Therapeutic perspectives for melatonin agonists and antagonists.

Melatonin is a neurohormone synthesized in the pineal gland during the dark period in all species, including humans. The diversity and differences in melatonin receptor distribution in the brain and extracerebral organs suggest multiple functional roles for melatonin. Administration of melatonin agonists reduces neophobia and treatment with a melatonin antagonist during the dark period reverses the anxiolytic-like effect of endogenous melatonin. Chronic treatment with agonists prevents various perturbations induced by chronic mild stress. Melatonin in vivo directly constricts cerebral arterioles in rats and decreases the lower limit of cerebral blood flow autoregulation, suggesting that melatonin may diminish the risk of hypoperfusion-induced cerebral ischemia. At the extracerebral level, melatonin regulates intestinal motility in rats. The intestinal postprandial motor response is shorter in the dark phase than in the light phase and this reduction is reversed in animals pretreated with a melatonin antagonist. Moreover, melatonin reduces the duration of cholecystokinin excitomotor effect. Endogenous melatonin may modulate intestinal motility to coordinate intestinal functions such as digestion and transit and control the metabolism of the animal. An adipocyte melatonin binding site may also participate in this control. Melatonin is involved in a wide range of physiological functions. The question remains as to whether evolution, adaptation and diurnal life have modified the physiological role of melatonin in humans. Moreover, the functional role of each of the receptor subtypes has to be characterized to design selective ligands to treat specific diseases.

5-substituted 3,4-dihydro-3-amino-2H-1-benzopyran derivatives: synthesis and interaction with serotoninergic receptors.

A new series of 3,4-dihydro-3-amino-2H-1-benzopyran derivatives (1 and 2) bearing various substituents on the 5-position was successfully prepared via palladium-mediated cross-coupling reactions. Some of the new compounds showed high affinity for 5-HT1A and 5-HT7 receptors. The best affinity for the 5-HT1A and 5-HT7 receptors was obtained for 2b (Ki = 0.3 nM for 5-HT1A and 3.1 nM for 5-HT7). The anxiolytic activity of compound 2b was evaluated.

Neurophysiological effects of an extract of Eschscholzia californica Cham. (Papaveraceae).

An aqueous alcohol extract of Eschscholzia californica (Ec) has been evaluated for benzodiazepine, neuroleptic, antidepressant, antihistaminic and analgesic properties, in order to complete the study of the sedative and anxiolytic effects previously demonstrated. The plant extract did not protect mice against the convulsant effects of pentylenetetrazol, and did not cause muscle relaxant effects but appeared to possess an affinity for the benzodiazepine receptor: thus, flumazenil, an antagonist of these receptors, suppressed the sedative and anxiolytic effects of the extract. The Ec extract induced peripheral analgesic effects in mice but did not possess antidepressant, neuroleptic or antihistaminic effects.

Reentrainment of the spontaneous locomotor activity rhythm to a daylight reversal in C57BL/6 and C3H/He mice: implication of melatonin.

The adaptation of the locomotor activity rhythm to a daylight reversal was previously found to be faster in C57BL/6 mice, which present a low level of melatonin, than in C3H/He mice, which exhibit a large nocturnal melatonin peak. Because pinealectomy has been shown to accelerate resynchronisation time in rats after a daylight reversal, we investigated the involvement of melatonin in the resynchronisation rate of locomotor activity rhythm in C57BL/6 and C3H/He strains. We first tested the effects of melatonin, administered at zeitgeber time (ZT) 20 (with ZT0 corresponding to light onset) for the 3 days preceding the daylight reversal, on the reentrainment of locomotor activity rhythm in both strains. Second, the effects of S-22153, a melatonin receptor antagonist, on the reentrainment of locomotor activity rhythm in C3H/He mice were examined. S-22153 was administered for the 3 days preceding the daylight reversal either at ZT12 or at ZT20, i.e., when endogenous melatonin levels are respectively low and high. Melatonin significantly delayed the resynchronisation of locomotor activity rhythm in C57BL/6 mice without affecting this parameter in C3H/He mice. S-22153 significantly accelerated the resynchronisation in C3H/He mice when administered at ZT20, but had no effect when administered at ZT12. These results support the hypothesis that the differences between C3H/He and C57BL/6 in the reentrainment of their locomotor activity rhythm depend, at least in part, on the interstrain differences in melatonin synthesis.

Anxiolytic-like properties of melatonin receptor agonists in mice: involvement of mt1 and/or MT2 receptors in the regulation of emotional responsiveness.

