Enhancing the rate of myelination and subsequent myelin repair could potentially arrest and reverse the disease progression of demyelinating disorders. To date, no such therapeutics with pro-remyelinating properties have been identified. We investigated the pro-myelinating properties of nefiractam , and whether it exerts these effects by modulating Wnt signalling, which is known to play a role in oligodendrocyte development (Feigenson et al, 2009). To investigate this, myelin formation, as assessed by immunofluorescent staining for MBP, was examined in organotypic hippocampal cultures from Wistar rats. When nefiracetam (1µM) was applied to the medium from the first day of culture (1DIV), the density of MBP staining was increased at 6DIV (209.2%±47.7, n=18, p<0.05, ANOVA) relative to untreated control (100%±14, n=18). Treatment with 10µM cardamonin, an inhibitor of β-catenin signalling did not cause a change in myelin density (109.2±30.3, n=18, p>0.05). However, both nefiracetam and cardamonin increased immunofluorescence of NG2, an oligodendrocyte precursor marker (p<0.001, Kolmogorov-Smirnov test, 2825≤n≤3622, fig.1). This suggests that, while modifying Wnt signalling may impact on the development of precursor cells, this does not correspond to an increase in mature myelinating cells. Therefore, the pro-myelinating effects of nefiracetam must involve some additional mechanism. We nest investigated the effect of nefiracetazm on spontaneous activity, which also modulates myelination (Demerens et al, 1996). Cultures were exposed to 250µg/ml lysolecithin (LPC) for 18 hours at 13DIV. After LPC exposure, cultures recovered in LPC-free medium ±1µM nefiracetam. After 24 hours, cultures were loaded with Fluo4-AM, and the number of cells displaying spontaneous calcium activity measured. Cultures not exposed to LPC displayed low levels of activity (1.7% of 5174 cells), which was unchanged by nefiracetam treatment (1.7% of 4819 cells, p=0.93, Χ2 test). LPC exposure caused an increase in activity (4.7% of 5608 cells, p<0.001), which was enhanced by nefiracetam treatment (12.9% of 4506 cells, p<0.001). When MBP density was measured in these cultures, LPC caused decreased MBP density (49.9%±5.3, n=27, P<0.05, ANOVA) compared to control (100%±11, n=27), but nefiracetam exposure post LPC caused a return to control levels (80%±10.7, n=24, p>0.05), suggesting that the increase in post LPC activity may be an adaptive response related to myelin repair, which is enhanced by nefiracetam. Together, these data suggest a complex mechanism of action for nefiracetam with respect to its action on myelin.