Impact of tobacco smoke constituents on nicotine-seeking behavior in adolescent and adult male rats.

Introduction: Given the rapid increase in teen vaping over recent years it is critical to understand mechanisms underlying addiction and relapse to tobacco use at this age. To evaluate the role of non-nicotine constituents in cigarette smoke, our lab has previously established a model of intravenous self-administration of aqueous cigarette smoke extract (CSE). We now compare the sensitivity of male adolescent and adult rats who have self-administered CSE or nicotine to reinstatement with the pharmacological stressor, yohimbine, with and without cues. Methods: Adolescents and adults, aged postnatal day (P) 34 and 84, were tested for the effect of yohimbine (0-2.5 mg/kg) on plasma corticosterone levels to establish a dose that was an effective stressor at both ages. Separate groups of animals were trained to lever press for food before beginning 1-hour drug self-administration sessions for nicotine or CSE (15 µg/kg/infusion nicotine content). Once stable responding was reached, drug was removed, and behavior extinguished. Drug-seeking behavior was reinstated with yohimbine, cues, or a combination of yohimbine and cues. Results: Although adolescents and adults showed different dose-responses for yohimbine-induced corticosterone release, a dose of 2.5 mg/kg increased stress hormone levels at both ages. Whereas both ages displayed similar responding for CSE and nicotine, adolescents self-administered more CSE and nicotine as compared to adults. Cues and cues + stress reinstated responding to a greater extent in animals that had self-administered CSE, regardless of age. Discussion: These findings suggest that non-nicotine tobacco smoke constituents influence later but not earlier stages of addiction in both adolescent and adult male rats.

Chronic exposure to cigarette smoke extract increases nicotine withdrawal symptoms in adult and adolescent male rats.

The aim of the current study was to determine whether non-nicotine constituents of cigarette smoke contribute to nicotine dependence in adolescent and adult male Sprague Dawley rats. For 10 days animals were given three times daily intravenous injections of nicotine (1.5 mg/kg/day) or cigarette smoke extract (CSE) containing an equivalent dose of nicotine. Both spontaneous and mecamylamine-precipitated withdrawal were then measured. Chronic treatment with CSE induced significantly greater somatic and affective withdrawal signs than nicotine in both adolescents and adults. Mecamylamine-precipitated somatic signs were similar at both ages. In contrast, animals spontaneously withdrawn from chronic drug treatment exhibited significant age differences: whereas adolescents chronically treated with nicotine did not show somatic signs, those treated with CSE showed similar physical withdrawal to those of adults. Mecamylamine did not precipitate anxiety-like behavior at either age. However, both adolescents and adults showed significant anxiety in a light-dark box test 18 h after spontaneous withdrawal. Anxiety-like behavior was still evident in an open field test 1 month after termination of drug treatment, with adolescents showing significantly greater affective symptoms than adults. Our findings indicate that non-nicotine constituents of cigarette smoke do contribute to dependence in both adolescents and adults and emphasize the importance of including smoke constituents with nicotine in animal models of tobacco dependence.

Chronic exposure to cigarette smoke extract upregulates nicotinic receptor binding in adult and adolescent rats.

Heavy smokers display increased radioligand binding of nicotinic acetylcholine receptors (nAChRs). This "upregulation" is thought to be a contributing factor to tobacco dependence. Although cigarette smoke contains thousands of constituents that can contribute to nicotine dependence, it is not well understood whether non-nicotine constituents contribute to nAChR upregulation. In this study, we used an aqueous cigarette smoke extract (CSE), which contains nicotine and soluble constituents of cigarette smoke, to induce nAChR upregulation in adult and adolescent rats. To do this, male rats were exposed to nicotine or CSE (1.5 mg/kg/day nicotine equivalent, intravenously) daily for ten days. This experimental procedure produces equivalent levels of brain and plasma nicotine in nicotine- and CSE-treated animals. We then assessed nAChR upregulation using quantitative autoradiography to measure changes in three nAChR types. Adolescents were found to have consistently greater α4ß2 nAChR binding than adults in many brain regions. Chronic nicotine exposure did not significantly increase nAChR binding in any brain region at either age. Chronic CSE exposure selectively increased α4ß2 nAChR binding in adolescent medial amygdala and α7 binding in adolescent central amygdala and lateral hypothalamus. CSE also increased α3ß4 nAChR binding in the medial habenula and interpeduncular nucleus, and α7 binding in the medial amygdala, independent of age. Overall, this work provides evidence that cigarette smoke constituents influence nAChR upregulation in an age-, nAChR type- and region-dependent manner.