The anxiolytic-like properties of melatonin have been established in rodents. The present study investigated the possible involvement of melatonin receptors/binding sites in the regulation of emotional responsiveness in mice, using an mt1/MT2 receptor specific agonist (S 23478) and two specific ligands of MT3 binding sites with agonistic properties (N-acetylserotonin (NAS) and 5-methoxycarbonylamino N-acetyltryptamine (5-MCA-NAT)). We examined the behavioural effects of these compounds in C3H/He mice confronted with two anxiety models: the free-exploratory test, in which C3H/He mice present neophobic reactions ("trait" anxiety), and the light/dark choice test, which is an unconditioned conflict test (inducing "state" anxiety). Melatonin and S 23478 decreased anxious reactions in both the free-exploratory test (5-25 mg/kg) and the light/dark choice test (melatonin: 20 mg/kg; S 23478: 10-20-40 mg/kg). NAS exerted anxiolytic-like effects only at a dose of 35 mg/kg in the free-exploratory test and at a dose of 40 mg/kg in the light/dark choice test. Finally, 5-MCA-NAT was devoid of anxiolytic-like effects in both tests. These results suggest that the anxiolytic properties of melatonin could involve the activation of mt1 and/or MT2 receptors rather than of the MT3 binding site.

Hypoexpression of benzodiazepine receptors in the amygdala of neophobic BALB/c mice compared to C57BL/6 mice.

The distribution of benzodiazepine receptors in the brain of neophobic BALB/c mice was studied by autoradiographic analysis using [3H]-diazepam and compared to that of the same receptors of the "nonemotional" C57BL/6 mice. This technique revealed no significant interstrain difference except for a lower density of diazepam binding sites in the amygdala of BALB/c mice. Therefore, the expression of benzodiazepine receptors in the amygdala of the two strains of mice were quantified by binding studies on brain membranes. The amygdala of BALB/c mice exhibited a fivefold decrease in the density of benzodiazepine receptors compared to C57BL/6 mice. These results suggest that the trait anxiety (neophobia) that characterizes BALB/c mice could be due, at least in part, to a genetic modulation of benzodiazepine receptor expression in the amygdala, a structure known to be strongly involved in fear behavior.

Antagonistic effects of S 22153, a new mt1 and MT2 receptor ligand, on the neophobia-reducing properties of melatonin in BALB/c mice.

When exposed to a free-exploratory situation, BALB/c mice are well known to exhibit strong avoidance responses toward unfamiliar places (neophobia). Because melatonin was found to significantly reduce neophobia in BALB/c mice, it seemed interesting to examine potential antagonistic effects of S 22153, a new melatonin mt1 and MT2 receptor ligand, on the neophobia-reducing properties of melatonin in BALB/c mice confronted with the free-exploratory paradigm. S 22153 was able to block, in a dose-dependent manner, the anxiolytic-like properties of melatonin when it was administered 5 min before melatonin. The antagonistic effects of S 22153 persisted when the drug was administered 2 or 4 h before melatonin, and were almost abolished when it was administered 6 h before melatonin. These results suggest that the anxiolytic-like effects of melatonin on the neophobic responses in BALB/c mice are mediated by mt1 and/or MT2 receptors.

Effects of melatonin on neophobic responses in different strains of mice.

Anxiolytic properties of melatonin in rodents had usually been examined in behavioral tests based on stressful situations, i.e., in animal models of "state" anxiety. However, no study reports effects of melatonin on emotionality of rodents submitted to situations devoid of stressful components as in the free-exploratory test, which gives to animals the opportunity to choose freely between familiar and unfamiliar places. This procedure has been proposed as a method for measuring an endogenous form of anxiety called "trait" anxiety. The present study first investigated the effects of melatonin on neophobic responses of male C57BL/6, C3H/He, and BALB/c mice submitted to a free-exploratory test. Results demonstrated that melatonin had no effect in C57BL/6 mice that presented very low neophobic responses, whereas it was effective in reducing neophobia of BALB/c and C3H/He mice that presented, respectively, strong and intermediate avoidance responses towards unfamiliarity. Indeed, mice of both latter strains treated with melatonin made fewer attempts to enter into the unfamiliar compartment, exhibited a lower latency of the first entry into the unfamiliar places, and spent more time in them. Thus, melatonin appeared to be equally effective in reducing "trait" anxiety in both BALB/c and C3H/He mice. Moreover, flumazenil was able to counteract, in a dose-dependent manner, the anxiolytic activity of melatonin in BALB/c, suggesting involvement of central GABAergic system in the pharmacological effects of melatonin.

Regulation of emotional behaviour by day length in mice: implication of melatonin.

