The Effects of Changes in Cigarette Menthol Content on Acute Nicotine Pharmacology and Smoking Topography.

INTRODUCTION: Menthol influences the appeal and addictiveness of cigarette smoking, however the data regarding menthol's effects on nicotine pharmacokinetics (PK) and smoking topography are inconsistent. This study investigated the impact of different cigarette menthol levels on nicotine pharmacology and smoking topography in current menthol smokers. AIMS AND METHODS: The study was a double-blind, randomized, four-period, crossover study to investigate the effects of smoking cigarettes with varying menthol content (0, 3, 6, and 12 mg menthol) on nicotine PK, smoking topography, and subjective effects in current menthol smokers. Each experimental session consisted of a prescribed use session, followed by 145 min of no smoking and a 1-h ad libitum smoking session. Serial blood samples were collected; smoking topography was recorded using CReSS Lab topography device. RESULTS: There was no significant effect of menthol on nicotine PK after prescribed smoking of cigarettes with varying menthol contents. During ad libitum smoking, there was significantly smaller total puff volume and puff duration in the 12 mg menthol condition compared to other menthol conditions. Subjective and sensory measures indicated significantly higher overall positive ratings for the 3 mg and 6 mg menthol cigarettes compared to the 0 mg menthol cigarette; the 12 mg menthol cigarette was less liked and harsher than the 3 mg condition. CONCLUSIONS: These findings suggest that menthol, at concentrations reflecting the marketplace (3-6 mg), contributes to positive subjective smoking experiences among menthol smokers, but does not have a significant effect on nicotine PK or smoking topography in an acute laboratory setting. IMPLICATIONS: While our data indicate that varying menthol content does not have a significant impact on nicotine's pharmacological effects under acute exposure conditions, these data highlight the contribution of menthol's flavor and sensory effects to product preference and positive smoking experiences, which facilitate repeated experimentation, progression to regular use, and subsequent dependence.

A Method to Amend Cigarettes With Menthol for Use in Research.

INTRODUCTION: Studies have evaluated the role of menthol cigarettes on various addiction-related outcomes; however, the effect of varying menthol content on these outcomes has not been evaluated. We developed a method to amend non-menthol SPECTRUM Research Cigarettes to contain menthol at four different levels. AIMS AND METHODS: SPECTRUM Research Cigarettes, NRC 600 (0.8 mg nicotine; 10 mg tar), were modified to contain target menthol amounts at 3, 6, and 12 mg/cigarette by injecting 25 µL ethanol/triacetin/menthol solutions of varying concentrations (120 mg menthol/mL, 240 mg/mL, and 480 mg/mL) into four distinct locations in the filter and tobacco rod. Menthol content was tested in triplicate in the whole cigarette and in the tobacco rod and filter at 1, 24, 48, and 72 hours for each target menthol level using an extraction solution of quinoline in methyl-tert-butyl ether and measured using gas chromatography with flame ionization detection. RESULTS: Injections into the filter and tobacco rod (12.5 µL each) yielded equal menthol distribution up to 72 hours. However, total menthol content decreased from an average of 90.3% of the target menthol concentration at 1 hour to 80.7% at 72 hours in cigarettes stored individually in glass tubes at room temperature. Analysis of urinary menthol glucuronide confirmed that amended cigarettes used within 24 hours of injection delivered dose-related menthol levels to participants in a clinical laboratory setting. CONCLUSION: This method can be used to modify cigarettes with a range of reliable menthol levels in both filter and tobacco rod for use in laboratory and clinical research. IMPLICATIONS: This study presents a technique for modifying cigarettes with different levels of menthol that can reliably deliver dose-related menthol levels to participants when smoked in a clinical study. The technique can be used to quickly amend cigarettes to examine the independent effects of varying flavor and additive levels on smoking behavior, nicotine pharmacokinetics, mainstream smoke emissions, and other laboratory or clinical research outcomes.

Sensory methods to evaluate perception of flavours in tobacco and other nicotine-containing products: a review.

