5.2.1. Known Receptors as Potential Targets for Endocannabinoids
Fan (92) has shown that cannabinoids including anandamide, inhibit 5-HT3 receptor-mediated currents. These data indicated that the 5-HT3 receptor ion channel is a site of action of cannabinoids and endocannabinoids. The direction of the effect is compatible with the antiemetic potential of cannabinoid agonists (93), since 5-HT3 receptor antagonist are well-established antiemetic medicinal drugs. Fan's observation went largely unchallenged but was also not supported further by experimental evidence. However, we have observed that the 5-HT3 antagonists MDL72222 and granisetron show cannabinoid-like profiles in the mouse tetrad (73). In assays for synaptosomal receptor binding, MDL72222 did not bind CB1 receptors, and vice versa, neither anandamide nor HU210 bound to 5-HT3 receptors (Fride et al., in preparation). Thus, the nature of the interaction between CB1 and 5-HT3 receptors needs to be clarified further.
The current state of knowledge is complex. Mice injected with the 5-HT2 receptor antagonist ketanserin displayed cannabimimetic effects on the tetrad, with potencies at least as high as those of anandamide (73). Anandamide(94) and oleamide (95) have been found to bind to 5-HT2 receptors, thus raising the possibility that endo-cannabinoids (and oleamide) may act by 5-HT2 receptor blockade. In another study, however, no 3H-ketanserin displacement at the 5-HT2 receptor by oleamide was found (96). Thus the nature and physiological significance of endocannabinoid-5-HT2 receptor interaction needs to be further clarified.
Anandamide (but not THC) was found to have dual effects on NMDA receptor activity (56). First, like THC, anandamide reduced calcium flux via CB1 receptors, this effect being reversed by SR141716A. Second, at low concentrations, anandamide but not THC stimulated calcium influx by directly modulating the NMDA receptor. More recently, inhibition of glutamatergic neurotransmission by the synthetic cannabinoid agonist WIN55,212-2 was reported in CB1 knockout mice (97). It remains to be determined whether this finding has relevance to the observations at the NMDA receptor.
220.127.116.11. Lysophosphatidic Acid (LPA)
LPA bears structural similarities to 2-AG. Contos et al. (98) have demonstrated that targeted deletion of the receptor gene for LPA resulted in a defective suckling response in the knockout mice. This phenomenon is strikingly similar to the mortality of SR141716A-treated pups, which also die within days after birth due to a lack of milk ingestion from birth (99). Therefore it is possible that LPA and cannabinoids crossreact with their respective receptors. The sparse data available thus far do not support such a hypothesis. Thus SR141716A-induced inhibition of cannabinoid-stimulated p38 mito-gen-activated protein kinases did not alter the effects of LPA on p38-MAPK phosphorylation (100). Moreover, whereas THC completely reversed the effects of neonatally applied SR141716A, LPA did not (99). In both these reports, oleoyl-sn-glycero-3-phos-phate were used. Thus it remains to be seen whether other LPA species, notably LPA from arachidonic acid, will display cross-reactivity with cannabinoids.
Vanilloid type 1 (VR1) receptors are found not only on sensory neurons where they are partly coexpressed with CB1 receptors (101), but also in several central nuclei including hypothalamus and basal ganglia, hippocampus, and cerebellum (102,103). In all these brain areas, CB1 receptors are found as well (104). Anandamide is a full agonist at VR1 receptors (101,105,106). Although still somewhat controversial (107,108,109), it appears now that sufficient amounts of anandamide are available in vivo to stimulate VR1 receptors under physiological conditions (103,108,109). Summing up the evidence available at present, Di Marzo et al. (103) have suggested that anandamide interacts with both receptors at binding sites that are situated extra-or intracellularly for CB1 and VR1, respectively (110). The specific dominant interaction depends on a number of factors such as ATP acting to enhance anandamide's effects at the VR1 receptor, levels of anandamide, tissue receptor distribution, and accessibility to the receptor (103,110). These observations suggest that anandamide may be not only an endocannnabinoid but also an "endovanilloid" (103).
5.2.2. Putative Novel Receptors for Endocannabinoids in the Central Nervous System
Several recent reports present evidence suggesting the existence of a new, unknown CB receptor in the brain. One report describes a reduction in amplitudes of excitatory postsynaptic currents by cannabinoids responsible for glutamatergic neurotransmisson in the hippocampus of wild type mice as well as CB1 -/- knockout mice (97). Further, Di Marzo and colleagues (87) showed that anandamide effectively produced major aspects of the tetrad and stimulated GTP-S binding in CB -/- mice; these effects were not inhibited by SR141716A. These findings were elaborated by Breivogel et al. (111), who observed that the putative receptor is not distributed in the brain in a fashion similar to that of CB1 receptors. Thus, anandamide and WIN55212-1 bound to some brain regions of CB1-/— knockout mice such as cortex, hippocampus, and brainstem, but not in the basal ganglia and cerebella of these mice. It possible that, due to unknown compensatory mechanisms and/or other changes in the knockout mice, receptor types that are physiologically irrelevant in the normal organism may become overexpressed in CB -/- mice. Hence evidence for new CB receptors would be strengthened greatly by experiments using normal tissue or animals. It is of benefit, however, that the two CB1 receptor knockout models are of different genetic backgrounds (83,84), thereby allowing for some degree of generalizaton. Indeed, very recently, WIN55,212-2 was also shown to stimulate GTP S binding in Ledent et al.'s knockout mice. The regional distribution where this was observed, however (cerebellum, not hippocampus) (112), was different from Breivogel et al.'s findings, where, for example, WIN55,212-2 stimulated GTP S binding in hippocampal, but not cerebellar tissue of CB-/- mice (111). Future studies will have to determine whether the putative CB receptors in the different models are the same or different entities.
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