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Background: Alcohol and tobacco are among the leading substances that are misused together and shared genetic vulnerability is likely. Increased susceptibility to nicotine self-administration has been shown in alcohol-preferring rat-lines. However, a nicotine-preferring (nP) rat-line has not been studied for alcohol preference. Objectives: To evaluate alcohol preference and intake in male and female nP rats. We hypothesized that nP rats and females would drink more ethanol than control rats and males, respectively. Methods: nP rats are being selectively outbred for high oral nicotine intake at Ege University. Seventeen nP (18(th) generation) and 20 naive female and male SD rats, not previously exposed to alcohol or nicotine, were used. Twelve-week-old rats were given intermittent access to 20% ethanol in a 2-bottle-choice-procedure for six weeks. After one week withdrawal, six weeks of oral nicotine self-administration was applied. Results: nP rats drank significantly more ethanol than controls and their preference for ethanol over water was higher. Female rats' ethanol intake was higher than males'. the nP rats' nicotine preference and intake were higher than controls, and they gained less weight. Conclusion: We have shown for the first time that nP rats also have high alcohol intake. Our results support the hypothesis that shared genetic factors may underlie concurrent addiction to nicotine and alcohol and have translational value in understanding their misuse. Considering the increased vulnerability for alcohol use disorder in smokers and sex differences observed, early preventive measures in families with a history of tobacco addiction, specifically targeting female members, could have public health benefits., Ege University Research FundEge University [17 TIP 071], This work was supported by Ege University Research Fund, Number: 17 TIP 071.

Smoking is the major cause of preventable deaths worldwide, and although there is a decline in overall smoking prevalence in developed countries, the decline in women is less pronounced than in men. Women become dependent faster and experience greater difficulties in quitting. Similar trends have been observed in animal models of nicotine/tobacco addiction. Individual differences in vulnerability to drug abuse are also observed in nicotine/tobacco addiction and point to the importance of sex differences. This Review, summarizes findings from three experimental approaches used to depict nicotine preference in animal models, intravenous and oral nicotine self-administration and nicotine-induced conditioned place preference. Nicotine preference is considered to be reflected in the animal's motivation to administer the drug (intravenously or orally) or to prefer an environment paired with the presence of the drug (conditioned place preference). These approaches all point to the importance of sex and age of the subjects; the preference of females and adolescents appear to be more pronounced than that of males and adults, respectively. A closer look at these factors will help us understand the mechanisms that underlie nicotine addiction and develop strategies to cope. Ignoring sex differences and reaching conclusions based only on studies using male subjects has resulted in erroneous generalizations in the past. Sex differences in nicotine preference have been clearly documented, and awareness on this aspect of nicotine dependence will significantly impact our success in translational research. © 2016 Wiley Periodicals, Inc.

Background: Nicotine exerts its central actions through nicotinic acetylcholine receptors (nAChRs), which in turn regulate major neurotransmitter systems including dopamine. Nicotinic and dopaminergic systems play significant roles in physiological functions, neuropsychiatric disorders, and addiction. Objectives: To evaluate possible differences in the expression of nAChR subunit and dopamine receptor (DR) mRNAs following voluntary nicotine intake. Methods: Male and female rats (n = 67) were exposed to long-term free-choice oral nicotine (24 hours/day, 6 weeks); rats with maximum and minimum nicotine preference/intake were selected. The mRNA levels of genes encoding α4,β2,α5, and α7 nAChR subunits and DR Drd1and Drd2 subtypes were evaluated in the striatum (STR), prefrontal cortex (PFC), and hippocampus using quantitative real-time polymerase chain reaction in selected rats (n = 30) and their control groups (n = 15). Results: In addition to baseline differences, expression changes were observed in the mRNA levels of evaluated genes in rats exposed to voluntary oral nicotine in a brain region-, sex-, and preference-related manner. Nicotine intake is correlated negatively with Chrnb2, Chrna7 and positively with Drd1 expression. In the cholinergic system, regional differences in Chnrb2 and Chrna5, sex differences in Chrna4 and Chrna5, and nicotine preference effects in the expression of all subunits except α4 were observed. Chrna5 was lower in maximum than in minimum preferring, and in male than female rats, supporting the inhibitory role of the α5 subunit in nicotine dependence. Nicotine increased Drd2 mRNA expression only in minimum preferring female rats in STR and PFC. Conclusion: Modulation of nAChR and DR gene expression by nicotine may have clinical implications and aid drug development. Pharmaceuticals targeting the nicotinic cholinergic and dopaminergic systems might be expected to have differential efficacy that varies with the patient’s sex or smoking status.

Background We have generated high- and low-nicotine preferring (high-NP, low-NP) rat lines using voluntary oral nicotine intake as the selection criterion. After nine generations, the estimated realized heritability for high intake was 0.26. The aim of the current study is to compare how nicotine withdrawal varies between these two lines. This new analysis would help elucidate if nicotine withdrawal and intake share common genetic mechanisms. Methods After exposing male and female Sprague Dawley rats (F 8 generation) to six weeks of nicotine exposure, nicotine was withdrawn. Somatic signs of withdrawal, locomotor activity, and weight were measured at 16 and 40 h. One week after withdrawal, resumption of nicotine intake was determined. Results The High-NP line had higher nicotine intake before and after withdrawal than the Low-NP line. High-NP rats were more active than Low-NP rats, and locomotor activity decreased during withdrawal; this decrease was more pronounced in the High-NP line. High-NP rats gained more weight during withdrawal than Low-NP rats. Escape attempts decreased during withdrawal in all groups, but overall females demonstrated more escape attempts than males. The other somatic signs of withdrawal were higher during withdrawal compared to baseline and more pronounced in females. Conclusions Selection for nicotine preference affected nicotine intake, locomotion and weight, suggesting the heritability of these traits. However, despite differences in nicotine preference and intake, high-NP and low-NP rats showed similar withdrawal responses: escape attempts decreased and somatic signs increased. Withdrawal responses of females were more pronounced than males suggesting sex differences in the negative affect induced by nicotine withdrawal. The major finding of this novel analysis is showing that nicotine preference does not predict withdrawal symptoms. This finding, together with sex differences observed during withdrawal, may contribute to a better understanding of nicotine dependence and have translational value in developing more effective strategies for smoking cessation.

