[Cannabis in France, new insights]. / La situation actuelle du cannabis en France.

France holds the record for cannabis use in Europe, especially among adolescents. This drug of abuse is thus mainly used during a very sensitive period of brain development, education, vehicle driving and development of life projects. In addition, synthetic derivatives of tetrahydrocannabinol (THC), which are more noxious than cannabis itself are now appearing on the market. Traficking and cultivation for personnal use have intensified; products proposed for sale are richer in THC; and some methods of consumption (e-cigarettes, vaporizers, water pipes) increase the supply of THC to the lungs and thence to the body and brain. It is in this context that attempts are being made to legalize this drug of abuse. Other attempts are made to disguise it as a medication. Meanwhile, the list of its psychic as well as physical damages grows longer, with some very severe cases of major injuries. This evolution takes place in spite of numerous warnings expressed by the French Academy of Medicine. Subsequently, it is prompted to carefully and vigorously denounce these events. This will be the aim of this thematic session.

[Neurobiology of endocannabinoids and central effects of tetrahydrocannabinol contained in indian hemp]. / Neurobiologie des endocannabinoïdes-- mise en relation avec les effets du tétrahydrocannabinol du chanvre indien.

Tetrahydrocannabinol, the main psychotropic component of Cannabis indica, is an addictive drug with multiple effects including both peripheral and central damages. All these effects are due to interference with endocannabinoidergic transmission. This endocannabinoid system subtly regulates many physiologicalfunctions. This regulation involves various ligands derived from arachidonic acid (anandamide, di-arachidonoylglycerol, virodhamin, noladin ether, N arachidonoyl dopamine, etc.) which stimulate two main types of receptor CB1 in the central nervous system and CB2 in the periphery. CB1 receptors are very numerous and ubiquitous in the brain. They influence various important functions (awakening, attention, delirium, hallucinations, memory, cognition, anxiety, humor stability, motor coordination, brain maturation, etc.). Far from mimicking endocannabinoids, THC caricatures their effects. It affects all brain structures, simultaneously, intensely and durably, inducing down-regulation of CB1 receptors and thereby reducing the effects of their physiological ligands. On account of its exceptional lipophilia, THC accumulates for days and even weeks in the brain. It is not a soft drug but rather a slow drug: its abuse induces long-lasting modifications and deterioration of brain function, potentially leading to various mental and psychiatric disorders.

The COMT Val158Met polymorphism affects the response to entacapone in Parkinson's disease: a randomized crossover clinical trial.

OBJECTIVE: In Parkinson disease (PD), the selective C-O-methyltransferase (COMT) inhibitor entacapone prolongs the effect of levodopa on motor symptoms (ON time) by increasing its bioavailability. The COMT Val158Met polymorphism is equally distributed in PD patients and modulates COMT activity, which can be high (Val/Val, COMT(HH) ), intermediate (Val/Met, COMT(HL) ), or low (Met/Met, COMT(LL) ). The objective of this study was to determine the response to entacapone in COMT(HH) and COMT(LL) PD patients. METHODS: Thirty-three PD patients, homozygous for the COMT alleles COMT(HH) (n = 17) and COMT(LL) (n = 16), were randomized in a double-blind crossover trial consisting of 2 successive acute levodopa challenges associated with 200mg entacapone or placebo. The primary endpoint was the gain in the best ON time. Secondary endpoints were levodopa pharmacokinetics and COMT activity in red blood cells. RESULTS: The gain in the best ON time was higher in COMT(HH) than in COMT(LL) patients (39 ± 10 vs 9 ± 9 minutes, p = 0.04, interaction between treatment and genotype). Area under the concentration over time curve of levodopa increased more after entacapone in COMT(HH) than in COMT(LL) patients (+62 ± 6% vs +34 ± 8%, p = 0.01). COMT inhibition by entacapone was higher in COMT(HH) than in COMT(LL) patients (-0.54 ± 0.07 vs -0.31 ± 0.06 pmol/min/mg protein, p = 0.02). INTERPRETATION: The COMT(HH) genotype in PD patients enhances the effect of entacapone on the pharmacodynamics and pharmacokinetics of levodopa. The response to entacapone after repeated administrations and in heterozygous patients remains to be determined.

