Structure‐activity relationship for the endogenous cannabinoid, anandamide, and certain of its analogues at vanilloid receptors in transfected cells and vas deferens

Abstract

1 This study was directed at exploring the structure-activity relationship for anandamide and certain of its analogues at the rat VR1 receptor in transfected cells and at investigating the relative extent to which anandamide interacts with CB 1 and vanilloid receptors in the mouse vas deferens. 2 pK i values for displacement of [ 3 H]-resiniferatoxin from membranes of rVR1 transfected CHO cells were signi®cantly less for anandamide (5.78) than for its structural analogues N-(4-hydroxyphenyl)-arachidonylamide (AM404; 6.18) and N-(3-methoxy-4-hydroxy)benzyl-arachidonylamide (arvanil; 6.77). 3 pEC 50 values for stimulating 45 Ca2+ uptake into rVR1 transfected CHO cells were signi®cantly less for anandamide (5.80) than for AM404 (6.32) or arvanil (9.29). Arvanil was also signi®cantly more potent than capsaicin (pEC 50 =7.37), a compound with the same substituted benzyl polar head group as arvanil. 4 In the mouse vas deferens, resiniferatoxin was 218 times more potent than capsaicin as an inhibitor of electrically-evoked contractions. Both drugs were antagonized to a similar extent by capsazepine (pK B =6.93 and 7.18 respectively) but were not antagonized by SR141716A (1 mM). Anandamide was less susceptible than capsaicin to antagonism by capsazepine (pK B =6.02) and less susceptible to antagonism by SR141716A (pK B =8.66) than methanandamide (pK B =9.56). WIN55212 was antagonized by SR141716A (pK B =9.02) but not by capsazepine (10 mM). 5 In conclusion, anandamide and certain of its analogues have anity and ecacy at the rat VR1 receptor. In the mouse vas deferens, which seems to express vanilloid and CB 1 receptors, both receptor types appear to contribute to anandamide-induced inhibition of evoked contractions.

Effects of piperine, the pungent component of black pepper, at the human vanilloid receptor (TRPV1)

“…Although the efficacy of many TRPV1 agonists such as resiniferatoxin, capsiate and gingerols have been characterised in detail in direct functional assays employing TRPV1 (Caterina et al, 1997;Dedov et al, 2002;Iida et al, 2003), none have been shown to have maximal efficacy greater than that of capsaicin. Similar findings have also been reported for anandamide and related analogues, PPAHV, and N-arachidonoyldopamine as determined by calcium imaging or calcium uptake Ross et al, 2001;Huang et al, 2002). There are some reports, however, that RTX can produce a greater increase in intracellular [Ca 2 þ ] than capsaicin although this may be due to either an interaction with internal Ca 2 þ stores (Marshall et al, 2003) or the ability of this compound to interact with protein kinase C pathways as a result of its phorbol ester activity (Harvey et al, 1995).…”

Molecular targets for cannabidiol and its synthetic analogues: effect on vanilloid VR1 receptors and on the cellular uptake and enzymatic hydrolysis of anandamide

“…These data suggest that the C-1 methyl and the aromatic`A' ring, which is chemically similar to the vanillyl moiety of capsaicin (Figure 1), are more important than the chiral part of CBD for its interaction with VR1. That CBD binds to the same site as capsaicin is suggested by the ®nding that both compounds displace [ 3 H]-resiniferatoxin from its speci®c binding sites in membranes from cells over-expressing VR1 receptors (this study and Ross et al, 2001). However, while capsaicin exhibits higher potency than a nity for vanilloid receptors (Szallasi & Blumberg, 1999), CBD is as active in the [ 3 H]-resiniferatoxin binding assay as in the hVR1 functional assay.…”

“…CBD desensitized VR1 to the action of capsaicin, thus opening the possibility that this cannabinoid exerts an antiin¯ammatory action in part by desensitization of sensory nociceptors. Future studies with capsazepine (which antagonizes capsaicin e ects in rats (Di Marzo et al, 2001) but not always in mice (Di Marzo et al, 2000b), and VR1`knockout' mice Caterina et al, 2000), should test the involvement of VR1 in the pharmacological actions of CBD.…”

“…The a nity of CBD and (+)-CBD for human VR1 receptors was assessed by means of displacement assays carried out with membranes (50 mg tube 71 ) from HEKhVR1 cells, prepared as described previously (Ross et al, 2001), and the high a nity VR1 ligand [ 3 H]-resiniferatoxin (48 Ci mmol 71 , NEN-Dupont), using the incubation conditions described previously (Ross et al, 2001 Cannabinoid CB 1 and CB 2 receptor binding assays These methods have been described previously by Devane et al (1992a) for CB 1 , and Bayewitch et al (1996) for CB 2 . For CB 1 receptor binding assays synaptosomal membranes from rat brains were used.…”

Identification of Species-specific Determinants of the Action of the Antagonist Capsazepine and the Agonist PPAHV on TRPV1

“…RTX Displacement Assay-RTX saturation and displacement binding were carried out as described elsewhere (20 …”