Pineal melatonin secretion occurs at night in all vertebrates and the duration of its secretion is negatively correlated with day length. As short-day exposure was previously shown to decrease emotional behaviour of mice toward an unfamiliar environment, the present study was designed to determine whether such behavioural changes could be mediated by melatonin. In a first experiment, the effects of a 3-week exposure to various day lengths (18h-6h, 12h-12h and 6h-18h light-dark conditions) on neophobic behaviour (free-exploratory paradigm) were examined in both BALB/c mice, which exhibit a very transitory melatonin peak of low amplitude in a 12h light-12h dark cycle, and C3H/He mice, which present a clear melatonin rise during the night-time. A second experiment was designed to determine if the decrease of emotional reactivity induced by a short-day exposure (6h-18h light-dark cycle during 3 weeks) in C3H/He mice could be counteracted by a daily treatment with a melatonin antagonist, S 22153 (1, 5 and 25 mg/kg/day). The short-day exposure was found to decrease neophobic reactions in both C3H/He and BALB/c mice. In contrast, the long-day exposure enhanced neophobia in C3H/He mice only. S 22153 was found to counteract, in a dose-dependent manner, the anxiolytic-like effects induced by the short-day exposure in C3H/He mice. The present results provide evidence that the modulation of circulating melatonin could be involved in the emotional changes related to day-length variations. Further studies are needed to investigate whether pinealectomy could counteract the photoperiod-related changes in anxiety.

The effects of melatonin on the behavioural disturbances induced by chronic mild stress in C3H/He mice.

In rodents, exposure to chronic mild stress (CMS) is known to induce unresponsiveness to environmental stimuli, as well as sleep disturbances, suggesting some analogies between this syndrome and human depression. Furthermore, numerous studies reported a decrease in nocturnal melatonin concentration in depressed patients, compared with controls. The present study was conducted to test a possible preventative action of daily treatment with melatonin on behavioural alterations induced in C3H/He mice by CMS exposure. In addition to daily spontaneous locomotor activity and preference for sucrose solution, the emotional behaviour of mice was examined in a stressful situation (light/dark choice test), as well as in a situation devoid of constraining components (free-exploratory paradigm), after three weeks of CMS. The results showed that the behaviour of C3H/He mice was disrupted after CMS. Stressed mice exhibited blunted emotional reactivity in both the light/dark choice test and the free-exploratory situation. While unstressed mice presented no variation in their preference for a sucrose solution, stressed mice presented a decrease in such preference towards the end of the CMS exposure. Furthermore, daily spontaneous locomotor activity of the mice was reduced after CMS. Daily treatment of stressed mice with melatonin was able to prevent several CMS-induced disturbances, except in the light/dark choice test, where melatonin was ineffective. Compared to the effects of 10 mg/kg of fluoxetine, which completely prevented CMS-induced dysregulation of behaviour, melatonin was less effective. The present results support the idea that melatonin may be implicated in an homeostatic system which protects animals from disruptions induced by chronic stress.

Comparative study of emotional behaviour in three inbred strains of mice.

Anxious behaviour constitutes one of the markers that best differentiated the 'non-emotional' C57BL/6 (C57) strain from the 'emotional' BALB/c (BALB) strain. Interestingly, C3H/He (C3H) mice, which possess a common genetic background to BALB, have also been found to be 'emotional' mice. The present study first illustrates that BALB and C3H mice exhibit a higher emotional level both in non-constraining situation (free-exploratory paradigm) and in a stressful one (light/dark choice test). Second, the present results confirmed the hypothesis that trait anxiety, modelled neophobic reactions in the free-exploratory paradigm, is related to GABA-benzodiazepine system rather than to the serotoninergic one, in contrast to state anxiety, modelled by the light/dark choice test. Like in BALB mice, chlordiazepoxide was able to counteract the neophobic responses in C3H mice confronted with the free-exploratory paradigm, in contrast to 8-OH-DPAT which did not modify C3H behaviour in this test. Conversely, both chlordiazepoxide and 8-OH-DPAT reduced anxious reactions of BALB and C3H mice in the light/dark choice test, showing that BALB and C3H mice are sensitive to serotonin-interacting drugs only when confronted with a stressful situation. Furthermore, chlordiazepoxide and 8-OH-DPAT were found to be devoid of anxiolytic-like effect in C57 mice. The lack of anxiolytic-like activity of these compounds in C57 mice can be related to the low basal level of emotional reactions of this strain. Finally, the present results showed that chronic stress was able to blunt general emotional level of mice in both experimental situations and that both BALB and C3H mice were more sensitive to chronic stress than C57 ones. In conclusion, BALB and C3H may be two strains of choice for testing potential anti-anxiety treatments which might be of great long-term benefit in the chronically anxious patients. Furthermore, these strains constitute interesting material for determining hereditary factors which presumably underlie the development of some chronic anxiety disorders. Finally, since C3H and BALB mice appeared particularly susceptible to chronic stress, it can be proposed that these strains may be a useful tool to examine the possible common origins of anxiety disorders and chronic stress-related disorders, such as certain forms of depression.

