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31 décembre 2003
 J Neurosci
2000; 20; 9; 3401-3407
Reversal of dopamine D2 receptor responses by an anandamide transport inhibitor
M Beltramo, de Fonseca, et al
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Cannabinoid receptors, the target of the marijuana constituent d9-tetrahydrocannabinol (Pertwee, 1997), are densely expressed in basal ganglia and cortex, regions of the CNS that are critical forthe control of cognition, motivation, and movement (Herkenhamet al., 1990; Matsuda et al., 1993; Tsou et al., 1998). This distribution provides multiple opportunities for functional interactions between endogenous cannabinoid substances, such as anandamide (Devane et al., 1992; Di Marzo et al., 1994), and ascending dopamine pathways.
That these interactions may occur in vivo isindicated by several observations. First, in the striatum of freely moving rats, anandamide release is greatly increased after activation of dopamine D2 family receptors with the selective agonist quinpirole (Giuffrida et al., 1999). Second, pretreatment with theCB1 cannabinoid antagonist SR141716A enhances the stimulation of motor behavior elicited by systemic administration ofquinpirole (Giuffrida et al., 1999), although it has little effect perse on basal motor activity (Rinaldi-Carmona et al., 1994; Comptonet al., 1996; Navarro et al., 1997). Third, injection of D2 family agonists into basal ganglia nuclei opposes the behavioral response to locally administered CB1 receptor agonists (Sanudo-Pena etal., 1996, 1998; Sanudo-Pena and Walker, 1998).
Finally, chronic treatment with D2 family antagonists results in upregulated expressionof CB1 receptor mRNA in striatum (Mailleux and Vanderhaeghen, 1993). Together, these findings suggest that one of the functions of anandamide in the CNS may be to modulate dopamine D2 receptor-induced facilitation of psychomotor activity.In agreement with this possibility, anandamide and other CB1agonists inhibit movement, produce catalepsy, and attenuated-amphetamine-induced hyperactivity and stereotypy (Pryor etal., 1978; Gorriti et al., 1999), whereas disruption of the CB1receptor gene profoundly affects movement control (Ledent etal., 1999; Zimmer et al., 1999).
When anandamide is administered as a drug, its effects arecurtailed by a two-step mechanism consisting of transport intocells, mediated by a high-affinity carrier system (Beltramo et al.,1997b; Hillard et al., 1997; Piomelli et al., 1999), followed by intracellular hydrolysis, catalyzed by a relatively nonselective amidohydrolase enzyme (Deutsch and Chin, 1993; Desarnaud et al., 1995; Cravatt et al., 1996). Consequently, the anandamide transport inhibitor N-(4-hydroxyphenyl)-arachidonamide (AM404) prolongs and enhances several responses to exogenous anandamide,including analgesia (Beltramo et al., 1997b) and vasodilatation (Calignano et al., 1997a).
We hypothesized that blockade of anandamide transport, by causing this lipid to accumulate at itssites of release, may help uncover a participation of anandamide in the control of dopamine neurotransmission and might offer a pharmacological strategy to correct pathological conditions characterized by dopaminergic dysfunction. To test this hypothesis, we investigated the pharmacological properties of AM404 in the rat CNS and examined the effects of this drug on behavioral responses elicited by the activation of D2 family receptors. [...]
DISCUSSION There is both experimental and medical interest in developingmolecules that selectively interfere with anandamide transport. Anandamide transport inhibitors may be used experimentally to uncover the functions of the endocannabinoid system, which are still essentially uncharacterized (for review, see Piomelli et al.,1998). Furthermore, anandamide transport inhibitors may offer a rational approach to a variety of disease conditions in which elevation of anandamide levels at its release sites may result in a more selective pharmacological response than direct activation of CB1 receptors by agonist drugs.
In the present study, we used the anandamide transport inhibitor AM404 to investigate functional interactions between anandamide and dopamine in the control of motor activity. The existence of such interactions was suggested by four key observations.
First, in the striatum of freely moving rats, activation of D2 family receptors stimulates anandamide release (Giuffrida et al.,1999). Second, blockade of CB1 cannabinoid receptors enhances the stimulation of motor behavior elicited by D2 agonists (Giuffridaet al., 1999). Third, CB1 agonists and D2 family agonists exert opposing behavioral effects when they are administered by local injection into individual basal ganglia nuclei (San˜udo-Pena et al. 1996, 1998; SanudoPena and Walker, 1998). Finally, treatment with D2 family antagonists causes an upregulation of CB1receptor expression in striatum (Mailleux and Vanderhaeghen,1993).
