Changes in behavioral and neuronal parameters by alcohol, cigarette, or their combined use in rats.

Few studies have explored the effects of the combined use of alcohol and cigarette in humans, despite its prevalence. Here we evaluated the effect of isolated and combined use on behaviors and neuronal parameters in rats. Male adult rats were divided into alcohol (AL, 2 g/kg, by oral gavage), cigarette smoke (TB, six cigarettes, by inhalation), combined use (ALTB), or control (CT, water by oral gavage and environmental air) groups, treated twice a day (09.00 and 14.00 h). After 4 weeks, the rats were tested in the open field for behavioral analysis and euthanized for brain volume estimation and counting of neurons in the hippocampus. All treatments increased locomotion, and this behavior was higher in the ALTB than TB group. Latency to exit from the central area was lower in the ALTB than in the AL or CT groups. Rearing behavior increased in TB and decreased in AL and ALTB rats. Combined ALTB rats significantly increased their grooming behavior. Only the AL group showed decreased neuron counts and increased brain volume. Our results show that the isolated and combined uses of alcohol and cigarette smoke have diverse effects on behavioral and neuronal parameters in rats after long-term treatment.

N-acetylcysteine Prevents Alcohol Related Neuroinflammation in Rats.

Alcoholism has been characterized as a systemic pro-inflammatory condition and alcohol withdrawal has been linked to various changes in the brain homeostasis, including oxidative stress and glutamate hyperactivity. N-acetylcysteine (NAC) is an anti-inflammatory and antioxidant multi-target drug with promising results in psychiatry, including drug addiction. We assessed the effects of NAC on the serum and brain inflammatory cytokines after cessation of chronic alcohol treatment in rats. Male Wistar rats received 2 g/kg alcohol or vehicle twice a day by oral gavage for 30 days. Rats were treated, from day 31 to 34, with NAC (60 or 90 mg/kg) or saline, intraperitoneally, once daily. Rats were sacrificed at day 35, trunk blood was collected and the frontal cortex and hippocampus dissected for assessment of TNF-α, IL-1ß, IL-6, IL-18, IL-10. NAC prevented the increase of pro-inflammatory cytokines and the decrease of anti-inflammatory cytokine in the frontal cortex and hippocampus. No changes were observed on serum cytokines. We conclude that NAC protects against inflammation induced by chronic (30 days) alcohol ingestion followed by 5 days cessation in two rat brain areas. Because inflammation has been documented and associated with craving and relapse in alcoholics, the data revealed by this study points to the validity of NAC clinical evaluation in the context of alcohol detoxification and withdrawal.

Effect of Alcohol and Tobacco Smoke on Long-Term Memory and Cell Proliferation in the Hippocampus of Rats.

INTRODUCTION: Alcohol is frequently used in combination with tobacco and few studies explore interactions between these two drugs of abuse. Here, we evaluated the effect of chronic alcohol administration and concomitant exposure to tobacco smoke on long-term memory and on cell proliferation in the hippocampus of rats. METHODS: Forty male Wistar rats were assigned to four groups and treated with alcohol (2g/kg by gavage) and/or exposed to tobacco smoke (from six cigarettes, by inhalation) twice a day (at 9:00 AM and 2:00 PM) for 30 days. Long-term memory was evaluated in the inhibitory avoidance test and hippocampal cell proliferation was analyzed for bromodeoxyuridine immunohistochemistry. RESULTS: Our results showed that alcohol, tobacco smoke, or their combination improved the long-term memory evaluated by the memory index in rats. Moreover, alcohol and tobacco coadministration decreased bromodeoxyuridine-labeled cells by 60% when compared to the control group, while alcohol treatment decreased labeled cells by 40%. The tobacco group showed a nonsignificant 26% decrease in labeled cells compared to the control group. CONCLUSIONS: Chronic alcohol and tobacco coadministration improves the long-term memory in rats in the inhibitory avoidance test. However, coadministration decreases the cell proliferation in the hippocampus of rats, suggesting a deleterious effect by the combined use of these drugs of abuse.