Effects of S-21007, a potent 5-HT3 partial agonist, in mouse anxiety.

AIM: To study the effect of S-21007, a 5-HT3 partial agonist in different animal models of anxiety in mice. METHODS: S-21007 effects were evaluated in the behavior tests after intraperitoneal and oral acute treatment or in the light/dark test after both acute and chronic treatments. RESULTS: S-21007 presented anxiolytic-like properties after acute administration in the light/dark box test, the mirrored chamber test, and the elevated plus-maze at low doses 10 ng.kg(-1)-100 micrograms.kg-1, 1-100 micrograms.kg-1 and 10-100 micrograms.kg-1, respectively. In the light/dark box test, S-21007 was active orally after acute treatment at 100 ng.kg(-1)-10 mg.kg-1 and after chronic treatment (14 d) at 1-10 micrograms.kg-1. S-21007 was devoid of sedative or stimulatory effects. CONCLUSION: S-21007 exhibited anxiolytic-like properties. The mechanism of action may be a desensitization of 5-HT3 receptor or an antagonist activity on the 5-HT3 receptors.

Paternal alcohol exposure: developmental and behavioral effects on the offspring of rats.

The effect of paternal alcohol exposure on neurochemical and behavioral parameters was investigated using as a model system glial cells derived from newborn rat brain and cultured for 4 weeks. The total brain neurochemical parameters from rats born to mothers sired by an alcohol treated father were also investigated. Enzymatic markers of nerve cell development (enolase isoenzymes and glutamine synthetase) and the defense system (superoxide dismutase) against free radicals formed during alcohol degradation were measured in order to evaluate nerve cell damage. Behavioral locomotor tests (open-field, novelty-seeking, light/dark) were carried out to show long-lasting effects of paternal alcoholization on the offspring. Behavioral and developmental alterations were found until 1 year of age in the offspring and a significant growth retardation was observed in the males. Our results suggest that paternal alcohol exposure produces developmental and behavioral effects in the offspring. The consequence of either alcohol withdrawal during stage one spermatogenesis, or maternal diet supplementation with manganese during pregnancy were investigated. It was observed that some of the effects of paternal alcohol exposure on the offspring may be reversed by these treatments.

Effects of a daylight cycle reversal on locomotor activity in several inbred strains of mice.

There is some evidence of melatonin implication in the nycthemeral regulation of running activity rhythm in rodents. Because some inbred strains of mice such as C57BL/6 and BALB/c have been generally found to present no nocturnal melatonin peak, in contrast to others such as C3H/He and CBA mice, the aim of this study was to examine the adaptation of daily locomotor activity to a light/dark cycle phase shift in these four strains. An apparatus consisting of two boxes connected by a tunnel was used to record spontaneous locomotor activity, defined as the number of transitions between the two boxes. Locomotor activity was monitored continuously during 3 days before and 14 days after a 12-h phase delay of the light/dark cycle. Results essentially showed that the adaptation of the locomotor activity rhythm to the phase shift was faster in C57BL/6 and BALB/c mice than in C3H/He and CBA mice. This could be related, at least in part, to the differences in melatonin synthesis between the former strains and the latter ones. Although melatonin nocturnal peak is not necessary to a daylight regulation of circadian functions in rodents, it could be considered as an endocrine message that takes part in the anticipation of the following light/dark cycle.

Novel and selective partial agonists of 5-HT3 receptors. 2. Synthesis and biological evaluation of piperazinopyridopyrrolopyrazines, piperazinopyrroloquinoxalines, and piperazinopyridopyrroloquinoxalines.

In continuation of our previous work on piperazinopyrrolothienopyrazine derivatives, three series of piperazinopyridopyrrolopyrazines, piperazinopyrroloquinoxalines, and piperazinopyridopyrroloquinoxalines were prepared and evaluated as 5-HT3 receptor ligands. The chemical modifications performed within these new series led to structure-activity relationships regarding both high affinity and selectivity for the 5-HT3 receptors that are in agreement with those established previously for the pyrrolothienopyrazine series. The best compound (8a) obtained in these new series is in the picomolar range of affinity for 5-HT3 receptors with a selectivity higher than 10(6). Four of the high-affinity 5-HT3 ligands (8a, 15a,b, and 16d) were selected in both the pyridopyrrolopyrazine and the pyrroloquinoxaline series and were characterized in vitro and in vivo as agonists or partial agonists. Compound 8a was also evaluated in the light/dark test where it showed potential anxiolytic-like activity at very low doses per os.

