The rostromedial tegmental nucleus RMTg is not a critical site for ethanol-induced motor activation in rats.

RATIONALE: Opioid drugs indirectly activate dopamine (DA) neurons in the ventral tegmental area (VTA) through a disinhibition mechanism mediated by mu opioid receptors (MORs) present both on the GABA projection neurons located in the medial tegmental nucleus/tail of the VTA (RMTg/tVTA) and on the VTA GABA interneurons. It is well demonstrated that ethanol, like opioid drugs, provokes VTA DA neuron disinhibition by interacting (through its secondary metabolite, salsolinol) with MORs present in VTA GABA interneurons, but it is not known whether ethanol could disinhibit VTA DA neurons through the MORs present in the RMTg/tVTA. OBJECTIVES: The objective of the present study was to determine whether ethanol, directly microinjected into the tVTA/RMTg, is also able to induce VTA DA neurons disinhibition. METHODS: Disinhibition of VTA DA neurons was indirectly assessed through the analysis of the motor activity of rats. Cannulae were placed into the tVTA/RMTg to perform microinjections of DAMGO (0.13 nmol), ethanol (150 or 300 nmol) or acetaldehyde (250 nmol) in animals pre-treated with either aCSF or the irreversible antagonist of MORs, beta-funaltrexamine (beta-FNA; 2.5 nmol). After injections, spontaneous activity was monitored for 30 min. RESULTS: Neither ethanol nor acetaldehyde directly administered into the RMTg/tVTA were able to increase the locomotor activity of rats at doses that, in previous studies performed in the posterior VTA, were effective in increasing motor activities. However, microinjections of 0.13 nmol of DAMGO into the tVTA/RMTg significantly increased the locomotor activity of rats. These activating effects were reduced by local pre-treatment of rats with beta-FNA (2.5 nmol). CONCLUSIONS: The tVTA/RMTg does not appear to be a key brain region for the disinhibiting action of ethanol on VTA DA neurons. The absence of dopamine in the tVTA/RMTg extracellular medium, the lack of local ethanol metabolism or both could explain the present results.

N-Acetylcysteine normalizes brain oxidative stress and neuroinflammation observed after protracted ethanol abstinence: a preclinical study in long-term ethanol-experienced male rats.

RATIONALE: Using a preclinical model based on the Alcohol Deprivation Effect (ADE), we have reported that N-Acetylcysteine (NAC) can prevent the relapse-like drinking behaviour in long-term ethanol-experienced male rats. OBJECTIVES: To investigate if chronic ethanol intake and protracted abstinence affect several glutamate transporters and whether NAC, administered during the withdrawal period, could restore the ethanol-induced brain potential dysfunctions. Furthermore, the antioxidant and anti-inflammatory effects of NAC during abstinence in rats under the ADE paradigm were also explored. METHODS: The expression of GLT1, GLAST and xCT in nucleus accumbens (Nacc) and dorsal striatum (DS) of male Wistar was analysed after water and chronic ethanol intake. We used the model based on the ADE within another cohort of male Wistar rats. During the fourth abstinence period, rats were treated for 9 days with vehicle or NAC (60, 100 mg/kg; s.c.). The effects of NAC treatment on (i) glutamate transporters expression in the Nacc and DS, (ii) the oxidative status in the hippocampus (Hip) and amygdala (AMG) and (iii) some neuroinflammatory markers in prefrontal cortex (PFC) were tested. RESULTS: NAC chronic administration during protracted abstinence restored oxidative stress markers (GSSG and GGSH/GSH) in the Hip. Furthermore, NAC was able to normalize some neuroinflammation markers in PFC without normalizing the observed downregulation of GLT1 and GLAST in Nacc. CONCLUSIONS: NAC restores brain oxidative stress and neuroinflammation that we previously observed after protracted ethanol abstinence in long-term ethanol-experienced male rats. This NAC effect could be a plausible mechanism for its anti-relapse effect. Also, brain oxidative stress and neuroinflammation could represent and identify plausible targets for searching new anti-relapse pharmacotherapies.