[Treatment of insomnia. Pharmacological approaches and their limitations]. / Le traitement de i'insomnie. Les possibilités pharmacologiques et leurs limites.

Far more people complain of inadequate sleep than of true insomnia warranting prescription of a hypnotic drug. The number of available hypnotics has fallen markedly in recent years. Numerous brain areas, transmitters and receptors are involved in sleep. Currently, the main hypnotics (benzodiazepine derivatives and the so-called 4Z group. Zolpidem, Zopiclone, EsZopiclone and Zaleplon) increase GABAergic transmission by acting on components of chloride channels, thereby inducing Cl entry to neurons and resulting in their hyperpolarisation. This pre-eminence of GABAergic transmission should not make us ignore other important transmitters and their receptors as potential targets for new hypnotic drugs; these include histamine (and H1 receptors), dopamine (D1 and D2), norepinephrine (alpha1), serotonin (5HT2), glutamate (NMDA), acetylcholine (nicotinic), hypocretin (OX1 and OX2), melatonin (MLT1 and MLT2), prostaglandin E2 (EP), prostaglandin D2 (DP1), and endocannabinoids (CB1). Knowledge of the pharmacodynamic, pharmacokinetic and clinical characteristics of current hypnotic drugs has allowed us to establish the profile of an ideal hypnotic for the future.

Regulation of feeding and anxiety by alpha-MSH reactive autoantibodies.

alpha-Melanocyte-stimulating hormone (alpha-MSH) is a stress-related neuropeptide involved in the regulation of motivated behavior, appetite and emotion including stimulation of satiety and anxiety. Although autoantibodies (autoAbs) reactive with alpha-MSH have been identified in human subjects and in rats, it remained unknown if these autoAbs are involved in the regulation of feeding and anxiety and if their production is related to stress. Here we show that repeated exposure of rats to anxiolytic mild stress by handling increases the levels and affinity of alpha-MSH reactive IgG autoAbs and that these changes are associated with adaptive feeding and anxiety responses during exposure of rats to a strong stress by food restriction. Importantly, an increase in affinity of alpha-MSH reactive autoAbs was associated with changes of their functional roles from stimulation to inhibition of alpha-MSH-mediated behavioural responses, suggesting that these autoAbs can be a carrier or a neutralizing molecule of alpha-MSH peptide, respectively. Using a model of passive transfer into the brain, we show that alpha-MSH autoAbs affinity purified from blood of rats exposed to repeated mild stress, but not from control rats, are able to increase acutely food intake, suppress anxiety and modify gene expression of hypothalamic neuropeptides in naïve rats. These data provide the first evidence that autoAbs reactive with alpha-MSH are involved in the physiological regulation of feeding and mood, supporting a further role of the immune system in the control of motivated behavior and adaptation to stress.

Synthesis, radiosynthesis and biological evaluation of 1,4-dihydroquinoline derivatives as new carriers for specific brain delivery.

In spite of numerous reports dealing with the use of 1,4-dihydropyridines as carriers to deliver biological active compounds to the brain, this chemical delivery system (CDS) suffers from poor stability of the 1,4-dihydropyridine derivatives towards oxidation and hydration reactions seriously limiting further investigations in vivo. In an attempt to overcome these limitations, we report herein the first biological evaluation of more stable annellated NADH models in the quinoline series as relevant neuroactive drug-carrier candidates. The radiolabeled 1,4-dihydroquinoline [(11)C] was prepared to be subsequently peripherally injected in rats. The injected animals were sacrificed and brains were collected. The radioactivity measured in rat brain indicated a rapid penetration of the carrier [(11)C] into the CNS. HPLC analysis of brain homogenates showed that oxidation of [(11)C] into the corresponding quinolinium salt [(11)C] was completed in less than 5 min. An in vivo evaluation in mice is also reported to illustrate the potential of such 1,4-dihydroquinoline derivatives to transport a neuroactive drug in the CNS. For this purpose, gamma-aminobutyric acid (GABA), well known to poorly cross the brain blood barrier (BBB) was connected to this 1,4-dihydroquinoline-type carrier. After i.p. injection of 1,4-dihydroquinoline-GABA derivative in mice, a significant alteration of locomotor activity (LMA) was observed presumably resulting from an enhancement of central GABAergic activity. These encouraging results give strong evidence for the capacity of carrier-GABA derivative to cross the BBB and exert a pharmacological effect on the CNS. This study paves the way for further progress in designing new redox chemical delivery systems.

