Alcohol self-administration and nicotine withdrawal alter biomarkers of stress and inflammation and prefrontal cortex changes in Gß subunits.

Background: Although alcohol and nicotine are often used together, the biological consequences of these substances are not well understood. Identifying shared targets will inform cessation pharmacotherapies and provide a deeper understanding of how co-use of alcohol and nicotine impacts health, including biomarkers of stress and inflammation.Objective: We examined the effects of nicotine exposure and withdrawal on alcohol self-administration (SA), stress and inflammatory biomarkers, and a G-protein coupled receptor subunit (Gß) in brain areas associated with drug use.Methods: Male rats were trained to SA alcohol and then received a nicotine pump (n = 7-8 per group). We assessed alcohol intake for 12 days during nicotine exposure and then following pump removal to elicit withdrawal. After the behavioral studies, we assessed plasma leptin, corticosterone, and interleukin-1ß (IL-1ß), and Gß protein expression in the amygdala, nucleus accumbens (NAc), and prefrontal cortex (PFC).Results: Nicotine exposure or withdrawal did not alter alcohol intake (p > .05). Alcohol and nicotine withdrawal elevated corticosterone levels (p = .015) and decreased Gß levels in the PFC (p = .004). In the absence of nicotine, alcohol SA suppressed IL-1ß levels (p = .039). Chronic exposure to nicotine or withdrawal during alcohol SA did not alter leptin levels or Gß expression in the amygdala or NAc (p's > .05).Conclusions: The combination of alcohol SA and nicotine withdrawal produced a persistent increase in stress biomarkers and a suppression in Gß expression in the PFC, providing an important first step toward understanding the common biological mechanisms of alcohol/nicotine misuse.

Amino acid systems in the interpeduncular nucleus are altered in a sex-dependent manner during nicotine withdrawal.

Prior work in male rodents established that the medial habenula-interpeduncular nucleus (MHb-IPN) pathway modulates nicotine withdrawal. Specifically, withdrawal severity has been closely associated with inhibitory tone in the IPN via interneurons that release γ-aminobutyric acid (GABA). Inhibitory tone in the IPN is regulated by projections from the MHb that co-release glutamate and acetylcholine. Within the IPN, inhibitory tone is also regulated via corticotropin-releasing factor type 1 (CRF1) receptors that control GABA release from local interneurons. This study extends previous work by comparing sex differences in GABA, glutamate, as well serotonin levels in the IPN during precipitated nicotine withdrawal. Sex differences in withdrawal-induced neurochemical effects were also compared following systemic administration of a CRF1 receptor antagonist. The results revealed that there were no group differences in serotonin levels in the IPN. A major finding was that females displayed a larger withdrawal-induced increases in GABA levels in the IPN than males. Also, withdrawal increased IPN glutamate levels in a similar manner in females and males. Blockade of CRF1 receptors produced a larger suppression of the withdrawal-induced increases in GABA levels in the IPN of females versus males, an effect that was likely related to the robust increase in glutamate following administration of the CRF1 receptor antagonist in females. These data suggest that amino acid systems in the IPN modulate sex differences in the behavioral effects of nicotine withdrawal. Furthermore, our data imply that medications that target stress-induced activation of the IPN may reduce withdrawal severity, particularly in females.

Exposure to Nicotine Vapor Produced by an Electronic Nicotine Delivery System Causes Short-Term Increases in Impulsive Choice in Adult Male Rats.

INTRODUCTION: Traditional cigarette use influences cost-benefit decision making by promoting impulsive choice. However, the impact of nicotine exposure via electronic nicotine delivery systems on impulsivity remains unclear. Hence, the present study examined the short- and long-term effects of nicotine vapor on impulsive choice. METHODS: Twenty-four adult male rats were trained in the delay discounting task to choose between small immediate food rewards and large delayed food rewards. After 24 days of training in the task rats were exposed to vapor containing either 0, 12, or 24 mg/mL of nicotine for 10 days. To validate inhalation of nicotine vapor serum cotinine levels were analyzed on exposure days 1, 5, and 10 using enzyme-linked immunosorbent assay. Following vapor exposure, rats were retrained in the discounting task until rats displayed stable responding and the effects of nicotine vapor on choice preference were assessed. RESULTS: Rats exposed to 12 and 24 mg/mL nicotine vapor displayed higher serum cotinine levels than control rats exposed to 0 mg/mL vapor. There were no differences in impulsive choice between any vapor exposure groups when tested 15 days after exposure, across 6 days of stable responding, suggesting that nicotine vapor does not have long lasting effects on impulsive choice. Interestingly, a subsequent nicotine vapor challenge revealed short-term increases in impulsive choice immediately following a single exposure to 24 mg/mL nicotine vapor, relative to choice preference immediately following exposure to 0 mg/mL vapor. CONCLUSIONS: These results suggest that exposure to nicotine vapor causes immediate, short-term increases in impulsive choice. IMPLICATIONS: E-cigarette use is increasing at an alarming rate, particularly among adolescents and young adults. This is concerning given the lack of research into the effects of nicotine vapor exposure on the brain and behavior. The present study describes a viable rodent model of human e-cigarette use and suggests that exposure to nicotine vapor produces short-term increases in impulsive choice.

