Effects of cannabinoids and cannabinoid-enriched Cannabis extracts on TRP channels and endocannabinoid metabolic enzymes

Abstract

Background and purpose: Cannabidiol (CBD) and Δ(9) -tetrahydrocannabinol (THC) interact with transient receptor potential (TRP) channels and enzymes of the endocannabinoid system.

Experimental approach: The effects of 11 pure cannabinoids and botanical extracts [botanical drug substance (BDS)] from Cannabis varieties selected to contain a more abundant cannabinoid, on TRPV1, TRPV2, TRPM8, TRPA1, human recombinant diacylglycerol lipase α (DAGLα), rat brain fatty acid amide hydrolase (FAAH), COS cell monoacylglycerol lipase (MAGL), human recombinant N-acylethanolamine acid amide hydrolase (NAAA) and anandamide cellular uptake (ACU) by RBL-2H3 cells, were studied using fluorescence-based calcium assays in transfected cells and radiolabelled substrate-based enzymatic assays. Cannabinol (CBN), cannabichromene (CBC), the acids (CBDA, CBGA, THCA) and propyl homologues (CBDV, CBGV, THCV) of CBD, cannabigerol (CBG) and THC, and tetrahydrocannabivarin acid (THCVA) were also tested.

Key results: CBD, CBG, CBGV and THCV stimulated and desensitized human TRPV1. CBC, CBD and CBN were potent rat TRPA1 agonists and desensitizers, but THCV-BDS was the most potent compound at this target. CBG-BDS and THCV-BDS were the most potent rat TRPM8 antagonists. All non-acid cannabinoids, except CBC and CBN, potently activated and desensitized rat TRPV2. CBDV and all the acids inhibited DAGLα. Some BDS, but not the pure compounds, inhibited MAGL. CBD was the only compound to inhibit FAAH, whereas the BDS of CBC > CBG > CBGV inhibited NAAA. CBC = CBG > CBD inhibited ACU, as did the BDS of THCVA, CBGV, CBDA and THCA, but the latter extracts were more potent inhibitors.

Conclusions and implications: These results are relevant to the analgesic, anti-inflammatory and anti-cancer effects of cannabinoids and Cannabis extracts.

© 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

Figures

Figure 1

Inhibition of TRPM8-mediated elevation of intracellular calcium by cannabigerol (CBG), a CBG-botanical drug substance rendered free of CBG [denoted as botanical extract (BDS)] and the reconstituted CBG-BDS (denoted as CBG-BDS reconstituted). Experiments were carried out in HEK-293 cells stably over-expressing the rat recombinant TRPM8 channel and are means of at least n = 3 separate experiments. ‘CBG-BDS reconstituted’ was obtained by mixing the amount of CBG present in a given amount of CBG-BDS and the remaining amount of the ‘CBG-free’ BDS. The concentrations tested of ‘CBG-BDS reconstituted’ were calculated to contain equimolar concentrations of CBG as in pure CBG. The potency of ‘CBG-BDS reconstituted’ was slightly lower than that shown in Table 2 for CBG-BDS but still higher than that of CBG. SEM are not shown for the sake of clarity and were never higher than 10% of the means.

Figure 2

(A) Dose-dependent effects of cannabigivarin (CBGV), Δ9-tetrahydrocannabiverin (THCV) and THCV-botanical drug substance (THCV-BDS) on TRPV1-mediated elevation of intracellular calcium in HEK-293 cells stably over-expressing the human recombinant TRP channel of vanilloid type-1 (TRPV1) channel (HEK-293-TRPV1 cells). (B) Dose-dependent inhibitory effects of a 5 min pre-incubation of HEK-293-TRPV1 cells with several pure cannabinoids on the TRPV1-mediated elevation of intracellular calcium induced by capsaicin (0.1 µM). (C) Dose-dependent inhibitory effects of a 5 min pre-incubation of HEK-293-TRPV1 cells with THCV and THCA on the TRPV1-mediated elevation of intracellular calcium induced by capsaicin (0.1 µM). Data are means of at least n = 3 separate experiments. SEM are not shown for the sake of clarity and were never higher than 10% of the means. CBC, cannabichromene; CBD, cannabidiol; CBG, cannabigerol; CBGV, cannabigivarin; CBN, cannabinol; THC, Δ9-tetrahydrocannabinol; THCV, propyl analogue of THC or tetrahydrocannabivarin.

Figure 3

Dose-dependent effects of several pure cannabinoids on lysophosphatidylcholine (LPC, 3 µM)-induced, TRP channel of vanilloid type-2 (TRPV2)-mediated elevation of intracellular calcium in HEK-293 cells stably over-expressing the rat recombinant TRPV2 channel. Compounds were given to cells 5 min prior to LPC. Data are means of at least n = 3 separate experiments. SEM are not shown for the sake of clarity and were never higher than 10% of the means. CBC, cannabichromene; CBD, cannabidiol; CBG, cannabigerol; CBGV, cannabigivarin; THC, Δ9-tetrahydrocannabinol; THCV, propyl analogue of THC or tetrahydrocannabivarin.

Figure 4

Inhibition of the effect of MAGL by cannabigerol (CBG), a CBG-botanical drug substance (CBG-BDS) as such or rendered free of CBG (denoted as BDS), and the reconstituted CBG-BDS (denoted as CBG-BDS reconstituted). Experiments were carried out in homogenates from COS cells and are means ± SEM of at least n = 3 separate experiments. ‘CBG-BDS reconstituted’ was obtained by mixing the amount of CBG present in a given amount of CBG-BDS and the remaining amount of the ‘CBG-free’ BDS. The concentrations tested of ‘CBG-BDS reconstituted’ were calculated to contain equimolar concentrations of CBG as in pure CBG. BDS, botanical drug substance; CBG, cannabigerol; MAGL, monoacylglycerol lipase.