Behavioural effects of Passiflora incarnata L. and its indole alkaloid and flavonoid derivatives and maltol in the mouse.

Lyophilised hydroalcoholic and aqueous extracts of the aerial parts of Passiflora incarnata L. (Passifloraceae) (Passion-flower), as well as chemical constituents of the plant, indole alkaloids (harman, harmin, harmalin, harmol, and harmalol) maltol and flavonoids (orientin, isoorientin, vitexin and isovitexin) were assessed for behavioral effects in mice. The accordance with the traditional use of P. incarnata, psychotropic properties were confirmed by some behavioral tests in mice. The anxiolytic properties of hydroalcoholic extract were confirmed at 400 mg/kg by the increase of rears and steps climbed in the staircase test (non-familiar environmental test), and the increase in locomotion and time spent in light side in the light/dark box choice test (non-familiar environmental test). The sedative properties of aqueous extract were confirmed at 400 g/kg by decrease of rears and steps climbed in the staircase test and the decrease of rears and locomotion in the free exploratory test. Moreover, the aqueous extract induced sleep in mice after treatment with a sub-hypnotic dose of pentobarbital.

Effects of day-length variations on emotional responses towards unfamiliarity in Swiss mice.

Pineal melatonin secretion occurs at night in all vertebrates and the duration of its secretion is negatively correlated with day length. As an anxiolytic activity of melatonin has been shown in rats and mice, this study examined possible changes of emotional reactivity in response to day length variations in Swiss mice. Three groups of mice were observed in a free-exploratory test: a group submitted to a short-day exposure (6:18 h light-dark cycle) for 2 weeks, a group submitted to a long-day exposure (18:6 h light-dark cycle) for 2 weeks and a control group which was maintained in housing 12:12 h light-dark cycle. The short-day exposed group of mice exhibited significantly fewer attempts to enter into the unfamiliar enclosure, spent significantly more time in it and presented significantly more rears than controls whereas the long-day exposed group of mice made more attempts than controls. These results suggest a decreased emotional level in short-day exposed mice and an increased level in long-day exposed mice. This could be interpreted as confirming the idea of anxiolytic-like properties of melatonin; however, the specific role of this hormone in the changes of anxiety related to day length must be assessed by further measures of potential variations of circulating melatonin.

5-HT1B receptor knock out--behavioral consequences.

Serotonin is a neuromodulator that is involved in a number of mood disorders such as depression, anxiety and impulsive violence. In an attempt to dissect the contribution of individual 5-HT receptor subtypes to behavior, we have generated by homologous recombination, mutant mice lacking the 5-HT1B receptor. These mice did not exhibit any obvious developmental or behavioral defect. However, the hyperlocomotor effect of the 5-HT1A/1B agonist, RU 24969 was completely absent in mutant mice, indicating that this effect is mediated by 5-HT1B receptors. Moreover, when confronted with an intruder, isolated mutant mice attacked the intruder faster and more intensely than wild-type mice, suggesting an involvement of 5-HT1B receptors in the modulation of aggressive behavior. These data might be related to the fact that a class of 5-HT1 agonists, termed serenics, have anti-aggressive properties, and with the findings that certain impulsive aggressive behaviors are associated with deficits in central serotonin.

Influence of dystrophin-gene mutation on mdx mouse behavior. I. Retention deficits at long delays in spontaneous alternation and bar-pressing tasks.

X-linked Duchenne muscular dystrophy (DMD) is frequently associated with a nonprogressive, cognitive defect attributed to the absence of dystrophin in the brain of DMD patients. The mutant mdx mouse, lacking in 427-kDa dystrophin in both muscle and brain tissues, is considered to be a valuable model of human DMD. In the present study, we compared mdx and C57BL/10 control mice and showed that mdx mice had impaired retention in a T-maze, delayed spontaneous alternation task 24 h, but not 6 h, after acquisition. mdx mice were not impaired in acquisition of a bar-pressing task on 4 consecutive days but showed poor retention 22 days after the last training session. Mutants and controls showed similar behavioral responses in free exploration and light/dark choice situations and did not differ in spontaneous locomotor activity or motor coordination. Retention impairments at long delays in mdx mice suggest a role of dystrophin in long-term consolidation processes.