Insulin restores the neurochemical effects of nicotine in the mesolimbic pathway of diabetic rats.

This study examined whether insulin modulates the neurochemical effects of nicotine in the mesolimbic pathway of diabetic rats. The rats received vehicle or streptozotocin (STZ) to induce hypoinsulinemia. A subset of STZ-treated rats was implanted with insulin pellets that rapidly normalized glucose levels. Two-weeks later, dialysis probes were implanted into the nucleus accumbens (NAc) and ipsilateral ventral tegmental area (VTA). The next day, dialysate samples were collected during baseline and then following systemic administration of nicotine. Samples were also collected following intra-VTA administration of the gamma-aminobutyric acid (GABA)A receptor antagonist, bicuculline. Dopamine, GABA, glutamate, and acetylcholine (ACh) levels were assessed using liquid chromatography/mass spectrometry (LC/MS). The results revealed that vehicle-treated rats displayed a nicotine-induced increase in NAc dopamine levels. In contrast, STZ-treated rats did not display any changes in NAc dopamine following nicotine administration, an effect that was likely related to a concomitant increase in GABA and decrease in glutamate levels in both the NAc and VTA. Intra-VTA administration of bicuculline increased NAc dopamine in vehicle-treated rats, and this effect was absent in STZ-treated rats. Vehicle-treated rats displayed a nicotine-induced increase in ACh levels in the NAc (but not VTA), an effect that was lower in the NAc of STZ-treated rats. Insulin supplementation normalized the neurochemical effects of nicotine in the NAc and VTA of STZ-treated rats, suggesting that insulin modulates the neurochemical effects of nicotine in the mesolimbic pathway of diabetic rats.

Amino acid modulation of dopamine in the nucleus accumbens mediates sex differences in nicotine withdrawal.

The aversive effect of nicotine withdrawal is greater in female versus male rats, and we postulate that this sex difference is mediated in the nucleus accumbens (NAc). Nicotine withdrawal induces decreases in NAc dopamine and increases in acetylcholine (ACh) levels in male rats. To our knowledge, these neurochemical markers of nicotine withdrawal have not been compared in female versus male rats. Given the role of amino acids in modulating NAc dopaminergic and cholinergic transmission, concomitant measures of gamma-aminobutyric acid (GABA) and glutamate levels were also compared across sex. Rats received continuous nicotine exposure for 14 days, and then NAc dialysate was collected during baseline and following administration of the nicotinic receptor antagonist mecamylamine to precipitate withdrawal. Chronic nicotine exposure was associated with larger increases in baseline dopamine, GABA and glutamate levels in the NAc of female versus male rats, whereas baseline ACh was only increased in male rats. During withdrawal, both sexes displayed equivalent increases in NAc ACh levels. As expected, male rats displayed decreases in dopamine, coupled with increases in GABA and decreases in glutamate levels, suggesting the possibility of increased inhibitory tone in the NAc during withdrawal. Relative to males, female rats displayed larger decreases in NAc dopamine and related increases in GABAergic transmission. As female rats also showed elevated glutamate levels that persist during withdrawal, it is suggested that sex differences may arise from increased glutamatergic drive of inhibitory tone in the NAc. The findings provide a potential mechanism whereby the aversive effects of nicotine withdrawal are enhanced in female rats.

Nicotine withdrawal increases stress-associated genes in the nucleus accumbens of female rats in a hormone-dependent manner.