Differences in mechanisms underlying reinstatement of cigarette smoke extract- and nicotine-seeking behavior in rats.

Despite extensive research, current therapies for smoking cessation are largely ineffective at maintaining abstinence for more than a year. Whereas most preclinical studies use nicotine alone, the goal of the present study was to evaluate whether inclusion of non-nicotine tobacco constituents provides better face validity for the development of new pharmacological therapies for smoking cessation. Here, we trained adult male rats to self-administer nicotine alone or cigarette smoke extract (CSE), which contains nicotine and other aqueous constituents of cigarette smoke. After stable self-administration behavior was established, animals underwent extinction training followed by drug and cue primed reinstatement testing. We show that animals that self-administered CSE had significant reinstatement in all drug and drug + cue stimulus conditions whereas animals that self-administered nicotine only showed significant reinstatement in the drug + cue conditions. AT-1001, an α3ß4 nicotinic acetylcholine receptor (nAChR) functional antagonist, attenuated drug + cue-primed reinstatement of both CSE- and nicotine-seeking behavior. However, AT-1001 was less potent in blocking drug-primed reinstatement in animals that had self-administered CSE than in those that had self-administered nicotine alone. This was the case even when nicotine was used to prime reinstatement in animals that had self-administered CSE, suggesting that prior CSE exposure had altered the functional role of α3ß4-containing nAChRs in drug-seeking behavior. These findings confirm the importance of non-nicotine tobacco constituents and α3ß4* nAChRs in cue- and nicotine-primed craving. They also suggest that tests using CSE may be more valid models to study tobacco dependence than use of nicotine alone.

Prenatal nicotine sex-dependently alters adolescent dopamine system development.

Despite persistent public health initiatives, many women continue to smoke during pregnancy. Since maternal smoking has been linked to persisting sex-dependent neurobehavioral deficits in offspring, some consider nicotine to be a safer alternative to tobacco during pregnancy, and the use of electronic nicotine delivery systems is on the rise. We presently show, however, that sustained exposure to low doses of nicotine during fetal development, approximating plasma levels seen clinically with the nicotine patch, produces substantial changes in developing corticostriatal dopamine systems in adolescence. Briefly, pregnant dams were implanted on gestational day 4 with an osmotic minipump that delivered either saline (GS) or nicotine (3 mg/kg/day) (GN) for two weeks. At birth, pups were cross-fostered with treatment naïve dams and were handled daily. Biochemical analyses, signaling assays, and behavioral responses to cocaine were assessed on postnatal day 32, representative of adolescence in the rodent. GN treatment had both sex-dependent and sex-independent effects on prefrontal dopamine systems, altering Catechol-O-methyl transferase (COMT)-dependent dopamine turnover in males and norepinephrine transporter (NET) binding expression in both sexes. GN enhanced cocaine-induced locomotor activity in females, concomitant with GN-induced reductions in striatal dopamine transporter (DAT) binding. GN enhanced ventral striatal D2-like receptor expression and G-protein coupling, while altering the roles of D2 and D3 receptors in cocaine-induced behaviors. These data show that low-dose prenatal nicotine treatment sex-dependently alters corticostriatal dopamine system development, which may underlie clinical deficits seen in adolescents exposed to tobacco or nicotine in utero.

The α3ß4 nAChR partial agonist AT-1001 attenuates stress-induced reinstatement of nicotine seeking in a rat model of relapse and induces minimal withdrawal in dependent rats.

