Simultaneous wireless and high-resolution detection of nucleus accumbens shell ethanol concentrations and free motion of rats upon voluntary ethanol intake.

Highly sensitive detection of ethanol concentrations in discrete brain regions of rats voluntarily accessing ethanol, with high temporal resolution, would represent a source of greatly desirable data in studies devoted to understanding the kinetics of the neurobiological basis of ethanol's ability to impact behavior. In the present study, we present a series of experiments aiming to validate and apply an original high-tech implantable device, consisting of the coupling, for the first time, of an amperometric biosensor for brain ethanol detection, with a sensor for detecting the microvibrations of the animal. This device allows the real-time comparison between the ethanol intake, its cerebral concentrations, and their effect on the motion when the animal is in the condition of voluntary drinking. To this end, we assessed in vitro the efficiency of three different biosensor designs loading diverse alcohol oxidase enzymes (AOx) obtained from three different AOx-donor strains: Hansenula polymorpha, Candida boidinii, and Pichia pastoris. In vitro data disclosed that the devices loading H. polymorpha and C. boidinii were similarly efficient (respectively, linear region slope [LRS]: 1.98 ± 0.07 and 1.38 ± 0.04 nA/mM) but significantly less than the P. pastoris-loaded one (LRS: 7.57 ± 0.12 nA/mM). The in vivo results indicate that this last biosensor design detected the rise of ethanol in the nucleus accumbens shell (AcbSh) after 15 minutes of voluntary 10% ethanol solution intake. At the same time, the microvibration sensor detected a significant increase in the rat's motion signal. Notably, both the biosensor and microvibration sensor described similar and parallel time-dependent U-shaped curves, thus providing a highly sensitive and time-locked high-resolution detection of the neurochemical and behavioral kinetics upon voluntary ethanol intake. The results overall indicate that such a dual telemetry unit represents a powerful device which, implanted in different brain areas, may boost further investigations on the neurobiological mechanisms that underlie ethanol-induced motor activity and reward.

Acetaldehyde mediates alcohol activation of the mesolimbic dopamine system.

Ethanol (EtOH), the main psychoactive ingredient of alcoholic drinks, is widely considered to be responsible for alcohol abuse and alcoholism through its positive motivational properties, which depend, at least partially, on the activation of the mesolimbic dopaminergic system. However, acetaldehyde (ACD), the first metabolite of EtOH, has been classically considered to be aversive and useful in the pharmacological therapy of alcoholics. Here we show that EtOH-derived ACD is necessary for EtOH-induced place preference, a pre-clinical test with high predictive validity for reward liability. We also found that ACD is essential for EtOH-increased microdialysate dopamine (DA) levels in the rat nucleus accumbens and that this effect is mimicked by intra-ventral tegmental area (VTA) ACD administration. Furthermore, in vitro, ACD enhances VTA DA neuronal firing through action on two ionic currents: reduction of the A-type K+ current and activation of the hyperpolarization-activated inward current. EtOH-stimulating properties on DA neurons are prevented by pharmacological blockade of local catalase, the main metabolic step for biotransformation of EtOH into ACD in the central nervous system. These results provide in-vivo and in-vitro evidence for a key role of ACD in the motivational properties of EtOH and its activation of the mesolimbic DA system. Additionally, these observations suggest that ACD, by increasing VTA DA neuronal activity, would oppose its well-known peripherally originating aversive properties. Careful consideration of these findings could help in devising new effective pharmacological therapies aimed at reducing EtOH intake in alcoholics.

Effect of simulated microgravity on PGE2-induced edema and hyperalgesia in rat paws: pharmacological data and biochemical correlates.

The aim of the present study has been to investigate the effect of conditions of modeled microgravity using a three-dimensional clinostat (Random Positioning Machine, RPM) on edema and thermal hyperalgesia induced by prostaglandin E2 (PGE2) in the hind paw of rat. Our results showed that RPM reduced PGE2-evoked edema associated to a significant decrease in hyperalgesia compared to ground control animals. To further characterize the mechanisms by which RPM induces anti-inflammatory and anti-hyperalgesic action, we performed biochemical assays of PGE2 and Western immunoblot experiments to assess whether the intraplantar administration of exogenous PGE2 modifies the expression of the iNOS. These results showed that RPM diminished the levels of PGE2 in exudates of paws previously treated with PGE2, but did not influence the iNOS expression.

