EFFECTS OF SLEEP DEPRIVATION IN THE PROCESS OF TAU PROTEIN DEPHOSPHORYLATION FOLLOWING SYNTHETIC TORPOR IN THE RAT

Introduction:One of the main hallmarks of tauopathies is the hyperphosphorylation of neuronal Tau protein. Interestingly, the accumulation of hyperphosphorylated Tau (PPTau) also characterizes “synthetic torpor” (ST), a torpor-like condition that can be pharmacologically induced in rats (Cerri et al., 2013). However, after few hours of the recovery from ST, PPTau levels reverse to control (Luppi et al., 2019). Since during the first hours of the recovery from ST rats show a strong sleep-pressure, aim of the present study has been to investigate the role of sleep in the dephosphorylation of PPTau following ST. Materials and Methods:Twelve Sprague-Dawley male rats (250-350g), adapted to an ambient temperature (Ta) of 24±0.5 C and to a 12h:12h Light-Dark cycle, were implanted under general anesthesia with a microcannula in the Raphe Pallidus (RP). After one-week of recovery, ST was induced for 6 hours according to the protocol described by Cerri et al. (2013), by the repeated injection (one injection/h) in the RP of the GABA-A agonist muscimol (100 nL - 1 mM). Soon after their return to normothermia, animals were either sleep deprived by gentle handling for 3 (n¼3) or 6 (n¼3) hours (R3SD and R6SD, respectively) or allowed to sleep (normal sleep, NS) for 3 (n¼3) or 6 (n¼3) hours (R3NS and R6NS, respectively). Soon after animals’ euthanasia, fresh sample of parietal cortex (P-Cx) were collected in order to evaluate byWestern Blot the levels of the following proteins and enzymes: AT8 (p[Ser202/Thr205]-Tau, phosphorylated Tau form), Tau-1 (unphosphorylated Tau form), p[Thr205]-Tau (a neuroprotective form of phosphorylated Tau; Ittner et al., 2016); p[Ser9]-GSK3b (the inhibited form of the main kinase targeting Tau); PP2A (the main phosphatase targeting Tau) and p[Ser473]-Akt2 (active anti-apoptotic factor and GSK3b inhibitor). Moreover, the plasma levels of melatonin were determined by ELISA.Results:Overall, AT8 levels were reduced in the SD groups compared to the NS ones (p<0.05), while Tau-1 levels were not significantly affected. A clear trend towards higher p[Thr205]-Tau levels was also observed after SD. The decrease in PPTau induced by SDwas accompanied by an increase in both p [Ser9]-GSK3b and p[Ser473]-Akt2 levels, although statistical significance was reached only for the latter (p<0.05). Also, PP2A levels were lower in R3SD vs. R3NS (p< 0.05). Finally, melatonin levels were higher in R3SD vs. R3 (p <0.05). Conclusions:The present results indicate that SD soon after ST enhances PPTau dephosphorylation, coherently with the observed concomitant increase of p[Ser9]-GSK3b and p[Ser473]-Akt2. This molecular pattern is known as being neuroprotective, and may be mediated by melatonin that can activate Akt2 and, consequently, inhibit GSK3b by acting on the PI3K/ Akt2/mTOR antiapoptotic pathway (Risso et al., 2015). These findings open interestingly translational perspectives in the use of sleep deprivation in patients suffering from hypothermia-induced brain PPTau formation due to general anesthesia (Whittington et al., 2013). Acknowledgements: The authors wish to thank Fondazione Carisbo that supported this work.