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Cannabinoid receptors and their endogenous ligands have been recently identified in the brain as potent inhibitors of neurotransmitter release. Here we show that, in a rat model of Parkinson's disease induced by unilateral nigral lesion with 6-hydroxydopamine (6-OHDA), the striatal levels of anandamide, but not that of the other endocannabinoid 2-arachidonoylglycerol, were increased. Moreover, we observed a decreased activity of the anandamide membrane transporter (AMT) and of the anandamide hydrolase [fatty acid amide hydrolase (FAAH)], whereas the binding of anandamide to cannabinoid receptors was unaffected. Spontaneous glutamatergic activity recorded from striatal spiny neurons was higher in 6-OHDA-lesioned rats. Inhibition of AMT by N-(4-hydroxyphenyl)-arachidonoylamide (AM-404) or by VDM11, or stimulation of the cannabinoid CB1 receptor by HU-210 reduced glutamatergic spontaneous activity in both naive and 6-OHDA-lesioned animals to a similar extent. Conversely, the FAAH inhibitors phenylmethylsulfonyl fluoride and methyl-arachidonoyl fluorophosphonate were much more effective in 6-OHDA-lesioned animals. The present study shows that inhibition of anandamide hydrolysis might represent a possible target to decrease the abnormal cortical glutamatergic drive in Parkinson's disease.

Anandamide (arachidonoylethanolamide, AEA) 1 belongs to an emerging class of endogenous lipids including amides and esters of long chain polyunsaturated fatty acids and is collectively termed "endocannabinoids" (1, 2). In fact, AEA has been isolated and characterized as an endogenous ligand for both CB 1 and, to a lesser extent, CB 2 cannabinoid receptor subtypes, and has been shown to mimic the psychotropic, antiemetic, and analgesic effects of cannabinoids (3). Recently, attention has been focused on the cardiovascular actions of AEA and their potential role in human shock conditions (4). In particular, a role for AEA has been proposed in both endothelium-dependent and -independent relaxations of vascular tissues, which involve several mechanisms including hyperpolarization of the smooth muscle cell membrane (5-7). A similar mechanism has been attributed in the past to a diffusible endothelium-derived hyperpolarizing factor (EDHF) different from nitric oxide (NO), whose chemical nature is still a matter of speculation (8). In fact, AEA has been proposed as an EDHF (5), though this hypothesis is still under debate (9, 10), and recent data strongly support the theory that EDHF is a cytochrome P450 metabolite (11). Whether or not an EDHF, AEA is likely to play an important role in the control of vascular tone (for reviews see Refs. 4 and 12), as suggested also by the observation that both endothelial cells and macrophages release this as well as the other endocannabinoid, 2-arachidonoyl-glycerol (2-AG) (1, 13-16).The pharmacological effects of AEA on CB 1 and CB 2 receptors depend, as for any other extracellular transmitter, on its life span in the extracellular space, which is limited by a two-step process: (i) its rapid and selective uptake by cells through the action of a membrane transporter and (ii) intracellular degradation. In particular, AEA is hydrolyzed to ethanolamine and arachidonic acid by the enzyme fatty acid amide hydrolase (FAAH) (17,18). Both components of this inactivation process of AEA are the objects of active investigations. Recent data seem to indicate that the uptake process is the rate-limiting step in AEA degradation (19 -23). There is pharmacological evidence suggesting that also the hypotensive action of AEA in vivo is limited by its re-uptake (24). However, the existence of the AEA membrane transporter in endothelial cells has never been investigated.Although cannabinoid receptor activation was recently shown to lead to AEA biosynthesis (25,26), the possibility of a functional link between CB 1 and CB 2 receptors and the AEA transporter has not been tested. Such a functional coupling might trigger self-elimination of AEA following activation by this lipid of cannabinoid receptor-dependent signaling pathways and would represent a regulatory loop critical for the manifold actions of this compound. A possible mechanism for this coupling may be suggested by findings that AEA binding to

Of the endocannabinoids (eCBs), anandamide (AEA) and 2-arachidonoylglycerol (2-AG) have received the most study. A functional interaction between these molecules has never been described. Using mouse brain slices, we found that stimulation of metabotropic glutamate 5 receptors by 3,5-dihydroxyphenylglycine (DHPG) depressed inhibitory transmission in the striatum through selective involvement of 2-AG metabolism and stimulation of presynaptic CB1 receptors. Elevation of AEA concentrations by pharmacological or genetic inhibition of AEA degradation reduced the levels, metabolism and physiological effects of 2-AG. Exogenous AEA and the stable AEA analog methanandamide inhibited basal and DHPG-stimulated 2-AG production, confirming that AEA is responsible for the downregulation of the other eCB. AEA is an endovanilloid substance, and the stimulation of transient receptor potential vanilloid 1 (TRPV1) channels mimicked the effects of endogenous AEA on 2-AG metabolism through a previously unknown glutathione-dependent pathway. Consistently, the interaction between AEA and 2-AG was lost after pharmacological and genetic inactivation of TRPV1 channels.

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