Background The hypothalamus harbors high levels of cholinergic neurons and axon terminals. Nicotinic acetylcholine receptors, which play an important role in cholinergic neurotransmission, are expressed abundantly in the hypothalamus. Accumulating evidence reveals a regulatory role for nicotine in the regulation of the stress responses. The present review will discuss the hypothalamic neuropeptides and their interaction with the nicotinic cholinergic system. The anatomical distribution of the cholinergic neurons, axon terminals and nicotinic receptors in discrete hypothalamic nuclei will be described. The effect of nicotinic cholinergic neurotransmission and nicotine exposure on hypothalamic-pituitaryadrenal (HPA) axis regulation at the hypothalamic level will be analyzed in view of the different neuropeptides involved. Methods Published research related to nicotinic cholinergic regulation of the HPA axis activity at the hypothalamic level is reviewed. Results The nicotinic cholinergic system is one of the major modulators of the HPA axis activity. There is substantial evidence supporting the regulation of hypothalamic neuropeptides by nicotinic acetylcholine receptors. However, most of the studies showing the nicotinic regulation of hypothalamic neuropeptides have employed systemic administration of nicotine. Additionally, we know little about the nicotinic receptor distribution on neuropeptide-synthesizing neurons in the hypothalamus and the physiological responses they trigger in these neurons. Conclusion Disturbed functioning of the HPA axis and hypothalamic neuropeptides results in pathologies such as depression, anxiety disorders and obesity, which are common and significant health problems. A better understanding of the nicotinic regulation of hypothalamic neuropeptides will aid in drug development and provide means to cope with these diseases. Considering that nicotine is also an abused substance, a better understanding of the role of the nicotinic cholinergic system on the HPA axis will aid in developing improved therapeutic strategies for smoking cessation.

Nicotine addiction is a serious health problem resulting in millions of preventable deaths worldwide. The gas messenger molecule nitric oxide (NO) plays a critical role in addiction, and nicotine increases nitric oxide metabolites (NOx) in the brain. Understanding the factors which underlie individual differences in nicotine preference and intake is important for developing effective therapeutic strategies for smoking cessation. The present study aimed to assess NO activity, by measuring its stable metabolites, in three brain regions that express high levels of nicotinic acetylcholine receptors in rats preselected for nicotine preference. Rats (n=88) were exposed to two-bottle, free choice of oral nicotine/water starting either as adolescents or adults; control animals received only water under identical conditions. Following 12 or six weeks of exposure, levels of NOx (nitrite+nitrate), were determined in the hippocampus, frontal cortex, and amygdala. Since the rats were singly housed during oral nicotine treatment, naïve rats were also included in the study to evaluate the effect of isolation stress. Isolation stress increased NOx in the hippocampus. Nicotine preference did not have a significant effect on NO activity, but rats with adolescent exposure had higher NOx levels in the frontal cortex compared to adult-onset rats. Our findings suggest that nicotine exposure during adolescence, regardless of the amount of nicotine consumed, results in higher NO activity in the frontal cortex of rats, which persists through adulthood.

Nicotine is a highly addictive drug and exerts its effect partially through causing dopamine release, thereby increasing intrasynaptic dopamine levels in the brain reward systems. Dopaine D1 receptor (DRD1) mRNAs and receptors are localized in reward-related brain regions, which receive cholinergic input. The aim of this study is to evaluate whether nicotine administration affects the expression of DRD1s, and if so, whether epigenetic mechanisms, such as histone acetylation, are involved. Twenty Male Sprague Dawley rats received nicotine (0.4 mg/kg/day, s.c.) or saline injections for 15 days. After nicotine/saline treatment, rats were perfused with saline; prefrontal cortex (PFC), corpus striatum (STR), and ventral tegmental area (VTA) were dissected. Homogenates were divided into two parts for total RNA isolation and histone H4 acetylation studies. DRD1 mRNA expression was significantly higher in the PFC of the nicotine-treated group compared with controls; similar trends were observed in the VTA and STR. To study epigenetic regulation, the 2kb upstream region of the DRD1 gene promoter was investigated for histone H4 acetylation in PFC samples. After chromatin immunoprecipitation with anti-acetyl histone H4 antibody, we found an increase in histone acetylation by two different primer pairs which amplified the −1365 to −1202 (P

WOS: 000310414300006, PubMed ID: 22960117, Cocaine and amphetamine regulated transcript (CART) mRNA and peptides are highly expressed in the paraventricular (PVN), dorsomedial (DMH) and arcuate (ARC) nuclei of the hypothalamus. It has been suggested that these nuclei regulate the hypothalamic-pituitary-adrenal (HPA) axis, autonomic nervous system activity, and feeding behavior. Our previous studies showed that forced swim stress augmented CART peptide expression significantly in whole hypothalamus of male rats. In another study, forced swim stress increased the number of CART-immunoreactive cells in female PVN, whereas no effect was observed in male PVN or in the ARC nucleus of either sex. In the present study, we evaluated the effect of forced swim stress on CART mRNA expression in PVN, DMH and ARC nuclei in both male and female rats. Twelve male (stressed and controls, n=6 each) and 12 female (stressed and controls, n=6 each) Sprague-Dawley rats were used. Control animals were only handled, whereas forced swim stress procedure was applied to the stressed groups. Brains were dissected and brain sections containing PVN, DMH and ARC nuclei were prepared. CART mRNA levels were determined by in situ hybridization. In male rats, forced swim stress upregulated CART mRNA expression in DMH and downregulated it in the ARC. In female rats, forced swim stress increased CART mRNA expression in PVN and DMH, whereas a decrease was observed in the ARC nucleus. Our results show that forced swim stress elicits region- and sex-specific changes in CART mRNA expression in rat hypothalamus that may help in explaining some of the effects of stress. (C) 2012 Elsevier B.V. All rights reserved., NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [3 R01 DA010732-05S1]; National Center for Research ResourcesUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Center for Research Resources (NCRR) [P51RR165]; Office of Research Infrastructure Programs/OD [P51OD11132], This study was supported by NIH Grant no. 3 R01 DA010732-05S1 and by the National Center for Research Resources (P51RR165) and is currently supported by the Office of Research Infrastructure Programs/OD (P51OD11132).