The Effects of N-Acetylcysteine on the Rat Mesocorticolimbic Pathway: Role of mGluR5 Receptors and Interaction with Ethanol.

N-acetylcysteine (NAC) is a prodrug that is marketed as a mucolytic agent and used for the treatment of acetaminophen overdose. Over the last few decades, evidence has been gathered that suggests the potential use of NAC as a new pharmacotherapy for alcohol use disorder (AUD), although its mechanism of action is already being debated. In this paper, we set out to assess both the potential involvement of the glutamate metabotropic receptors (mGluR) in the possible dual effect of NAC administered at two different doses and NAC's effect on ethanol-induced activation. To this aim, 30 or 120 mg/kg of NAC was intraperitoneally administered to rats with the presence or absence of the negative allosteric modulator of mGluR5 (MTEP 0.1 mg/kg). Thereafter, the cFOS IR-cell expression was analyzed. Secondly, we explored the effect of 120 mg/kg of NAC on the neurochemical and behavioral activation induced by intra-VTA ethanol administration (150 nmol). Our results showed that the high NAC dose stimulated cFOS expression in the NAcc, and that this effect was suppressed in the presence of MTEP, thus suggesting the implication of mGluR5. Additionally, high doses could attenuate the ethanol-induced increase in cFOS-expression in the NAcc, probably due to a phenomenon based on the long-term depression of the MSNs. Additional experiments are required to corroborate our hypothesis.

Efficacy of N-acetylcysteine in the prevention of alcohol relapse-like drinking: Study in long-term ethanol-experienced male rats.

Alcohol use disorders are chronic and highly relapsing disorders, thus alcoholic patients have a high rate of recidivism for drug use even after long periods of abstinence. The literature points to the potential usefulness of N-acetylcysteine (NAC) in the management of several substance use disorders probably due to its capacity to restore brain homeostasis of the glutamate system disrupted in addiction. However, there is little evidence in the case of alcohol. The aim of this study was to explore the potential anti-relapse efficacy of NAC using the alcohol deprivation effect (ADE) model in long-term experienced rats. Two experiments were performed in male Wistar rats to: (a) test the efficacy of NAC to prevent relapse and (b) discriminate the best administration schedule (intermittent vs. continuous) for NAC. In the first experiment, animals were implanted with mini-osmotic pumps delivering 0 or 1 mg/hr NAC during 14 days. In a second experiment, rats received 0, 60, or 100 mg/kg once daily by subcutaneous injection. The efficacy to prevent ADE was evaluated in both experiments. NAC subcutaneously administered, either by continuous infusion or by intermittent injections regimen, is able to block the ADE. The best results were obtained after using 60 mg/kg NAC dose. Our findings support the hypothesis that NAC may represent a valuable therapy in the management of alcohol relapse.

Antidepressant efficacy of a selective organic cation transporter blocker in a mouse model of depression.

Current antidepressants act principally by blocking monoamine reuptake by high-affinity transporters in the brain. However, these antidepressants show important shortcomings such as slow action onset and limited efficacy in nearly a third of patients with major depression disorder. Here, we report the development of a prodrug targeting organic cation transporters (OCT), atypical monoamine transporters recently implicated in the regulation of mood. Using molecular modeling, we designed a selective OCT2 blocker, which was modified to increase brain penetration. This compound, H2-cyanome, was tested in a rodent model of chronic depression induced by 7-week corticosterone exposure. In male mice, prolonged administration of H2-cyanome induced positive effects on several behaviors mimicking symptoms of depression, including anhedonia, anxiety, social withdrawal, and memory impairment. Importantly, in this validated model, H2-cyanome compared favorably with the classical antidepressant fluoxetine, with a faster action on anhedonia and better anxiolytic effects. Integrated Z-scoring across these depression-like variables revealed a lower depression score for mice treated with H2-cyanome than for mice treated with fluoxetine for 3 weeks. Repeated H2-cyanome administration increased ventral tegmental area dopaminergic neuron firing, which may underlie its rapid action on anhedonia. H2-cyanome, like fluoxetine, also modulated several intracellular signaling pathways previously involved in antidepressant response. Our findings provide proof-of-concept of antidepressant efficacy of an OCT blocker, and a mechanistic framework for the development of new classes of antidepressants and therapeutic alternatives for resistant depression and other psychiatric disturbances such as anxiety.