Rational design of central selective acetylcholinesterase inhibitors by means of a "bio-oxidisable prodrug" strategy.

This work deals with the design of a bio-oxidisable prodrug strategy for the development of new central selective acetylcholinesterase inhibitors. This prodrug approach is expected to reduce peripheral anticholinesterase activity responsible for various side effects observed with presently marketed AChE inhibitors. The design of these new AChE inhibitors in quinoline series is roughly based on cyclic analogues of rivastigmine. The key activation step of the prodrug involves an oxidation of an N-alkyl-1,4-dihydroquinoline 1 to the corresponding quinolinium salt 2 unmasking the positive charge required for binding to the catalytic anionic site of the enzyme. The synthesis of a set of 1,4-dihydroquinolines 1 and their corresponding quinolinium salts 2 is presented. An in vitro biological evaluation revealed that while all reduced forms 1 were unable to exhibit any anticholinesterase activity (IC50 > 10(6) nM), most of the quinolinium salts 2 displayed high AChE inhibitory activity (IC50 ranging from 6 microM to 7 nM). These preliminary in vitro assays validate the use of these cyclic analogues of rivastigmine in quinoline series as appealing chemical tools for further in vivo development of this bio-oxidisable prodrug approach.

Evidence for the involvement of the adenosine A(2A) receptor in the lowered susceptibility to pentylenetetrazol-induced seizures produced in mice by long-term treatment with caffeine.

Long-term caffeine intake has been reported to decrease the susceptibility to convulsants in mice. Occurrence of seizures following long-term oral administration of caffeine (0.3g/l) was investigated using adenosine A(2A) receptor knockout (A(2A)R KO) and control (A(2A)R WT) mice. Clonic seizures induced by acute pentylenetetrazol (PTZ, 50mg/kg i.p.) were significantly attenuated in adenosine A(2A)R KO mice drinking only water and reduced by a 14-day caffeine treatment in adenosine A(2A)R WT mice. In addition we showed a protecting effect of a 21-day caffeine treatment in A(2A)R WT mice against kindled seizures induced by PTZ in an increasing dose schedule. Summing up, these protective effects against PTZ-induced seizures occurring when adenosine A(2A)R is absent or chronically blocked by a relevant dose of caffeine may be related to a decreased neuronal excitability.

Synthesis and biological effects of c(Lys-Lys-Pro-Tyr-Ile-Leu-Lys-Lys-Pro-Tyr-Ile-Leu) (JMV2012), a new analogue of neurotensin that crosses the blood-brain barrier.

The central administration of neurotensin (NT) or of its C-terminal hexapeptide fragment NT(8-13), produces strong analgesic effects in tests evaluating acute pain. The use of NT-derived peptides as pharmaceutical agents to relief severe pain in patients could be of great interest. Unfortunately, peptides do not readily penetrate the blood-brain barrier. We have observed that the cyclic NT(8-13) analogue, c(Lys-Lys-Pro-Tyr-Ile-Leu-Lys-Lys-Pro-Tyr-Ile-Leu) (JMV2012, compound 1), when peripherally administered to mice produced analgesic and hypothermic effects, suggesting the peptide penetrates the blood-brain barrier and functions effectively like a drug. Moreover, dimeric compounds show increased potency compared to their corresponding monomer. We present the synthesis of the cyclic dimer compound 1 (JMV2012). In mice, compound 1 induced a profound hypothermia and a potent analgesia, even when peripherally administered. Compound 1 appears to be an ideal lead compound for the development of bioactive NT analogues as novel analgesics drugs.