OBJECTIVES: Sensory methods use human senses to evaluate product attributes. This review provides an overview of the types of sensory methods used to evaluate the perception of flavour in tobacco and other nicotine-containing (ToNic) products and to discuss how sensory data could inform flavoured ToNic product policy. DATA SOURCES: PubMed, Embase and Web of Science. STUDY SELECTION: All peer-reviewed studies evaluating ToNic products using a sensory method published before 23 May 2020. DATA EXTRACTION: Two independent coders completed title/abstract and full-text screening to choose articles for inclusion (Cohen's kappa=0.85, strong agreement). Each coder completed data extraction on half the articles, recording relevant information (eg, sensory methods used, results). The coders categorised sensory methods and generated overarching themes. DATA SYNTHESIS: Of 110 articles identified, we included 29 articles containing 35 studies that used sensory methods to investigate ToNic products. The sensory methods included analytic methods such as discrimination and descriptive tests and hedonic methods such as liking tests. Six themes emerged regarding how sensory methods can be used to understand consumer perception and liking of ToNic products and to inform ToNic product policy. CONCLUSIONS: The identified studies highlight that sensory data can inform ToNic product policy. Analytic and sensory hedonic ratings can be used to assess a ToNic product's ability to promote addiction in the user (ie, abuse liability). Lastly, hedonic ratings can provide information to assess potential use behaviours.

Transitions in frequency of hookah smoking among youth and adults: findings from waves 1 and 2 of the Population Assessment of Tobacco and Health (PATH) study, 2013-15.

BACKGROUND AND AIMS: The prevalence of hookah smoking has increased in the United States since at least 2010, especially among youth and young adults. This study assessed self-reported reasons for hookah smoking cessation and transition to or maintenance of high-frequency hookah smoking among current hookah smokers. DESIGN: Separately analyzed data from the Population Assessment of Tobacco and Health (PATH) study, a longitudinal cohort study. Frequency of and reasons for hookah smoking were ascertained at wave 1 (2013-14); frequency of hookah smoking and past-year cessation were ascertained at wave 2 (2014-15). Weighted multivariable logistic and ordinal logistic regression models were fitted to predict hookah smoking cessation and frequency of hookah smoking at wave 2, respectively, accounting for demographic and behavioral risk factors, reasons for hookah smoking and frequency of hookah smoking at wave 1. SETTING: United States. PARTICIPANTS: A total of 693 youth and 4400 adult past-year hookah smokers. MEASUREMENTS: Self-reported tobacco-use patterns and associated health behaviors were measured via audio computer-assisted self-interviews (ACASI). FINDINGS: At wave 1, 5.9% of youth and 7.5% of adults were past-year hookah smokers. Among all age groups, the leading reasons for hookah smoking were enjoyment of socializing while smoking, the availability of appealing flavors and believing that it was less harmful than cigarette smoking. The odds of cessation were lower for adults who liked hookah flavors [adjusted odds ratio (aOR) = 0.40; 95% confidence interval (CI) = 0.26-0.62] compared with adults who did not like hookah flavors. The odds of transitioning to, or maintaining, monthly or more frequent hookah smoking at wave 2, compared with cessation or less than monthly smoking, were higher for adults who liked hookah flavors [adjusted proportional odds ratio (aPOR) = 2.10; 95% CI = 1.48-2.99 and enjoyed socializing while smoking hookah (aPOR = 1.82; 95% CI =1.13-2.94) compared with adults who did not like hookah flavors or socializing. CONCLUSION: The availability of appealing flavors, affordability and socialization while smoking hookah in the United States are associated with reduced likelihood of cessation and increased likelihood of high-frequency hookah smoking.

Analysis of Nicotine and Non-nicotine Tobacco Constituents in Aqueous Smoke/Aerosol Extracts by UHPLC and Ultraperformance Convergence Chromatography-Tandem Mass Spectrometry.

The non-nicotine constituents of tobacco may alter the reinforcing effects of nicotine, but the quantitative and qualitative profiles of these chemicals in tobacco products such as electronic cigarettes (e-cigarettes), cigars, and waterpipe tobacco are not well characterized. The objective of this work was to develop and validate analytical methods to utilize saline both as an extraction solvent for smoke condensates from cigarettes, little cigars, and waterpipe tobacco and aerosols from e-cigarettes and as a delivery vehicle of nicotine and non-nicotine constitents for nonclinical pharmacological studies. Ultrahigh-performance liquid chromatography was used to analyze nicotine and acetaldehyde, and a novel ultraperformance convergence chromatography-tandem mass spectrometry method was developed to analyze anabasine, anatabine, cotinine, myosmine, nornicotine, harmane, and norharmane. Linearity was confirmed for each standard curve with correlation coefficients (r) ≥ 0.99, and relative errors (RE) for the standards were ≤±10% over the calibration ranges. Method validation was performed by preparing triplicate samples in saline to mimic the composition and concentration of each analyte in the smoke or aerosol condensate and were used to determine method accuracy and precision. Relative standard deviation values were ≤15% and mean RE ≤15% for each analyte at each concentration level. Selectivity of the methods was demonstrated by the absence of peaks in blank vehicle or diluent samples. Storage stability was assessed over ∼45 days. Precision (%RSD ≤ 13) and recovery (percent of day 0 ≥ 80%) indicated that the saline formulations of all four products could be considered stable for up to ∼45 days at 4-8 °C. Therefore, the use of saline both as an extraction solvent and as a delivery vehicle adds versatility and improved performance in the study of the pharmacological effects of constituents from mainstream smoke and aerosols generated from cigarettes, little cigars, waterpipes, and e-cigarettes.