Activation of MORs in the VTA induces changes on cFos expression in different projecting regions: Effect of inflammatory pain.

Chronic pain is a worldwide major health problem and many pain-suffering patients are under opioid based therapy. Epidemiological data show that pain intensity correlates with the risk of misuse of prescription opioids, and other drugs of abuse including alcohol. This increased vulnerability to suffer Substance Use Disorders could be, in part, caused by functional changes that occur over the mesocorticolimbic system, a brain pathway involved in reward processing and addiction. Previous data in rats revealed that inflammatory pain desensitizes mu opioid receptors (MORs) in the ventral tegmental area (VTA). As a consequence, pain alters dopamine release in the nucleus accumbens (NAc) derived from MOR activation in the VTA and also increases intake of high doses of heroine. Given that the VTA neurons target different brain regions, in the present study we first analyzed changes induced by inflammatory pain in the MOR dependent activation pattern of the main VTA projecting areas. To do that, we administered two doses (7 or 14 ng) of DAMGO (MORs agonist) or artificial cerebrospinal fluid (aCSF) focally into the VTA of rats and measured the activation in projection areas by cFos immunohistochemistry. Our results show that focal injections of DAMGO in the VTA increases cFos expression in the majority of its projecting areas, namely NAc, basolateral amygdala (BLA), cingulate cortex (ACC) and bed nucleus of the stria terminalis (BNST), as compared to aCSF. Second, we analyzed whether inflammatory pain would affect to cFos expression using a group of rats injected with CFA in the hind paw. In this case, we found that cFos expression was not significantly different between DAMGO and aCSF administered rats in BLA, ACC and BNST. Our results confirm that inflammatory pain induces desensitization of VTA MORs in a region dependent manner which can be very relevant for addictive behaviours.

Pre-Clinical Studies with D-Penicillamine as a Novel Pharmacological Strategy to Treat Alcoholism: Updated Evidences.

Ethanol, as other drugs of abuse, is able to activate the ventral tegmental area dopamine (VTA-DA) neurons leading to positively motivational alcohol-seeking behavior and use, and, ultimately to ethanol addiction. In the last decades, the involvement of brain-derived acetaldehyde (ACD) in the ethanol actions in the mesolimbic pathway has been widely demonstrated. Consistent published results have provided a mechanistic support to the use of ACD inactivating agents to block the motivational and reinforcing properties of ethanol. Hence, in the last years, several pre-clinical studies have been performed in order to analyze the effects of the sequestering ACD agents in the prevention of ethanol relapse-like drinking behavior as well as in chronic alcohol consumption. In this sense, one of the most explored interventions has been the administration of D-Penicillamine (DP). These pre-clinical studies, that we critically summarize in this article, are considered a critical step for the potential development of a novel pharmacotherapeutic strategy for alcohol addiction treatment that could improve the outcomes of current ones. Thus, on one hand, several experimental findings provide the rationale for using DP as a novel therapeutic intervention alone and/or in combination to prevent relapse into alcohol seeking and consumption. On the other hand, its effectiveness in reducing voluntary ethanol consumption in long-term experienced animals still remains unclear. Finally, this drug offers the additional advantage that has already been approved for use in humans, hence it could be easily implemented as a new therapeutic intervention for relapse prevention in alcoholism.

Glutamate and Opioid Antagonists Modulate Dopamine Levels Evoked by Innately Attractive Male Chemosignals in the Nucleus Accumbens of Female Rats.