Behavioral actions of urotensin-II.

Urotensin-II (U-II) and urotensin-II-related peptide (URP) have been identified as the endogenous ligands of the orphan G-protein-coupled receptor GPR14 now renamed UT. The occurrence of U-II and URP in the central nervous system, and the widespread distribution of UT in the brain suggest that U-II and URP may play various behavioral activities. Studies conducted in rodents have shown that central administration of U-II stimulates locomotion, provokes anxiety- and depressive-like states, enhances feeding activity and increases the duration of paradoxical sleep episodes. These observations indicate that, besides the endocrine/paracrine activities of U-II and URP on cardiovascular and kidney functions, these peptides may act as neurotransmitters and/or neuromodulators to regulate various neurobiological activities.

[Dmt1, d-1-Nal3]morphiceptin, a novel opioid peptide analog with high analgesic activity.

The morphiceptin-derived peptide [Dmt1, d-1-Nal3]morphiceptin, labeled mu-opioid receptor (MOP) with very high affinity and selectivity in the receptor binding assays. In the mouse hot plate test, [Dmt1, d-1-Nal3]morphiceptin given intracerebroventricularly (i.c.v.) produced profound supraspinal analgesia, being approximately 100-fold more potent than the endogenous MOP receptor ligand, endomorphin-2. The antinociceptive effect of this new analog lasted up to 120min. Thus, [Dmt1, d-1-Nal3]morphiceptin is an interesting and extraordinarily potent analgesic, raising the possibility of novel approaches in the design of clinically useful drugs for pain treatment.

Locomotor effects of morphine or alcohol in mice after a repeated treatment with the cannabinoid agonist HU 210.

The consequences of the consumption of cannabinoids with other drugs of abuse are of particular medical relevance. Several studies investigated the ability of cannabinoids to induce a locomotor cross-sensitization to other addictive drugs, but results remain inconsistent. Therefore, we investigated in mice the consequences of a repeated treatment with the cannabinoid agonist HU 210 on motor effects of morphine or alcohol. In mice receiving a daily injection of HU 210 (12.5 to 200 microg/kg) during 7 days, no hetero-sensitization to the stimulation induced by either morphine (7.5 mg/kg) or alcohol (1 or 1.5 g/kg) emerged, from 1 day up to 35 days after the end of the sub-chronic treatment with HU 210. Even a chronic treatment with a high dose of HU 210 (14 days, 200 microg/kg) induced no subsequent enhancement of the stimulant effects of morphine or alcohol. In fact, the motor stimulant effect of morphine or alcohol in chronically HU 210 pre-treated mice was even abolished until the 3rd day of abstinence. This reduction was presumably due to residual HU 210 since this effect was prevented by the cannabinoid antagonist rimonabant. Afterwards, chronically cannabinoid pre-treated mice remained less active than vehicle pre-treated mice from the 7th day up to the 35th day after the end of the 14-day treatment with HU 210. In conclusion, we failed to detect any hetero-sensitization whatever the pre-treatment regimen. However, only after the 14-day regimen, HU 210 pre-treated mice displayed a long-lasting decrease in activity, suggesting that some neuronal adaptive changes may have occurred.

Novel highly potent mu-opioid receptor antagonist based on endomorphin-2 structure.