The strong reinforcing effects of nicotine and the negative symptoms such as anxiety experienced during a quit attempt often lead to relapse and low success rates for smoking cessation. Treatments that not only block the reinforcing effects of nicotine but also attenuate the motivation to relapse are needed to improve cessation rates. Recent genetic and preclinical studies have highlighted the involvement of the α3, ß4, and α5 nicotinic acetylcholine receptor (nAChR) subunits and the α3ß4 nAChR subtype in nicotine dependence and withdrawal. However, the involvement of these nAChR in relapse is not fully understood. We previously reported that the α3ß4 nAChR partial agonist AT-1001 selectively decreases nicotine self-administration in rats without affecting food responding. In the present experiments, we examined the efficacy of AT-1001 in attenuating reinstatement of nicotine-seeking behavior in a model of stress-induced relapse. Rats extinguished from nicotine self-administration were treated with the pharmacological stressor yohimbine prior to AT-1001 treatment and reinstatement testing. We also examined whether AT-1001 produced any withdrawal-related effects when administered to nicotine-dependent rats. We found that AT-1001 dose-dependently reduced yohimbine stress-induced reinstatement of nicotine seeking. When administered to nicotine-dependent rats at the dose that significantly blocked nicotine reinstatement, AT-1001 elicited minimal somatic withdrawal signs in comparison to the nicotinic antagonist mecamylamine, which is known to produce robust withdrawal. Our data suggest that α3ß4 nAChR-targeted compounds may be a promising approach for nicotine addiction treatment because they can not only block nicotine's reinforcing effects, but also decrease motivation to relapse without producing significant withdrawal effects.

Nicotine Increases Alcohol Intake in Adolescent Male Rats.

Background: Use of alcohol and tobacco, the two most concurrently abused drugs, typically first occurs during adolescence. Yet, there have been no systematic analyses of ethanol (EtOH) and nicotine (Nic) interactions during adolescence. Recent animal studies report that kappa-opioid (KOR) receptor activation mediates age differences in drug reinforcement. Our hypothesis is that concurrent self-administration of EtOH and Nic will be greater in adolescent rats because of age differences in KOR function. Furthermore, exposure to alcohol and nicotine during adolescence has been reported to increase EtOH intake in adulthood. We performed a longitudinal animal study and hypothesized adolescent rats allowed to self-administer nicotine would drink more alcohol as adults. Methods: Adolescent, postnatal day (P)32, and adult (P90) male and female Sprague-Dawley rats were allowed to self-administer EtOH, Nic, or a combination of both, EtOH+Nic, in an intravenous self-administration paradigm. The role of KOR was pharmacologically evaluated with the KOR antagonist, norbinaltorphamine (norBNI) and with the KOR agonist, U50,488H. Alcohol drinking was subsequently evaluated with male rats in a drinking in the dark (DID), 2-bottle choice test. Results: Concurrent Nic increased EtOH intake in adolescent males, but not in adults or females. Pharmacological blockade of KOR with norBNI robustly increased EtOH+Nic self-administration in adult male rats, but had no effect with female rats. Lastly, in our longitudinal study with male rats, we found prior self-administration of Nic or EtOH+Nic during adolescence increased subsequent oral EtOH intake, whereas prior self-administration of EtOH alone in adults increased subsequent EtOH drinking. Conclusions: There are major age- and sex-differences in the reinforcing effects of EtOH+Nic. Adolescent males are sensitive to the reinforcing interactions of the two drugs, whereas this effect is inhibited by KOR activation in male adults. Nicotine self-administration in adolescent males also increased subsequent oral EtOH intake. These findings suggest that brain mechanisms underlying the reinforcing effects of EtOH and nicotine are both age- and sex-dependent, and that tobacco or e-cigarette use may increase the vulnerability of teenage boys to alcohol abuse.

Self-administration of nicotine and cigarette smoke extract in adolescent and adult rats.

