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. 2009 Nov;331(2):591-7.
doi: 10.1124/jpet.109.158162. Epub 2009 Aug 7.

Blockade of 2-arachidonoylglycerol Hydrolysis by Selective Monoacylglycerol Lipase Inhibitor 4-nitrophenyl 4-(dibenzo[d][1,3]dioxol-5-yl(hydroxy)methyl)piperidine-1-carboxylate (JZL184) Enhances Retrograde Endocannabinoid Signaling

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Free PMC article

Blockade of 2-arachidonoylglycerol Hydrolysis by Selective Monoacylglycerol Lipase Inhibitor 4-nitrophenyl 4-(dibenzo[d][1,3]dioxol-5-yl(hydroxy)methyl)piperidine-1-carboxylate (JZL184) Enhances Retrograde Endocannabinoid Signaling

Bin Pan et al. J Pharmacol Exp Ther. .
Free PMC article

Abstract

Endocannabinoid (eCB) signaling mediates depolarization-induced suppression of excitation (DSE) and inhibition (DSI), two prominent forms of retrograde synaptic depression. N-Arachidonoylethanolamine (AEA) and 2-arachidonoylglycerol (2-AG), two known eCBs, are degraded by fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL), respectively. Selective blockade of FAAH and MAGL is critical for determining the roles of the eCBs in DSE/DSI and understanding how their action is regulated. 4-Nitrophenyl 4-(dibenzo[d][1,3]dioxol-5-yl(hydroxy)methyl)piperidine-1-carboxylate (JZL184) is a recently developed, highly selective, and potent MAGL inhibitor that increases 2-AG but not AEA concentrations in mouse brain. Here, we report that JZL184 prolongs DSE in Purkinje neurons in cerebellar slices and DSI in CA1 pyramidal neurons in hippocampal slices. The effect of JZL184 on DSE/DSI is mimicked by the nonselective MAGL inhibitor methyl arachidonyl fluorophosphonate. In contrast, neither the selective FAAH inhibitor cyclohexylcarbamic acid 3'-carbomoylbiphenyl-3-yl ester (URB597) nor FAAH knockout has a significant effect on DSE/DSI. JZL184 produces greater enhancement of DSE/DSI in mouse neurons than that in rat neurons. The latter finding is consistent with biochemical studies showing that JZL184 is more potent in inhibiting mouse MAGL than rat MAGL. These results indicate that the degradation of 2-AG by MAGL is the rate-limiting step that determines the time course of DSE/DSI and that JZL184 is a useful tool for the study of 2-AG-mediated signaling.

Figures

Fig. 1.
Fig. 1.
MAGL inhibitor JZL184 (JZL) potentiates DSE mouse cerebellar Purkinje neurons and DSI in mouse hippocampal pyramidal neurons. A, Left, examples of EPSCs in Purkinje neurons 4 s before (−4 s), 4 s, and 60 s after the depolarization (1 s from −60 to 0 mV), in the presence of the solvent (DMSO, control) or JZL184. Right, time course of averaged DSE in slices treated with DMSO (control, n = 8) and different concentrations of JZL184 (JZL, 10 nM, n = 8; 100 nM, n = 6). The lines are single exponential fitting curves of the decay of DSE. B, JZL184 caused a dose-dependent increase in the decay time constant (τ) of DSE (n = 4–8 for each point). C, JZL184 (1 μM) dramatically prolonged DSE when longer depolarization (5 s from −60 to 0 mV) was used to induce DSE (n = 6–8). D, Left, examples of IPSCs in hippocampal pyramidal neurons 4 s before (−4 s), 0 s, and 24 s after the depolarization (5 s from −60 to 0 mV), in the presence of the solvent (DMSO) or JZL184. Right, time course of averaged DSI in slices treated with DMSO (control, n = 10) and 100 nM JZL184 (n = 9).
Fig. 2.
Fig. 2.
MAGL inhibitor JZL184 selectively amplifies the effect of 2-AG, whereas FAAH inhibitor URB597 amplifies the effect of AEA. A, bath application of 2-AG (10 μM) depressed EPSCs in mouse cerebellar Purkinje neurons (n = 5). This depression was enhanced by JZL184 (JZL, 100 nM, n = 5), but not by URB597 (1 μM, n = 3). The 2-AG-induced depression of EPSCs was blocked by AM251 (2 μM). B, bath application of AEA (25 μM) depressed EPSCs in mouse cerebellar Purkinje neurons (n = 4). This depression was enhanced by URB597 (1 μM, n = 3), but not by JZL184 (JZL, 100 nM, n = 4). The AEA-induced depression of EPSCs was blocked by AM251 (2 μM).
Fig. 3.
Fig. 3.
Pharmacological blockade and genetic knockout of FAAH did not affect DSE and DSI. A, URB597 (1 μM) did not affect DSE in cerebellar Purkinje neurons (n = 5 each group). B, URB597 (1 μM) did not affect DSI in CA1 pyramidal neurons (n = 9–10). C and D, cerebellar DSE (C) and hippocampal DSI (D) were not significantly different in slices prepared from FAAH−/− and FAAH+/+ mice (n = 7–9). E, JZL184 (JZL, 100 nM) prolonged DSE to a similar extent in cerebellar slices from FAAH−/− and FAAH+/+ mice (n = 7 each group). F, URB597 did not affect DSE in slices from FAAH−/− and FAAH+/+ mice (n = 7 each group).
Fig. 4.
Fig. 4.
Effects of MAGL inhibitor JZL184 (JZL) and FAAH inhibitor URB597 on DSE in rat neurons. A and B, JZL (1 μM) significantly prolonged DSE in rat cerebellar Purkinje neurons (A) and rat hippocampal pyramidal neurons (B). C and D, URB597 (1 μM) had no significant effect on DSE in rat cerebellar Purkinje neurons (C) and rat hippocampal pyramidal neurons (D).
Fig. 5.
Fig. 5.
The effect of nonselective MAGL inhibitor MAFP on DSE and DSI. A and B, MAFP (500 nM) prolonged DSE in mouse (A) and rat (B) cerebellar Purkinje neurons. C and D, MAFP (500 nM) prolonged DSI in mouse (C) and rat (D) hippocampal pyramidal neurons.

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