The mu-opioid agonists endomorphin-1 (Tyr-Pro-Trp-Phe-NH(2)) and endomorphin-2 (Tyr-Pro-Phe-Phe-NH(2)) exhibit an extremely high selectivity for the mu-opioid receptor and thus represent a potential framework for modification into mu-antagonists. Here we report on the synthesis and biological evaluation of novel [d-2-Nal(4)]endomorphin-2 analogs, [Sar(2),d-2-Nal(4)]endomorphin-2 and [Dmt(1),Sar(2),d-2-Nal(4)]endomorphin-2 (Dmt=2'6'-dimethyltyrosine; Sar=N-methylglycine, sarcosine; d-2-Nal=3-(2-naphthyl)-d-alanine). [Dmt(1),Sar(2),d-2-Nal(4)]endomorphin-2 possessed very high affinity for the mu-opioid receptor (IC(50)=0.01+/-0.001 nM) and turned out to be a potent and extremely selective mu-opioid receptor antagonist, as judged by the in vitro aequorin luminescence-based calcium assay (pA(2)=9.19). However, in the in vivo hot plate test in mice this analog was less potent than our earlier mu-opioid receptor antagonist, [Dmt(1),d-2-Nal(4)]endomorphin-2 (antanal-2). The exceptional mu-opioid receptor in vitro activity and selectivity of [Dmt(1), Sar(2),d-2-Nal(4)]endomorphin-2 makes this analog a valuable pharmacological tool, but further modifications are needed to improve its in vivo profile.

Antidepressant-like effect of endomorphin-1 and endomorphin-2 in mice.

Endomorphin-1 (Tyr-Pro-Trp-Phe-NH(2)) and endomorphin-2 (Tyr-Pro-Phe-Phe-NH(2)) are two recently isolated mu-opioid selective peptides with a potent antinociceptive activity, involved in a number of physiological processes, including food intake, vasomotricity, sexual behavior, as well as neuroendocrine and cardiorespiratory functions. The neuroanatomical distribution of endomorphins prompted us to study their antidepressant activity in two animal behavioral models of depression: forced-swimming and tail-suspension tests. In both tests, the intracerebroventricular (i.c.v.) injection of either endomorphin-1 or endomorphin-2 significantly decreased the duration of immobility, interpreted as an expression of 'behavioral despair', which could be related to the depression syndrome. These effects of endomorphins did not result from the stimulation of the animal motor activity. We have also demonstrated that the antidepressant-like effect of endomorphins was antagonized by the universal opioid antagonist, naloxone and the mu-opioid receptor selective antagonist, beta-funaltrexamine. In contrast, this effect was not antagonized by delta- and kappa-opioid receptor selective antagonists, naltrindole and nor-binaltorphimine, respectively. The results of the present study demonstrate that endomorphin-1 and endomorphin-2 produce potent antidepressant-like effects after i.c.v. injection in mice. We may suggest that endomorphins and the mu-opioid receptors might be involved in the physiopathology of depressive disorders, and that the endomorphinergic system could serve as a novel target for the development of antidepressant drugs.

Pharmacological characterization of the receptor mediating the anorexigenic action of the octadecaneuropeptide: evidence for an endozepinergic tone regulating food intake.

Peptides of the endozepine family, including diazepam-binding inhibitor, the triakontatetraneuropeptide, and the octadecaneuropeptide (ODN), act through three types of receptors, that is, central-type benzodiazepine receptors (CBR), peripheral-type (mitochondrial) benzodiazepine receptors (PBR) and a metabotropic receptor positively coupled to phospholipase C via a pertussis toxin-sensitive G protein. We have previously reported that ODN exerts a potent anorexigenic effect in rat and we have found that the action of ODN is not affected by the mixed CBR/PBR agonist diazepam. In the present report, we have tested the possible involvement of the metabotropic receptor in the anorexigenic activity of ODN. Intracerebroventricular administration of the C-terminal octapeptide (OP) and its head-to-tail cyclic analog cyclo(1-8)OP (cOP) at a dose of 100 ng mimicked the inhibitory effect of ODN on food intake in food-deprived mice. The specific CBR antagonist flumazenil and the PBR antagonist PK11195 did not prevent the effect of ODN, OP, and cOP on food consumption. In contrast, the selective metabotropic endozepine receptor antagonist cyclo(1-8)[DLeu(5)]OP (100-1000 ng; cDLOP) suppressed the anorexigenic effect of ODN, OP, and cOP. At the highest concentration tested (1000 ng), cDLOP provoked by itself a significant increase in food intake. Taken together, the present results indicate that the anorexigenic effect of ODN and OP is mediated through activation of the metabotropic receptor recently characterized in astrocytes. The data also suggest that endogenous ODN, acting via this receptor, exerts an inhibitory tone on feeding behavior.