Sexual chemosignals detected by vomeronasal and olfactory systems mediate intersexual attraction in rodents, and act as a natural reinforcer to them. The mesolimbic pathway processes natural rewards, and the nucleus accumbens receives olfactory information via glutamatergic projections from the amygdala. Thus, the aim of this study was to investigate the involvement of the mesolimbic pathway in the attraction toward sexual chemosignals. Our data show that female rats with no previous experience with males or their chemosignals display an innate preference for male-soiled bedding. Focal administration of the opioid antagonist ß-funaltrexamine into the posterior ventral tegmental area does not affect preference for male chemosignals. Nevertheless, exposure to male-soiled bedding elicits an increase in dopamine efflux in the nucleus accumbens shell and core, measured by microdialysis. Infusion of the opioid antagonist naltrexone in the accumbens core does not significantly affect dopamine efflux during exposure to male chemosignals, although it enhances dopamine levels 40 min after withdrawal of the stimuli. By contrast, infusion of the glutamate antagonist kynurenic acid in the accumbens shell inhibits the release of dopamine and reduces the time that females spend investigating male-soiled bedding. These data are in agreement with previous reports in male rats showing that exposure to opposite-sex odors elicits dopamine release in the accumbens, and with data in female mice showing that the behavioral preference for male chemosignals is not affected by opioidergic antagonists. We hypothesize that glutamatergic projections from the amygdala into the accumbens might be important to modulate the neurochemical and behavioral responses elicited by sexual chemosignals in rats.

Acetaldehyde sequestration by D-penicillamine prevents ethanol relapse-like drinking in rats: evidence from an operant self-administration paradigm.

RATIONALE: Previous experiments in our laboratory have shown that D-penicillamine (DP) (acetaldehyde sequestering agent) is able to block the increase in ethanol consumption observed after a period of imposed deprivation (the so-called alcohol deprivation effect (ADE)), using a non-operant paradigm in Wistar rats. OBJECTIVES: This study is aimed at investigating the robustness and reproducibility of our previous data using an operant paradigm, which is considered to be a valid and reliable model of human drug consumption, and the ADE, probably the most often used measure of ethanol relapse-drinking behaviour in rats. METHODS: Male Wistar rats with a limited (30-min sessions), intermittent and extended background of ethanol operant self-administration were used. In order to evaluate the efficacy of several DP doses (6.25, 12.5 and 25 mg/kg i.p.) in preventing alcohol relapse, we set up a protocol based on the ADE. In a separate experiment, the effect of DP on spontaneous motor activity of rats was also tested. RESULTS: A significant ADE was observed in animals treated with saline. DP treatment blocked the increase in ethanol responses following the imposed abstinence period. The higher dose suppressed the ADE and provoked a significant reduction in ethanol consumption with respect to the baseline conditions. Basal motor activity was not altered after DP treatment. CONCLUSION: Our positive results with DP, using two different paradigms that evaluate relapse of ethanol drinking, will help to increase the positive predictive value of pre-clinical experiments and offer a solid base to inspire human studies with DP.

Disposition of d-penicillamine, a promising drug for preventing alcohol-relapse. Influence of dose, chronic alcohol consumption and age: studies in rats.

Pharmacokinetic studies concerning d-penicillamine (an acetaldehyde sequestering agent) are scarce and have not evaluated the influence of chronic ethanol consumption and age on its disposition. Since recent preclinical studies propose d-penicillamine as a promising treatment for alcohol relapse, the main aim of the present work was to evaluate the influence of these two factors on d-penicillamine disposition in order to guide future clinical studies on the anti-relapse efficacy of this drug in alcoholism. Additionally, the effect of the administered dose was also evaluated. To this end, three studies were carried out. Study 1 assessed the influence of dose on d-penicillamine disposition, whereas studies 2 and 3 evaluated, respectively, the influence of chronic alcohol consumption and age. Rapid intravenous administrations of 2, 10 and 30 mg/kg of d-penicillamine were performed using young or adult ethanol-naïve rats or adult ethanol-experienced (subjected to a long-term ethanol self-administration protocol) rats. Pharmacokinetic parameters were derived from the biexponential model. Statistical analysis of CL, normalized AUC0 (∞) , V1 and k10 revealed that disposition, in the range plasma concentrations assayed, is non-linear both in young ethanol-naïve and in adult ethanol-experienced rats. Notably, no significant changes in t1/2 were detected. Chronic ethanol consumption significantly reduced CL values by 35% without affecting t1/2 . d-Penicillamine disposition was equivalent in young and adult animals. In conclusion, although DP pharmacokinetics is non-linear, the lack of significant alterations of the t1/2 would potentially simplify the clinical use of this drug. Chronic consumption of ethanol also alters d-penicillamine disposition but, again, does not modify t1/2.

