and i), thus confirming the involvement of the inflammasome pathway. Since the inflammasome-activated pathway is known to interplay with autophagy [6, 8, 9], we also evaluated the distribution of LC3B (a marker of the autophagosome membrane [26]) puncta in NPCs. Interestingly, the number of LC3B puncta was significantly increased in cells that were exposed to ethanol compared to control cells (Fig. 2f and j), suggesting that ethanol may also induce LC3B lipidation, an indication of involvement of the autophagy pathway. Finally, to determine whether ethanol could exert a pro-apoptotic effect on NPCs that could potentially impair neurogenesis [27], we determined the expression of Casp3. Though not significant, we detected an increased number of Casp3+ cells in ethanol exposed cultures compared to controls (Fig. 2f and k), suggesting that ethanol may condition long-term survival of progenitors and/or differentiating cells without an acute effect, and that pathways other than canonical apoptosis might be involved in AUD.