Trends and Associations of Menthol Cigarette Smoking Among US Middle and High School Students-National Youth Tobacco Survey, 2011-2018.

INTRODUCTION: Youth cigarette smoking decreased significantly over the last two decades in the United States. This study provides estimates and trends from 2011 to 2018 and factors associated with youth menthol and non-menthol smoking from 2016 to 2018. METHODS: Using data from the 2011-2018 National Youth Tobacco Surveys, past 30-day (current) menthol and non-menthol cigarette smoking were estimated for all youth (prevalence) and youth smokers (proportions). Trends were examined using Joinpoint regression, calculating the annual percent change (APC). Multivariate logistic regression analyses identified factors associated with menthol smoking. RESULTS: From 2011 to 2018, menthol cigarette smoking among current youth cigarette smokers significantly decreased from 57.3% to 45.7% (APC: -3.0%), while non-menthol (38.2% to 47.3% [APC: 2.9%]) and unknown menthol status (not sure\missing) (4.5% to 7.0% [APC: 7.1%]) significantly increased. Menthol cigarette smoking among high school, male, female, and non-Hispanic white current cigarette smokers decreased, but remained unchanged among middle school, non-Hispanic black, and Hispanic smokers. Significantly higher proportions of menthol cigarette smokers smoked on ≥20 days, ≥2 cigarettes per day, and ≥100 cigarettes in their lifetime compared to non-menthol smokers. Among current cigarette smokers, non-Hispanic blacks, Hispanics, flavored non-cigarette tobacco users, frequent smokers (≥20 days), those smoking 2-5 cigarettes per day, and those living with someone who uses tobacco had higher odds of menthol cigarette smoking. CONCLUSIONS: In 2018, nearly half of current youth cigarette smokers smoked menthol cigarettes. While menthol cigarette smoking declined from 2011 to 2018 among all youth and among youth smokers, there was no change in menthol cigarette smoking among non-Hispanic black, Hispanic, and middle school cigarette smokers. IMPLICATIONS: This study finds that overall cigarette and menthol cigarette smoking declined in youth from 2011 to 2018. However, menthol cigarette smoking among non-Hispanic black, Hispanic, and middle school youth cigarette smokers did not change. Information from this study can help inform efforts to reduce menthol cigarette smoking among US youth, particularly racial/ethnic minority populations.

Examining the role of menthol cigarettes in progression to established smoking among youth.

BACKGROUND: Menthol, a flavoring compound added to cigarettes, makes cigarettes more appealing to youth and inexperienced smokers and increases cigarettes' abuse liability. However, limited studies are available on menthol's role in smoking progression. METHODS: To assess the association between menthol in cigarettes and progression to established smoking, we used five waves of data from the Evaluation of Public Education Campaign on Teen Tobacco Cohort Study, a nationally representative longitudinal survey of U.S. youth conducted as part of "The Real Cost" evaluation. We used discrete time survival analysis to model the occurrence of two event outcomes-progression to established, current smoking and progression to established, frequent smoking-using a logit model with a menthol use indicator as the key explanatory variable. Based on this framework, we estimated the effect of prior menthol use on the odds of smoking progression. RESULTS: In the progression to established, current smoking model, prior menthol use was significantly associated with progression [adjusted odds ratio (aOR) = 1.80, p < .05, confidence interval (CI) = (1.03-3.16)]. While results were in a similar direction for the model of progression to established, frequent smoking, the association between prior menthol use and this progression model did not reach significance [aOR = 1.56, CI = (0.80-3.03)]. CONCLUSION: The results suggest a relationship between using menthol cigarettes and progression from experimental to established, current smoking among youth. This study adds to a growing literature base that supports that menthol cigarettes, compared to nonmenthol cigarettes, put youth at increased risk for regular cigarette use.