Improved effect of the combination naltrexone/D-penicillamine in the prevention of alcohol relapse-like drinking in rats.

Opioid antagonists are licensed drugs for treating alcohol use disorders; nonetheless, clinical studies have evidenced their limited effectiveness. Preclinical findings indicate that opioid receptor (OR) antagonists, such as naltrexone (NTX), reduce the alcohol deprivation effect (ADE). However, a detailed analysis of published data shows the existence of a delayed increase in ethanol consumption after continuous OR blockade, a phenomenon originally called as 'delayed ADE'. We have recently reported that D-penicillamine (DP) is able to prevent ADE through a mechanism dependent on the inactivation of acetaldehyde, the main metabolite of ethanol. Hypothetically, OR activation would be triggered by acetaldehyde after ethanol consumption. Hence, we conjecture that the combination of NTX and DP, due to their distinct but complementary mechanisms to impede OR activation, may be more efficacious in the prevention of the ADE and, specifically, the 'delayed ADE'. Herein, we compare the effects of the combination NTX/DP (NTX: 2×5 mg/kg SC injection daily/DP: SC infusion (0.25 mg/h)) versus NTX on the ADE in long-term ethanol-experienced rats. As expected, NTX-treated animals displayed a delayed ADE. However, NTX/DP treatment prevented this delayed effect. Our present data indicate that this combination therapy shows an adequate anti-relapse preclinical efficacy being able to overcome the preclinical limitations of NTX alone.

Opposite motor responses elicited by ethanol in the posterior VTA: the role of acetaldehyde and the non-metabolized fraction of ethanol.

Recent electrophysiological evidence suggests that ethanol simultaneously exerts opposite effects on the activity of dopamine (DA) neurons in the ventral tegmental area (VTA) through two parallel mechanisms, one promoting and the other reducing the GABA release onto VTA DA neurons. Here we explore the possible behavioural implications of these findings by investigating the role displayed by acetaldehyde (the main metabolite of ethanol) and the non-metabolized fraction of ethanol in motor activity of rats. We analyse the appearance of motor activation or depression after intra-VTA administration of ethanol in rats subjected to different pharmacological pre-treatments designed to preferentially test either the effects of acetaldehyde or the non-metabolized ethanol. Motor activity was evaluated after intra-VTA administration of 35 nmol of ethanol, an apparently ineffective dose that does not modify the motor activity of animals. Pharmacological pre-treatments were used in order to either increase (cyanamide, 10 mg/kg, ip) or decrease (D-penicillamine, 50 mg/kg, ip and sodium azide, 7 mg/kg, ip) acetaldehyde levels in the VTA. Pre-treatments aimed to augment acetaldehyde, increased motor activity of rats. Otherwise, pre-treatments intended to decrease local acetaldehyde levels evoked significant reductions in motor activity that were prevented by the local blockade (bicuculline, 17.5 pmol) of the GABAA receptors. Our findings suggest that the brain-generated acetaldehyde is involved in the stimulant effects of ethanol, whereas the non-biotransformed fraction of ethanol, acting through the GABAA receptors, would account for the depressant effects. The present behavioural findings suggest that ethanol dually modulates the activity of DA neurons.

Efficacy of D-penicillamine, a sequestering acetaldehyde agent, in the prevention of alcohol relapse-like drinking in rats.