CART mRNA and peptides are highly expressed in the anatomical structures composing the hypothalamo-pituitary-adrenal (HPA) axis and sympatho-adrenal system. Anatomical and functional studies suggest that CART peptides may have a role in the regulation of the neuroendocrine and autonomic responses during stress. Our previous study showed that CART peptides increased significantly in the male hypothalamus and amygdala 10 min after the forced swim stress. The present study aimed to examine the effect of forced swim stress on CART peptide expression in selected brain regions, including those where CART peptide expression has not been reported before (frontal cortex, pons, medulla oblongata), as well as in endocrine glands related to stress in male Sprague Dawley rats. A total of 16 (n = 8) animals were used, including control groups. Rats were subjected to forced swim on two consecutive days, and sacrificed on the second day, 2 h after the termination of the stress procedure. Frontal cortex, pons, medulla oblongata, hypothalamus, pituitary and adrenal glands were dissected and homogenized. CART peptide expression in these tissues was measured by Western Blotting and six different CART peptide fragments were identified. Our results showed that forced swim stress elicited region-specific changes in CART peptide expression. CART was upregulated in the frontal cortex, hypothalamus, medulla oblongata and adrenal gland while there was no change in the pons and pituitary. Enhanced CART peptide fragments in these brain regions and adrenal glands may have a role in the regulation of the HPA and sympatho-adrenal axis activity during stress response.

An increasing number of studies suggest that nicotine/tobacco addiction is modulated by ovarian hormones. The levels of estrogen and progesterone appear to be important in the success of quit attempts and smoking cessation. In women smokers with the diagnosis or risk of breast cancer, the estrogen receptor modulator tamoxifen (TAM) is widely used, and even though the detrimental health effects of smoking are known, this vulnerable group has difficulty quitting and continues to smoke. The current study tested the effect of the estrogen receptor modulator TAM and the progesterone receptor antagonist mifepriston (RU486) on nicotine-induced conditioned place preference (CPP) in adult female rats. A three chambered CPP apparatus was used and nicotine was paired with the initially non-preferred chamber. Rats received nicotine or saline and hormone receptor modulators (vehicle, TAM, RU486) in a 2 × 3 experimental design. We have previously shown that nicotine induces CPP in male Sprague–Dawley rats but not in females. Our results show that while nicotine alone does not induce CPP in female rats, rats treated with TAM exhibit nicotine-induced CPP. Although RU486 has an aversive effect when applied alone, this is ameliorated by nicotine. These results confirm the role of ovarian hormone receptors in nicotine-induced CPP and may have clinical implications for developing more efficient smoking cessation approaches in women smokers.

To elucidate sex differences in nicotine addiction and the underlying mechanisms of the conditioning aspects of nicotine, nicotine-induced conditioned place preference (CPP) was evaluated in male and female Sprague Dawley rats using a three-chambered CPP apparatus and a biased design. In a series of experiments, the dose-response curve was obtained, pairings between the drug and initially non-preferred versus preferred compartments were compared, and the involvement of mGluR5 receptors in nicotine-induced CPP was evaluated. Modulation of nicotine-induced CPP with mGluR5 inhibition was obtained by MPEP (2-methyl-6-(phenylethynyl)-pyridine hydrochloride). Our results show that nicotine induces CPP dose-dependently in male rats but not in female rats. The comparison of the biased protocol, pairing nicotine with the initially preferred and non-preferred chambers, indicated that nicotine-induced CPP in male rats under both conditions, but the effect was stronger when nicotine was paired with the initially non-preferred side. The selective mGluR5 antagonist MPEP inhibited nicotine-induced CPP in male rats. In conclusion, the results of the current study in rats demonstrate that the conditioning effect of nicotine is more important in males than in females. Furthermore, in line with reported findings, our results suggest that mGluR5 antagonism may be therapeutically useful in smoking cessation during the maintenance of smoking behavior when conditioning plays an important role, notwithstanding the fact that this effect is observed only in male rats, not in females.

WOS: 000252342800015, PubMed ID: 18188754, Results from recent studies suggest that chronic cigarette smoking is associated with increased white matter volume in the brain as determined by in vivo neuroimaging. We used diffusion tensor imaging to examine the microstructural integrity of the white matter in 10 chronic smokers and 10 nonsmokers. All individuals were healthy, without histories of medical or psychiatric illness. Fractional anisotropy (FA) and trace were measured in the genu, body, and splenium of the corpus callosum. FA provides a measure of directional versus nondirectional water diffusion, whereas trace provides a measure of nondirectional water diffusion. Lower FA and higher trace values are considered to reflect less brain integrity. Voxel-based morphometry was used to define volumes in each of these regions of the corpus callosum. Chronic smokers exhibited significantly higher FA in the body and whole corpus callosum and a strong trend for higher FA in the splenium compared with nonsmokers. FA did not differ between groups in the genu, and neither trace nor white matter volumes differed between groups in any of the regions of interest. When subdivided by Fagerstrm score (low vs. high), the low Fagerstrm group exhibited significantly higher FA in the body of the corpus callosum compared with the high Fagerstrm group and the nonsmokers. These results suggest that, among healthy adults, lower exposure to cigarette smoking is associated with increased microstructural integrity of the white matter compared with either no exposure or higher exposure. Additional studies are needed to further explore differences in white matter integrity between smokers and nonsmokers., NCI NIH HHSUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [CA84718, P50 CA084718]; NIBIB NIH HHSUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of Biomedical Imaging & Bioengineering (NIBIB) [R01 EB004155, EB4155]; NIDA NIH HHSUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute on Drug Abuse (NIDA) [K08 DA014276-03, DA14276, K08 DA014276, K08 DA014276-05, K08 DA014276-02, K08 DA014276-04, K08 DA014276-01A2]