Although smoking initiation typically occurs during adolescence, most preclinical studies of tobacco use involve adult animals. Furthermore, their focus is largely on nicotine alone, even though cigarette smoke contains thousands of constituents. The present study therefore aimed to determine whether aqueous constituents in cigarette smoke affect acquisition of nicotine self-administration during adolescence in rats. Adolescent and adult male rats, aged postnatal day (P) 25 and 85, respectively, were food trained on a fixed ratio 1 (FR1) schedule, then allowed to self-administer one of 5 doses of nicotine (0, 3.75, 7.5, 15, or 30 µg/kg) or aqueous cigarette smoke extract (CSE) with equivalent nicotine content. Three progressively more difficult schedules of reinforcement, FR1, FR2, and FR5, were used. Both adolescent and adult rats acquired self-administration of nicotine and CSE. Nicotine and CSE similarly increased non-reinforced responding in adolescents, leading to enhanced overall drug intake as compared to adults. When data were corrected for age-dependent alterations in non-reinforced responding, adolescents responded more for low doses of nicotine and CSE than adults at the FR1 reinforcement schedule. No differences in adolescent responding for the two drugs were seen at this schedule, whereas adults had fewer responses for CSE than for nicotine. However, when the reinforcement schedule was increased to FR5, animals dose-dependently self-administered both nicotine and CSE, but no drug or age differences were observed. These data suggest that non-nicotine tobacco smoke constituents do not influence the reinforcing effect of nicotine in adolescents.

Reinforcing and neural activating effects of norharmane, a non-nicotine tobacco constituent, alone and in combination with nicotine.

Tobacco use is the leading cause of preventable death. Although the health risks are well known, cessation rates remain low. Whereas behavioral and neuroanatomical studies on tobacco addiction conventionally use nicotine, there is evidence that other constituents, such as monoamine oxidase inhibitors, may be important factors for modeling smoking. The aims of the present study were therefore to determine whether norharmane, a tobacco constituent and monoamine oxidase inhibitor, is self-administered alone and/or in combination with nicotine, and to evaluate the neural mechanisms underlying acquisition of self-administration of the two drugs. Sprague-Dawley rats were catheterized and allowed to intravenously self-administer either saline, nicotine (7.5 µg/kg/inj), norharmane (0.25 or 2.5 µg/kg/inj), alone or combined together (7.5+2.5 µg/kg/inj) for five days at fixed ratio (FR)1, two days each at FR2 and FR5, and one day at progressive ratio. Animals acquired self-administration of norharmane alone (2.5 µg/kg/inj), and the reinforcing effects of nicotine and norharmane were additive. For neuroanatomical analyses, rats self-administered the same treatments for six days at FR1, then brains were collected and processed by in situ hybridization for cfos mRNA expression. Treatment-specific profiles of regional cfos expression and correlations between cfos mRNA levels and behavioral responding were observed. Thus, not only was norharmane behaviorally reinforcing but, when combined with nicotine, resulted in patterns of neural activation distinct from that of norharmane or nicotine alone. This suggests that non-nicotine constituents can have central activating effects independent of nicotine, further substantiating the need for their inclusion in preclinical investigations of tobacco dependence.

Comparison of the reinforcing properties of nicotine and cigarette smoke extract in rats.

Tobacco dependence is difficult to treat, with the vast majority of those who try to quit relapsing within the first year. Improvements in smoking cessation therapies may be achieved by improving current preclinical research methods. However, most experimental tests in animals use nicotine alone, ignoring the 8000 other constituents found in tobacco smoke. To improve on this model, we have used self-administration to test the reinforcing properties of aqueous cigarette smoke extract (CSE) in rats, made by bubbling cigarette smoke through a saline solution. CSE is more potent than nicotine alone in both the acquisition and maintenance of self-administration, but did not exhibit higher progressive ratio responding. Mecamylamine and varenicline had similar potencies to block nicotine and CSE self-administration, indicating the involvement of nicotinic receptors in CSE reinforcement. Following extinction of responding, reinstatement was triggered by exposing animals to a pharmacological stressor, yohimbine (2.5 mg/kg, i.p.), alone and in combination with cues. Animals that self-administered CSE were significantly more sensitive to stress-induced reinstatement than those that self-administered nicotine. Ligand binding autoradiography studies showed nicotine and CSE to have similar affinities for different nicotinic receptor types. CSE significantly reduced MAO-A and MAO-B activities in vitro, whereas nicotine did not. Although CSE inhibition of MAO-A activity in vitro was found to be partially irreversible, irreversible inhibition was not observed in vivo. These experiments show that CSE is an effective reinforcer acting via nicotinic receptors. Furthermore, it better models MAO inhibition and is more sensitive to stress-induced reinstatement than nicotine alone, which is a potent trigger for relapse in smokers.