RATIONALE: Nowadays, very few approved anti-relapse treatments for alcoholism exist, and their overall efficacy can be considered moderate. An exciting rationale drug development opportunity for the treatment of chronic alcoholism is the use of acetaldehyde sequestering agents. Although these compounds are able to attenuate or prevent most of the behavioral and neurochemical effects of ethanol, the efficacy of acetaldehyde sequestration, by using agents such as D-penicillamine (DP), in relapse prevention has not been studied yet. OBJECTIVES: The aim of this study was to analyze the effects of DP treatment on the alcohol deprivation effect (ADE) in long-term ethanol-experienced rats as a model of relapse behavior and measure drug plasma and brain levels during treatment. METHODS: Rats were subcutaneously implanted with mini-osmotic pumps delivering 0, 0.25, or 1 mg/h of DP during 1 week. The efficacy to prevent ADE was determined. DP plasma and brain levels achieved during its subcutaneous administration were measured. In a second experiment, animals received bilateral infusions of 0 or 1.5 µg/h of DP directly into pVTA, and the appearance of ADE was evaluated. RESULTS: One milligram per hour, but not 0.25 mg/h, DP infusion prevented ADE and reduced the total ethanol preference in animals. DP plasma concentrations associated with ADE suppression were around 3-4 µg/ml, and brain DP levels in these conditions were about 2-3 % of those found in plasma. Intra-pVTA DP administration also suppressed ADE. CONCLUSION: DP is able to prevent alcohol-relapse-like drinking in rats suggesting that this drug may be a useful new tool in the management of relapse in alcohol-dependent patients.

Systemic administration of D-penicillamine prevents the locomotor activation after intra-VTA ethanol administration in rats.

Although recently published studies seem to confirm the important role displayed by acetaldehyde (ACH), the main metabolite of ethanol, in the behavioral effects of ethanol, the origin of ACH is still a matter of debate. While some authors confer more importance to the central (brain metabolism) origin of ACH, others indicate that the hepatic origin could be more relevant. In this study we have addressed this topic using an experimental approach that combines local microinjections of ethanol into the ventral tegmental area (VTA) (which guarantees the brain origin of the ACH) to induce motor activation in rats together with systemic administration (i.p.) of several doses (0, 12.5, 25 and 50 mg/kg) of D-penicillamine (DP), a sequestering agent of ACH with contrasted efficiency to abolish the behavioral effects of the drug. Our results clearly show that DP prevented in a dose-dependent manner the motor activation induced by intra-VTA ethanol, being the 50 mg/kg dose the most efficient. DP per se did not affect the basal activity of the rats. In order to determine the specificity of the DP action, we also studied the effects of DP 50 mg/kg on the DAMGO-induced motor activation after the intra-VTA administration of this mu-opioid receptors agonist. DP did not significantly modify the motor activation induced by DAMGO thus confirming the specificity of the DP effects. Our results clearly suggest that the brain-derived ACH is necessary to manifest the activating effects resulting from ethanol administration.

Locomotor stimulant effects of acute and repeated intrategmental injections of salsolinol in rats: role of mu-opioid receptors.

RATIONALE: Microinjections of ethanol and acetaldehyde into ventral tegmental area (VTA) produce locomotor activation in rats through mechanisms dependent on the mu-opioid receptors. However, it is not clear how these drugs can interact with these receptors. It has been hypothesized that salsolinol could be the responsible for this interaction. OBJECTIVES: The aim of the study was to investigate the ability of salsolinol to induce both motor activation and motor sensitization in rats after repeated intra-VTA administration. MATERIALS: Rats received one microinjection into the posterior VTA of artificial cerebrospinal fluid (aCSF; 200 nL), salsolinol (0.3-3,000.0 pmol/200 nL), or salsolinol (30.0 pmol/200 nL) with either naltrexone (13.2 nmol/200 nL) or with the antagonist of the mu-opioid receptors, beta-funaltrexamine (beta-FNA; 2.5 nmol/300 nL). In the sensitization experiments, four microinjections of salsolinol (30.0 pmol/200 nL) or aCSF (200 nL) were performed over a 2-week period. This period was followed by a single challenge session, in which 0.3 pmol of salsolinol was microinjected to rats. Spontaneous activity was always monitored postinjection. RESULTS: Intra-VTA salsolinol administration induces an increase of the spontaneous motor activity of the rats with the maximal effect at the dose of 30.0 pmol/200 nL. Salsolinol effects were blocked by the treatment with naltrexone or beta-FNA. Moreover, repeated injections of salsolinol produced locomotor sensitization. CONCLUSIONS: Salsolinol induces locomotor activity and motor sensitization after intra-VTA administration. Moreover, the implication of the mu-opioid receptors was shown since the treatment with naltrexone or beta-FNA was able to suppress the salsolinol effects.