Comparison of cigarette, little cigar, and waterpipe tobacco smoke condensate and e-cigarette aerosol condensate in a self-administration model.

The pharmacological effects of tobacco products are primarily mediated by nicotine; however, research suggests that several non-nicotine tobacco constituents may alter the reinforcing effects of nicotine. This study evaluated the reinforcing effects of aqueous solutions of smoke/aerosol condensate from cigarettes, little cigars, electronic cigarettes (e-cigarettes), and waterpipe tobacco in a self-administration procedure to determine if abuse liability of these tobacco products differed. Adult male Sprague-Dawley rats (n = 64 total) were trained to self-administer intravenous nicotine (30 µg/kg/infusion) on a fixed ratio 5 schedule of reinforcement. Following nicotine dose-effect assessment (1, 7.5, 15, and 30 µg/kg/infusion), rats were given access to smoke/aerosol condensate derived from their assigned tobacco product. Rats responded for smoke/aerosol condensate containing 1, 7.5, 15, and 30 µg/kg/infusion nicotine, with the ratio of nicotine:non-nicotine constituents held constant across doses for each tobacco product. Responding for nicotine or smoke/aerosol condensate was also assessed on a progressive ratio schedule of reinforcement. Cigarette, little cigar, and e-cigarette smoke/aerosol condensates shifted the nicotine dose-effect curve leftward, whereas waterpipe tobacco smoke condensate shifted the dose-effect curve rightward. Smoke/aerosol condensate from all tobacco products produced similar levels of responding compared to nicotine alone during the progressive ratio phase. Results suggest that non-nicotine constituents in cigarettes, little cigars, and e-cigarettes differentially enhance nicotine's reinforcing potency. In contrast, waterpipe tobacco blunted nicotine's reinforcing potency, suggesting that it may contain unique constituents that dampen nicotine's reinforcing effects.

Characteristics of Hookah Tobacco Smoking Sessions and Correlates of Use Frequency Among US Adults: Findings From Wave 1 of the Population Assessment of Tobacco and Health (PATH) Study.

Introduction: Hookah tobacco smoking has increased in the United States. However, information on hookah use frequency and other characteristics of hookah use is limited. Methods: Investigators analyzed data from Wave 1 of the Population Assessment of Tobacco and Health (PATH) Study, a nationally representative, longitudinal cohort study of US youth and adults. Our analysis draws on baseline data from adult (ages ≥18 years) ever (N = 10 624) and past year (n = 3947) hookah users. Bivariate and regression analyses were conducted to identify associations between demographics, use characteristics, and hookah use frequency. Results: Overall, 16.4% of adults reported ever smoking tobacco from a hookah. Of those, 31.9% reported smoking hookah within the past year. Among 3947 past-year hookah tobacco smokers, 10.7% were daily/weekly users, 13.7% were monthly users, 42.1% smoked every couple of months, and 33.5% smoked about once a year. Among daily/weekly hookah users, 66% were young adults (ages 18-24 years). When comparing daily/weekly hookah users to those who smoked every couple of months, more frequent hookah use was associated with younger age, male gender, a greater number of times the hookah is refilled during a session, fewer people sharing, and hookah ownership. Conclusion: Although there were few demographic differences between daily/weekly users and less frequent hookah tobacco smokers, some notable differences in use behaviors exist among use frequency groups. Unlike other tobacco products, hookah is often smoked communally, over an extended time period. A detailed understanding of hookah user characteristics and experiences could inform hookah-specific measures, messaging, and regulations. Implications: To our knowledge, this is one of the first studies to characterize hookah use frequency, session length, and other hookah use experiences using a nationally representative sample of US adult hookah smokers. Understanding characteristics of hookah tobacco smokers, their use experiences, and patterns of hookah use frequency would be useful in assessing risks for hookah tobacco dependence at the population level. Additionally, identifying variation in use behaviors of hookah tobacco smokers could be used to inform the development of strategies to reduce and prevent hookah tobacco use, such as tobacco regulations or public education efforts, particularly among young adults.

Co-occurrence of tobacco product use, substance use, and mental health problems among youth: Findings from wave 1 (2013-2014) of the population assessment of tobacco and health (PATH) study.