INTRODUCTION: Previous work led to our hypothesis that sex differences produced by nicotine withdrawal are modulated by stress and dopamine systems in the nucleus accumbens (NAcc). We investigated our hypothesis by studying intact females to determine whether the mechanisms that promote withdrawal are ovarian-hormone mediated. METHODS: Female rats were ovariectomized (OVX) or received sham surgery (intact) on postnatal day (PND 45-46). On PND 60, they received sham surgery (controls) or were prepared with nicotine pumps. Fourteen days later, half of the rats had their pumps removed (nicotine withdrawal) and the other half received sham surgery (nicotine exposure). Twenty-four hours later, the rats were tested for anxiety-like behavior using the elevated plus maze and light/dark transfer procedures. The NAcc was then dissected for analysis of several genes related to stress (CRF, UCN, CRF-R1, CRF-R2, CRF-BP, and Arrb2) or receptors for dopamine (Drd1 and Drd2) and estradiol (Esr2). RESULTS: During withdrawal, intact females displayed an increase in anxiety-like behavior in both tests and CRF, UCN, and Drd1 gene expression. During nicotine exposure, intact females displayed a decrease in CRF-R1, CRF-R2, Drd3, and Esr2 gene expression and an increase in CRF-BP. This pattern of results was absent in OVX females. CONCLUSIONS: Nicotine withdrawal produced an increase in anxiety-like behavior and stress-associated genes in intact females that is distinct from changes produced by nicotine exposure. The latter effects were absent in OVX females, suggesting that stress produced by withdrawal is ovarian-hormone mediated. These findings have important implications towards understanding tobacco use liability among females.

Cholinergic transmission during nicotine withdrawal is influenced by age and pre-exposure to nicotine: implications for teenage smoking.

Adolescence is a unique period of development characterized by enhanced tobacco use and long-term vulnerability to neurochemical changes produced by adolescent nicotine exposure. In order to understand the underlying mechanisms that contribute to developmental differences in tobacco use, this study compared changes in cholinergic transmission during nicotine exposure and withdrawal in naïve adult rats compared to (1) adolescent rats and (2) adult rats that were pre-exposed to nicotine during adolescence. The first study compared extracellular levels of acetylcholine (ACh) in the nucleus accumbens (NAc) during nicotine exposure and precipitated withdrawal using microdialysis procedures. Adolescent (postnatal day, PND, 28-42) and adult rats (PND60-74) were prepared with osmotic pumps that delivered nicotine for 14 days (adolescents 4.7 mg/kg/day; adults 3.2 mg/kg/day; expressed as base). Another group of adults was exposed to nicotine during adolescence and then again in adulthood (pre-exposed adults) using similar methods. Control rats received a sham surgery. Following 13 days of nicotine exposure, the rats were implanted with microdialysis probes in the NAc. The following day, dialysis samples were collected during baseline and following systemic administration of the nicotinic receptor antagonist mecamylamine (1.5 and 3.0 mg/kg, i.p.) to precipitate withdrawal. A second study compared various metabolic differences in cholinergic transmission using the same treatment procedures as the first study. Following 14 days of nicotine exposure, the NAc was dissected and acetylcholinesterase (AChE) activity was compared across groups. In order to examine potential group differences in nicotine metabolism, blood plasma levels of cotinine (a nicotine metabolite) were also compared following 14 days of nicotine exposure. The results from the first study revealed that nicotine exposure increased baseline ACh levels to a greater extent in adolescent versus adult rats. During nicotine withdrawal, ACh levels in the NAc were increased in a similar manner in adolescent versus adult rats. However, the increase in ACh that was observed in adult rats experiencing nicotine withdrawal was blunted in pre-exposed adults. These neurochemical effects do not appear to be related to nicotine metabolism, as plasma cotinine levels were similar across all groups. The second study revealed that nicotine exposure increased AChE activity in the NAc to a greater extent in adolescent versus adult rats. There was no difference in AChE activity in pre-exposed versus naïve adult rats. In conclusion, our results suggest that nicotine exposure during adolescence enhances baseline ACh in the NAc. However, the finding that ACh levels were similar during withdrawal in adolescent and adult rats suggests that the enhanced vulnerability to tobacco use during adolescence is not related to age differences in withdrawal-induced increases in cholinergic transmission. Our results also suggest that exposure to nicotine during adolescence suppresses withdrawal-induced increases in cholinergic responses during withdrawal. Taken together, this report illustrates important short- and long-term changes within cholinergic systems that may contribute to the enhanced susceptibility to tobacco use during adolescence.