Low-cost apparatus for cigarette smoke exposure in rats.

BACKGROUND: The effects of tobacco smoke on the central nervous system are usually studied with isolated nicotine, ignoring other compounds present in cigarette smoke. The few studies that use in vivo whole-body cigarette smoke exposure are usually performed in expensive commercial apparatus. NEW METHOD: We presented a feasible, safe, and low-cost apparatus for cigarette smoke exposure in rodents. RESULTS: Rats exposed to cigarette smoke in this apparatus showed cotinine levels similar to human active smokers. Additional results showed that cigarette smoke exposure increased glutamate and aspartic acid levels and decreased leucine, isoleucine, ornithine, phenylalanine, and tryptophan levels in the cerebrospinal fluid of rats. COMPARISON WITH EXISTING METHOD(S): Our apparatus is feasible, safe, and costs 67-fold less than a commercial automatized smoking machine. Beyond the low cost, it does not require specialized knowledge for building or maintenance. CONCLUSIONS: We concluded that our low-cost apparatus is reliable and reproduces cigarette smoke use in humans.

Hepatic and renal damage by alcohol and cigarette smoking in rats.

Chronic use of alcohol and tobacco cigarettes is associated to millions of deaths per year, either by direct or indirect causes. However, few studies have explored the additional risks of the combined use of these drugs. Here we assessed the effect of the combined use of alcohol and cigarette smoke on liver or kidney morphology, and on biochemical parameters in chronically treated rats. Male Wistar rats were allocated to receive 2 g/kg alcohol orally, which was followed by the inhalation of smoke from six cigarettes during 2 h (ALTB group) for 28 days. Other groups received alcohol alone (AL) or were exposed to cigarette smoke (TB) alone and were compared to control (CT) rats, which received water followed by ambient air. On day 29, rats were euthanized and blood samples were collected for aminotransferase enzymes (AST and ALT), creatinine, and urea analysis. Liver and kidney were weighted, dissected, fixed, and stained with hematoxylin and eosin for morphological analysis. Our results showed that necrosis was elevated in the AL, TB, and mainly the ALTB group in both liver and kidney of rats. Serum levels of AST and ALT were reduced by cigarette smoke exposure, independently of alcohol use. Serum creatinine levels increased after tobacco smoke exposure. On the other hand, TB and AL groups decreased serum urea levels, and their association restored that decrease. Absolute liver and kidney weights were lower in the cigarette smoke exposure rats. Lastly, body weight gain was lower in TB group and combined use restored it. Thus, we may infer that the use of alcohol, exposure to tobacco cigarette smoke or, mainly, their association promotes liver and kidney injuries, and this damage is related with biochemical changes in rats.

Combined use of alcohol and cigarette increases locomotion and glutamate levels in the cerebrospinal fluid without changes on GABAA or NMDA receptor subunit mRNA expression in the hippocampus of rats.

Interactions on neurotransmitter systems in the reward pathways may explain the high frequency of combined use of alcohol and cigarettes in humans. In this study, we evaluated some behavioral and neurochemical changes promoted by chronic exposure to alcohol and cigarette smoke in rats. Adult rats were administered with 2 g/kg alcohol (v.o.) or/and inhaled the smoke from 6 cigarettes, twice/day, for 30 days. Behavioral tests were performed 3 h after the alcohol administration and 1 h after the last exposure to cigarette smoke in the morning. Cerebrospinal fluid was collected for glutamate determination and the hippocampus was dissected for GABAA and NMDA receptor subunits mRNA expression determination. Results showed that the combined use of alcohol and cigarette smoke (ALTB) in rats increased the locomotor activity and all interventions decreased anxiety-like behaviors. Despite being on a short-term withdrawal, the cigarette smoke exposure decreased the percentage of open arm entries in the elevated plus maze test, which was prevented by combined use with alcohol. Even though GABAA and glutamate receptor subunits expression did not change in the hippocampus, glutamate levels were significantly higher in the cerebrospinal fluid from ALTB rats. Therefore, we showed that the combined use of alcohol and cigarette maintained a psychostimulant effect after a short-term withdrawal that was associated with the elevated glutamatergic activity. The combined use also prevented anxiety-like signs in cigarette smoke exposure rats, decreasing an adverse effect caused by nicotine withdrawal. These results could explain, in part, the elevated frequency of combined use of these two drugs of abuse in humans.