INTRODUCTION: Cigarette use is associated with substance use and mental health problems among youth, but associations are unknown for non-cigarette tobacco product use, as well as the increasingly common poly-tobacco use. METHODS: The current study examined co-occurrence of substance use and mental health problems across tobacco products among 13,617 youth aged 12-17years from Wave 1 (2013-2014) of the nationally representative Population Assessment of Tobacco and Health (PATH) Study. Participants self-reported ever cigarette, e-cigarette, smokeless tobacco, traditional cigar, cigarillo, filtered cigar, hookah, and other tobacco product use; alcohol, marijuana, and other drugs; and lifetime substance use, internalizing and externalizing problems. RESULTS: In multivariable regression analyses, use of each tobacco product was associated with substance use, particularly cigarillos and marijuana (AOR=18.9, 95% CI: 15.3-23.4). Cigarette (AOR=14.7, 95% CI: 11.8-18.2) and cigarillo (AOR=8.1, 95% CI: 6.3-10.3) use were strongly associated with substance use problems and tobacco users were more likely to report internalizing (AOR=1.6, 95% CI: 1.4-1.8) and externalizing (AOR=1.4, 95% CI: 1.3-1.6) problems. Female tobacco users were more likely to have internalizing problems than male tobacco users. Poly-tobacco users were more likely than exclusive users to use substances (AOR=3.4, 95% CI: 2.7-4.3) and have mental health (AOR=1.2, 95% CI: 1.0-1.5) and substance use (AOR=4.7, 95% CI: 3.4-6.6) problems. CONCLUSIONS: Regardless of the tobacco product used, findings reveal high co-occurrence of substance use and mental health problems among youth tobacco users, especially poly-tobacco users. These findings suggest the need to address comorbidities among high risk youth in prevention and treatment settings.

Tobacco's minor alkaloids: Effects on place conditioning and nucleus accumbens dopamine release in adult and adolescent rats.

Tobacco products are some of the most commonly used psychoactive drugs worldwide. Besides nicotine, alkaloids in tobacco include cotinine, myosmine, and anatabine. Scientific investigation of these constituents and their contribution to tobacco dependence is less well developed than for nicotine. The present study evaluated the nucleus accumbens dopamine-releasing properties and rewarding and/or aversive properties of nicotine (0.2-0.8mg/kg), cotinine (0.5-5.0mg/kg), anatabine (0.5-5.0mg/kg), and myosmine (5.0-20.0mg/kg) through in vivo microdialysis and place conditioning, respectively, in adult and adolescent male rats. Nicotine increased dopamine release at both ages, and anatabine and myosmine increased dopamine release in adults, but not adolescents. The dopamine release results were not related to place conditioning, as nicotine and cotinine had no effect on place conditioning, whereas anatabine and myosmine produced aversion in both ages. While the nucleus accumbens shell is hypothesized to play a role in strengthening drug-context associations following initiation of drug use, it may have little involvement in the motivational effects of tobacco constituents once these associations have been acquired. Effects of myosmine and anatabine on dopamine release may require a fully developed dopamine system, since no effects of these tobacco alkaloids were observed during adolescence. In summary, while anatabine and myosmine-induced dopamine release in nucleus accumbens may play a role in tobacco dependence in adults, the nature of that role remains to be elucidated.

Co-occurrence of tobacco product use, substance use, and mental health problems among adults: Findings from Wave 1 (2013-2014) of the Population Assessment of Tobacco and Health (PATH) Study.

BACKGROUND: Although non-cigarette tobacco product use is increasing among U.S. adults, their associations with substance use and mental health problems are unclear. This study examined co-occurrence of tobacco use, substance use, and mental health problems, and its moderation by gender, among 32,202U.S. adults from Wave 1 (2013-2014) of the nationally representative longitudinal Population Assessment of Tobacco and Health (PATH) Study. METHODS: Participants self-reported current cigarette, e-cigarette, traditional cigar, cigarillo, filtered cigar, hookah, smokeless tobacco and other tobacco product use; past year alcohol, marijuana, and other drug use; and past year substance use, internalizing and externalizing problems. RESULTS: Compared to non-current tobacco users, current users were more likely to report alcohol or drug use (adjusted odds ratio (AOR)=2.6; 95% confidence interval (CI): 2.3, 2.9), with the strongest associations observed for cigarillo and hookah users. Across all tobacco product groups, users were more likely to report internalizing (AOR=1.9; 95% CI: 1.7, 2.1), externalizing (AOR=1.6; 95% CI: 1.5, 1.8), and substance use (AOR=3.4; 95% CI: 2.9, 4.1) problems than non-users. Gender moderated many of these associations and, of these, all non-cigarette tobacco product associations were stronger among females. CONCLUSIONS: This nationally representative study of U.S. adults is the first to comprehensively document tobacco use, substance use, and mental health comorbidities across the range of currently available tobacco products, while also demonstrating that female tobacco users are at increased risk for substance use and mental health problems. These findings may point to gender differences in vulnerability and suggest that interventions incorporate gender-specific approaches.