Female rats display enhanced rewarding effects of ethanol that are hormone dependent.

BACKGROUND: Ethanol (EtOH) abuse is a major health and economic concern, particularly for females who appear to be more sensitive to the rewarding effects of EtOH. This study compared sex differences to the rewarding and aversive effects of EtOH using place-conditioning procedures in rats. METHODS: Separate groups of adult (male, female, ovariectomized [OVX] female) and adolescent (male and female) rats received EtOH (0, 0.5, 1.0, 2.0, or 2.5 g/kg, intraperitoneal) and were confined to their initially nonpreferred side of our conditioning apparatus for 30 minutes. On alternate days, they received saline and were confined to the other side. Following 5 drug pairings, the rats were retested for preference behavior. Separate cohorts of the same groups of rats were injected with a similar dose range of EtOH, and blood EtOH levels (BELs) were compared 30 minutes later. RESULTS: EtOH produced rewarding or aversive effects in a dose-dependent manner. An intermediate dose of EtOH (1.0 g/kg) produced rewarding effects in adult female, but not in male or OVX female rats, suggesting that ovarian hormones facilitate the rewarding effects of EtOH. Similarly, this intermediate dose of EtOH produced rewarding effects in adolescent female, but not in male rats. The highest dose of EtOH (2.5 g/kg) produced aversive effects that were similar across all adult groups. However, the aversive effects of EtOH were lower in adolescents than adults, suggesting that adolescents are less sensitive to the aversive effects of EtOH. The aversive effects of EtOH did not vary across the estrous cycle in intact adult females. There were also no group differences in BELs, suggesting that our results are not related to EtOH metabolism. CONCLUSION: Our results in rats suggest that human females may be more vulnerable to EtOH abuse due to enhanced rewarding effects of this drug that are mediated by the presence of ovarian hormones.

Enhanced nicotine self-administration and suppressed dopaminergic systems in a rat model of diabetes.

Patients with diabetes display a heightened propensity to use tobacco; however, it is unclear whether they experience enhanced rewarding effects of nicotine. Thus, this study examined the reinforcing effects of nicotine in a rodent model of diabetes involving administration of streptozotocin (STZ), a drug that is toxic to pancreatic insulin-producing cells. The first study compared STZ- and vehicle-treated rats that had 23-hour access to intravenous self-administration (IVSA) of nicotine or saline and concomitant access to food and water. In order to examine the contribution of dopamine to our behavioral effects, dopamine transporter (DAT), D1 and D2 receptor levels were compared in the nucleus accumbens (NAc) following 10 days of nicotine or saline IVSA. Dopamine levels in the NAc were also compared following nicotine administration. Lastly, nicotine metabolism and dose-dependent effects of nicotine IVSA were assessed. The results revealed that STZ-treated rats displayed enhanced nicotine intake and a robust increase in food and water intake relative to controls. Protein analysis revealed an increase in DAT and a decrease in D1 receptor levels in the NAc of STZ- versus vehicle-treated rats regardless of IVSA condition. STZ-treated rats also displayed suppressed NAc dopamine levels during baseline and in response to nicotine. STZ treatment did not alter our assessment of nicotine metabolism. Furthermore, STZ treatment increased nicotine IVSA in a dose-dependent manner. Our findings suggest that STZ-treatment increased the rewarding effects of nicotine. This suggests that strong reinforcing effects of nicotine may contribute to greater tobacco use in patients with diabetes.

RECORD, a high-throughput, customizable system that unveils behavioral strategies leveraged by rodents during foraging-like decision-making.

Translational studies benefit from experimental designs where laboratory organisms use human-relevant behaviors. One such behavior is decision-making, however studying complex decision-making in rodents is labor-intensive and typically restricted to two levels of cost/reward. We design a fully automated, inexpensive, high-throughput framework to study decision-making across multiple levels of rewards and costs: the REward-COst in Rodent Decision-making (RECORD) system. RECORD integrates three components: 1) 3D-printed arenas, 2) custom electronic hardware, and 3) software. We validated four behavioral protocols without employing any food or water restriction, highlighting the versatility of our system. RECORD data exposes heterogeneity in decision-making both within and across individuals that is quantifiably constrained. Using oxycodone self-administration and alcohol-consumption as test cases, we reveal how analytic approaches that incorporate behavioral heterogeneity are sensitive to detecting perturbations in decision-making. RECORD is a powerful approach to studying decision-making in rodents, with features that facilitate translational studies of decision-making in psychiatric disorders.