In keeping with these results, we found that AM404 counteracts two characteristic responses mediated by activation of D2 family receptors: apomorphine-induced yawning and quinpirole induced stimulation of motor behaviors. These effects are achieved at doses of AM404 that may elicit only a mild hypokinesia when the drug is administered alone and may selectively inhibit anandamide transport in vitro.
In addition, doses of AM404 identical to those used in the present study are able to produce a time-dependent increase in the levels of anandamide in peripheral blood (A. Giuffrida, F. Rodrġguez de Fonseca, F.Nava, J. Belluzzi, and D. Piorrelli, in preparation). Thus, our results are consistent with the hypothesis that anandamide released by stimulation of D2 family receptors participates in th econtrol of dopamine-induced psychomotor activation.
CB1 receptor agonists elicit a broad spectrum of behavioral responses that include catalepsy, analgesia, reduced movement and hypothermia (Pertwee, 1997). The finding that AM404 evokes only a moderate slow-onset hypokinesia when it is administered alone demarcates the pharmacological profile of this anandamide transport inhibitor from those of direct-acting cannabimimetic drugs. This distinction may result from the ability of AM404 to enhance anandamide signaling in an activity dependent manner by causing anandamide to accumulate in discrete regions of the CNS only when release of this endocannabinoid substance is triggered by appropriate stimuli.
In the absence of such stimuli, tonic anandamide release may be very low, accountingfor the weak and slow-developing motor effects of AM404 in naive animals. We considered that the pharmacological profile of AM404 could offer an original strategy to correct behavioral abnormalities that are generally associated with dysfunction in dopamine neurotransmission. As an initial test of this hypothesis, we examined the effects of AM404 in SHR, a rat line in which hyperactivityand attention deficits have been linked to a defective regulation of mesocorticolimbic dopamine pathways (Esposito et al.,1999; Russell, 2000; Sadile, 2000). We found that administration of a low systemic dose of AM404 (1 mg/kg) normalizes motoractivity in SHR with no overt motor effect in WKY controls, the strain from which SHR originate (Okamoto, 1969).
These results suggest that pharmacological inhibition of anandamide inactivation may alleviate hyperactivity in SHR. Additional experiments are needed to determine whether this effect is mediated by an elevation of anandamide levels in brain regions involved in the control of movement and attention.The multiple physiological functions served by dopamine in the control of psychomotor activity and the lack of animal models that capture the complexities of psychiatric diseases make it difficult to extrapolate from rodent models to human syndromes.Yet, the spectrum of pharmacological properties displayed by AM404 and the ability of this drug to counteract potential manifestations of dopamine dysregulation suggest that anandamide transport may be a valuable target for the development of novelneuropsychiatric medicines.
The endogenous cannabinoid, anandamide, activates the hypothalamo-pituitary-adrenal axis in CB1 cannabinoid receptor knockout mice.
Wenger T, Ledent C, Tramu G
Department of Human Morphology and Developmental Embryology, Semmelweis University, Budapest, Hungary.
Neuroendocrinology 2003; 78; 6; 294-300

The purpose of this study was to investigate the effects of the endogenous cannabinoid arachidonoyl-ethanolamide, anandamide (AEA), on the activity of the hypothalamo-pituitary-adrenal (HPA) axis in cannabinoid receptor (CB(1) receptor) inactivated (KO) mice. A low dose (0.01 mg/kg i.p.) of AEA significantly increased plasma corticotropin (ACTH) and corticosterone concentrations in both wild-type (+/+) and in mutant (-/-) animals. In each case, hormone levels reached their peaks at 90 min after AEA administration. In a parallel experiment, AEA administration was preceded by the injection of SR 141716A (1.0 mg/kg), a selective and potent CB(1) receptor antagonist, or of capsazepine (5.0 mg/kg), a potent vanilloid receptor of type 1 (VR1) antagonist. The latter drugs did not prevent the effects of AEA on the HPA axis. Using Fos protein immunohistochemistry, we observed that the parvocellular part of the hypothalamic paraventricular nucleus (PVN) was activated as early as 45 min after AEA injection and reached peak levels after 60 min in both +/+ and -/- mice. Furthermore, the CB(1) and VR1 receptor antagonists did not block the effects of AEA on Fos immunoreactivity. The results strongly support the view that activation of the HPA axis produced by AEA possibly occurs via a currently unknown (CB(x)) cannabinoid receptor present in PVN.