Motor stimulant effects of ethanol and acetaldehyde injected into the posterior ventral tegmental area of rats: role of opioid receptors.

RATIONALE: A recently published study has shown that microinjections of ethanol, or its metabolite, acetaldehyde into the substantia nigra pars reticulata, are able to produce behavioral activation in rats. Another brain site that could participate in such effects is the ventral tegmental area (VTA). OBJECTIVES: We have investigated the locomotor-activating effects of local microinjections of ethanol and acetaldehyde into the posterior VTA of rats and the role of opioid receptors in such effects. MATERIALS: Cannulae were placed into the posterior VTA to perform microinjections of ethanol (75 or 150 nmol) or acetaldehyde (25 or 250 nmol) in animals not previously microinjected or microinjected with either the nonselective opioid antagonist naltrexone (13.2 nmol) or the irreversible antagonist of the micro-opioid receptors beta-funaltrexamine (beta-FNA; 2.5 nmol). After injections, spontaneous activity was monitored for 60 min. RESULTS: Injections of ethanol or acetaldehyde into the VTA increased the locomotor activity of rats with maximal effects at doses of 150 nmol for ethanol and 250 nmol for acetaldehyde. These locomotor-activating effects were reduced by previously administering naltrexone (13.2 nmol) or beta-FNA (2.5 nmol) into the VTA. CONCLUSIONS: The posterior VTA is another brain region involved in the locomotor activation after the intracerebroventricular administration of ethanol or acetaldehyde. Our data indicate that opioid receptors, particularly the micro-opioid receptors, could be the target of the actions of these compounds in the VTA. These results are consistent with the hypothesis that acetaldehyde could be a mediator of some ethanol effects.

Intestinal absorption enhancement via the paracellular route by fatty acids, chitosans and others: a target for drug delivery.

Peroral delivery of hydrophilic drugs is one of the greatest challenges in biopharmaceutical research. Hydrophilic drugs usually present low bioavailability after oral administration. One of the causes of this low bioavailability is their poor intestinal permeation through the paracellular pathway. This pathway is actually restricted by the presence of tight junctions at the apical side of the enterocytes. In the last few years, great interest has been focused on the structure and cellular regulation of tight junctions, materializing in more in-depth knowledge of this intestinal barrier. Simultaneously, and on the basis of this understanding, continuous efforts are being made to develop agents that can modulate tight junctions and magnify the paracellular permeability of hydrophilic compounds without causing significant intestinal damage. This review focuses on strategies to improve the paracellular permeation of poorly absorbed drugs as a way to enhance their bioavailability after oral administration. Most of the research on this subject has been carried out using in vitro models (mainly Caco-2 cell monolayers), which yield useful information on the potential effects and mechanisms of action of absorption-enhancing compounds. However, in vivo studies, which are much more scarce, are needed to confirm the effects of potential enhancers and to evaluate the suitability of including these compounds as excipients in drug formulation. We review the in vitro and in situ studies involving the most promising paracellular permeation enhancers (e.g., medium chain fatty acids and chitosan and its derivatives), analyzing the degree of drug absorption enhancement achieved, as well as the potential associated toxicity. The few studies performed in vivo are also presented. In addition, the findings of recent absorption enhancers, such as zonula occludens toxin or thiolated polymers, are reviewed.

Hippocampal dopamine receptors modulate cFos expression in the rat nucleus accumbens evoked by chemical stimulation of the ventral hippocampus.