Effectiveness of pharmacotherapies for diabetes on nicotine, food, and water intake in insulin-resistant rats.

The intersectionality between diabetes medications and nicotine consumption was assessed in female and male rats. Briefly, the rats were fed a high-fat diet (HFD) or regular diet (RD) for 4 weeks. Then separate groups received vehicle or a low dose of streptozotocin (STZ; 25 mg/kg). Three days later, insulin resistance was assessed by measuring plasma glucose levels for 180 min following an injection of insulin (0.75 U/kg). The rats were then prepared with jugular catheters, and they were given 23 h access to nicotine intravenous self-administration (IVSA) in 4 days cycles with 3 days of forced abstinence in their home cages where they consumed their respective diet. During the IVSA sessions, operant responses for food and water and changes in body weight were recorded. Prior to administration of the pharmacotherapies, the rats were given access to two doses of nicotine (0.015 then 0.03 mg/kg for the remainder of the study). Then, daily injections of the pharmacotherapies were given at the onset of dark cycle (6 p.m.) in the following order: 1) dapagliflozin (3.0 then 10.0 mg/kg), 2) insulin (0.75 U/kg twice), and 3) bromocriptine (3.0 then 10.0 mg/kg). The results suggest that our HFD+STZ regiment induced insulin resistance in female and male rats. Also, the HFD-fed rats displayed higher nicotine intake than RD controls, regardless of sex. Administration of insulin, but not dapagliflozin or bromocriptine, normalized nicotine intake in HFD-fed rats to control levels. These results have clinical implications regarding the potential efficacy of insulin to control excessive nicotine intake in persons with diabetes.

Sex and diet-dependent gene alterations in human and rat brains with a history of nicotine exposure.

Introduction: Chronic nicotine exposure induces changes in the expression of key regulatory genes associated with metabolic function and neuronal alterations in the brain. Many bioregulatory genes have been associated with exposure to nicotine, but the modulating effects of sex and diet on gene expression in nicotine-exposed brains have been largely unexplored. Both humans and rodents display motivation for nicotine use and the emergence of withdrawal symptoms during abstinence. Research comparing pre-clinical models with human subjects provides an important opportunity to understand common biomarkers of the harmful effects of nicotine as well as information that may help guide the development of more effective interventions for nicotine cessation. Methods: Human postmortem dorsolateral prefrontal cortex (dLPFC) tissue BA9 was collected from female and male subjects, smokers and non-smokers (N = 12 per group). Rat frontal lobes were collected from female and male rats that received a regular diet (RD) or a high-fat diet (HFD) (N = 12 per group) for 14 days following implantation of a osmotic mini-pump (Alzet) that delivered nicotine continuously. Controls (control-s) received a sham surgical procedure. RNA was extracted from tissue from human and rat samples and reversed-transcribed to cDNA. Gene expression of CHRNA10 (Cholinergic receptor nicotinic alpha 10), CERKL (Ceramide Kinase-Like), SMYD1 (SET and MYD Domin Containing 1), and FA2H (Fatty Acid 2-Hydrolase) in humans was compared to rats in each subset of groups and quantified by qPCR methods. Additionally, protein expression of FA2H was analyzed by immunohistochemistry (IHC) in human dLPFC. Results: Humans with a history of smoking displayed decreased CHRNA10 (p = 0.0005), CERKL (p ≤ 0.0001), and SMYD1 (p = 0.0005) expression and increased FA2H (p = 0.0097) expression compared to non-smokers (p < 0.05). Similar patterns of results were observed in nicotine exposed vs. control rats. Interestingly, sex-related differences in gene expression for CERKL and FA2H were observed. In addition, ANCOVA analysis showed a significant effect of nicotine in a sex-different manner, including an increase in CERKL in male and female rats with RD or HFD. In rats exposed to an HFD, FA2H gene expression was lower in nicotine-treated rats compared to RD rats treated with nicotine. Protein expression of FA2H (p = 0.001) by IHC was significantly higher in smokers compared to non-smokers. Conclusion: These results suggest that a history of long-term nicotine exposure in humans alters the expression of sphingolipid metabolism-related (CERKL, SMYD1, and FA2H) and neuronal (CHRNA10) marker genes similarly as compared to rats. Sex- and diet-dependent differences appear in nicotine-exposed rats, critical in regulating sphingolipid metabolism and nicotinic acetylcholine receptors. This research enhances the construct validity of rat models of nicotine usage by showing a similar pattern of changes in gene expression in human subjects with a smoking history.

