Sex Differences in X-ray-Induced Endothelial Damage: Effect of Taurine and N-Acetylcysteine.

Ionizing radiation (IR) can induce some associated pathological conditions due to numerous cell damages. The influence of sex is scarcely known, and even less known is whether the effect of antioxidants is sex-dependent. Given the increased use of IR, we investigated whether male human umbilical vein endothelial cells (MHUVECs) and female human umbilical vein endothelial cells (FHUVECs) respond differently to IR exposure and whether the antioxidants 10 mM taurine (TAU) and 5 mM N-acetylcysteine (NAC) can prevent IR-induced damage in a sex-dependent way. In untreated cells, sex differences were observed only during autophagy, which was higher in FHUVECs. In non-irradiated cells, preincubation with TAU and NAC did not modify viability, lactate dehydrogenase (LDH) release, migration, or autophagy, whereas only NAC increased malondialdehyde (MDA) levels in FHUVECs. X-ray irradiation increased LDH release and reduced viability and migration in a sex-independent manner. TAU and NAC did not affect viability while reduced LDH release in irradiated cells: they have the same protective effect in FHUVECs, while, TAU was more protective than NAC in male cells.. Moreover, TAU and NAC significantly promoted the closure of wounds in both sexes in irradiated cells, but NAC was more effective at doing this in FHUVECs. In irradiated cells, TAU did not change autophagy, while NAC attenuated the differences between the sexes. Finally, NAC significantly decreased MDA in MHUVECs and increased MDA in FHUVECs. In conclusion, FHUVECs appear to be more susceptible to IR damage, and the effects of the two antioxidants present some sex differences, suggesting the need to study the influence of sex in radiation mitigators.

Further characterization of the effect of the prototypical antidepressant imipramine on the microstructure of licking for sucrose.

We previously reported that treatment with the prototypical antidepressant imipramine induced a dose-dependent reduction of the ingestion of a 10% sucrose solution, due to reduction of the licking burst number, thus suggesting reduced motivation and/or increased satiation. Importantly, the experimental sessions were performed in an alternate order, either 1-h or 24-h after imipramine administration. The observation that imipramine effect was more pronounced in the "1-h after-treatment" sessions, i.e. at the time of the brain drug Cmax, led us to suggest that it was likely related to brain drug levels at testing time. However, such an experimental design does not allow to rule out the alternative possibility that the observed effect might be due to post-session administration, as previously observed with memantine. To determine whether imipramine-induced decrease of sucrose ingestion could be observed even in absence of post-session administration, we examined the effect of a daily 22 day treatment with imipramine (5, 10 and 20 mg/kg). In the first half of the treatment period all behavioural tests were performed 1-h after administration. In the second half of the treatment period, tests were performed alternatively either 1-h or 24-h after imipramine administration. The results confirm that imipramine reduces sucrose ingestion due to a reduction of the licking burst number. Most importantly, these results demonstrate that this effect does not require imipramine post-session administration, since it was present before the beginning of post-session administrations. This supports the interpretation of the reduction of sucrose ingestion as a consequence of reduced motivation and/or increased satiation. Thus, these findings, taken together with the results of our previous study, might be relevant in explaining the effects of imipramine in models of drug-seeking and in body weight gain reduction in rats, but not in accounting for the antidepressant therapeutic effect. At variance with the results of our previous study, an increase in burst size was present in the first half of the treatment period, which might be interpreted as a prohedonic effect and/or as a compensatory effect.

Memantine effects on ingestion microstructure and the effect of administration time: A within-subject study.

