Background A growing body of research suggests the therapeutic use of cannabis may affect the use of other substances, including reducing the use of alcohol, tobacco and prescription drugs such as opioid analgesics. However, most of the evidence stems from small, cross-sectional surveys or population-level studies, both of which have significant limitations, including the inability to conclusively determine causality for behavioural changes. Furthermore, very little detail has been gathered on the factors that potentially impact substitution, including patient characteristics and patterns of cannabis use (e.g., X, Y, Z). Additionally, despite consistent calls by physicians, academics, patients and policy-makers around the globe citing the need for high quality studies to identify the risks and benefits of cannabis in both medical and non-medical applications, there are many pre-existing and ongoing challenges to conducting such research. These include shifting regulatory policies that may be impacting access to cannabis for both medical and non-medical use, and that could ultimately be affecting patient retention in prospective medical cannabis studies. In the interest of learning more about how the use of cannabis effects the use of alcohol, tobacco and other substances, and to better understand factors that may be impacting retention in prospective cannabis research, I designed and conducted two studies: 1. The Canadian Cannabis Patient Survey 2019 (CCPS 2019) was a national cross-sectional survey of 2102 Canadian medical cannabis patients that examined demographics, patient patterns of cannabis use, and self-reported changes in the use of alcohol, tobacco, prescription drugs and illicit substances following medical cannabis initiation. 2. The Tilray Observational Patient Survey (TOPS) was a prospective, multi-site, observational study examining the impact of medical cannabis products on quality of life and the use of prescription drugs of 1145 patients over a 6 month period, which provided an opportunity to conduct a survival analysis and other analyses to assess variables potentially impacting retention in longitudinal cannabis studies. Methods This dissertation includes three analyses of the data resulting from these studies in the form of one published and two submitted manuscripts. The first paper provides an overview of research to date examining the impact of cannabis and cannabinoids on alcohol use, followed by an analysis of the 973 CCPS 2019 participants who either previously or currently use alcohol. The questionnaire gathered a detailed inventory of alcohol use prior and post medical cannabis initiation using two separate but related measures: drinking days per month, and standard drinks per week. The analyses used descriptive statistics as well as univariate and multivariate regression analyses to explore patient characteristics and other variables potentially associated with changes in alcohol use post medical cannabis, including assessing the impact of “intent” to use medical cannabis to reduce alcohol use, as well as participation in other substance use treatment modalities. Findings suggest that medical cannabis initiation is associated with significant reductions in alcohol use, and that younger age (<55 years of age), specific intent to use medical cannabis to reduce alcohol use, and greater patterns of alcohol use prior to medical cannabis initiation were associated with greater odds of reducing alcohol. The second paper follows a similar methodology and format as the first paper, but with a focus on tobacco/nicotine (T/N) use. In this case, 650 survey participants reported past or current T/N use, and the analysis focused on assessing patient characteristics and other variables associated with changes in T/N uses per day, with the primary outcome of interest being no use in the 30 days prior to the survey, which was considered to be complete cessation of T/N use. The findings suggest that odds of T/N cessation were greater amongst those who were age 55 or older or that reported >25 T/N uses per day prior to initiating medical cannabis use, and that specific intent to use medical cannabis in T/N reduction/cessation efforts resulted in significantly greater odds of reducing T/N use, while involvement with traditional T/N cessation treatments (pharmacological or psychobehavioral) was negatively associated with T/N cessation. The third paper addresses the challenge of retaining patients in prospective observational medical cannabis studies at a time when there are major policy changes disrupting the legal supply while also increasing access options for adults who use cannabis. The Tilray Observational Patient Study (TOPS) was one of the largest national prospective medical cannabis studies ever conducted, taking place at 21 medical clinics in five provinces. The study was designed to assess the impact of medical cannabis on quality of life and prescription drug use over a six month period. However, initial data analysis on 1145 patients enrolled at/before Oct 15, 2018 that had completed at least one post baseline visit highlighted a significant rate of patients that were lost to follow up (LTFU). This high drop out rate, coupled with a compensation scheme that provided credits to help cover the cost of medical cannabis led to concerns of potential retention bias limited the conclusions that could be drawn from this data. However, the study and resulting data provided a unique opportunity to examine baseline patient characteristics that may have been protective of LTFU, so a survival analysis was conducted on this cohort. Additionally, since the study took place during the official launch of the legalization of adult non-medical use of cannabis in Canada on Oct. 17th, 2018, the potential impact of this significant increase in legal access options on the odds of study retention was the subject of additional analyses. The survival analysis found that baseline use of antidepressants or antiseizure medications, citing no preference for either THC or CBD, and inhalation as a primary method of use were associated with increased probability of survival/retention in the study at six months. Additionally, while the legalization of non-medical adult cannabis use in October 2018 resulted in more than three times the odds of participants being LTFU at six months, being under 55 years old, having a preference for THC, or citing inhalation as a primary method of use was partially protective of LTFU following legalization. Discussion The studies in this dissertation presented an opportunity to gather subjective and objective data on naturalistic patterns of medical cannabis use from large, heterogeneous cohorts of patients, and to explore associated impacts on the use of alcohol, tobacco and other substances. The results of these studies provide a more comprehensive understanding of the public health risks and benefits associated with the medical use of cannabis, and could subsequently inform policy decisions affecting access to cannabis vis-à-vis other drugs, private and public payer considerations related to cost-coverage for medical cannabis, and potentially lead to the development of novel alcohol and tobacco cessation strategies. Additionally, the survival analysis conducted on TOPS participants highlights some of the challenges of conducting medical cannabis research at a time when patients have a multitude of cannabis access options, including legal adult dispensaries and a still robust illicit market. Future longitudinal medical cannabis studies should consider the potential impact of policy changes effecting cannabis access on study retention/survival, and may want to focus on patient populations with characteristics associated with lower odds of LTFU. / Graduate / 2022-03-09