Anti-inflammatory activity of linalool and linalyl acetate constituents of essential oils.

Linalool and linalyl acetate are the principal components of many essential oils known to possess several biological activities, attributable to these monoterpene compounds. In this work, we evaluated individually the anti-inflammatory properties of (-) linalool, that is, the natural occurring enantiomer, and its racemate form, present in various amounts in distilled or extracted essential oils. Because in the linalool-containing essential oils, linalyl acetate, is frequently present, we also examined the anti-inflammatory action of this monoterpene ester. Carrageenin-induced edema in rats was used as a model of inflammation. The experimental data indicate that both the pure enantiomer and its racemate induced, after systemic administration, a reduction of edema. Moreover, the pure enantiomer, at a dose of 25 mg/kg, elicited a delayed and more prolonged effect, while the racemate form induced a significant reduction of the edema only one hour after carrageenin administration. At higher doses, no differences were observed between the (-) enantiomer and the racemate; a further increase in the dose of both forms did not result in an increased effect at any time of observation. The effects of equi-molar doses of linalyl acetate on local edema were less relevant and more delayed than that of the corresponding alcohol. These finding suggest a typical pro-drug behavior of linalyl acetate. The results obtained indicate that linalool and the corresponding acetate play a major role in the anti-inflammatory activity displayed by the essential oils containing them, and provide further evidence suggesting that linalool and linalyl acetate-producing species are potentially anti-inflammatory agents.

Effects of simulated microgravity conditions on carrageenin-induced oedema in rat.

The effects of simulated microgravity conditions, using a three-dimensional clinostat (Random Positioning Machine, RPM), on carrageenin-induced paw oedema in rats as a model of local inflammation were evaluated. RPM-exposed animals showed a significant reduction of oedema and a more pronounced decrease in body weight with respect to control groups. Moreover, aspirin (ASA) treatment, an anti-inflammatory agent, on RPM-exposed rats did not exhibit any activity after carrageenin challenge with respect to RPM control animals on the ground. ASA activity on RPM could be prevented by RPM-induced anti-oedematous effect. RPM-induced anti-oedematous effect did not reversed by pre-treatment with the non-selective glucocorticoid receptor antagonist, mifepristone ruling out the supposed influence of an of cortisol release during the RPM treatment.

Carbamazepine prevents imipramine-induced behavioural sensitization to the dopamine D(2)-like receptor agonist quinpirole.

Chronic treatment with antidepressants potentiates the behavioural sensitivity to the administration of dopamine receptor agonists. Such supersensitivity might be involved in the mechanism of action of antidepressant drugs, but it has also been suggested to play a role in the mechanisms underlying antidepressant treatment-related mania (i.e. antidepressant-induced mood switch and rapid cycling). Consistently to this hypothesis, we have recently shown that lithium salts, which are poorly effective in antidepressant-related mania, fail to prevent the development of imipramine-induced supersensitivity to the locomotor effect of the dopamine D(2)-like receptor agonist quinpirole. In the present paper, we report the ability of carbamazepine, an anticonvulsant with antimanic and mood stabiliser properties, to prevent the development of supersensitivity to the locomotor response to quinpirole induced by chronic treatment with imipramine. The present results, together with the results of our previous study, might contribute to explain the different responsiveness to lithium and carbamazepine observed in some manic patients, and are consistent with the clinical data suggesting that carbamazepine might be more effective than lithium in antidepressant-related mania.

Synthesis and analgesic-antiinflammatory activities of novel acylarylhydrazones with a 5-phenyl-4-R-3-pyrrolyl-acyl moiety.