Recently, we have shown that D1 and D2 receptors in the ventral hippocampus (VH) modulate both the locomotor activation and the increase in dopamine (DA) levels in the rat nucleus accumbens (NAc) induced by NMDA stimulation of the VH. In the present study we analyze the possible role of VH D1 and D2 receptors in the modulation of the cFos expression in NAc (core and shell subregions) and in dorsal striatum. This was assessed by immunohistochemical analysis of cFos expression in the rat brains after retro-dialysis application of NMDA (50mM, 10 min) into VH, in absence and in presence of either the D1/D5 receptor antagonist SCH 23390 (100 and 250 microM, 60 min) or the D2 receptor antagonist raclopride (100 and 250 microM, 60 min). NMDA induced a robust increase in the cFos expression in the NAc shell, both in the ipsilateral and contralateral side. No statistically significant increases were observed in the NAc core and in the dorsal striatum. Simultaneous application of SCH 23390 and NMDA into the VH attenuated the NMDA-evoked cFos expression in NAc shell. In contrast, raclopride had no significant effect. Our present results show that the NMDA receptor mediated effects in the VH require D1 receptors and suggest that DA in VH strongly modulates the excitatory outputs from this brain area.

Hippocampal dopamine receptors modulate the motor activation and the increase in dopamine levels in the rat nucleus accumbens evoked by chemical stimulation of the ventral hippocampus.

A number of studies have shown that chemical stimulation (using N-methyl-D-aspartate (NMDA) infusions) or electrical stimulation of the ventral hippocampus (VH) elicits locomotor activation and sustained increases in nucleus accumbens (NAc) dopamine (DA) levels in rodents. How DA neurotransmission in NAc is involved in these effects has also been well established. However, the modulatory role of the DA receptors located in VH is not yet fully understood. The purpose of this study was to characterize the role played by VH D1 and D2 subtype receptors in both the locomotor activation and NAc DA increases induced by NMDA stimulation of the VH. This was assessed by studying how retrodialysis application of NMDA (50 mM, 10 min) affects motor activity and NAc DA levels during simultaneous retrodialysis administration of the D1/D5 receptor antagonist SCH 23390 (100 and 250 microM, 60 min) or the D2 receptor antagonist raclopride (100 and 250 microM, 60 min). SCH 23390 attenuated or completely abolished NMDA-evoked locomotor activation and the concurrent increase in NAc DA levels. On the other hand, raclopride was initially able to attenuate the effects of VH NMDA stimulation. However, in the last phase of the experiments, animals showed an important increase in clonic seizure activity with a simultaneous and dramatic increase in NAc DA levels. Our results show that the NMDA receptor-mediated effects in the VH require both D1 and, probably, D2 receptors and suggest that DA in VH strongly modulates the excitatory outputs from this brain area.

Assessment and modulation of acamprosate intestinal absorption: comparative studies using in situ, in vitro (CACO-2 cell monolayers) and in vivo models.

The purpose of this study was to explore the intestinal absorption mechanism of acamprosate and to attempt to improve the bioavailability (BA) of the drug through modulation of its intestinal absorption using two enhancers (polysorbate 80 and sodium caprate) based on in situ, in vitro and in vivo models and comparing the results obtained. Intestinal transport of the drug, in the absence and in presence of polysorbate 80 (0.06, 0.28 and 9.6 mM) or sodium caprate (13 and 16 mM) was measured by using an in situ rat gut technique and Caco-2 cell monolayers. Additionally, the effect of sodium caprate on drug oral bioavailability, measured as urinary recovery, was quantified by performing in vivo experiments with the rat as animal model. Only sodium caprate was able to increase the absorption rate constant (ka) of acamprosate in the mid-intestine of the rats from 0.29 +/- 0.07 h-1 in the absence of the promoter to 0.51 +/- 0.19 h-1 in the presence of C10 16 mM, along with the apparent permeability (Papp) obtained in Caco-2 cells (around two-fold). However, the drug bioavailability in rats (around 20%) did not improve in the presence of any of the concentrations tested (13, 16 and 50 mM). It is concluded that acamprosate absorption likely occurs via paracellular pathway and can be enhanced by sodium caprate in situ and in vitro but not in vivo-thus suggesting that although in situ and in vitro studies could be useful in early screening to select a potential promoter, in vivo studies in animal models are necessary to confirm the utility of the enhancer and to determine the influence of physiological variables.