Age-dependent alterations in reward-seeking behavior after brief nicotine exposure.

RATIONALE: Adolescence is a developmental period that coincides with the onset of tobacco use. Teen smokers are also more likely to abuse other drugs compared to nonsmokers. Previous studies with rats have shown that low-dose nicotine pretreatment enhances initial acquisition of cocaine self-administration when given during early adolescence, but not at later ages. The aim of the present study was to determine whether these nicotine pretreatment effects extend to extinction and reinstatement of reward-seeking behavior. METHODS: Adolescent [postnatal day (P)28] and adult rats (P86) were pretreated for 4 days with nicotine (60 µg/kg, i.v.) or saline. Following pretreatment, rats were allowed to nose poke for cocaine (500 µg/kg/infusion) or sucrose pellets for at least 12 days or until meeting acquisition criterion. Responding was then extinguished for at least 7 days or until extinction criterion was met. The following day, the rats were reinstated with either a priming injection of cocaine (10 mg/kg, i.p.) or sucrose pellets. RESULTS: Nicotine markedly enhanced extinction of cocaine self-administration in adolescent rats, but not adults. Pretreatment also enhanced the acquisition of cocaine self-administration in adolescents, while reducing discrimination for the reinforced hole in adults. There were no pretreatment or age effects on cocaine-induced reinstatement. In contrast, nicotine induced only minor enhancement of sucrose-taking behavior in adolescents, with no significant impact on extinction or reinstatement at either age. CONCLUSIONS: Nicotine pretreatment affects reward-related behavior in both an age- and reward-dependent manner. These findings show that brief nicotine exposure during early adolescence enhances drug-related learning.

Nicotine alters limbic function in adolescent rat by a 5-HT1A receptor mechanism.

Epidemiological studies have shown that adolescent smoking is associated with health risk behaviors, including high-risk sexual activity and illicit drug use. Using rat as an animal model, we evaluated the behavioral and biochemical effects of a 4-day, low-dose nicotine pretreatment (60 µg/kg; intravenous) during adolescence and adulthood. Nicotine pretreatment significantly increased initial acquisition of cocaine self-administration, quinpirole-induced locomotor activity, and penile erection in adolescent rats, aged postnatal day (P)32. These effects were long lasting, remaining evident 10 days after the last nicotine treatment, and were observed when nicotine pretreatment was administered during early adolescence (P28-31), but not late adolescence (P38-41) or adulthood (P86-89). Neurochemical analyses of c-fos mRNA expression, and of monoamine transmitter and transporter levels, showed that forebrain limbic systems are continuing to develop during early adolescence, and that this maturation is critically altered by brief nicotine exposure. Nicotine selectively increased c-fos mRNA expression in the nucleus accumbens shell and basolateral amygdala in adolescent, but not adult animals, and altered serotonin markers in these regions as well as the prefrontal cortex. Nicotine enhancement of cocaine self-administration and quinpirole-induced locomotor activity was blocked by co-administration of WAY 100 635 (N-{2-[4-(2-methoxyphenyl)-1-piperazinyl] ethyl}-N-(2-pyridinyl)cyclohexanecarboxamide), a selective serotonin 1A (5-HT1A) receptor antagonist. Early adolescent pretreatment with the mixed autoreceptor/heteroceptor 5-HT1A receptor agonist, 8-OH-DPAT, but not the autoreceptor-selective agonist, S-15535, also enhanced quinpirole-induced locomotor activation. Nicotine enhancement of quinpirole-induced penile erection was not blocked by WAY 100 635 nor mimicked by 8-OH-DPAT. These findings indicate that early adolescent nicotine exposure uniquely alters limbic function by both 5-HT1A and non-5-HT1A receptor mechanisms.

The monoamine oxidase (MAO) inhibitor tranylcypromine enhances nicotine self-administration in rats through a mechanism independent of MAO inhibition.