Withdrawal from repeated nicotine vapor exposure increases somatic signs of physical dependence, anxiety-like behavior, and brain reward thresholds in adult male rats.

Nicotine vapor consumption via electronic nicotine delivery systems has increased over the last decade. While prior work has shed light on the health effects of nicotine vapor inhalation, its unique effects on the brain and behavior have not been thoroughly explored. In this study we assessed markers of withdrawal following 14 days of nicotine vapor exposure. For Experiment 1, 21 adult male rats were exposed to ambient air or 6, 12, or 24 mg/mL nicotine vapor for 14 consecutive days. Following exposure on day 14, rats were injected with the nicotinic receptor antagonist mecamylamine (3.0 mg/mL) and assessed for somatic withdrawal signs and anxiety-like behavior in the elevated plus maze. For Experiment 2, 12 adult male rats were tested for intracranial self-stimulation (ICSS) immediately following exposure to vehicle vapor (50%/50%, vegetable glycerin/propylene glycol) or 24 mg/mL nicotine vapor, for 14 consecutive days. ICSS behavior was assessed for an additional 14 days, following cessation of repeated vapor exposure. Results reveal that rats with repeated nicotine vapor exposure display an increase in behavioral indicators of withdrawal following injection of mecamylamine (precipitated withdrawal). Additionally, increases in ICSS stimulation thresholds, indicative of reduced brain reward sensitivity, persist following cessation of repeated nicotine vapor exposure (spontaneous withdrawal). These data suggest that repeated e-cigarette use leads to nicotine dependence and withdrawal that affects behavior and brain reward function. Further characterization of the health effects of nicotine vapor is necessary to improve treatment strategies for nicotine use disorder and public health policies related to novel nicotine delivery systems.

Adolescent rats are resistant to adaptations in excitatory and inhibitory mechanisms that modulate mesolimbic dopamine during nicotine withdrawal.

Adolescent smokers report enhanced positive responses to tobacco and fewer negative effects of withdrawal from this drug than adults, and this is believed to propel higher tobacco use during adolescence. Differential dopaminergic responses to nicotine are thought to underlie these age-related effects, as adolescent rats experience lower withdrawal-related deficits in nucleus accumbens (NAcc) dopamine versus adults. This study examined whether age differences in NAcc dopamine during withdrawal are mediated by excitatory or inhibitory transmission in the ventral tegmental area (VTA) dopamine cell body region. In vivo microdialysis was used to monitor extracellular levels of glutamate and gamma-aminobutyric acid (GABA) in the VTA of adolescent and adult rats experiencing nicotine withdrawal. In adults, nicotine withdrawal produced decreases in VTA glutamate levels (44% decrease) and increases in VTA GABA levels (38% increase). In contrast, adolescents did not exhibit changes in either of these measures. Naïve controls of both ages did not display changes in NAcc dopamine, VTA glutamate, or VTA GABA following mecamylamine. These results indicate that adolescents display resistance to withdrawal-related neurochemical processes that inhibit mesolimbic dopamine function in adults experiencing nicotine withdrawal. Our findings provide a potential mechanism involving VTA amino acid neurotransmission that modulates age differences during withdrawal.

Sex and age differences in approach behavior toward a port that delivers nicotine vapor.

The goal of our laboratory is to study the mechanisms that promote nicotine use, particularly in vulnerable populations. To more closely mimic human use patterns, the present study employed nicotine vapor methods involving passive exposure for 14 days in adolescent and adult female and male rats. Age and sex differences in approach behavior (nosepokes) were assessed in a port that delivered nicotine plumes on Day 1 and 14 of our exposure regimen. Controls received ambient air in exposure chambers. After the final session, rats received a nicotinic receptor antagonist to precipitate withdrawal. Then, physical signs, anxiety-like behavior, and plasma levels of cotinine (a nicotine metabolite) were assessed. Over time, females displayed a larger increase in approach behavior to the nicotine port than males, an effect that was larger in adolescents. Nosepoke responses in adolescent females were correlated with anxiety-like behavior, but not physical signs of withdrawal. Adolescents gained more weight than adults regardless of treatment, and the weight gain was larger in male adolescents. Female adolescents also displayed the highest levels of cotinine than all other groups. These findings suggest that nicotine vapor produces greater motivational effects in adolescent females as compared to their adult and male counterparts.

