Nicotinic Acetylcholine Receptors as Targets for Tobacco Cessation Therapeutics: Cutting-Edge Methodologies to Understand Receptor Assembly and Trafficking.
ABSTRACT: Tobacco dependence is a chronic relapsing disorder and nicotine, the primary alkaloid in tobacco, acts at nicotinic receptors to stimulate dopamine release in brain, which is responsible for the reinforcing properties of nicotine, leading to addiction. Although the majority of tobacco users express the desire to quit, only a small percentage of those attempting to quit are successful using the currently available pharmacotherapies. Nicotine upregulates the number of specific nicotinic receptors on the neuronal cell surface. An increase in receptor trafficking or preferential stoichiometric assembly of receptor subunits involves changes in assembly, endoplasmic reticulum export, vesicle transport, decreased degradation, desensitization, enhanced maturation of functional pentamers, and pharmacological chaperoning. Understanding these changes on a mechanistic level is important to the development of nicotinic receptors as drug targets. For this reason, cutting-edge methodologies are being developed and employed to pinpoint distinct changes in localization, assembly, export, vesicle trafficking, and stoichiometry in order to further understand the physiology of these receptors and to evaluate the action of novel therapeutics for smoking cessation.
Project description:Neuronal nicotinic acetylcholine receptors are activated by both endogenous acetylcholine and exogenous nicotine, making sequence variations in these receptors likely candidates for association with tobacco phenotypes. Previous studies have found evidence for significant association between single nucleotide polymorphisms (SNPs) in the genomic region containing the CHRNA6 and CHRNB3 genes and tobacco behaviors. In this study, we provide support for an association between these genes and tobacco dependence in the National Youth Survey Family Study wave 10, a nationally representative sample of households. Eight SNPs in the CHRNA6 and CHRNB3 genomic region were genotyped in 1051 subjects, approximately half of whom are members of sibling pairs. Genetic association with DSM-IV dependence was assessed using a family-based approach as implemented in the statistical package PBAT. Individual SNPs were tested for association with quit attempts and overall dependence. Variation in CHRNA6 was found to be associated with tobacco dependence (p=0.007 in Caucasians). SNPs in CHRNB3 were found to be associated with the number of quit attempts (p=0.0024). Together these results further implicate the region downstream of CHRNA6 and the region upstream of CHRNB3 in risk of nicotine dependence.
Project description:Diseases associated with tobacco use constitute a major health problem worldwide. Upon cessation of tobacco use, an unpleasant withdrawal syndrome occurs in dependent individuals. Avoidance of the negative state produced by nicotine withdrawal represents a motivational component that promotes continued tobacco use and relapse after smoking cessation. With the modest success rate of currently available smoking cessation therapies, understanding mechanisms involved in the nicotine withdrawal syndrome are crucial for developing successful treatments. Animal models provide a useful tool for examining neuroadaptative mechanisms and factors influencing nicotine withdrawal, including sex, age, and genetic factors. Such research has also identified an important role for nicotinic receptor subtypes in different aspects of the nicotine withdrawal syndrome (e.g., physical vs. affective signs). In addition to nicotinic receptors, the opioid and endocannabinoid systems, various signal transduction pathways, neurotransmitters, and neuropeptides have been implicated in the nicotine withdrawal syndrome. Animal studies have informed human studies of genetic variants and potential targets for smoking cessation therapies. Overall, the available literature indicates that the nicotine withdrawal syndrome is complex, and involves a range of neurobiological mechanisms. As research in nicotine withdrawal progresses, new pharmacological options for smokers attempting to quit can be identified, and treatments with fewer side effects that are better tailored to the unique characteristics of patients may become available. This article is part of the Special Issue entitled 'The Nicotinic Acetylcholine Receptor: From Molecular Biology to Cognition'.
Project description: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.
Project description:Nicotine addiction is initiated by its binding to high-affinity nicotinic receptors in brain composed primarily of alpha4 and beta2 subunits. For nicotinic receptors expressed in vivo or heterologously, nicotine exposure over hours to days increases or "upregulates" high-affinity nicotine binding to receptors through a posttranslational mechanism thought to increase receptor numbers. Using heterologous expression, we find nicotine exposure causes a fourfold to sixfold higher binding to alpha4beta2 receptors that does not correspond with any significant change in the number of surface receptors or a change in the assembly, trafficking, or cell-surface turnover of the receptors. However, upregulation does alter the functional state of the receptor, slowing desensitization and enhancing sensitivity to acetylcholine. Based on these findings, we propose an alternative mechanism to explain nicotine-induced upregulation in which nicotine exposure slowly stabilizes alpha4beta2 receptors in a high-affinity state that is more easily activated, thereby providing a memory for nicotine exposure.
Project description:Nicotine is the main tobacco component responsible for tobacco addiction and is used extensively in smoking and smoking cessation therapies. However, little is known about its effects on the immune system. We confirmed that multiple nicotinic receptors are expressed on mouse and human cytotoxic T lymphocytes (CTLs) and demonstrated that nicotinic receptors on mouse CTLs are regulated during activation. Acute nicotine presence during activation increases primary CTL expansion in vitro, but impairs in vivo expansion after transfer and subsequent memory CTL differentiation, which reduces protection against subsequent pathogen challenges. Furthermore, nicotine abolishes the regulatory effect of rapamycin on memory CTL programming, which can be attributed to the fact that rapamycin enhances expression of nicotinic receptors. Interestingly, naïve CTLs from chronic nicotine-treated mice have normal memory programming, which is impaired by nicotine during activation in vitro. In conclusion, simultaneous exposure to nicotine and antigen during CTL activation negatively affects memory development.