In a between-subject comparison of two memantine administration schedules we observed that treatment with the NMDA receptor antagonist memantine before testing sessions reduced ingestion of a 10% sucrose solution in rats, due to reduced licking burst size, thus suggesting a blunted hedonic response. Conversely, daily post-session administration reduced burst number, indicating a reduced level of behavioural activation, likely due to the development of conditioned taste aversion (CTA). In this study, the effect of pre-session and post-session memantine administration was investigated within-subjects. Memantine was administered in daily intraperitoneal injections for 13 days, on alternate days, either 1-h before-"before testing" sessions-or immediately after a 30-min session-"after testing" sessions. The effects on the microstructure of licking for a 10% sucrose solution were examined in the course of treatment and for 21 days after treatment discontinuation. The results show reduced burst size in the "before testing" sessions, without effects on the intra-burst lick rate, an index of motoric effects. Moreover, burst number was reduced since the third session of both administration conditions until the end of treatment. Interestingly, the effect of memantine of reducing the activation of ingestive behaviour was less pronounced in this study with respect to that observed with the previous study post-session administration schedule, in spite of the longer treatment. This apparent paradox might be explained if one considers these effects as instances of a memory-related effect, such as the development of CTA. In the framework of this hypothesis, the "before testing" sessions, not being followed by memantine administration, can be considered as extinction sessions performed every other day. Moreover, the animals treated with memantine at the highest dose failed to recover to pre-treatment ingestion levels 21 days after treatment discontinuation, while the animals treated after testing sessions in the previously published study showed a complete recovery well before the 15th day test. Within the same interpretative framework, this might depend by the reduced number and frequency of the extinction trials-i.e. the number of the sessions run after treatment discontinuation-in the present study. These results provide further support to the conclusion that memantine administration before sessions reduce burst size, an effect which is likely due to blockade of NMDA receptors occurring during behavioural testing. The observation that this effect can be obtained even in absence of a reduced intra-burst lick rate, which rules out the involvement of motor impairment, provides an important piece of evidence in support to the interpretation of this effect as a blunted hedonic response. Moreover, these results provide further evidence that burst number reduction is due to a memory-related effect induced by memantine administration after sessions.

Microstructure analysis of sucrose ingestion in the course of chronic treatment with imipramine.

The analysis of licking microstructure provides measures which might be interpreted in terms of psychological constructs, such as pleasure and motivation, relevant for the interpretation of the effects of antidepressant drugs. The aim of this study was to characterise the effect of the prototypical antidepressant imipramine on the microstructure of licking for a 10% sucrose solution. In particular, ten 30-min sessions were performed in the course of a daily 21 day treatment with imipramine - 5, 10 and 20 mg/kg/die administered intraperitoneally. To interpret drug effects in relation to the presumed concentration of imipramine and its active metabolite desipramine, the experimental sessions were performed in an alternate order either 1-h or 24-h after imipramine administration. In the sessions performed 1-h after drug administration, the results showed a dose-dependent reduction of sucrose ingestion, accounted for by a reduction of the licking burst number. Moreover, reduced intra-burst lick rate and increased latency to lick were observed with the highest doses. Imipramine effect in the sessions performed 24-h after drug administration was similar but less pronounced. These results are consistent with the hypothesis that the reduction of sucrose ingestion might be due to reduced motivation and/or to a potentiation of satiety signals. These effects appear to be related, at least in part, to brain drug levels at testing time, and do not seem related to the mechanisms underlying the antidepressant therapeutic effect. However, these results might be relevant in explaining the effects of imipramine in models of drug-seeking and on body weight.

Daily memantine treatment blunts hedonic response to sucrose in rats.

RATIONALE: Preclinical and clinical studies suggest the potential use of memantine in the treatment of binge eating disorder. The aim of this study was to further investigate the mechanisms by which memantine influences the motivational aspects of ingestion through the analysis of licking microstructure. To interpret treatment effects in relation to drug action at specific functionally relevant times, we compared the effect of two different administration schedules. METHODS: Memantine was administered daily for a week, either 1 h before or immediately after a 30-min daily session. The effects on the microstructure of licking for a 10% sucrose solution in rats were examined in the course of treatment and for 15 days after treatment discontinuation. RESULTS: Treatment before testing reduced ingestion due to reduced burst size and increased latency in the first session. However, a progressive increase in burst number across sessions led to a full recovery of ingestion levels by the end of treatment. Daily post-session administration induced a dramatic decrease of activation of licking behaviour, indicated by reduced burst number, accompanied to reduced burst size. A slow recovery of ingestion took place after treatment discontinuation. CONCLUSION: These results suggest a reduced hedonic/reward evaluation response, an effect likely due to NMDA receptor blockade occurring during the testing time and support the hypothesis that memantine interferes with the hedonic/non-homeostatic mechanisms regulating food intake and food-seeking. The effect of post-session administration might be explained by the development of conditioned taste aversion.

Role of dopamine D1-like and D2-like receptors in the activation of ingestive behaviour in thirsty rats licking for water.