Mouth Level Exposure and Similarity to Machine-smoked Constituent Yields.

OBJECTIVES: The goal of this review was to evaluate which standard machine-smoking regimen may be most appropriate to inform tobacco product regulation based on the fraction of cigarette smoke yields that best represents the range of human smoke exposures. METHODS: We searched PubMed and Web of Science to identify peer-reviewed studies that reported percentages of smokers who smoked more or less like a particular machine-smoking regimen based on human mouth level exposure (MLE) tobacco constituent yields. RESULTS: Three studies met our inclusion criteria and were reviewed. Studies found that Canadian Intense (CI) yields were equal to or greater than 86% to 97% of smokers' nicotine and tar MLE yields. CONCLUSIONS: MLE yields indicate that a small percentage of individuals (less than 14%) are exposed to nicotine and tar yields equal to or greater than those measured by the CI regimen. Whereas no machine-smoking regimen reflects human puffing behavior with complete accuracy, based on MLE data, CI constituent yields constitute the best representation of exposure that encompasses the majority of smokers, and may be the most informative for regulatory purposes.

Neuronal calcium/calmodulin-dependent protein kinase II mediates nicotine reward in the conditioned place preference test in mice.

Several recent studies have indicated the involvement of calcium-dependent mechanisms, in particular the abundant calcium-activated kinase, calcium/calmodulin-dependent kinase II (CaMKII), in behaviors associated with nicotine dependence in mice. Behavioral and biochemical studies have shown that CaMKII is involved in acute and chronic nicotine behaviors and nicotine withdrawal; however, evidence of a role for CaMKII in nicotine reward is lacking. Thus, the goal of the current study was to examine the role of CaMKII in nicotine reward. Using pharmacological and genetic tools, we tested nicotine conditioned place preference (CPP) in C57Bl/6 mice after administration of CaMKII antagonists and in α-CaMKII wild-type (+/+) and heterozygote (±) mice. CaMKII antagonists blocked expression of nicotine CPP, and the preference score was significantly reduced in α-CaMKII ± mice compared with their +/+ counterparts. Further, we assessed CaMKII activity in the ventral tegmental area (VTA), nucleus accumbens (NAc), prefrontal cortex, and hippocampus after nicotine CPP and found significant increases in CaMKII activity in the mouse VTA and NAc that were blocked by CaMKII antagonists. The findings from this study show that CaMKII mediates nicotine reward and suggest that increases in CaMKII activity in the VTA and NAc are relevant to nicotine reward behaviors.

Effects of orally-bioavailable short-acting kappa opioid receptor-selective antagonist LY2456302 on nicotine withdrawal in mice.

Kappa opioid receptor (KOR) signaling has been implicated in mediating behavioral and biochemical effects associated with drug dependence. The most commonly used KOR antagonists, norbinaltorphimine (norBNI) and (3R)-7-Hydroxy-N{(1S)-1-{[(3R,4R)-4-(3-hydroxyphenyl)-3,4-dimethyl-1-piperidinyl]methyl}-2-methylpropyl}-1,2,3,4-tetrahydro-3-isoquinoline-carboxamide (JDTic), have provided a wealth of information in this area; however, the delayed onset and long-lasting effects of these antagonists complicate experimental design and interpretation of results, and make them less than ideal for clinical studies. Initial studies with the recently developed KOR antagonist, LY2456302, show that the compound is a short acting, high-affinity, selective KOR antagonist with therapeutic potential for mood disorders and ethanol use in animal models, and is well tolerated in humans. The goal of the current study was to evaluate the effectiveness of LY2456302 in alleviating the nicotine withdrawal syndrome in mice. Mice were chronically treated with nicotine for 14 days and physical and affective nicotine withdrawal signs were measured using a spontaneous nicotine withdrawal model and conditioned place aversion (CPA) following pre-treatment with LY2456302, administered orally. Vehicle treated nicotine withdrawn mice displayed significant anxiety-related behavior, somatic signs, hyperalgesia, and CPA. Similar to previous studies with norBNI and JDTic, LY2456302 alleviated the nicotine withdrawal syndrome, as evidenced by decreased expression of nicotine withdrawal induced anxiety-related behavior, somatic signs, and CPA, and increased hotplate latency in nicotine withdrawn mice following pre-treatment. Given the current results, and with its favorable pharmacokinetic and pharmacodynamic profile, LY2456302 may be a useful therapeutic agent for treatment of multiple aspects of the nicotine withdrawal syndrome.