Rationale. Nicotine is often considered a ‘gateway’ drug because people typically experiment with tobacco before illicit drugs such as cocaine and amphetamine. We have shown that nicotine increases approach to reward-associated stimuli, this is referred to as ‘sign-tracking’, and that this effect persists after nicotine is discontinued. Individuals who are high in sign-tracking also show increased cocaine self-administration. Objectives. The goal of this experiment was to determine whether nicotine enhanced sign tracking could result in greater cocaine self-administration. Method. Rats were randomly assigned to one of 2 groups (NIC or SAL), and injected with their assigned solution (0.4 mg/kg base or placebo, respectively) 15 min before conditioning sessions. During conditioning sessions, a lever/light stimulus was inserted into the chamber for 15 s and immediately followed by sucrose delivery. Approach to the sucrose receptacle was recorded by monitoring head entries and defined as goal tracking. Contact with the lever was recorded and defined as ‘sign-tracking’. After 29 conditioning sessions, the rats were instrumented for cocaine self-administration and were shaped to respond for cocaine on the same lever that served as the CS. After 10 days of acquisition of cocaine self-administration (0.16 mg/inf), demand for cocaine was tested over 6 days using a within session procedure that increased cocaine price every 10 min. Results. We showed increased sign-tracking, but not goal tracking in the NIC group relative to the SAL group. The NIC group also showed increased demand for cocaine during the price manipulation, but the essential value of cocaine did not differ, relative to the SAL group. Conclusion. Our results support a gateway interpretation of substance use – when both the gateway drug (nicotine) and drug-associated rewards (the lever/light) occur together, they can promote future self-administration of illicit drugs such as cocaine.