Correlations between subunits of GABAA and NMDA receptors after chronic alcohol treatment or withdrawal, and the effect of taurine in the hippocampus of rats.

Chronic use of alcohol and its withdrawal impairs the delicate balance between GABAergic and glutamatergic systems. This imbalance includes changes in GABA receptors - importantly in GABAA subtypes - and glutamate receptors, especially in NMDA subtypes. A better comprehension of the different roles of GABAAR and NMDAR subunits could be helpful to define new strategies to counteract the deleterious effects observed during alcohol withdrawal. Taurine, a sulfonated amino acid, has been proposed to attenuate alcohol withdrawal symptoms due to its neuromodulatory properties. In this study, we evaluated the correlations between GABAAR and NMDAR subunits in the hippocampus of rats chronically treated with alcohol or in alcohol withdrawal, and the effects of taurine treatment on these parameters. Male Wistar rats received alcohol (2 g/kg) or water by oral gavage (control), 2 × /day, for 28 days. From day 29 to day 33, withdrawal rats received water instead of alcohol and all groups were reallocated to receive 100 mg/kg taurine or saline intraperitoneally (i.p.), once a day. On day 34, rats were euthanized and the hippocampus was dissected for GABAAR α1, α4, δ, and γ2 and NMDAR GluN2A and GluN2B subunits mRNA expression determination by RT qPCR. There were no differences between groups in the studied GABAAR and NMDA subunits. However, we observed a correlation of α1 and γ2 subunits induced by taurine, while in the alcohol group there was a correlation between α4 and GluN2A. In the group treated with alcohol and taurine, we observed an extra correlation, between α1 and GluN2A. After 5 days of withdrawal, a correlation observed in the control group, between δ and GluN2A, was reestablished. The correlation found between subunits suggests a neuroadaptation of GABAergic and glutamatergic systems in withdrawal rats. Results from this study contribute to the elucidation of the mechanisms beyond neuroadaptations observed in alcohol use and withdrawal.

Taurine enhances voluntary alcohol intake and promotes anxiolytic-like behaviors in rats.

Taurine is an amino acid usually added to energy drinks. In rodents, acute taurine administration decreases voluntary alcohol intake, and subchronic administration restores different behavioral features impaired by alcohol withdrawal. In the present study, we evaluated the effects of chronic taurine treatment on voluntary alcohol consumption and changes in behavioral parameters in rats. Adult male Wistar rats were divided into two groups and were allowed to choose from two bottles containing 20% alcohol or 0.08% saccharin (vehicle solution), or two bottles containing vehicle, 24 h per day, for 5 weeks. After 3 weeks, rats received 100 mg/kg taurine (TAU) or saline (SAL) intraperitoneally once a day for 2 weeks, and daily alcohol consumption was monitored. On days 22 and 33, rats were tested in the open-field, and on day 34, they were exposed to the light/dark task (LDT). Our results show for the first time that chronic taurine treatment enhanced voluntary alcohol intake and preference in rats, and that these changes were accompanied by an anxiolytic-like phenotype in alcohol-treated rats, possibly due to its synergistic effect with alcohol on the dopaminergic and GABAergic systems.

Cigarette smoking and antioxidant defences in packed red blood cells prior to storage.