Our current study aims to evaluate the mechanisms of tranylcypromine (TCP)-mediated enhancement of nicotine self-administration. We replicated our previous findings which demonstrate that 1 h pretreatment with TCP (3 mg/kg, i.p.) enhances nicotine self-administration (7.5 µg/kg/inj, i.v.) when compared with vehicle-treated rodents. We tested whether TCP-mediated enhancement of nicotine self-administration was due to MAO inhibition or off-target effects by (i) extending the TCP pretreatment time from 1 to 20 h, and (ii) evaluating the role of the individual TCP stereoisomers in nicotine self-administration studies. While 20 h and (-)TCP pretreatment induced significant inhibition of MAO (60-90%), animals found nicotine only weakly reinforcing. Furthermore, while both (+) and (±)TCP treatment induced nearly 100% MAO inhibition, (+)TCP pretreated animals took longer to acquire nicotine self-administration compared to (±)TCP pretreated animals. Stable nicotine self-administration in (+)TCP pretreated animals was influenced by nicotinic receptor activation but not nicotine-paired cues. The opposite was found in (±)TCP pretreated animals. Treatment with (-) or (±)TCP increased dopamine and serotonin overflow, while the (+) and (±)TCP treatment enhanced monoamine overflow subsequent to nicotine. Together, our data suggests TCP enhancement of nicotine self-administration are mediated through mechanisms independent of MAO inhibition, including nicotine-paired cues and monoamine uptake inhibition.

Serotonergic mechanism underlying tranylcypromine enhancement of nicotine self-administration.

Although nicotine is generally considered to be the main psychoactive component of tobacco, growing evidence highlights the importance of nonnicotine compounds in smoking reinforcement. Monoamine oxidase (MAO) inhibition is a major consequence of smoking and MAO inhibitors, such as tranylcypromine, increase nicotine reinforcement. Tranylcypromine has multiple pharmacological effects, increasing monoamine release for a few hours immediately after its administration and blocking MAO activity for several days. To assess the relative role of these two actions, adult male rats were tested in consecutive daily 3-h sessions for self-administration of nicotine (3 µg kg⁻¹) inj⁻¹, i.v.) either 20 or 1 h following administration of tranylcypromine (3 mg kg⁻¹). Both paradigms were shown to produce highly significant inhibition of MAO activity. However, whereas animals readily acquired self-administration when pretreated with tranylcypromine 1 h prior to testing, they did not with the longer pretreatment interval. Such animals did immediately acquire nicotine self-administration when the tranylcypromine pretreatment interval was switched to 1 h prior to testing on Day 4, indicating that an acute effect of the MAO inhibitor was responsible for enhanced nicotine reinforcement. Several lines of evidence implicate serotonin (5-HT) as the mediator of this enhancement: (1) Tranyclypromine-enhanced nicotine reinforcement was blocked by the 5-HT2 receptor antagonists, ritanserin and ketanserin; (2) parachloroamphetamine (PCA), a 5-HT releaser, also enhanced nicotine self-administration in animals in which MAO activity was inhibited; (3) pretreatment with tranylcypromine increased PCA-induced 5-HT overflow in the nucleus accumbens. These findings suggest that MAO inhibition enhances serotonergic transmission, which serves a critical role in the reinforcing effects of nicotine.

Locomotor and stress responses to nicotine differ in adolescent and adult rats.

Since adolescence is a critical period for the initiation of tobacco use, we have systematically compared behavioral and endocrine responses to nicotine in Sprague-Dawley rats of both sexes at early adolescence (postnatal day (P) 28), mid- adolescence (P38) and adulthood (P90). Locomotion and center time in a novel open field were evaluated for 30min following intravenous injection of saline or nicotine (60microg/kg), followed by measurement of plasma corticosterone. Complex age and sex differences in behavioral and endocrine response were observed, which were dependent on the functional endpoint examined. Whereas there were age differences in nicotine effects on all functional measures, sex differences were largely restricted to adult stress-related corticosterone and center-time responses. Although significant drug effects were detected at P28 and P90, there was no effect of nicotine at P38 on any measure examined. In saline-treated males, but not females, there were significant positive correlations across age between initial ambulatory counts and both initial vertical counts and total center time. Nicotine treatment increased correlations in both sexes, and yielded a significant negative interaction between initial ambulatory counts and plasma corticosterone. The unique responses of adolescents to nicotine are consistent with an immature function of nicotinic acetylcholine receptors at this age.