Determination of behaviorally effective tobacco constituent doses in rats.

INTRODUCTION: While nicotine has been established as the primary addictive drug that promotes tobacco use, recent peer-reviewed studies suggest that tobacco smoke contains additional chemical constituents that may have addictive potential. Additional research is necessary to determine the addictive potential of these tobacco constituents individually and in combination with tobacco smoke condensate; however, the behaviorally effective constituent doses necessary to conduct such studies are unclear. The primary objective of this study was to conduct behavioral studies in adult rats to determine the relevant behaviorally effective doses of the tobacco constituents, cotinine, myosmine, and anatabine to be used in future studies assessing the addictive potential of these compounds. METHODS: Separate groups of adult male Sprague Dawley rats were treated with vehicle, nicotine, or various doses of cotinine, mysomine, or anatabine. Effects on locomotor activity were measured in 10-min bins for 60min. RESULTS: Nicotine (0.8mg/kg) produced a biphasic effect on locomotor activity, with hypoactivity during the first 10min and hyperactivity at 40-50min. In contrast, cotinine (0.1mg/kg) and myosmine (10-50mg/kg) decreased activity without a later increase. Anatabine significantly increased locomotor activity at 1mg/kg, but decreased it at 10mg/kg. Prominent effects on overt behavior were observed at anatabine doses of 10mg/kg and above. CONCLUSION: Nicotine, cotinine, myosmine, and anatabine produced distinct time- and dose-dependent patterns of effects on locomotor activity. Results from the study will aid in the selection of relevant doses for future studies assessing the addictive potential of these non-nicotine tobacco constituents.

Youth tobacco cessation: quitting intentions and past-year quit attempts.

BACKGROUND: Despite declining use of conventional tobacco products, youth use of non-cigarette tobacco has become prevalent; however, quitting behaviors remain largely unexplored. PURPOSE: To examine nationally representative data on quit intentions and past-year attempts to quit all tobacco use among current youth tobacco users. METHODS: In 2013, data were analyzed from the 2012 National Youth Tobacco Survey (NYTS). Weighted prevalence estimates of quit intentions and past-year quit attempts for current youth tobacco users are presented. RESULTS: Prevalence of quit intentions and past-year attempts to quit all tobacco use were 52.8% and 51.5%, respectively, among current youth tobacco users. Among non-mutually exclusive groups, current cigarette smokers had the highest prevalence of quit intentions (56.8%) and past-year quit attempts (52.5%), whereas current hookah users had the lowest prevalence of quit intentions (41.5%) and past-year quit attempts (43.7%). Quit intentions among black, non-Hispanics (65.0%) and Hispanics (60.4%) were significantly higher versus white, non-Hispanics (47.5%). Youth reporting parental advice against tobacco had significantly higher prevalence of quit intentions (56.7%) and past-year quit attempts (55.0%) than those not reporting parental advice. Youth who agreed all tobacco products are dangerous (58.5%) had significantly higher prevalence of quit intentions than those who disagreed (37.0%). CONCLUSIONS: Continued efforts are needed to better understand youth motivation for quitting all tobacco products. Public health messaging about the dangers of all tobacco and cessation efforts should be aimed at the full range of tobacco products, not just cigarettes, and tailored to meet the needs of youth polytobacco users.

Calcium/calmodulin-dependent protein kinase IV mediates acute nicotine-induced antinociception in acute thermal pain tests.