Background Nicotine is the major addictive component in tobacco, and despite well-established adverse health effects of tobacco addiction, some smokers have difficulty quitting. The acute cognitive enhancement and/or the amelioration of the cognitive disruption during withdrawal that some smokers experience after smoking are among important factors that hinder quit attempts. The animal model presented in the current study is comparable to the human smoking condition although nicotine intake routes are different. Rats were exposed to a free choice of oral nicotine starting at adolescence, and given a water maze (WM) task as adults. This design allowed us to see if rats alter their nicotine intake during the WM task and if nicotine preference and intake modify abilities and strategies rats use for problem solving. Methods Male and female rats were exposed to a free choice of oral nicotine/water for 24 weeks, starting at five weeks of age. After this period, they were selected based on their nicotine intake and, together with control animals that received only water, were subjected to a place-learning task in the WM. Free-choice nicotine exposure continued during WM testing. Following acquisition, the probe trial presented the rats with a choice between using two different strategies for problem solving. Results Nicotine supported acquisition and rats increased their nicotine intake during WM testing; this effect was more pronounced in male rats with minimum nicotine preference and intake. Furthermore, nicotine modified the “female type” strategy in solving the place-learning task and nicotine treated female rats, unlike control females, behaved like males. Conclusions The increase in nicotine intake during mental engagement, and the sexually dimorphic effect of nicotine on problem solving strategies that we have observed in rats, may suggest that implementing sex-specific smoking cessation approaches, especially under stressful and cognitively demanding conditions, may be useful in helping smokers quit.

CART is expressed abundantly in the hypothalamic paraventricular nucleus and locus coeruleus, major corticotropin releasing factor (CRF) and noradrenaline sources, respectively. There is a bidirectional relation between CART and hypothalamo-pituitary-adrenal axis activity. CART stimulates CRF, adrenocorticotropic hormone and glucocorticoid secretion, whereas CRF and glucocorticoids increase the transcriptional activity of the CART gene; adrenalectomy declines CART expression in the hypothalamus. Stress exposure modulates CART expression in hypothalamus and amygdala in rat brain in a region and sex specific manner. CART may be a mediator peptide in the interaction between stress, drug abuse, and feeding. The review discusses the established role of CART as it relates to the stress response.

Previous studies that have examined the impact of cigarette smoking on cognition have revealed mixed results; some studies report no impact and others report detrimental effects, especially in older individuals. Few studies, however, have examined the effects of cigarette smoking on both young and old healthy individuals using highly robust and standardized methods of cognitive assessment. This study draws on an international database to contrast cognitive differences between younger and older individuals who regularly smoke cigarettes and non-smokers. Data were sampled from 1000 highly screened healthy individuals free of medical or psychiatric health complications. A cohort of 62 regular smokers (n = 4545 years of age; n = 1745 years) with a Fagerstrom nicotine dependency score of 1 or more were identified and matched to a cohort of 62 healthy nonsmokers (n = 4345 years; n = 1945 years) on demographic variables and estimated intelligence. Performances on cognitive measures of attention, reaction time, cognitive flexibility, psychomotor speed, and memory were considered for analysis. As a group, smokers performed more poorly than nonsmokers on one measure of executive function. A significant age and smoking status interaction was identified with older smokers performing more poorly than older nonsmokers and younger smokers on a measure of long-delayed recall of new information. Cigarette smoking is associated with isolated and subtle cognitive difficulties among very healthy individuals.

Cholinergic and dopaminergic systems are involved in spatial memory and are modulated by nitric oxide (NO); NO has well documented effects on place learning in rodents. The aim of the present study was to investigate the effect of NOS inhibition on place learning in the water maze and to evaluate the relationships between NOS inhibition, learning performance, dopamine (DA) D2 and muscarinic acetylcholine (mACh) receptors. Male Sprague–Dawley rats received the NOS inhibitor Nω-Nitro- l -Arginine ( l -NA), or saline and were trained in the water maze. Rats that were not trained, but received the same treatments were also included. Following treatments with or without water maze training, [ 3 H]-QNB and [ 3 H]-spiperone binding in cortex, striatum and hippocampus were determined to assess the effects of NOS inhibition and/or learning on DA D2 and mACh receptor regulation. The overall results of the present study showed that: (1) NOS inhibition impairs performance in the MWM; (2) NOS inhibition does not affect specific binding to DA D2 (striatum and hippocampus) and mACh (cortex and hippocampus) receptors; (3) MWM training lowers D2 and mACh receptor binding in cortical regions.

In order to test for glucocorticoid regulation of CART in the arcuate nucleus, adrenalectomies (ADX) and hormone replacements (HRs) were carried out in groups of rats. CART mRNA levels were determined by in situ hybridization and CART peptide levels by immunocytochemistry. ADX caused a lowering of CART mRNA and peptides levels in the arcuate and this was reversed by HR. These results indicate a glucocorticoid regulation of CART in the arcuate. The regulation could be direct through an action of glucocorticoid receptors or indirectly through ADX-induced changes in leptin levels. These findings suggest a role for CART in the stress response.

After the recognition of nitric oxide (NO) as a messenger molecule in the nervous system, carbon monoxide (CO) has received attention with similar properties. The present study aims to elucidate the effects of CO on synaptosomal dopamine ( 3 H-DA) and glutamate ( 3 H-Glu) uptake and on cGMP levels; possible interaction between NO and CO systems was also evaluated. Our results provide evidence for the inhibition of DA and Glu uptake by CO in a time-, dose-, and temperature-dependent manner in rat striatum and hippocampus, respectively; the inhibition observed was sexually dimorphic with more pronounced effects in females. Basal cGMP levels were higher in female rats than males in the striatum and exogenous CO increased striatal cGMP levels only in males; no effect of CO was observed in the hippocampus. In vivo nitric oxide synthase (NOS) inhibition increased DA and Glu uptake; however, CO was still effective in inhibiting uptake following NOS inhibiton. Taken together, these findings suggest a role for CO in trans-synaptic regulation through modulation of DA and Glu transporters and of cGMP levels; the effect on cGMP levels is independent of NOS activity and appears to be sexually dimorphic and region specific. Neuropsychopharmacology (2003) 28, 1176–1181, advance online publication, 9 April 2003; doi:10.1038/sj.npp.1300132