A new series of arylidene 5-phenyl-4-R-pyrrole-3-carbohydrazides 1a-j were prepared and evaluated for their analgesic-antiinflammatory activities. All synthesized compounds showed a significant analgesic action in mice after intraperitoneal administration at a dose of 100 microM/kg. Two of these, 1b, (4'-methylbenzylidene)-5-phenyl-1H-pyrrole-3-carbohydrazide, and 1d, (4'-chlorobenzylidene)-5-phenyl-1H-pyrrole-3-carbohydrazide, were found to be more potent as antinociceptive agents respect to dipyrone and indometacin, used as reference drugs. Among compounds 1, only 1b showed a moderate antiinflammatory effect in rats while 1d proved to be a potent non antiinflammatory analgesic.

In vitro release and antiinflammatory activity of topical formulations of ketoprofen.

Ketoprofen (KP) is a potent nonsteroidal anti-inflammatory drug (NSAID) widely used in clinical practice for the control of acute and chronic pain of soft tissues and skeletal muscle system. The importance of KP in the therapeutic field, has stimulated the development of topical dosage forms to improve its percutaneous absorption through the application site. Moreover they could provide relatively consistent drug levels for prolonged periods and avoid gastric irritation, typical side effect of NSAID oral administration. Since the topical formulation efficiency depends on vehicle characteristics, some different ointments, at 1% and 5% concentrations of KP, were evaluated by in vitro and in vivo studies. Among tested ointments, 1% Carbopol cream and 5% Carbopol gel showed the best fluxes of drug through regenerated cellulose membrane. The in vivo percutaneous absorption of KP, evaluated by carrageenan-induced paw edema in rats, showed a good correlation with the in vitro results about considered creams, but the gels in vivo activity was not in according to their in vitro behaviour. The extemporaneous Carbopol cream was able to produce a better edema inhibition than the commercial one, taken as a reference and widely utilized as a topical therapeutic item. About gels, the obtained results were nearly the maximum response considered possible for a topical antiinflammatory drug.

Exploratory behaviour and grooming after repeated restraint and chronic mild stress: effect of desipramine.

In a previous study, we have recently shown that chronic treatment with desipramine either reduced or potentiated the locomotor response to the dopamine D(2)-like receptor agonist quinpirole, a behavioural response mediated by the mesolimbic dopamine system, depending on whether the animals were subjected, respectively, to repeated restraint or to chronic mild stress (different stressors randomly presented). In this study, we examined the interaction between prolonged exposure to either repeated restraint stress or chronic mild stress with the chronic administration of the antidepressant desipramine on two spontaneous behaviours, in which an involvement of the mesolimbic dopamine system has been suggested: novelty-induced exploratory activity and grooming. Exploratory activity in the open field was reduced by chronic mild stress regardless of the drug treatment, while it was not influenced by restraint stress. Desipramine reduced exploratory activity in rats subjected to restraint stress. Restraint stress increased grooming and desipramine reversed this effect, while increasing grooming in the chronic mild stress group. These findings suggest that antidepressants exert their effect by opposing the modifications induced by stress. The available experimental evidence is consistent with the hypothesis that an important role in the observed behavioural changes is played by the mesolimbic dopamine system.

In vitro permeation through porcine buccal mucosa of Salvia desoleana Atzei & Picci essential oil from topical formulations.

In the light of recent studies, which have shown that the essential oil derived from some Lamiaceae species has appreciable anti-inflammatory activity, moderate anti-microbial action and the ability to inhibit induced hyperalgesia, an assessment of the diffusion and permeation of Salvia desoleana Atzei & Picci (S. desoleana) essential oil through porcine buccal mucosa was considered useful for a possible application in the stomatological field. Topical formulations (microemulsions, hydrogels and microemulsion-hydrogels) were prepared for application to the buccal mucosa. The mucosa permeation of the oil from the formulations was evaluated using Franz cells, with porcine buccal mucosa as septum between the formulations (donor compartment) and the receptor phase chambers. The study also aimed at optimising the permeability of the S. desoleana essential oil by means of an enhancer, the diethylene glycol monoethyl ether Transcutol. The diffusion of the oil through the membrane was determined by evaluating the amount of essential oil components present in the receiving solution, the flux and the permeation coefficient (at the steady state) in the different formulations at set intervals. Qualitative and quantitative determinations were done by gas chromatographic analysis. All the formulations allow a high permeability coefficient in comparison with the pure essential oil. In particular, the components with a terpenic structure (beta-pinene, cineole, alpha-terpineol and linalool) have the highest capacity to pass through the porcine buccal mucosa when compared to the other components (linalyl acetate and alpha-terpinil acetate). Moreover, the enhancer, diethylene glycol monoethyl ether largely increases the permeation of the essential oil components in relation to the concentration.

