The synthetic cannabinoid WIN 55212-2 reduces the frequency of miniature inhibitory postsynaptic currents (mIPSCs) in brain slices, but the identity of the endogenous messenger underpinning DSI (despolarisation-induced suppression of inhibition) is unknown (Wilson & Nicoll, 2001). Here we compare the effects of synthetic CB1 agonists with endocannabinoid candidates on mIPSCs in cultured monolayers in the absence of permeability and metabolic barriers.

Culture techniques and saline composition have been published previously (Nicholson et al. 2001). Cells were whole-cell patch-clamped at 22-24 ^oC. All solutions contained 0.1 % dimethylsulphoxide (vehicle) and 0.1 % bovine serum albumin. To isolate mIPSCs, 200 nM tetrodotoxin, 2 mM CNQX and 10 mM D-AP5, 2 mM Mg²⁺ and 0.5 mM Ca²⁺ were included. Intracellular saline contained (mM) CsCl₂ (132), CaCl₂ (1), MgCl₂ (2), Hepes (10) and EGTA (11); pH 7.4. Data were analysed using Spike 2 and Prism (Graphpad) software. Data (presented as means ± S.E.M.) were analysed using a Friedman ANOVA, with a Dunn’s post test. P values < 0.05 were considered significant.

In physiological saline, at a holding potential of -60 mV, WIN 55212-2 significantly reduced the frequency of mIPSCs (control, 1.54 ± 0.23 Hz; 0.8 mM WIN 55212-2, 1.03 ± 0.17 Hz; wash, 1.62 ± 0.28 Hz, P < 0.05, n = 5), consistent with effects in slices (Wilson & Nicoll, 2001). Anandamide (ANA) irreversibly mimicked this effect (control, 1.70 ± 0.41; 32 mM anandamide, 0.76 ± 0.26; wash, 0.88 ± 0.24, P < 0.01, n = 6). The CB2 ligand palmitoylethanolamide (PEA) did not depress the frequency of mIPSCs (control, 2.62 ± 0.64; 10 mM PEA, 2.49 ± 0.69; wash, 2.54 ± 0.63, P > 0.05). In the presence of the CB1 antagonist, AM251, the depressant effect of WIN 55212-2 was prevented (control, 1.17 ± 0.32 Hz; 0.8 mM WIN 55212-2, 1.12 ± 0.28 Hz; wash, 1.15 ± 0.27 Hz, P > 0.05, n = 6). The depressant effect of the endogenous cannabinoid, anandamide was also prevented in the presence of the CB1 antagonist AM251 (control, 0.79 ± 0.18 Hz; 32 mM ANA, 0.83 ± 0.24 Hz; wash, 0.78 ± 0.18 Hz, P > 0.05, n = 5). The sleep hormone oleamide causes an increase in mIPSC frequency (control, 0.86 ± 0.22 Hz; CoA, 2.13 ± 0.64 Hz; wash, 0.73 ± 0.15 Hz; P < 0.05, n = 4).

ANA but not PEA mimicks synthetic CB1 ligands in reducing the frequency of mIPSCs. These effects are prevented by the selective CB1 antagonist, AM251. The sleep hormone oleamide does not apear to be cannabamimetic.

Thanks to The Wellcome Trust for financial support and to David Hills for the cell cultures.

All procedures accord with UK legislation