BACKGROUND: Red blood cells from smoking donors can have more lesions from oxidative stress, decreasing the benefits of blood transfusion. We aimed to explore the effect of cigarette smoking on the oxidative status of packed red blood cells (PRBCs) prior to storage. MATERIALS AND METHODS: We compared serum vitamin C, plasmatic malondialdehyde (MDA), and non-protein thiol groups (GSH) levels in PRBCs, as well glutathione peroxidase (GPx) and glutathione s-transferase (GST) activity in PRBCs from smoking (n=36) and non-smoking (n=36) donors. We also correlated urinary cotinine levels with these parameters. RESULTS: Cigarette smoking was associated with decreased serum levels of vitamin C and GPx, and increased GST activity in PRBCs. We found negative correlations between cotinine, GPx activity and vitamin C levels, and a positive correlation between cotinine and GST activity. DISCUSSION: Cigarette smoking changed antioxidant defences of PRBCs prior to storage and these parameters are correlated with cotinine levels. Increased RBC antioxidants such as GST may reflect an exposure to oxidants during erythropoiesis. Because of the inability of mature RBCs to resynthesise antioxidants, PRBCs from smokers may have higher risk of storage lesions than those from non-smoker donors.

Comparative study on the effects of cigarette smoke exposure, ethanol consumption and association: Behavioral parameters, apoptosis, glial fibrillary acid protein and S100ß immunoreactivity in different regions of the rat hippocampus.

Exposure to cigarette smoke and ethanol are proposed to trigger neurotoxicity, apoptosis, and to impair neuronal signaling. However, it is little known how the combination of both might trigger astrogliosis and the morphological changes capable of affecting a differential susceptibility of hippocampal regions to these licit drugs. The present study investigated the chronic effects of exposure to cigarette smoke and/or ethanol on behavioral parameters, apoptosis, and alteration in immunoreactivity of glial fibrillary acid protein (GFAP) and S100ß in the CA1, CA3, and dentate gyrus (DG) of the rat hippocampus. Adult male Wistar rats (n = 32) were divided into four groups: vehicle (VE, glucose 3% in water, 10 mL/kg), cigarette smoke (TOB, total 12 cigarettes per day), ethanol (ethanol, 2 g/kg), and cigarette smoke plus ethanol (TOB plus ethanol, total 12 cigarettes per day plus ethanol 2 g/kg) for 54 days. The groups were submitted to tail-flick, open-field, and inhibitory avoidance tasks. The results showed that ethanol per se worsened the short-term memory. The association between TOB and ethanol increased the immunoreactivity of cleaved caspase-3 in the CA3 and DG regions. The TOB plus ethanol group showed a lower immunoreactivity to GFAP in all regions of the hippocampus. In addition, ethanol and TOB per se also reduced the immunoreactivity for GFAP in the DG. Ethanol increased S100ß immunoreactivity only in the DG. In conclusion, this study showed that only ethanol worsened short-term memory, and the DG became more susceptible to changes in the markers investigated. This evidence suggests that DG is more sensitive to neurotoxicity induced by cigarette smoke and ethanol.

Combined Exposure to Alcohol and Tobacco Smoke Changes Oxidative, Inflammatory, and Neurotrophic Parameters in Different Areas of the Brains of Rats.

Devastating effects of exposure to alcohol and tobacco smoke on health are extensively reported in the literature. However, few studies have attempted to elucidate the consequences of their combined use on the central nervous system. Here we studied the effect of this combined use on some oxidative, inflammatory, and neurotrophic parameters in the hippocampus, striatum, and frontal cortex of rats. Adult Wistar rats were allocated into control (CT), alcohol (AL), tobacco smoke (TB), or combined (ALTB) groups. Rats were exposed to environmental air (CT and AL groups) or to the smoke from six cigarettes (TB and ALTB groups) immediately after tap water (CT and TB) or 2 g of alcohol/kg (AL and ALTB) oral gavage administration, twice a day, for 4 weeks. On day 28, rats were euthanized and areas of the brain were dissected to evaluate some cellular redox parameters, pro-inflammatory cytokine levels, and brain-derived neurotrophic factor (BDNF) levels. A one-way analysis of variance showed that the ALTB combined treatment significantly increased oxidative stress levels in the hippocampus. ALTB also increased interleukin-1ß levels in the striatum and frontal cortex and tumoral necrosis factor-α levels in the frontal cortex compared with those of AL, TB, and CT rats. Combined treatment also decreased the BDNF levels in the frontal cortex of rats. Oxidative damage was found, more importantly, in the hippocampus, and inflammatory parameters were extended to all areas of the brain that were studied. Our results showed an interaction between alcohol and tobacco smoke according to the area of the brain, suggesting an additional risk of neural damage in alcoholics who smoke.