Early-life adversity increases anxiety-like behavior and modifies synaptic protein expression in a region-specific manner.

Early-life adversity (ELA) can induce persistent neurological changes and increase the risk for developing affective or substance use disorders. Disruptions to the reward circuitry of the brain and pathways serving motivation and emotion have been implicated in the link between ELA and altered adult behavior. The molecular mechanisms that mediate the long-term effects of ELA, however, are not fully understood. We examined whether ELA in the form of neonatal maternal separation (MatSep) modifies behavior and synaptic protein expression in adults. We hypothesized that MatSep would affect dopaminergic and glutamatergic signaling and enhance sensitivity to methamphetamine (Meth) reward or increase anxiety. Male Wistar rats were subjected to MatSep for 180 min/d on postnatal days (PND) 2-14 and allowed to grow to adulthood (PND 60) with no further manipulation. The hippocampus (Hipp), medial prefrontal cortex (mPFC), nucleus accumbens (NAc), and caudate putamen (CPu) were isolated from one subgroup of animals and subjected to Western blot and protein quantitation for tyrosine hydroxylase (TH), α-synuclein (ALPHA), NMDA receptor (NMDAR), dopamine receptor-1 (D1) and -2 (D2), dopamine transporter (DAT), and postsynaptic density 95 (PSD95). Separate group of animals were tested for anxiety-like behavior and conditioned place preference (CPP) to Meth at 0.0, 0.1, and 1.0 mg/kg doses. MatSep rats displayed an increase in basal levels of anxiety-like behavior compared to control animals. MatSep rats also demonstrated CPP to Meth, but their responses did not differ significantly from controls at any drug dose. Increased NMDAR, D2, and ALPHA expression was observed in the NAc and CPu following MatSep; D2 and ALPHA levels were also elevated in the mPFC, along with DAT. MatSep rats had reduced D1 expression in the mPFC and Hipp, with the Hipp also showing a reduction in D2. Only the CPu showed elevated TH and decreased DAT expression levels. No significant changes were found in PSD95 expression in MatSep rats. In conclusion, ELA is associated with long-lasting and region-specific changes in synaptic protein expression that diminish dopamine neurotransmission and increase anxiety-like behavior in adults. These findings illustrate potential mechanisms through which ELA may increase vulnerability to stress-related illness.

Converging vulnerability factors for compulsive food and drug use.

Highly palatable foods and substance of abuse have intersecting neurobiological, metabolic and behavioral effects relevant for understanding vulnerability to conditions related to food (e.g., obesity, binge eating disorder) and drug (e.g., substance use disorder) misuse. Here, we review data from animal models, clinical populations and epidemiological evidence in behavioral, genetic, pathophysiologic and therapeutic domains. Results suggest that consumption of highly palatable food and drugs of abuse both impact and conversely are regulated by metabolic hormones and metabolic status. Palatable foods high in fat and/or sugar can elicit adaptation in brain reward and withdrawal circuitry akin to substances of abuse. Intake of or withdrawal from palatable food can impact behavioral sensitivity to drugs of abuse and vice versa. A robust literature suggests common substrates and roles for negative reinforcement, negative affect, negative urgency, and impulse control deficits, with both highly palatable foods and substances of abuse. Candidate genetic risk loci shared by obesity and alcohol use disorders have been identified in molecules classically associated with both metabolic and motivational functions. Finally, certain drugs may have overlapping therapeutic potential to treat obesity, diabetes, binge-related eating disorders and substance use disorders. Taken together, data are consistent with the hypotheses that compulsive food and substance use share overlapping, interacting substrates at neurobiological and metabolic levels and that motivated behavior associated with feeding or substance use might constitute vulnerability factors for one another. This article is part of the special issue on 'Vulnerabilities to Substance Abuse'.

Female rats display greater nicotine withdrawal-induced cellular activation of a central portion of the interpeduncular nucleus versus males: A study of Fos immunoreactivity within provisionally assigned interpeduncular subnuclei.