Systemic kainic acid (KA) administration to rats triggers wet dog shakes (WDS) followed by epileptic seizures. Although WDS are often associated with the occurrence of seizures, we have recently shown that following nitric oxide (NO) synthesis inhibition, the number of WDS decreased; subsequently the onset of seizure activity was shortened, and the number of convulsions was increased. Somatostatin (SS), whose release appears to be controlled by NO, inhibits seizure activity. There are sex differences in seizure susceptibility as well as in SS and NO activities in brain. The present study was undertaken to assess the effect of octreotide (OC), a stable SS analogue, on KA-induced WDS and seizures in rats, with emphasis on possible sex differences. WDS and seizures were induced by KA in male and female (proestrus) Sprague Dawley rats; OC or saline was injected 30 min before KA and the behavior was monitored for 120 min after KA. Octreotide increased the number of WDS and decreased the number of convulsions; this effect was more pronounced in males. Onset of KA-induced seizure activity was earlier in females than males; however, there was no effect of OC on seizure latency. Seizure activity started after the termination of WDS. These results show OC has opposite effects on WDS and convulsions, in that it stimulates the former and inhibits the latter. These results support our previous findings that WDS and seizure activity involve separate mechanisms and suggest that WDS may have an inhibitory effect on limbic seizures.

Electrical stimulation of limbic structures, pharmacological interventions, and getting wet induces wet dog shakes (WDS) in rats. WDS are often associated with the occurrence of seizures. In this study, we evaluated the effects of reduced NO production on physiologically (wetting)- or pharmacologically (kainic acid; KA)-induced WDS and KA-triggered seizures. Following wetting, naive and saline-treated rats displayed more WDS than rats treated with NO synthase (NOS) inhibitor, N omega-nitro-L-arginine (L-NA). In another experiment, WDS and seizures were monitored after KA treatment alone or in combination with L-NA. Again, NOS inhibition reduced the number of KA-triggered WDS but augmented the number and severity of seizures. Our results suggest that not only do physiologically- and kainate-induced WDS share a common mechanism that includes NO, but that there is also an antagonism between WDS and convulsions.

Nicotine affects sensory pathways and an interaction between taste and nicotine preference is likely. In addition to pharmacologic effects, orosensory factors are important in nicotine dependence. Recent evidence suggests a link between taste (notably bitter) receptor genes and nicotine addiction.To explore the possible interaction between taste and nicotine preference in rats, including sex as a factor.Adult male and female Sprague Dawley rats (n = 82) were used in free choice oral intake experiments. In Experiment 1 rats received water from one bottle and one of the taste substances (quinine, sucrose, or saccharine) from the other bottle for 12 days. Following a wash-out period, Experiment 2a was initiated in the same rats. Rats received water from one bottle and nicotine (10 and 20 mg/l) from the other for 12 days. In Experiment 2b, nicotine exposure was continued for four more weeks. Liquid intake and weight were measured at four-day (Experiments 1 and 2a) and one week (Experiment 2b) periods.In female rats, quinine and subsequent nicotine intake were positively correlated and quinine intake and weight gain were negatively correlated. No association was depicted between nicotine consumption and sweet tastants in either female or male animals.The results suggest that bitter taste and nicotine preference are related, but only in female rats. This finding is parallel to observations in human smokers. Our study may be a preliminary step in the search for common genes that underlie nicotine dependence and taste preference.

CART peptide is a neurotransmitter involved in various physiological processes including feeding, sensory processing, development, addiction, and stress. Substantial amounts of CART mRNA and CART peptide expression have been demonstrated in the hypothalamic periventricular area, the paraventricular nucleus of hypothalamus, the anterior lobe of the pituitary gland and the adrenal gland in addition to many other brain areas. This localization defines the HPA axis, responsible for the stress response. The aim of the present study was to assess the possible mediation of the CART peptides in the stress response by testing for changes in CART in adrenalectomized animals. Three groups of male Sprague–Dawley rats were used for the study: sham operated, adrenalectomized (ADX), and ADX+hormone replacement (corticosterone, 30 μg/ml in drinking water/5 days). All rats were perfused 7 days after the surgery, brains were removed and serial coronal sections were prepared. Immunohistochemistry was used to assess CART peptide expression in paraventricular and supraoptic cells. ADX lowered both the number and percentage of CART-positive cells compared to the sham-operated group, and hormone replacement partially restored the decrease in the CART cell numbers in ADX animals. There were no significant changes in the supraoptic nucleus. Our results suggest a role for CART peptides in the stress response.

Advances in our biological understanding and control of cognitive functioning have not been matched in the behavioral realm. The bio-behavioral approach outlined in this paper helps to close this bio-behavioral gap by aiming for a taxonomic precision in psychology that is comparable to that in physiology. The approach is applied to the phenomenon of group sex differences, and is illustrated by referring to three sets of recently reported findings: (a) interactive effects of nicotine and sex on the trait of cognitive style; (b) sexually dimorphic cognitive style in rats and the involvement of nitric oxide as a potential physiological mechanism and (c) sexually dimorphic phasic arousal change induced by smoking a cigarette following deprivation.

Nicotine exerts its central actions by regulating cationic fluxes through nicotinic acetylcholine receptors (nAChRs). By this effect, the drug likely also modifies events occurring beyond the nAChR, including the regulation of nitric oxide (NO) synthesis. The present study was undertaken to assess the effects of acute and chronic nicotine administration (0.4 mg/kg, s.c.) on levels of NO(-)(2)+NO(-)(3), stable metabolites of NO, in brain regions of male and female rats. Nicotine increased levels of the metabolites, and therefore presumably of NO, with sex differences in the degree of stimulation, the brain regions affected, and the variance between the effects of acute and chronic administration. Prior inhibition of NO synthase eliminated the effect of nicotine in all regions studied. While nicotine appeared to increase NO indirectly via glutamate receptors in the cortex and hippocampus, this was not true of the corpus striatum, where blocking NMDA-type glutamate receptors with MK-801 had no effect. The findings support the view that NO is likely involved in some of the central effects of nicotine.