Different effect of desipramine on locomotor activity in quinpirole-treated rats after repeated restraint and chronic mild stress.

We have studied the effect of chronic treatment with the tricyclic antidepressant drug desipramine on locomotor activity in rats challenged with the administration of the D2-like dopamine agonist quinpirole, after prolonged exposure to two different stress regimens, repeated restraint stress and chronic mild stress (different stressors randomly presented). These stress schedules have been previously reported to influence in opposite ways the sensitivity to the locomotor response mediated by the stimulation of mesolimbic dopamine receptors. In particular, repeated restraint has been reported to induce an increased response to the locomotor effect of amphetamine, while chronic mild stress has been reported to induce a decreased locomotor response to quinpirole. In the present study, repeated restraint stress failed to influence the locomotor activity after challenge with quinpirole, while chronic mild stress reduced this response. Chronic treatment with desipramine failed to influence this response in the control group, but exerted opposite effects in the two stressed groups. In particular, chronic desipramine reduced locomotor activity in quinpirole-treated rats in the restraint stress group, and increased it in the chronic mild stress group, thus preventing the subsensitivity induced by this stress regimen. The present results, taken together with results from earlier studies, are consistent with the hypothesis that the effect of antidepressants on the sensitivity of the mesolimbic dopamine receptors mediating the locomotor behavioural response tends to be opposite with respect to that exerted by stress, regardless of its direction. However, since we failed to show an increased locomotor activity after quinpirole challenge in the repeated restraint group, this hypothesis remains to be demonstrated. The two stress schedules reduced body weight gain in a similar way, therefore their different effects do not seem to be due to a difference in stress severity. Thus, the observation that both stress schedules reduced body weight gain in a similar way, but only chronic mild stress reduced the sensitivity to the locomotor response to quinpirole, shows that this effect is not an artefact of body weight decrease.

Chemical composition and antimicrobial action of the essential oils of Salvia desoleana and S. sclarea.

The chemical composition of the essential oils of Salvia desoleana Atzei & Picci and Salvia sclarea L. from Sardinia (Italy) was analysed by GC and GC/MS. S. desoleana oil had a high content of monoterpenic esters (linalyl acetate and alpha-terpinyl acetate) and a lower amount of the corresponding alcohols while S. sclarea oil was characterised by a higher content of alcohols and lower quantity of esters. We studied the antimicrobial activity of these oils concerning their use in pharmaceutical preparations for local application. Only weak microbiostatic inhibitory activity was seen against S. aureus, E. coli, S. epidermidis and C. albicans, but since inhibition increased progressively with contact time, better results could be obtained by using these oils in bioadhesive formulations that would also have anti-inflammatory and peripheral analgesic action at a local level, as demonstrated in experimental animals following systemic application.

Anti-inflammatory activity of aqueous extracts and steroidal sapogenins of Agave americana.

Lyophilized aqueous extracts obtained from Agave americana L (Agavaceae) collected in the north of Sardinia were characterized with regard to their steroidal sapogenin content. Extracts of A. americana and genins isolated from them were evaluated for anti-inflammatory properties by testing their effects on carrageenin-induced edema. The effect of orally administered genins on gastric mucous membranes was also assessed. Lyophilized extracts administered by the intraperitoneal route at doses equivalent to 200 and 300 mg/kg of fresh plant starting material, showed good anti-inflammatory activity. Doses of genins (total steroidal sapogenins, hecogenin and tigogenin) equivalent to the amount in the lyophilized extracts produced an antiedentatous effect which was much stronger and more efficacious than that obtained with an i.p. administration of 5 mg/kg of indomethacin or dexamethasone 21-phosphate at a dose equivalent to the molar content of hecogenin administered. At the doses used to evaluate the anti-inflammatory activity, the genins did not have any harmful effect on the gastric mucous membranes. Lesions occurred when significantly higher doses of hecogenin were given, but gastric damage was still less than that caused by the drugs used for comparative purposes.