RATIONALE: Analysis of lick pattern for sucrose and NaCl and of the forced swimming response after dopamine antagonist administration led us to suggest that dopamine on D1-like receptors is involved in behavioural activation, and the level of activation is "reboosted" on the basis of an evaluation process involving D2-like receptors. Although some studies investigated licking microstructure for water after dopamine antagonists, the within-session time course of their effect was never investigated. OBJECTIVES: The aims of this study were to further investigate the role of dopamine receptors in the mechanisms governing water ingestion, focussing on the within-session time course of the microstructure parameters, and to test the proposed hypothesis. MATERIALS AND METHODS: The effects of the dopamine D1-like receptor antagonist SCH 23390 (0.01-0.04 mg/kg) and of the dopamine D2-like receptor antagonist raclopride (0.025-0.25 mg/kg) on licking microstructure for water were examined in 20-h water-deprived rats in 30-min sessions. RESULTS: As previously observed with sucrose and NaCl, SCH 23390 reduced licking by reducing burst number, suggesting reduced behavioural activation. Moreover, it resulted in an increased burst size. Raclopride reduced the size of licking bursts, while their number was either increased or decreased depending on the dose. CONCLUSION: The results support the suggestion that D1 receptors are involved in behavioural activation and D2 receptors are involved in a related evaluation process. Within the framework of the proposed hypothesis, the increased burst size after D1-like receptor blockade might be interpreted as a pro-hedonic effect consequent to the increased cost of the activation of the licking response.

Imipramine administered before the first of two forced swim sessions results in reduced immobility in the second session 24 h later.

A previous study investigating the effects of dopamine receptor antagonists administered before the first of two 24-h apart forced swim test (FST) sessions, provided evidence suggesting that evaluation of response efficacy - dependent on dopamine D2-like receptors - might play a role in setting the balance between active behaviours and immobility in this test. Regardless of the underlying mechanisms, the observation that the effects of drugs in the first session have consequences in the second session might be relevant for a better understanding of the FST in behavioural/functional terms. Thus, the first objective of this study was to investigate the consequences in the second session of the administration of the prototypic antidepressant drug imipramine before the first of two sessions. A second objective was to investigate the effect of dopamine D1-like and D2-like receptor blockade on the effects of imipramine. Imipramine (20 mg/kg) was administered 24-h, 6-h and 30-min before the first of two FST sessions performed 24-h apart. SCH 23390 (0.01, 0.04 mg/kg) or raclopride (0.0125, 0.25 mg/kg) were administered 30-min before the first session. Imipramine increased active behaviours both in the first and in the second session. Raclopride attenuated and SCH 23390 potentiated imipramine effects only in the first session and to a limited extent. These results show that imipramine administration before the first of two FST sessions induces an increase in active behaviours in the second session, and suggest that this effect is the consequence of the behavioural effects of imipramine in the first session.

Within-session decrement of the emission of licking bursts following reward devaluation in rats licking for sucrose.

We previously observed that dopamine D2-like receptor blockade in rats licking for sucrose produced a within-session decrement of the emission of licking bursts similar to the effect of either reward devaluation, or neuroleptics, on operant responding for different rewards, which, accordingly, we interpreted as an extinction-like effect. This implies that exposing animals to reward devaluation would result in a drop of burst number taking place only after the contact with the devalued reward. To test this prediction, we compared the difference in the within-session time course of burst number in response to high (10%) versus low (2%) concentration sucrose solutions, either in a condition of reward devaluation (exposure to 2% after daily 10%), or in a condition which does not involve changes in the reward value (two groups of subjects each repeatedly exposed to only one of the two concentrations). Reward devaluation resulted in a within-session decrement of the burst number, with the response rate dropping only after the contact with the devalued reward, as predicted. This response pattern was reliably observed only in subjects at their first devaluation experience. In contrast, exposure of separate groups of animals to the two different concentrations yielded lower levels of burst number in the low concentration group apparent since the beginning of the session, as previously observed with dopamine D1-like receptor blockade. These results show that the analysis of burst number, but not of burst size, reveals a specific activation pattern in response to reward devaluation, which differs from the pattern observed comparing the response to two different sucrose concentrations in separate groups of subjects, i.e. in a condition not involving reward devaluation. Finally, the characterisation of the experimental measures of the analysis of licking microstructure in behaviourally (and psychologically) meaningful functional terms, might be relevant for the investigation of the mechanisms underlying behavioural activation and the related evaluation processes.

Dopamine on D2-like receptors "reboosts" dopamine D1-like receptor-mediated behavioural activation in rats licking for a isotonic NaCl solution.

