0021 SLEEP DEPRIVATION ACTIVATES NLRP3 INFLAMMASOMES IN NEURONS AND GLIA

Abstract Introduction: The nucleotide-binding domain leucine rich family pyrin containing 3 (NLRP3) inflammasome is a protein complex that activates the somnogenic pro-inflammatory molecule interleukin-1 beta (IL-1β). Upon activation, NLRP3 recruits the apoptosis-associated speck-like protein containing a carboxyl-terminal caspase-recruitment domain (ASC) to activate the enzyme caspase-1, which converts the pro-form of IL-1β into its active form. We previously found that the gene expression of NLRP3 inflammasome components, caspase-1 activity, and IL-1β protein is enhanced in the cortex after sleep deprivation, although which particular cells are activated remains unknown. Thus, we examined the effects of sleep deprivation on the immuno-reactivity to inflammasome-related markers in neurons and glia. Methods: Male mice lacking NLRP3 and C57BL/6 wild-type control mice were sleep deprived for 6 h prior to dark onset [Zeitgeber (ZT) 12] using the gentle handling method; or were allowed to sleep ad libitum. Immediately after sleep deprivation (ZT 12), mice were perfused and the brains were processed for immunohistochemistry. Cells were double- and triple-labeled with inflammasome-related markers (i.e., anti-NLRP3, anti-ASC, and anti-IL-1β) and neuronal and glial-markers [i.e., anti-NeuN (neuron marker), anti-glial fibrillary acidic protein (GFAP; astrocyte marker), and anti-CD11b (microglia marker)]. We assessed the percent change of immuno-reactive cells to inflammasome-related markers in neurons and glia. Results: In C57BL/6 wild-type mice, we found significant enhancements in immunoreactivity to anti-NLRP3, anti-ASC, and anti-IL-1β in cells that were also immuno-reactive to anti-NeuN, anti-GFAP, and anti-CD11b after sleep deprivation compared to ad libitum sleep. However, the pattern of enhancement differed between cell types depending upon cortical and subcortical brain regions. Mice lacking NLRP3 did not show significant increases in immuno-reactivity to inflammasome-related antibodies, including IL-1β, in neurons and glia. Conclusion: These data indicate that sleep deprivation activates the NLRP3 inflammasome in neurons, astrocytes, and microglia, although this activation varies depending upon the brain area. Support (If Any): Veterans Affairs I01RX00928 (MRZ), I01BX001404 (RB), I01BX002774 (RES), National Institutes of Health MH016259 (MRZ), NS092926 (DG), JTM received partial salary compensation and funding from Merck (MISP), but no conflict of interest