Nicotine produces dose-dependent enhancement of performance in an active avoidance test, and also increases the levels of NO2- and NO3-, which are stable metabolites of nitric oxide (NO), in various brain regions of rats. On the basis of these two observations, we hypothesized that the beneficial effect of nicotine on learning could result from increased NO in relevant brain regions. We therefore tested active avoidance performance in rats given L-N-omega-nitroarginine (L-NA) to inhibit NO synthetase (NOS) prior to nicotine administration, Male Sprague-Dawley rats received L-NA (30 or 50 mg/kg), nicotine (0.4 mg/kg), saline or combinations of these treatments before learning trials. Rats were also tested on the inclined plane, to assess the possible effects due to impairment of motor function by drug treatments on active avoidance learning. L-NA treatment impaired the acquisition of active avoidance learning, and this defect was partially overcome by the co-administration of nicotine. Nicotine facilitated learning and significantly increased the number of correct responses. The threshold for the effect of NOS inhibition on performance exceeded 30 mg/kg L-NA, whereas 50 mg/kg impaired learning and also eliminated the nicotine-induced enhancement of learning. On the fifth day of learning trials, no facilitation of learning by nicotine was observed in rats receiving either dose of L-NA. Our results suggest that NO is involved in the facilitation of active avoidance learning by nicotine. (C) 2000 Lippincott Williams & Wilkins.

In a water maze (WM), rats employ different and sexually dimorphic behavioral strategies to solve a place-learning task, a test of cognitive/propositional ability. Puberty is the last step in brain development and marks an important phase with regard to sexually dimorphic cognitive performance and behavior. The present study assessed possible sex differences in cognitive style before and after puberty in a WM place-learning task. Since nitric oxide (NO) is implicated in spatial learning and hippocampal function, and since brain NO(-)(2) + NO(-)(3) levels (stable metabolites of NO) display region-specific sex differences in rat brain, NO(-)(2) + NO(-)(3) levels were determined after behavioral testing. The sex-related style difference emerged very clearly but only in the adult rats, which suggests that the female behavioral strategy in the WM place-learning task requires the presence of female sex hormones at puberty. Although NO(-)(2) + NO(-)(3) levels were higher in the adult rats and males compared to prepubertal and female rats, respectively, no significant correlations emerged between brain NO and behavior. The fact that the behavioral sexually dimorphic cognitive-style effect observed here and in previous studies appears to emerge only after puberty suggests that awareness of such postpubertal sex differences may also be important in human educational and therapeutic contexts.

Because of the reported presence of both CART peptide and NOS activity in the same hypothalamic nuclei, their colocalization was examined. Eighteen percent of the neurons in the supraoptic nuclei, and 16% of the neurons in the paraventricular nucleus contained both CART immunoreactivity and NOS activity. Many other neurons in these regions stained for only one marker although they were often close by. Thus, CART peptides and NO may interact in these regions.

Abnormal glutamate metabolism is implied in the pathogenesis of Amyotrophic Lateral Sclerosis (ALS) and cerebrospinal fluid (CSF) glutamate levels appear to be elevated. Since nitric oxide (NO) inhibits glutamate transport, excessive amounts of nitric oxide could underlie the glutamate induced neurotoxicity in ALS. Stable metabolites of NO (NO2- + NO3-) levels were determined in serum and CSF of sporadic ALS patients and control subjects. NO2- + NO3- levels were higher in ALS, in males and in serum samples compared to controls, females and CSF, respectively. Furthermore, while the difference between serum and CSF NO2- + NO3- levels was significant in males (higher in serum) no such difference was observed in females. Our results suggest that nitric oxide may be involved in the pathogenesis of ALS directly or indirectly and in a sexually dimorphic manner.

The effect of sex and nicotine on cognitive style was examined in rats using a water maze task that allows differentiation between cognitive ability and style. During the 12-day acquisition period with the platform in the same location (either visible or hidden) there were no effects or interactions attributable to nicotine and sex, either in terms of learning rate or asymptotic latency. On the final test day the platform was visible and shifted in its location, and on the first trial the new location was proximal to the rats starting position, in contrast to the more distal location of the platform during the previous acquisition days. This platform relocation presented the rats with a choice between two competing cognitive styles: using local visual (look-out) cues vs. navigational cues. Performance on the test day yielded a nicotine x sex interaction, such that only saline-treated female rats showed a clear preference for the perceptual-proximal look-out cognitive style by swimming straight to the newly-relocated visible platform with mean escape latency that approximated the limits of swimming speed. The other three groups did not differ from each other, and preferred navigational cues. The results show that male and female rats use different strategies in problem solving, and that nicotine shifts the female pattern to that of the male.

Francisco Aboitiz*, F. Xavier Castellanos and Ximena Carrasco Departamento de Psiquiatria, Facultad de Medicina, and Centro Interdisciplinario de Neurociencia, Pontificia Universidad Catolica de Chile, Santiago, Chile New York University Child Study Center, Nathan Kline Institute for Psychiatric Research, New York, NY, USA Servicio de Neurologia y Psiquiatria infantil, Hospital Luis Calvo Mackenna Facultad de Medicina, Universidad de Chile, Santiago, Chile

This study tested for sex differences in the effects of chronic nicotine administration and withdrawal on nicotinic acetylcholine receptor binding in brain. Rats received nicotine (0.6 mg/kg, s.c.) or saline once daily for 15 days, and were sacrificed 1 or 20 days after termination of treatment. Saturation studies of nAChR binding were performed using [3H]cytisine as the radioligand in whole brain minus cerebellum taken from animals in the chronic treatment groups and from naive rats. Male but not female rats that received chronic nicotine had higher receptor densities than corresponding control groups; up-regulation of nAChR was not seen 20 days after withdrawal. Furthermore, in groups that showed no up-regulation (controls and rats withdrawn for 20 days), nAChR densities were higher in female rats than males. The findings underscore the importance of sex differences in pharmacological responses as well as in basal neurochemical parameters.