RATIONALE: We recently suggested that dopamine on D1-like receptors is involved in the activation of goal-directed responses and the level of response activation is "reboosted" on the basis of an evaluation process involving D2-like receptors assessing "response efficacy". A main piece of evidence in support of this hypothesis was the observation of an "extinction mimicry" effect in the time course of licking bursts after dopamine D2-like receptor blockade in rats licking for sucrose. OBJECTIVES: The aim of this study was to determine whether the pattern of licking observed with sucrose as a reward could be reproduced in rats licking for a different reward (0.9% NaCl). MATERIALS AND METHODS: We investigated the effects of the dopamine D1-like receptor antagonist SCH 23390 (0.01-0.04 mg/kg) and of the dopamine D2-like receptor antagonist raclopride (0.025-0.25 mg/kg) on the microstructure of licking for a 0.9% NaCl solution in 12-h water-deprived rats in 30-min sessions. RESULTS: As previously observed with sucrose as a reward, raclopride reduced the size of licking bursts and produced on the burst number time course an "extinction mimicry" effect, while SCH 23390 reduced licking exclusively by reducing burst number. CONCLUSIONS: These results are consistent with the proposed hypothesis and provide support to the use of the study of licking microstructure as a valid model not only for the investigation of the mechanisms governing ingestive behaviour but also for the investigation of the mechanisms underlying behavioural activation and the related evaluation processes.

Glutamyl cycle in the rat liver appears to be sex-gender specific.

Numerous studies show sexually dimorphic responses of drug metabolizing enzymes in the liver, but it is less clear whether xenobiotic detoxification mediated by glutathione is sex-gender specific. Therefore, we investigated whether sex-gender differences exist in the biosynthesis and metabolism of GSH in the rat liver. Livers were obtained from Sprague-Dawley rats of both sexes for measurement of glutathione, its precursors and metabolites by capillary electrophoresis, whereas H(2)S and malondialdehyde were measured by colorimetric assays. The expression of glutamylcysteine ligase (GCL), the key enzyme in glutathione synthesis was detected by Western blotting and immunohistochemistry. It was observed that L-methionine, glutathione, taurine and malondialdehyde (a marker of lipid peroxidation) were similar in livers from both sexes, while L-cysteine levels were significantly higher and H(2)S was lower in female rat livers. Furthermore, L-methionine and L-cysteine, L-cysteine and glutathione, L-cysteine and taurine were positively associated only in male livers. Finally, the female liver expressed less GCL than the male liver. These data suggest that the glutamyl cycle in the liver is sexually dimorphic. This difference could be linked to the increased sensitivity of females to drugs and xenobiotics.

Effect of the dopamine D1-like receptor antagonist SCH 23390 on the microstructure of ingestive behaviour in water-deprived rats licking for water and NaCl solutions.

The analysis of licking microstructure provides measures, size and number of licking bouts, which might reveal, respectively, reward evaluation and behavioural activation. Based on the different effects of the dopamine D1-like and D2-like receptor antagonists SCH 23390 and raclopride on licking for sucrose, in particular the failure of the former to reduce bout size and the ability of the latter to induce a within-session decrement of bout number resembling either reward devaluation or neuroleptics on instrumental responding, we suggested that activation of reward-associated responses depends on dopamine D1-like receptor stimulation, and its level is updated on the basis of a dopamine D2-like receptor-mediated reward evaluation. Consistent results were obtained in a study examining the effect of dopamine D2-like receptor antagonism in rats licking for NaCl solutions and water. In this study, we examined the effects of the dopamine D1-like receptor antagonist SCH 23390 (0, 10, 20 and 40 µg/kg) on the microstructure of licking for water and sodium chloride solutions (0.075 M, 0.15 M, 0.3 M) in 12 h water deprived rats. Rats were exposed to each solution for 60 s either after the first lick or after 3 min that the animals were placed in the chambers. Bout size, but not bout number, was decreased at the highest NaCl concentration. SCH 23390 produced a decrease of bout number and of lick number mainly due to the decreased number of subjects engaging in licking behaviour, and failed to reduce bout size for Na Cl and water at a dose which increased the latency to the 1st lick but did not affect the intra-bout lick rate. In agreement with previous observations, these results suggest that dopamine D1-like receptors play an important role in the activation of reward-oriented responses.

Possible role of dopamine D1-like and D2-like receptors in behavioural activation and "contingent" reward evaluation in sodium-replete and sodium-depleted rats licking for NaCl solutions.