Age influences the effects of nicotine and monoamine oxidase inhibition on mood-related behaviors in rats.

RATIONALE: Epidemiological studies have demonstrated a comorbidity of smoking with depression and anxiety, particularly during adolescence. However, few animal studies have considered possible synergistic interactions between nicotine and other tobacco smoke constituents, such as monoamine oxidase (MAO) inhibitors, in the regulation of mood. OBJECTIVES: The aim of the study was to test the hypothesis that nicotine combined with the irreversible MAO inhibitor, tranylcypromine, will differentially affect depression- and anxiety-related behaviors in adolescent and adult rats. METHODS: Nicotine (0, 0.05, 0.2 mg/kg, s.c.) and tranylcypromine (3 mg/kg, i.p.) were tested separately, or together, on male rats aged postnatal days 30 and 68, in three mood-related behavioral tests: forced swim test (FST), elevated plus maze (EPM), and open field. RESULTS: Nicotine (0.2 mg/kg) in adults significantly decreased floating time in the FST and increased time spent in the open arm of the EPM, with no change in locomotor activity. Tranylcypromine pretreatment combined with nicotine (0.2 mg/kg) significantly increased locomotor activity and time spent in the center of the open field. Whereas nicotine alone had no significant effect on adolescents, it significantly increased locomotor activity and decreased floating time in the FST when combined with tranylcypromine pretreatment. CONCLUSIONS: There is an age-dependent effect of nicotine, alone and in combination with MAO inhibition, on mood-related behaviors. Whereas nicotine alone induces mood improvement in adults, it has no effect on adolescents. Nicotine combined with tranylcypromine has unique, age-dependent effects. Thus, experimental studies of smoking should consider both age and other tobacco constituents, such as MAO inhibitors, as critical factors.

Age-dependent effects of low-dose nicotine treatment on cocaine-induced behavioral plasticity in rats.

RATIONALE: Epidemiological evidence of early adolescent tobacco use, prior to that of marijuana and other illicit drugs, has led to the hypothesis that nicotine is a "gateway" drug that sensitizes reward pathways to the addictive effects of other psychostimulants. OBJECTIVE: To test this hypothesis, we have compared the effect of a brief, low-dose nicotine pretreatment of adolescent and adult rats on subsequent locomotor response to acute and chronic cocaine. MATERIALS AND METHODS: Adolescents, aged postnatal day (P) 28, and adults, aged P86, were given four daily injections of saline or nicotine (0.06 mg/kg, i.v.). At P32 and P90, rats were given acute injections of cocaine (0, 0.4 or 1.0 mg/kg, i.v.) and monitored for locomotor activity in either a habituated or novel test environment. To examine cocaine sensitization, rats were treated for 3 days with saline or cocaine (0.4 mg/kg, i.v.), and, after 1 day of withdrawal, were given a challenge dose of cocaine (0.4 mg/kg, i.v.). RESULTS: Nicotine pretreatment did not affect acute, drug-induced locomotor activity at either age. However, age differences in cocaine response were observed, with adolescent animals showing enhanced locomotor activity in the novel environment. Adolescent controls did not exhibit cocaine-induced locomotor sensitization, whereas adults did. Nicotine pretreatment during adolescence promoted the development and expression of a sensitized response to repeated cocaine exposure similar to that observed in saline-pretreated adult controls. CONCLUSIONS: These findings show that brief pretreatment with nicotine, in a low dose comparable to that inhaled in 2-4 cigarettes, enhances cocaine-induced behavioral plasticity in adolescent rats.

Prenatal nicotine exposure changes natural and drug-induced reinforcement in adolescent male rats.