WOS: 000323737700007, PubMed ID: 23912820, Previous animal studies have revealed significant involvement of genetics in nicotine intake; however, the extent of the genetic contribution to this behavior has not been well addressed. We report the first study of nine generations of selection for high and low voluntary nicotine intake in outbred Sprague-Dawley rats. Bidirectional mass selection resulted in progressively greater nicotine consumption in the high nicotine-preferring line but no decrease in nicotine intake in the low nicotine-preferring line across generations. Our estimated realized heritability for high voluntary nicotine intake is 0.26 vs close to zero for low voluntary nicotine intake. In contrast, we found no differences between the lines across generations for saccharine intake. These selected lines may provide useful animal models for identifying susceptibility and resistance genes and variants for controlling voluntary nicotine intake in rodents, although we recognize that more generations of selection of these two lines and independent replication of our selection for high and low nicotine-preferring lines are needed., Ege UniversityEge University [001 BAM 2006]; National Institutes of HealthUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [DA-012844], We are grateful to Dr. Gonca Mola, Merve Evren, and Tuna Nesil and Muzeyyen Ugur for their assistance in data collection and to Professor Qin Zhang of China Agricultural University for calculating inbreeding coefficients for the study. The animal-related study was funded by the Ege University Research Fund (Grant 001 BAM 2006). The analysis of data and preparation of this report were supported in part by National Institutes of Health grant DA-012844 to MDL.

Nicotine is a highly addictive drug and exerts its effect partially through causing dopamine release, thereby increasing intrasynaptic dopamine levels in the brain reward systems. Dopaine D1 receptor (DRD1) mRNAs and receptors are localized in reward-related brain regions, which receive cholinergic input. The aim of this study is to evaluate whether nicotine administration affects the expression of DRD1s, and if so, whether epigenetic mechanisms, such as histone acetylation, are involved. Twenty Male Sprague Dawley rats received nicotine (0.4 mg/kg/day, s.c.) or saline injections for 15 days. After nicotine/saline treatment, rats were perfused with saline; prefrontal cortex (PFC), corpus striatum (STR), and ventral tegmental area (VTA) were dissected. Homogenates were divided into two parts for total RNA isolation and histone H4 acetylation studies. DRD1 mRNA expression was significantly higher in the PFC of the nicotine-treated group compared with controls; similar trends were observed in the VTA and STR. To study epigenetic regulation, the 2kb upstream region of the DRD1 gene promoter was investigated for histone H4 acetylation in PFC samples. After chromatin immunoprecipitation with anti-acetyl histone H4 antibody, we found an increase in histone acetylation by two different primer pairs which amplified the -1365 to -1202 (P 0.005) and -170 to +12 (P 0.05) upstream regions of the DRD1 promoter. Our results suggest that intermittent subcutaneous nicotine administration increases the expression of DRD1 mRNA in the PFC of rats, and this increase may be due to changes in histone H4 acetylation of the 2kb promoter of the DRD1 gene.

To determine the effects of tamoxifen and hormone replacement therapy in order to assess their role in depressive behavior:Different protocols of hormone replacement therapies were administered to surgically ovariectomized rats. Intact rats were used for tamoxifen experiments. Properly assigned control groups were used and cognitive processes were studied on animal models of surgical menopause using the Porsolt forced swim test and locomotor activity experiments.In the tamoxifen experiments, an interaction between treatment and days did not reach statistical significance, but indicated a trend in this direction [F(1,26)=3.557, p = 0.071]. The number of repeated movements significantly decreased after the Porsolt test (F(1,44) = 8.483, P0.006) in the hormone replacement experiments. In the tamoxifen experiments, the number of repeated movements significantly decreased after the Porsolt test (F(1,26) = 74.410, P0.001).While sequential hormone replacement is found to be protective against depression, tamoxifen seems to augment behavioral despair

Nicotine addiction is a complex process that begins with self-administration. Consequently, this process has been studied extensively using animal models. A person is usually not called “smoker” if s/he has smoked for a week or a month in a lifetime; in general, a smoker has been smoking for many years. Furthermore, a smoker has free access to cigarettes and can smoke whenever she/he wants, provided there are no social/legal restraints. Subsequently, in an animal model of tobacco addiction, it will be desirable to expose the animal to free access nicotine for 24 hours/day for many weeks, starting at different stages of development. Oral nicotine self-administration studies in rodents present some important advantages and mimic human smoking nicely. For example, animals are not food deprived, are exposed to nicotine choice for up to 24 hours a day for extended periods, environmental cues and learning does not interfere with self-administration of nicotine. Oral alcohol selfadministration has been used in rodents for over five decades and has contributed significantly to the understanding of alcohol addiction. We provide a review of literature and compare oral alcohol and nicotine intake in rodents. Methodological issues, post ingestional and systemic effects, discrimination and the important influences of taste, genetic vulnerability, sex and age on intake are discussed. The review ends with recommendations for future research on oral self-administration of nicotine.

3H-glutamate (GLU) uptake was measured in hippocampal synaptosomes from rat brain. Addition of sodium nitroprusside (SNP) (Sodium nitroferricyanide), a generator of nitric oxide (NO), produced a time-, temperature-, and dose-dependent inhibition of 3H-GLU uptake. The inhibition was due to changes in both Kd and Vmax of GLU uptake, and it was at least partially reversible upon washing. Addition of reduced hemoglobin (Hb), a substance that binds NO, prevented the SNP-induced depression of uptake. Potassium ferricyanide, a compound similar to SNP, did not cause a reduction in 3H-GLU uptake. Utilization of another generator of NO, S-nitroso-N-acetylpenicillamine (SNAP), produced similar results as did NO itself. Decreases in uptake were also observed in the striatum and cerebellum. Similar treatments did not consistently affect 3H-norepinephrine (NE) uptake, suggesting some selectivity in the NO effect. Thus, the observed inhibition of 3H-GLU uptake appears to be produced by NO, and it may represent a novel type of transynaptic retrograde regulation of transport. If found in vivo, inhibition of uptake activity could also be involved in the toxic effects of NO, the neurotoxicity of glutamate, and other potential neuronal changes associated with NO such as hippocampal long-term potentiation. © 1994 Wiley-Liss, Inc.1

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system characterized by myelin breakdown. The free radical nitric oxide (NO), which is considered to be a major metabolite in immune function and in autoimmune disorders, is among the possible mediators causing the inflammatory reactions in MS. Consequently, NO has been implicated in the pathogenesis of MS and its animal model experimental allergic encephalomyelitis (EAE). In this study, stable metabolites of NO (NO2-+NO3-) levels were determined in sera of MS patients (n=23) and control subjects (n=16). NO2-+NO3- levels were higher in MS patients when compared to control subjects. However, there was not any correlation with serum NO2-+NO3- values and clinical features of the disease such as duration of sickness, the time elapsed from the last attack and EDSS values. Our results imply that nitric oxide may be involved in the pathogenesis of MS although further studies are required to elucidate underlying mechanisms. (C) 2002 Published by Elsevier Science Ltd.

