Figure 1

Effects of CBD on OPC cytotoxicity. (a) Although CBD did not induce cell death at low concentrations (0.1 and 1 μM), cytotoxic effects were observed at 2.5 and 5 μM. OPCs were exposed to CBD and cell death was quantified 48 h later using the LDH method. Data represent the mean±S.E.M. of n=3 cultures analyzed in triplicate. Statistical significance was determined by a one-way ANOVA: **P⩽0.01 and ***P⩽0.001 versus untreated cells. (b) CBD did not augment OPC proliferation evident as an increase in BrdU incorporation. OPCs were incubated with BrdU (10 μM) for 24 h and at least 10 000 cells were then quantified by immunocytochemistry. (c) Flow cytometry analysis revealed the CBD does not affect the cell cycle progression of OPCs. Freshly isolated OPCs were incubated for 24 h in the presence or absence of CBD (1 μM), and the cell cycle of a minimum of 10 000 cells was analyzed on a FACSAria flow cytometer following PI staining. (d, e) CBD did not alter intracellular Ca²⁺ levels under basal conditions. OPCs were loaded with the Ca²⁺-sensitive dye Fluo-4 and the fluorescence signal was monitored on a scanning confocal microscope under basal conditions and after adding CBD (1 μM: n=5 coverslips). (d) Representative pseudocolor images showing Ca²⁺ levels in cultured OPCs under basal conditions and in the presence of CBD (1 μM). Scale bar=50 μm. (e) Fluorescence traces showing the Ca²⁺ time course in individual OPCs (n=8) from the experiment shown in (d), both under basal conditions and in the presence of CBD.