Synthesis and characterization of potent and selective mu-opioid receptor antagonists, [Dmt(1), D-2-Nal(4)]endomorphin-1 (Antanal-1) and [Dmt(1), D-2-Nal(4)]endomorphin-2 (Antanal-2).

To synthesize potent antagonists of the mu-opioid receptor, we prepared a series of endomorphin-1 and endomorphin-2 analogues with 3-(1-naphthyl)-d-alanine (d-1-Nal) or 3-(2-naphthyl)-d-alanine (d-2-Nal) in position 4. Some of these analogues displayed weak antagonist properties. We tried to strengthen these properties by introducing the structurally modified tyrosine residue 2,6-dimethyltyrosine (Dmt) in place of Tyr1. Among the synthesized compounds, [Dmt1, d-2-Nal4]endomorphin-1, designated antanal-1, and [Dmt1, d-2-Nal4]endomorphin-2, designated antanal-2, turned out to be highly potent and selective mu-opioid receptor antagonists, as judged on the basis of two functional assays, the receptor binding assay and the hot plate test of analgesia. Interestingly, another analogue of this series, [Dmt1, d-1-Nal4]endomorphin-1, turned out to be a moderately potent mixed mu-agonist/delta-antagonist.

Modulation of morphine and alcohol motor stimulant effects by cannabinoid receptors ligands.

Previous studies evidenced in rats the suppression by cannabinoids of motor stimulant effects of various drugs of abuse. Here we investigated, in mice, the effects of an acute or a chronic administration with the cannabinoid agonist HU 210 on the motor stimulant effects of either morphine or alcohol. HU 210 (12.5-200 microg/kg), when acutely administered, antagonized the stimulant effects of morphine (7.5 mg/kg) or alcohol (1 or 1.5 g/kg). A tolerance to this antagonistic interaction with morphine and alcohol occurred after a 7-day or a 14-day HU 210 treatment, leading to the reappearance of morphine- and alcohol-induced stimulation. The CB1 receptor antagonist rimonabant (10 mg/kg) enhanced the stimulant effect induced by low doses of morphine (5 or 7.5 mg/kg). Rimonabant (3 or 10 mg/kg) altered the locomotor effect of alcohol in a biphasic manner. It enhanced the stimulant effect of low doses of alcohol (1 or 1.5 g/kg) while decreasing the locomotor activity of mice treated with a high dose (3 g/kg) of alcohol. Furthermore, rimonabant (3 and 10 mg/kg) enhanced the duration of alcohol-induced loss of righting reflex (4 g/kg), suggesting a dual implication of cannabinoidergic pathways in acute effects of alcohol.

Antidepressant-like effect of hyperfoliatin, a polyisoprenylated phloroglucinol derivative from Hypericum perfoliatum (Clusiaceae) is associated with an inhibition of neuronal monoamines uptake.

This study investigated, in mice, the antidepressant like effect of hyperfoliatin, a prenylated phloroglucinol derivative isolated from the aerial parts of Hypericum perfoliatum, as well as its action on monoaminergic systems. In the forced-swimming test, hyperfoliatin dose-dependently reduced immobility time. Immobility was interpreted as an expression of "behavioural despair", which could be a component of depression syndrome. The effect of hyperfoliatin did not result from the stimulation of animal motor activity. Hyperfoliatin inhibited, in a concentration-dependent manner, the [(3)H]-dopamine, [(3)H]-serotonin and [(3)H]-noradrenaline synaptosomal uptakes, but did not prevent the binding of specific ligands to the monoamine transporters. These data suggest that the antidepressant-like effect of hyperfoliatin on the forced-swimming test is probably associated to monoamine uptake inhibition, due to a mechanism of action different from that of known antidepressants.