Based on the different effects of the dopamine D1-like and D2-like receptor antagonists SCH 23390 and raclopride on the measures of licking microstructure in rats, we suggested that the level of activation of reward-associated responses depends on dopamine D1-like receptor stimulation, and is updated, or "reboosted", on the basis of a dopamine D2-like receptor-mediated reward evaluation. To further test this hypothesis, we examined the effects of the dopamine D2-like receptor antagonist raclopride (0, 25, 125, 250µg/kg) and of the dopamine D1-like receptor antagonist SCH 23390 (0, 10, 20 and 40µg/kg) on the microstructure of licking for two different NaCl solutions (0.9% and 2.7%) in rats in sodium-replete status and in the sodium-depleted status induced by the diuretic drug furosemide. Rats were exposed to each solution for 180 seconds after the first lick. Both in sodium-replete and in sodium-depleted status, SCH 23390 produced a decrease of burst number, a measure of behavioural activation, without affecting their size, a measure of reward evaluation. Raclopride reduced burst number but appeared also to exert some effects on burst size. Sodium depletion resulted in an increased intake for both NaCl solutions due to an increase in burst number and size, and in a reduced sensitivity to the effect of raclopride on lick number. These results are not in contrast with the proposed hypothesis and are consistent with previous evidence suggesting a role for dopamine D2-like receptors in the increased NaCl appetite induced by sodium depletion.

Possible role of dopamine D1-like and D2-like receptors in behavioural activation and evaluation of response efficacy in the forced swimming test.

Based on the different effects of the dopamine D1-like and D2-like receptor antagonists SCH 23390 and raclopride on the measures of licking microstructure in rats ingesting a sucrose solution, we suggested that the behavioural activation of reward-associated responses depends on dopamine D1-like receptor stimulation, and its level is updated, or "reboosted", on the basis of a dopamine D2-like receptor-mediated evaluation process. The aim of this study was to test this hypothesis on the forced swimming test response. The effects of the dopamine D1-like and D2-like receptor antagonists SCH 23390 (0.01-0.04 mg/kg) and raclopride (0.025-0.25 mg/kg) administered before a 15-min exposure to forced swimming, and the response to a second session performed 24 h later, were examined. SCH 23390 dose-dependently reduced climbing scores in the first session and increased them in the second session, but the within-session decline of this measure was similar to that observed in the control group in both sessions. Raclopride-treated subjects showed a slightly reduced level of climbing scores at the beginning of the first session, but persisted in emitting this costly behavioural response up to the end of the session, while no effects were observed in the second session. These results, along with our results examining licking for sucrose, are consistent with the hypothesis that behavioural activation and response effort allocation are directly mediated by dopamine D1-like receptor stimulation, but the level of this activation is updated, or "reboosted", on the basis of a dopamine D2-like receptor-mediated mechanism of response efficacy evaluation.

Clozapine increases reward evaluation but not overall ingestive behaviour in rats licking for sucrose.

RATIONALE: Clozapine and the "atypical" antipsychotics are less prone than neuroleptics to induce extrapyramidal motor effects, worsening of the negative symptoms of schizophrenia and dysphoria. This is paralleled by preclinical evidence showing reduced suppression of behaviours aimed at the pursuit of reward, with increased measures of reward efficacy. Serotonin 5-HT2 receptors seem to play a role in determining this profile. OBJECTIVE: We investigated the effects of clozapine on the microstructure of ingestive behaviour, which might reveal behavioural dimensions, such as reward evaluation and behavioural activation, which might be relevant in explaining its atypical profile. Moreover, we investigated the possibility that coadministration of the typical antipsychotic haloperidol and the 5-HT2A/2C receptor antagonist ritanserin might mimic clozapine effects. MATERIALS AND METHODS: The effects of clozapine (0.5, 1 and 5 mg/kg) and of the coadministration of haloperidol (0.05 mg/kg) and ritanserin (0.5 and 3 mg/kg) have been examined on the microstructure of licking for a 10% sucrose solution in rats. RESULTS: Clozapine failed to affect whole ingestion as revealed by the lack of effect on lick number. However, it increased reward evaluation at the dose of 1 mg/kg, as revealed by increased mean bout size. Haloperidol resulted in a decreased bout size. Ritanserin failed to exert any effects either alone or when coadministered with haloperidol. CONCLUSION: The ability of clozapine to increase reward evaluation might contribute to explain its atypical profile both in the clinical setting and in preclinical studies. These results suggest that 5-HT2A/2C receptors are not involved in the observed effect.