Schizophrenia has been associated with deficits in functional brain lateralization. According to some authors, the reduction of asymmetry could even promote this psychosis. At the same time, schizophrenia is accompanied by a high prevalence of nicotine dependency compared to any other population. This association is very interesting, because sex-dependent effects of smoking in auditory language asymmetries have been reported recently, and the verbal domain is also one major focus in cognitive deficit studies of schizophrenia. Thus, the altered laterality pattern in schizophrenia could, at least in part, result from secondary artefacts due to smoking rather than being a pure cause of the disease itself. To test this hypothesis, the present study examined auditory language lateralization in 67 schizophrenia patients and in 72 healthy controls in a phonemic and an emotional dichotic listening task. Our findings replicate previous research, in that smoking reduces language lateralization in men in phonemic dichotic listening. In addition, we show that smoking also reduces laterality in women in the emotional dichotic listening task. Thus, smoking alters phonemic and emotional language asymmetries differentially for men and women, with a stronger effect for men in the left hemisphere phonemic task, and a stronger effect for women in the right hemisphere emotional task. Together, these findings point towards an effect of smoking which is possibly independent of sex and hemisphere. Importantly, by testing equal numbers of smoking and non-smoking patients and controls, we found no schizophrenia-associated asymmetry effect. Possible neurobiological mechanisms with which smoking may alter auditory microcircuits and thereby diminish left-right differences are discussed.

WOS: 000292408200022, PubMed ID: 21504750, To study individual differences in nicotine preference and intake, male and female rats were given free access to a choice of oral nicotine (10 or 20 mg/L) or water for 24 h/day for periods of at least six weeks, starting at adolescence or adulthood. A total of 341 rats, were used in four different experiments; weight, nicotine intake and total liquid consumption were recorded weekly. Results show that rats can discriminate nicotine from water, can regulate their intake, and that there are readily detected individual differences in nicotine preference. Ward analyses indicated that the animals could be divided into minimum, median and maximum preferring subgroups in all experiments. The effect of saccharine on nicotine intake was also evaluated; although the addition of saccharine increased total intake, rats drank unsweetened nicotine solutions and those with higher preferences for nicotine, preferred nicotine over water with or without saccharine added. Nicotine reduced weight gain and the effect was more pronounced in females than males. The average nicotine consumption of adolescent rats was higher than adults and nicotine exposure during adolescence reduced nicotine intake in adult rats. About half of the rats which had access to nicotine as adolescents and also as adults had a persistent pattern of consumption; the behavior was very stable in the female minimum preferring groups and a much higher ratio of rats sustained their adolescent behavior as adults. The change in preference was more pronounced when there was an interval between adolescent and adult exposure; female rats showed a more stable behavior than males suggesting a greater role for environmental influences on males. In conclusion, marked individual differences were observed in oral nicotine intake as measured in a continuous access 2-bottle choice test. Age and sex of the subjects and previous exposure to nicotine are significant factors which affect preference in rats. (C) 2011 Elsevier Ltd. All rights reserved., Ege UniversityEge University [2006 TIP 008, 2006 BAM 001]; NIHUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USA [DA-03194], The authors would like to thank Dr. Gonca Mola, Merve Ulug Evren, and Tuna Nesil for their assistance in data collection and Dr. Hatice Uluer for statistical evaluation of the results. Supported by Ege University Research Fund Grants 2006 TIP 008, 2006 BAM 001 and NIH grant DA-03194.

NO (nitric oxide) produced in limbic brain regions has important roles in the regulation of autonomic nervous system and HPA axis activity, anxiety, fear learning, long-term memory formation, and depression. NO is synthesized from l-arginine in a reaction catalyzed by nitric oxide synthase (NOS). Neuronal nitric oxide synthase (nNOS), one of the three isoforms of NOS, is synthesized constitutively in nerve cells. Increasing evidence indicates that nNOS expression in the nervous system may be regulated by stress and nicotinic receptors. Furthermore, data obtained from several studies suggest that signaling pathways induced by stress and nicotinic receptors may converge on various signal transduction molecules to regulate nNOS expression in brain. In the present study, we used Western Blot analysis to test the effect of forced swim stress, chronic nicotine administration, and the combined effect of both procedures on nNOS expression in the hippocampus, amygdala and frontal cortex of the male and female rat brain. Basal nNOS levels of the three brain regions examined did not show sex differences. However, forced swim stress and chronic nicotine administration increased nNOS expression in the hippocampus of female rats. When stress and nicotine were applied together, no additional increment was observed. Stress and nicotine did not regulate nNOS expression in the amygdala and the frontal cortex of either sex. Data obtained from the present study indicate that the regulation of stress and nicotine induced-nNOS expression in rat hippocampus shows sexual dimorphism and nNOS expression in the female rat hippocampus increases by nicotine and stress.

Smoking affects a widespread network of neuronal functions by altering the properties of acetylcholinergic transmission. Recent studies show that nicotine consumption affects ascending auditory pathways and alters auditory attention, particularly in men. Here we show that smoking affects language lateralization in a sex-specific way. We assessed brain asymmetries of 90 healthy, right-handed participants using a classic consonant-vowel syllable dichotic listening paradigm in a 2×3 experimental design with sex (male, female) and smoking status (non-smoker, light smoker, heavy smoker) as between-subject factors. Our results revealed that male smokers had a significantly less lateralized response pattern compared to the other groups due to a decreased response rate of their right ear. This finding suggests a group-specific impairment of the speech dominant left hemisphere. In addition, decreased overall response accuracy was observed in male smokers compared to the other experimental groups. Similar adverse effects of smoking were not detected in women. Further, a significant negative correlation was detected between the severity of nicotine dependency and response accuracy in male but not in female smokers. Taken together, these results show that smoking modulates functional brain lateralization significantly and in a sexually dimorphic manner. Given that some psychiatric disorders have been associated with altered brain asymmetries and increased smoking prevalence, nicotinergic effects need to be specifically investigated in this context in future studies.