BACKGROUND: The interpeduncular nucleus (>1840) (IPN) has been shown to modulate the behavioral effects of nicotine withdrawal in male rodents. To date, the contribution of this brain structure to sex differences in withdrawal is largely unexplored. METHODS: This study compared neuronal activation, as reported by observable Fos expression in the IPN of nicotine-dependent female and male rats experiencing withdrawal. We provisionally localized the Fos-expressing cells to certain IPN subnuclei within Swanson's standardized brain atlas (2018). Adult female and male rats were prepared with a pump that delivered nicotine (3.2 mg/kg/day; base) continuously. Controls received a sham surgery. Fourteen days later, the rats received administration of saline or the nicotinic receptor antagonist, mecamylamine (3.0 mg/kg; salt), and physical signs and anxiety-like behavior were assessed. The rats were then euthanized and brain sections containing the IPN were processed for Fos immunofluorescence to infer the possible IPN subnuclei displaying differential activation between sexes. RESULTS: Both female and male rats displayed withdrawal-induced Fos expression within the IPN. Compared to males, female rats displayed greater numbers of withdrawal-induced Fos-positive cells within a circumscribed portion of the IPN that may fall within the cytoarchitectural boundaries of the central subnucleus (>1840) (IPNc). The withdrawal-induced activation of the IPN was correlated with negative affective states in females, but not males. CONCLUSION: These data suggest that a centrally located group of IPN cells, presumably situated partly or completely within the IPNc, play a role in modulating sex differences in negative affective states produced by withdrawal.

The emergence of insulin resistance following a chronic high-fat diet regimen coincides with an increase in the reinforcing effects of nicotine in a sex-dependent manner.

The present study assessed the sex-dependent effects of insulin resistance on the reinforcing effects of nicotine. Female and male rats received a chronic high-fat diet (HFD) or regular diet (RD) for 8 weeks. A subset of rats then received vehicle or a dose of streptozotocin (STZ; 25 mg/kg) that induces insulin resistance. To assess insulin resistance, glucose levels were measured 15, 30, 60, 120, and 180 min after an insulin injection (0.75 U/kg). Nine days later, the rats were given extended access to intravenous self-administration (IVSA) of nicotine (0.015, 0.03, 0.06 mg/kg) in an operant box where they consumed their respective diet ad libitum and performed responses for water deliveries. Each nicotine dose was delivered for 4 days with 3 intermittent days of abstinence in their home cage. The day after the last IVSA session, physical signs were compared following administration of mecamylamine (3.0 mg/kg) to precipitate nicotine withdrawal. The results revealed that there were no changes in insulin resistance or nicotine intake in HFD alone rats regardless of sex. Insulin resistance was observed in HFD-fed rats that received STZ, and the magnitude of this effect was greater in males versus females. Our major finding was that nicotine intake was greater among HFD + STZ female rats as compared to males. Lastly, the physical signs of withdrawal were similar across all groups. Our results suggest that females diagnosed with disorders that disrupt insulin signaling, such as diabetes may be at risk of greater vulnerability to nicotine use due to enhanced reinforcing effects of this drug.

Protracted withdrawal from alcohol and drugs of abuse impairs long-term potentiation of intrinsic excitability in the juxtacapsular bed nucleus of the stria terminalis.

The juxtacapsular bed nucleus of the stria terminalis (jcBNST) is activated in response to basolateral amygdala (BLA) inputs through the stria terminalis and projects back to the anterior BLA and to the central nucleus of the amygdala. Here we show a form of long-term potentiation of the intrinsic excitability (LTP-IE) of jcBNST neurons in response to high-frequency stimulation of the stria terminalis. This LTP-IE, which was characterized by a decrease in the firing threshold and increased temporal fidelity of firing, was impaired during protracted withdrawal from self-administration of alcohol, cocaine, and heroin. Such impairment was graded and was more pronounced in rats that self-administered amounts of the drugs sufficient to maintain dependence. Dysregulation of the corticotropin-releasing factor (CRF) system has been implicated in manifestation of protracted withdrawal from dependent drug use. Administration of the selective corticotropin-releasing factor receptor 1 (CRF(1)) antagonist R121919 [2,5-dimethyl-3-(6-dimethyl-4-methylpyridin-3-yl)-7-dipropylamino-pyrazolo[1,5-a]pyrimidine)], but not of the CRF(2) antagonist astressin(2)-B, normalized jcBNST LTP-IE in animals with a history of alcohol dependence; repeated, but not acute, administration of CRF itself produced a decreased jcBNST LTP-IE. Thus, changes in the intrinsic properties of jcBNST neurons mediated by chronic activation of the CRF system may contribute to the persistent emotional dysregulation associated with protracted withdrawal.