REGION-SPECIFIC CHANGES IN BRAIN PEPTIDOGLYCAN FOLLOWING SLEEP DEPRIVATION

Abstract Introduction Bacterial cell wall peptidoglycan (PG) and muramyl peptides (MPs), isolated from mammalian brains and urine following sleep deprivation (SD), promote non-rapid eye movement sleep. These PG/MPs likely originate from the host microbiome and have been quantified in neonatal murine brain. PG/MP amounts and dynamics in healthy, adult murine brain remain unknown. Methods Wildtype mice acclimated to standard lab conditions were sacrificed at Zeitgeber time (ZT) 3 or ZT15 with (treatment, N=8), or without (control, N=8) 3h of SD prior to time points. Hypothalamic (HT), somatosensory cortex (Sctx) and brain stem (BS) areas were dissected, homogenized in phosphate buffered saline and centrifuged. PG/MP contents in resultant supernatants were determined using an ELISA (MyBioSource), interpolating sample PG from the standard curve, and expressed as ng peptidoglycan per mg tissue wet weight (ng/mg). Results At ZT3 and ZT15, BS PG values were significantly higher than HT or Sctx values, while HT and Sctx values did not differ from each other. At ZT3, mean PG values from control mice were: 3.6 in HT, 3.7 in Sctx, and 8.6 ng/mg in BS. After SD, corresponding values were: 3.0, 4.8 (statistically significant increase, p<0.05), and 7.5 ng/mg. Further, within all 8 individual mice after SD prior to ZT3, PG levels in Sctx were higher than corresponding values in HT (p<0.001).At ZT15, PG control values were: 4.6 in HT, 4.6 in Sctx, and 8.9 ng/mg in BS. After SD, PG level at ZT15 was not significantly changed in any brain area assayed. However, PG values after SD at ZT15 compared to ZT3 SD values were significantly higher for HT and BS (p<0.0005 and p<0.005, respectively). In an independent experiment (see Dykstra-Aiello et. al. this volume) we confirmed PG values at ZT15 in BS were significantly higher than Sctx values. Conclusion Results indicate unique PG regulation by brain area, sleep loss, and time-of-day suggesting physiological roles for brain PG guiding host behaviors such as sleep. Thus, mammalian sleep-wake regulation and its various associated cognitive states, are the product of millions of years of co-evolutionary symbioses between microbes and their hosts. Support (If Any) Supported by the W.M. Keck Foundation and NIH (NS025378).