Rationale: Nicotine (NIC) potently increases operant responding for non-NIC reinforcers, and this effect may depend on drug-mediated increases in incentive motivation. According to this hypothesis, NIC should also potently increase approach to Pavlovian-conditioned stimuli associated with rewards. Objective: The present studies explored the effects of NIC on Pavlovian-conditioned approach responses. Method: To do so, liquid dippers were used to deliver an unconditioned stimulus (US; 0.1 ml sucrose) after presentation of a conditioned stimulus (CS; 30 s illumination of a stimulus light) - both the CS and US were presented in receptacles equipped to monitor head entries. Results: In experiment 1, the CS and US were presented in the same receptacle, but NIC pretreatment (0.4 mg/kg base) did not increase conditioned approach responses. Delivery of the sucrose US was then shifted to receptacle in a different location. All rats learned to approach the new US location (goal-tracking) at similar rates. Approach to the CS receptacle (sign-tracking) declined for saline-pretreated rats, but NIC pretreatment increased sign-tracking. In experiment 2, NIC pretreatment increased sign-tracking when the CS and US were spatially separated during acquisition. In experiment 3, NIC pretreatments were replaced with saline, but the effect of NIC persisted for an additional 24 test sessions. Conclusion: The findings suggest that NIC increases incentive motivation and that this effect is long-lasting, persisting beyond the pharmacological effects of NIC.

Rationale: Nicotine (NIC) potently increases operant responding for non-NIC reinforcers, and this effect may depend on drug-mediated increases in incentive motivation. According to this hypothesis, NIC should also potently increase approach to Pavlovian-conditioned stimuli associated with rewards. Objective: The present studies explored the effects of NIC on Pavlovian-conditioned approach responses. Method: To do so, liquid dippers were used to deliver an unconditioned stimulus (US; 0.1 ml sucrose) after presentation of a conditioned stimulus (CS; 30 s illumination of a stimulus light) - both the CS and US were presented in receptacles equipped to monitor head entries. Results: In experiment 1, the CS and US were presented in the same receptacle, but NIC pretreatment (0.4 mg/kg base) did not increase conditioned approach responses. Delivery of the sucrose US was then shifted to receptacle in a different location. All rats learned to approach the new US location (goal-tracking) at similar rates. Approach to the CS receptacle (sign-tracking) declined for saline-pretreated rats, but NIC pretreatment increased sign-tracking. In experiment 2, NIC pretreatment increased sign-tracking when the CS and US were spatially separated during acquisition. In experiment 3, NIC pretreatments were replaced with saline, but the effect of NIC persisted for an additional 24 test sessions. Conclusion: The findings suggest that NIC increases incentive motivation and that this effect is long-lasting, persisting beyond the pharmacological effects of NIC.

Rationale: Varenicline (VAR), a smoking cessation aid that is a partial agonist at nicotinic receptors, mimics the reinforcement-enhancing effects of nicotine. Varenicline, when accompanied by non-drug cues, is self-administered by rats, though it is unclear whether this results from varenicline acting as a primary reinforcer or a reinforcement enhancer of the cues. Objectives: This study sought to disentangle these two potential actions. Methods: Rats were allowed to self-administer intravenous nicotine, saline, or varenicline during 1-h sessions in operant chambers equipped with two levers. Five groups had concurrent access to drug infusions and a moderately reinforcing visual stimulus (VS) for responding on separate levers. Meeting the reinforcement schedule on one lever was reinforced with VAR (0.01, 0.06, 0.1 mg/kg/infusion), nicotine (0.06 mg/kg/infusion), or saline, while meeting the same schedule on the other lever delivered the VS. Additional groups were reinforced for pressing a single "active" lever and received VAR paired with the VS, the VS with response-independent infusions of VAR, or VAR alone (0.1 mg/kg/infusion). Results: Rats readily responded for VAR paired with VS on a single lever. However, when VAR was the only reinforcer contingent on a response, rats did not respond more than for saline. Conclusions: These findings show that VAR does not serve as a primary reinforcer in rats at doses that increase responding for non-drug reinforcers. These data are consistent with research showing that the primary reinforcing effects of VAR are weak, at best, and that the primary reinforcing and reinforcement-enhancing actions of nicotinic drugs are pharmacologically distinct.

