attenuation of GR in adrenal corticotropes (using Adcre‑GR floxed mice) will protect autophagic flux and DHEA levels despite sleep fragmentation, implicating GR‑mediated mTORC1 activation as the causal node.

Experimental Approach

• Model: C57BL/6 mice, 12 months old, exposed to 4 weeks of sleep fragmentation (gentle cage tapping every 2 min during the light phase). Controls receive undisturbed sleep.
• Readouts: adrenal corticosterone and DHEA measured by LC‑MS/MS at zeitgeber time 0 (awakening) and 6; immunoblot for LC3‑II, p62, phosphorylated AMPK, and phosphorylated S6K in microdissected zona reticularis vs fasciculata; electron microscopy for mitochondrial morphology; CAR analogue assessed via stress‑induced corticosterone surge after novel cage exposure.
• Intervention arms: (a) spermidine in drinking water (3 mM), (b) rapamycin (0.1 mg/kg i.p.) given at ZT18, (c) vehicle.
• Validation: repeat key measurements in Adcre‑GR floxed mice with and without sleep fragmentation.

Potential Outcomes

If predictions hold, the data will demonstrate that sleep‑dependent autophagy is a gatekeeper of adrenal steroidogenic fidelity, directly linking disturbed rest to the endocrine aging phenotype. Failure to observe autophagy changes or DHEA rescue would falsify the hypothesis, suggesting that nocturnal glucocorticoid excess acts through alternative pathways (e.g., transcriptional repression) or that systemic mechanisms dominate adrenal aging.