Synthesis and antiinflammatory activity of some imidazolinylpyrazole derivatives.

Some new imidazolinyl-pyrazoles (4) were synthesized by condensation of 2-imidazolylhydrazine hydroiodid (1a) with beta-ketoesters to afford the intermediate 3 that, as free base, ring closes only to 4. The attempt to obtain the imidazo-triazepinone 5, starting from 2-imidazolinyl-2-methylhydrazine, was accomplished, in low yield, only in reacting with ethyl acetoacetate. The pharmacological evaluation of antiinflammatory and analgesic activities of pyrazoles 4a-f revealed an appreciable antiinflammatory activity.

Choleretic activity of 3-[ring substituted benzotriazol-1(2)-yl]alkanoic and alkenoic acids.

In order to investigate the influence of structural modifications on the high choleretic activity of 3-(benzotriazol-1-yl)butanoic acid, a set of new benzotriazolyl alkanoic and alkenoic acids was prepared and, together with some other acids previously described, tested in rats by i.v. administration at the dose of 0.5 mmol/kg. Most of the tested compounds exhibited a good choleretic activity comparable with or higher than that of the model acid and of dehydrocholic acid (+56% mean increase of bile volume during 4 hours). Influence of nature and position of substituents was shown in some cases: a moderate decrease of activity was observed for methoxy derivatives and for the introduction of a methyl group in position 6, while a trifluoromethyl group in the same position enhanced the activity (10). Activity was maintained after the introduction of unsaturation in the chain (17,18), but was completely suppressed when unsaturation was associated with a shortening of the alkenoic chain (16). Moving the butanoic chain from position 1 to position 2 in the case of the nitroderivative (15) produced a striking increase of activity (from +42 to +118 mean variation of bile volume during 4 hours), while the same change in the unsubstituted acid 1 abolished the activity.

Synthesis and CNS activities of pyridopyrazinone and pyridodiazepinone derivatives.

New tricyclic derivatives with cyclocondensed pyrido-pyrazine 7,10 and pyrido-diazepine 20a,20b skeletons were synthetized and biologically investigated. The compounds, preliminarily tested on explorative, muscle relaxing, antinociceptive, spontaneous motor activities and influence on the narcotic effect of Evipan, revealed interesting CNS depressant and analgesic activities. The pyrido[2,3-e]pyrrolo[1,2-a]pyrazine structure of 7 appeared the most promising for analgesic and neuroleptic activities. The above compounds were assayed also for their capacity to inhibit DNA synthesis in Ehrlich ascites tumor cells; 20a appeared to be able of inducing a significant inhibition.

Cyclopenta[e] [1,5] benzodiazepin-10 (9H)-one derivatives as CNS depressant agents.

A series of dialkylaminoalkyl derivatives of cyclopenta[e] [1,5] benzodiazepin-10(9H)-one (E1-4) and its 6-chloro derivative (E5-8) was prepared to evaluate their CNS activity in comparison with that of isosteric pyridodiazepinones (A1-4) previously described. The results of the pharmacological screening show a significant depressant activity more remarkable in 6-chloro derivatives, which also revealed a high and lasting analgesic activity. The replacement of pyridine with benzene nucleus did not show any significant or homogeneous activity variation.

Synthesis and pharmacological activity of 4-carbamoyl-6-beta-thienyl-4,5-dihydropyridazin-3-(2H)ones.

A new series of 4-carbamoyl-6-beta-thienyl-4,5-dihydropyridazin-3-(2H)ones 4a-g have been synthesized and tested for their anti-inflammatory and analgesic properties. Among the tested compounds, only 4f at 1 mmole/Kg showed antiinflammatory activity that was comparable with that of indomethacin (5 mg/Kg) though of shorter duration. Compounds 4a, 4e and especially 4g at 0.2 mmoles/Kg displayed relevant analgesic activity, 4g being the most potent derivative in the writhing test. Compounds 4c and 4g were found to possess analgesic activity also in the hot plate test.