Project description:Genes coding for nicotinic acetylcholine receptors may influence response to nicotine replacement therapy for smoking cessation. We examined the association of a 3' untranslated region polymorphism (rs2072661) in the nicotinic acetylcholine receptor beta2 subunit (CHRNB2) gene with quitting success in response to nicotine versus placebo patch during a short-term test of patch effects. In a within-subjects cross-over design, smokers of European descent (n = 156) received 21 mg nicotine and placebo patch in counter-balanced order, during two separate 5-day simulated quit attempts, each preceded by a week of ad libitum smoking. Abstinence was assessed daily by CO < 5 ppm. Smokers with the CHRNB2 GG genotype had more days of abstinence during the nicotine versus placebo patch week compared with those with the AG or AA genotypes (P < 0.01). Moreover, nicotine patch increased the probability of quitting on the target quit day, quitting anytime during the patch week, and avoiding relapse among those with the GG genotype but not the AA/AG genotypes, although the nicotine x genotype interaction was significant only for quitting on the target quit day (P < 0.05). Regardless of patch condition, quitting on the target quit day was more likely in those with the GG genotype versus AA/AG genotypes (P < 0.05). Genetic associations were not observed for craving or withdrawal responses to nicotine versus placebo patch. These findings are consistent with previous evidence of association of this variant with smoking cessation and suggest that polymorphisms in the nicotinic acetylcholine receptor beta2 subunit gene may influence therapeutic responsiveness to cessation medications.
Project description:The long-term goal of this project is to establish whether and how chronic nicotine exposure affects nervous system function. The biological targets of nicotine action are diverse members of the superfamily of neurotransmitter-gated ion channels called nicotinic acetylcholine receptors (nAChR). nAChR play multiple, critical roles in chemical signaling throughout the brain and body. They also must be involved in nicotine dependence, which drives tobacco product use responsible for tremendous economic and personal costs. To define changes in gene expression induced by nicotine exposure in a model neuronal cell lines expressing at least two nicotinic receptor subtypes. Nicotine exposure exerst at least some of its effects on nervous system function by altering gene expression. Cells of the SH-SY5Y human neuroblastoma will be exposed to an efficacious dose of nicotine or to control medium for two different periods. Keywords: time-course
Project description:The acronym SePhaChARNS, for "selective pharmacological chaperoning of acetylcholine receptor number and stoichiometry," is introduced. We hypothesize that SePhaChARNS underlies classical observations that chronic exposure to nicotine causes "upregulation" of nicotinic receptors (nAChRs). If the hypothesis is proven, (1) SePhaChARNS is the molecular mechanism of the first step in neuroadaptation to chronic nicotine; and (2) nicotine addiction is partially a disease of excessive chaperoning. The chaperone is a pharmacological one, nicotine; and the chaperoned molecules are alpha4beta2* nAChRs. SePhaChARNS may also underlie two inadvertent therapeutic effects of tobacco use: (1) the inverse correlation between tobacco use and Parkinson's disease; and (2) the suppression of seizures by nicotine in autosomal dominant nocturnal frontal lobe epilepsy. SePhaChARNS arises from the thermodynamics of pharmacological chaperoning: ligand binding, especially at subunit interfaces, stabilizes AChRs during assembly and maturation, and this stabilization is most pronounced for the highest-affinity subunit compositions, stoichiometries, and functional states of receptors. Several chemical and pharmacokinetic characteristics render exogenous nicotine a more potent pharmacological chaperone than endogenous acetylcholine. SePhaChARNS is modified by desensitized states of nAChRs, by acid trapping of nicotine in organelles, and by other aspects of proteostasis. SePhaChARNS is selective at the cellular, and possibly subcellular, levels because of variations in the detailed nAChR subunit composition, as well as in expression of auxiliary proteins such as lynx. One important implication of the SePhaChARNS hypothesis is that therapeutically relevant nicotinic receptor drugs could be discovered by studying events in intracellular compartments rather than exclusively at the surface membrane.
Project description:Tobacco smoking is a leading preventable cause of death in the United States and produces a major health and economic burden. Although the majority of smokers want to quit, few are successful. These data highlight the need for additional research into the neurobiology of tobacco dependence. Addiction to nicotine, the main psychoactive component of tobacco, is influenced by multiple factors that include individual differences in genetic makeup. Twin studies have demonstrated that genetic factors can influence vulnerability to nicotine addiction, and subsequent research has identified genes that may alter sensitivity to nicotine. In humans, genome-wide and candidate gene association studies have demonstrated that genes encoding nicotinic acetylcholine receptor (nAChR) proteins are associated with multiple smoking phenotypes. Similarly, research in mice has provided evidence that naturally occurring variability in nAChR genes is associated with changes in nicotine sensitivity. Furthermore, the use of genetic knockout mice has allowed researchers to determine the nAChR genes that mediate the effects of nicotine, whereas research with knockin mice has demonstrated that changes to nAChR genes can dramatically alter nicotine sensitivity. This review will examine the genetic factors that alter susceptibility to nicotine addiction, with an emphasis on the genes that encode nAChR proteins.
Project description:The long-term goal of this project is to establish whether and how chronic nicotine exposure affects nervous system function. The biological targets of nicotine action are diverse members of the superfamily of neurotransmitter-gated ion channels called nicotinic acetylcholine receptors (nAChR). nAChR play multiple, critical roles in chemical signaling throughout the brain and body. They also must be involved in nicotine dependence, which drives tobacco product use responsible for tremendous economic and personal costs. To define changes in gene expression induced by nicotine exposure in a model neuronal cell lines expressing at least two nicotinic receptor subtypes. Nicotine exposure exerst at least some of its effects on nervous system function by altering gene expression. Cells of the SH-SY5Y human neuroblastoma will be exposed to an efficacious dose of nicotine or to control medium for two different periods.