Acute intraperitoneal administration of taurine decreases the glycemia and reduces food intake in type 1 diabetic rats.

Taurine, an amino acid with antioxidant and osmoregulatory properties, has been studied for its possible antidiabetic properties in type 1 and type 2 diabetic animals. In type 2 diabetic mice, taurine decreases blood glucose through increased insulin secretion and insulin receptor sensitization. However, insulin is absent in type 1 diabetic individuals. The aim of this study was to evaluate the effects of taurine on parameters related to the energy balance that could explain the metabolic action of this amino acid in type 1 diabetic rats. Control and streptozotocin-induced diabetic rats received saline or taurine (100 mg/kg/day), intraperitoneally, for 30 days. Parameters such as palatable food intake, gastrointestinal transit rate, serum glucose, insulin, leptin, and glucagon levels were measured 60 min after the last taurine administration. Liver, kidneys, heart, and retroperitoneal fat were dissected and weighted. Glycogen levels were measured in the liver and soleus muscle. Our results showed that acute taurine administration decreased glycemia. It also decreased food intake in diabetic rats, without affecting other metabolic parameters. Altogether, our results suggest that in type 1 diabetic rats, taurine decreases blood glucose by a non-insulin-dependent mechanism. Due to the safety profile of taurine, and its effect on glycemia, this amino acid may help to design new drugs to add benefit to insulin therapy in type 1 diabetic individuals.

Taurine restores the exploratory behavior following alcohol withdrawal and decreases BDNF mRNA expression in the frontal cortex of chronic alcohol-treated rats.

Alcohol use disorder is an alarming health problem, and the withdrawal symptoms increase the risk of relapse. We have hypothesized that taurine, a multitarget substance acting as a gamma-aminobutyric acid A receptor (GABAAR) positive modulator and a partial inhibitor of N-methyl-d-aspartate (NMDA) glutamate receptors, may reduce the withdrawal symptoms or modify behaviors when combined with alcohol. Therefore, we investigated the effects of taurine on behavior in the open field test (OFT), the GABAAR α2 subunit and BDNF mRNA expression in the frontal cortex of rats after chronic alcohol treatment or upon withdrawal. Rats received alcohol 2g/kg (alcohol and withdrawal groups) or water (control group) twice daily by oral gavage for 28days. On day 29, the withdrawal rats received water instead of alcohol, and all groups were reallocated to receive 100mg/kg taurine or vehicle intraperitoneally, once a day for 5days. On day 33, the rats were exposed to OFT; 18h later, they were euthanized, and the frontal cortex was dissected for GABAAR α2 subunit detection and BDNF mRNA expression determination by real-time quantitative PCR. Taurine administration restored rearing behavior to the control levels in the withdrawal rats. Taurine also showed anxiolytic-like effects in control rats and did not change the behaviors in the chronic alcohol group. Chronic alcohol treatment or withdrawal did not change the GABAAR α2 subunit or BDNF mRNA expression in the frontal cortex, but taurine decreased the α2 subunit level in control rats and to the BDNF levels in the alcohol rat group. We conclude that taurine restored exploratory behavior after alcohol withdrawal but that this effect was not related to the GABAAR α2 subunit or BDNF mRNA expression in the frontal cortex of the rats.