Clinical studies have demonstrated an increased incidence of substance misuse and obesity in adolescents whose mothers smoked during pregnancy. Although dopamine systems that mediate natural and drug-induced reinforcement have been shown in animal studies to be altered by gestational nicotine treatment, it is not clear whether there are concomitant changes in reinforcement sensitivity. To test whether prenatal nicotine exposure influences sensitivity to natural and drug rewards, timed pregnant rats were implanted with osmotic minipumps delivering saline or nicotine (3 mg/kg/day) from gestational day 4 to 18. Male offspring were tested as adolescents, on postnatal day 32, for operant responding maintained by sucrose pellets or i.v. cocaine (200 or 500 mug/kg per injection). Cocaine-induced stereotypy and c-fos mRNA expression in cortex and striatum were also examined. Complex changes in reward circuitry were observed in the offspring of nicotine-exposed dams. Nicotine-exposed adolescents did not self-administer the low dose of cocaine, but, at the higher dose, exhibited significantly greater cocaine intake and c-fos mRNA expression in nucleus accumbens than did controls. In contrast, control animals showed significantly greater drug-induced stereotypy at both cocaine doses. Operant responding maintained by sucrose was also influenced by gestational nicotine exposure. At a fixed ratio (FR) 1 schedule, although the number of pellets eaten by the two experimental groups was equivalent, more pellets were left uneaten by nicotine-exposed offspring. At FR2 and FR5 schedules, the responding maintained by sucrose pellets was lower in nicotine-exposed offspring. These findings suggest that nicotine exposure during gestation may induce changes in both natural and drug reward pathways.

Negative allosteric modulation of nicotinic acetylcholine receptors blocks nicotine self-administration in rats.

Drugs that antagonize nicotinic acetylcholine receptors (nAChRs) can be used to inhibit nicotine-induced behavior in both humans and animals. The aim of our experiments is to establish a proof-of-principle that antagonism of nAChRs by negative allosteric modulation can alter behavior in a relevant animal model of addiction, nicotine self-administration. We have identified a novel, negative allosteric modulator of nAChRs, UCI-30002 [N-(1,2,3,4-tetrahydro-1-naphthyl)-4-nitroaniline], with selectivity for the major neuronal nAChR subtypes over muscle-type nAChRs. After systemic administration, UCI-30002 significantly reduces nicotine self-administration in rats on both fixed ratio and progressive ratio schedules of reinforcement. The minimum effective dose that significantly alters nicotine self-administration corresponds to brain concentrations of UCI-30002 that produce at least 30% inhibition of the major neuronal nAChR subtypes measured in vitro. UCI-30002 has no effect on responding for food reinforcement in rats on either type of schedule, indicating that there is no effect on general responding or natural reward. UCI-30002 represents validation of the concept that negative allosteric modulators may have significant benefits as a strategy for treating nicotine addiction and encourages the development of subtype-selective modulators.

Gestational exposure to nicotine and monoamine oxidase inhibitors influences cocaine-induced locomotion in adolescent rats.

RATIONALE: Many pregnant women continue to smoke, despite a strong association between maternal smoking and neurobehavioral deficits in the offspring. Although gestational nicotine (GN) treatment in rodents is used as the primary animal model of maternal smoking, tobacco smoke contains more than 4,000 constituents, including monoamine oxidase inhibitors (MAOIs). OBJECTIVES: The aim of this study was to determine whether there are interactions between the effects of gestational exposure to nicotine and MAOIs on cocaine-induced locomotor sensitization in adolescent rats. MATERIALS AND METHODS: Pregnant rats were implanted on day 4 of gestation with osmotic minipumps delivering saline, nicotine (3 mg/kg per day), the MAOIs clorgyline and deprenyl (1 and 0.25 mg/kg per day, respectively), or nicotine/clorgyline/deprenyl (GMN). Adolescent female offspring were tested for cocaine-induced locomotor sensitization. Animals were treated with saline or cocaine (5 or 15 mg/kg, intraperitoneally) daily from postnatal (P) days 32-36 and challenged with cocaine (15 mg/kg) on P51 (day 20). RESULTS: Group differences were observed in chronic but not acute effects of cocaine. Whereas gestational MAOI treatment, with or without nicotine, increased ambulatory response to cocaine on day 5, the opposite was found for vertical activity. Different adaptive responses were observed on cocaine challenge day. GNM animals exhibited enhanced locomotor activity in the cocaine-associated environment before cocaine challenge on day 20. In contrast, only GN animals exhibited significant locomotor sensitization to the cocaine challenge. CONCLUSIONS: Gestational nicotine and MAOIs both influence brain development. Such interactions may sensitize adolescents to drug abuse and should be considered in animal models of maternal smoking.