Calcium-activated second messengers such as calcium/calmodulin-dependent protein kinase II have been implicated in drug-induced antinociception. The less abundant calcium-activated second messenger, calcium/calmodulin-dependent protein kinase IV (CaMKIV), mediates emotional responses to pain and tolerance to morphine analgesia but its role in nicotine-mediated antinociception is currently unknown. The goal of this study was to evaluate the role of CaMKIV in the acute effects of nicotine, primarily acute nicotine-induced antinociception. CaMKIV knockout (-/-), heterozygote (+/-), and wild-type (+/+) mice were injected with various doses of nicotine and evaluated in a battery of tests, including the tail-flick and hot-plate tests for antinociception, body temperature, and locomotor activity. Our results show a genotype-dependent reduction in tail-flick and hot-plate latency in CaMKIV (+/-) and (-/-) mice after acute nicotine treatment, whereas no difference was observed between genotypes in the body temperature and locomotor activity assessments. The results of this study support a role for CaMKIV in acute nicotine-induced spinal and supraspinal pain mechanisms, and further implicate involvement of calcium-dependent mechanisms in drug-induced antinociception.

The α3ß4* nicotinic acetylcholine receptor subtype mediates nicotine reward and physical nicotine withdrawal signs independently of the α5 subunit in the mouse.

The 15q25 gene cluster contains genes that code for the α5, α3, and ß4 nicotinic acetylcholine receptor (nAChRs) subunits, and in human genetic studies, has shown the most robust association with smoking behavior and nicotine dependence to date. The limited available animal studies implicate a role for the α5 and ß4 nAChR subunits in nicotine dependence and withdrawal; however studies focusing on the behavioral role of the α3ß4* nAChR receptor subtype in nicotine dependence are lacking. Because of the apparent role of the α3ß4* nAChR subtype in nicotine dependence, the goal of the current study was to better evaluate the involvement of this subtype in nicotine mediated behavioral responses. Using the selective α3ß4* nAChR antagonist, α-conotoxin AuIB, we assessed the role of α3ß4* nAChRs in acute nicotine, nicotine reward, and physical and affective nicotine withdrawal. Because α5 has also been implicated in nicotine dependence behaviors in mice and can form functional receptors with α3ß4*, we also evaluated the role of the α3ß4α5* nAChR subtype in nicotine reward and somatic nicotine withdrawal signs by blocking the α3ß4* nAChR subtype in α5 nAChR knockout mice with AuIB. AuIB had no significant effect on acute nicotine behaviors, but dose-dependently attenuated nicotine reward and physical withdrawal signs, with no significant effect in affective withdrawal measures. Interestingly, AuIB also attenuated nicotine reward and somatic signs in α5 nAChR knockout mice. This study shows that α3ß4* nAChRs mediate nicotine reward and physical nicotine withdrawal, but not acute nicotine behaviors or affective nicotine withdrawal signs in mice. The α5 subunit is not required in the receptor assembly to mediate these effects. Our findings suggest an important role for the α3ß4* nAChR subtype in nicotine reward and physical aspects of the nicotine withdrawal syndrome.

Beta2-containing nicotinic acetylcholine receptors mediate calcium/calmodulin-dependent protein kinase-II and synapsin I protein levels in the nucleus accumbens after nicotine withdrawal in mice.

Nicotinic acetylcholine receptors are calcium-permeable and the initial targets for nicotine. Studies suggest that calcium-dependent mechanisms mediate some behavioral responses to nicotine; however, the post-receptor calcium-dependent mechanisms associated with chronic nicotine and nicotine withdrawal remain unclear. The proteins calcium/calmodulin-dependent protein kinase II (CaMKII) and synapsin I are essential for neurotransmitter release and were shown to be involved in drug dependence. In the current study, using pharmacological techniques, we sought to (a) complement previously published behavioral findings from our lab indicating a role for calcium-dependent signaling in nicotine dependence and (b) expand on previously published acute biochemical and pharmacological findings indicating the relevance of calcium-dependent mechanisms in acute nicotine responses by evaluating the function of CaMKII and synapsin I after chronic nicotine and withdrawal in the nucleus accumbens, a brain region implicated in drug dependence. Male mice were chronically infused with nicotine for 14 days, and treated with the ß2-selective antagonist dihydro-ß-erythroidine (DHßE), or the α7 antagonist, methyllycaconitine citrate (MLA) 20min prior to dissection of the nucleus accumbens. Results show that phosphorylated and total CaMKII and synapsin I protein levels were significantly increased in the nucleus accumbens after chronic nicotine infusion, and reduced after treatment with DHßE, but not MLA. A spontaneous nicotine withdrawal assessment also revealed significant reductions in phosphorylated CaMKII and synapsin I levels 24h after cessation of nicotine treatment. Our findings suggest that post-receptor calcium-dependent mechanisms associated with nicotine withdrawal are mediated through ß2-containing nicotinic receptors.