Theories of the neurobiological basis of Attention-Deficit/Hyperactivity Disorder (ADHD) have largely focused on dysregulation of central dopaminergic function. However, other neurotransmitter systems may be implicated in specific cognitive deficits in ADHD. Interest in the potential involvement of nicotinic cholinergic systems in ADHD has arisen in part from the observation that adolescents and adults with ADHD smoke cigarettes at significantly higher rates than people without this disorder. In addition, several studies report that nicotine alleviates ADHD symptoms, and recent neuro-genetics studies indicate that cholinergic systems may be altered in persons with ADHD. In this review, we describe the evidence for a role of central nicotinic cholinergic systems in cognitive deficits in ADHD. We also propose mechanisms by which alterations in cholinergic function may contribute directly and/or indirectly to these deficits. Finally, we identify specific paradigms and models to guide future investigations into the specific involvement of nicotinic cholinergic systems in ADHD, possibly leading to the development of more effective pharmacotherapies for ADHD.

Cigarette smoke is suspected to cause diverse illnesses in smokers and people breathing second- and third-hand smoke. Although different studies have been done to elucidate the impact on health due to smoking, there is a lack of kinetic information regarding the degradation of nicotine under different environmental conditions. As a consequence, currently it is not possible to determine thoroughly the risk due to exposure to nicotine and the compounds derived from its decomposition. With the aim of contributing to clarify the different degradation paths followed by nicotine during and after the consumption of cigarettes, this work presents a theoretical study of the hydrogen atom abstraction reaction by hydroxyl radical at four sites in the nicotine molecule in a broad range of temperature, specifically be-tween 200-3000 K. The site-specific kinetic rate constants were computed by means of the multi-structural torsional variational transition state theory with small curvature tunneling contribution, performing ab initio calculations at the level M06-2X/aug-cc-pVQZ//M06-2X/cc-pVTZ. According to our computations, the dependence on temperature of the studied rate constants exhibited a non-Arrhenius fashion, with a minimum at 873 K. A negative temperature dependence was observed at temperatures lower than 873 K, indicating more prolonged exposure to nicotine in warmer environments. On the other hand, the opposite behavior was observed at higher temperatures; this non-Arrhenius be-havior results of interest in tobacco cigarette combustion, inducing different reaction mechanisms depending on the burning conditions of the different smoking devices. The results indicate that multi-structural and torsional anharmonicity is an im-portant factor in the computation of accurate rate constants, especially at high tem-peratures where the higher-energy conformers of the different species exert a larger influence. The anharmonicity factors suggest that disregarding the anharmonic de-viations leads to overestimation of the rate constant coefficients, by a factor between four and six. Our computed overall kinetic rate constant at 298 K exhibited very good agreement with the only experimental value meausred by Borduas et al. [1], af-fording certainty about our calculated site-specific rate constants, which are currently inaccessible to experiments. However, further experimental studies are necessary to validate our kinetic studies at other temperatures.

Indiana University-Purdue University Indianapolis (IUPUI) / Streptococcus mutans, an inhabitant of oral biofilm or dental plaque, adheres to the tooth surface via protein antigen I/II (PA I/II). Pathologic lesions of atherosclerosis (AT) and infective endocarditis (IE) harbor S. mutans. Serotypes f and k strains with collagen binding protein genes cbm and cnm are uncommon in the mouth, but these are the most prevalent S. mutans strains in AT and IE tissues and can invade endothelial cells (EC) in vitro. Tobacco use increases the risk for cardiovascular and oral diseases. Oral S. mutans encounter many substances including nicotine. Arginine is present in saliva and the EC glycocalyx that coats and protects ECs from shear forces of blood flow. Prior studies demonstrated arginine alters S. mutans biofilm. This work characterizes S. mutans serotype k strains and serotype c strains, the most prevalent in the mouth. The effects of nicotine and arginine on biofilm mass, metabolic activity and EC invasion were investigated. Biofilm production by serotypes c and k strains did not differ; there were no differences in responses to nicotine and arginine between these serotypes. Increased production of biofilm was associated with the cbm and cnm genes. Nicotine increased biofilm for all strains whereas arginine plus nicotine reduced bacteria and the extracellular polymeric substances. Previous EC invasion studies were performed with planktonic cultures of S. mutans; therefore, EC invasion by biofilm was evaluated. Significant factors for EC invasion by S. mutans are presence of the cbm gene and lack of PA I/II expression on the bacterial cell surface. Presence of the cnm gene increased EC invasion by biofilm but not planktonic cells. Planktonic cells of six strains invaded better than biofilm, whereas four strains showed increased invasion by biofilm cells. Neither nicotine nor arginine significantly altered the ability of S. mutans biofilm cells to invade ECs. Not all strains with cbm or cnm and no PA I/II expression invaded EC. A strain with PA I/II expression and without cbm and cnm genes invaded EC. While cbm, cnm and PA I/II expression are predictors of EC invasion, additional mechanisms for EC invasion by S. mutans remain to be revealed. / 2021-08-21

A growing body of research is targeted towards characterizing and explaining nicotine’s complex interactions with the ApoE E4 allele on brain responses underlying cognitive processes. However, when and how the ε4 allele modulates neuroelectric brain responses in the presence of nicotine versus nicotine abstinence in nicotine-dependent smokers is not well characterized. Being able to understand this modulation is potentially quite important given that recent research implies that, relative to non-ε4 carriers, young adult carriers of the ε4 allele exhibit greater cognitive benefits from the use of nicotine. In the present study, electroencephalography (EEG) and the oddball-related P3b event-related potential (ERP) were used to better characterize the potential moderating effects of ApoE on P3b ERP amplitude changes associated with overnight nicotine deprivation in dependent smokers. Results showed a significant interaction between ApoE genotype and nicotine use, as ε4 carriers, relative to noncarriers, demonstrated significantly greater decreases following overnight deprivation, relative to prequit baseline levels. Additionally, there was a main of effect of P3b ERP amplitude to target stimuli being greater in ε4 allele carriers than in noncarriers during nicotine use, but no main effect of APOE genotype during overnight nicotine deprivation. These results are consistent with findings that the ApoE genotype moderates the effects of nicotine and alters neuroelectric brain responses associated with selective attention to infrequent target stimuli.

Rationale: Psychotic disorders such as schizophrenia are often accompanied by high rates of cigarette smoking, reduced quit success, and high relapse rates, negatively affecting patient outcomes. However, the mechanisms underlying altered relapse-like behaviors in individuals diagnosed with psychosis are poorly understood. Objectives: The present study analyzed changes in extinction and reinstatement of nicotine conditioned place preference (CPP) and resulting changes in brain-derived neurotrophic factor (BDNF) in a novel heritable rodent model of psychosis, demonstrating increased dopamine D2 receptor sensitivity, to explore mechanisms contributing to changes in relapse-like behaviors. Methods: Male and female offspring of two neonatal quinpirole-treated (QQ) and two neonatal saline-treated (SS) Sprague-Dawley rats (F1 generation) were tested on an extended CPP paradigm to analyze extinction and nicotine-primed reinstatement. Brain tissue was analyzed 60 min after the last nicotine injection for BDNF response in the ventral tegmental area (VTA), the infralimbic (IfL) and prelimbic (PrL) cortices. Results: F1 generation QQ offspring demonstrated delayed extinction and more robust reinstatement compared to SS control animals. In addition, QQ animals demonstrated an enhanced BDNF response to nicotine in the VTA, IfL and Prl cortices compared to SS offspring. Conclusions: This study is the first to demonstrate altered relapse-like behavior in a heritable rodent model with relevance to comorbid drug abuse and psychosis. This altered pattern of behavior is hypothesized to be related to elevated activity-dependent BDNF in brain areas associated with drug reinforcement during conditioning that persists through the extinction phase, rendering aberrantly salient drug associations resistant to extinction and enhancing relapse vulnerability.