Molecular and Behavioral Analysis of Nicotine Dependence (Project 1) - Blendy/Gould
A major challenge in nicotine addiction research today is determining why smoking behaviors endure or can be reinstated after abstinence. While reward is clearly an integral part of the addictive nature of nicotine, relapse to smoking is often motivated by the desire to alleviate withdrawal symptoms, including deficits in attention, learning and memory and affect. Nicotine abstinence symptoms that promote smoking relapse are most evident in the first few days after quitting, suggesting this as an important period to investigate neural mechanisms that may contribute to smoking relapse. Project 1 examines the behavioral, cellular, and molecular mechanisms underlying nicotine dependence by focusing on behavioral changes that occur during the critical period immediately following nicotine cessation. Our ultimate goal is to identify therapeutic targets for smoking cessation through a comprehensive behavioral and molecular analysis in preclinical models. We will achieve this goal through the following specific aims:
1. To determine the effects of nicotine withdrawal on cognition and affective behaviors, and to evaluate reversal of these effects by nicotine re-exposure and medication (varenicline based on the "best in class" model). Nicotine deprivation effects on contextual conditioning (fear conditioning) have already been demonstrated by the Gould laboratory at 24 hours post cessation of nicotine treatment. Thus, we will assess a broader range of cognitive functions such as trace conditioning (working memory), spatial learning, and appetitive learning (to examine operant learning and impulsivity), and 5CSRTT (attention and impulsivity). Furthermore, we will evaluate effects of chronic nicotine on models of anxiety and depression to assess affective components of nicotine deprivation.
2. To identify alterations in neural activity underlying the effects of nicotine withdrawal. Chronic nicotine exposure and subsequent withdrawal likely produces a distinct neural signature in the brains of mice that have undergone behavioral tasks such as contextual conditioning and demonstrate cognitive deficits. Nicotine re-exposure and/or varenicline will alter this activation pattern. We will use the immediate early gene FOS and phosphorylation of the transcription factor CREB (P-CREB) as related, but independent measures of neuronal activity. Beta2 KO mice show normal contextual conditioning when withdrawn from chronic nicotine treatment, thus, changes in FOS and P-CREB will be examined in the KO and WT mice as an important control group. In addition we will use a retrograde neuronal tracer, injected into the hippocampus, to identify afferent brain regions to determine if these projections are altered following nicotine withdrawal and learning. Lastly, we will use brain site–specific varenicline drug delivery to dissect brain regions involved in the ameliorative effects of the drug on withdrawal-related deficits (including regions identified as clinically relevant in CIRNA project 2 (human neuroimaging).
3. To identify genes altered by nicotine withdrawal. Nicotine withdrawal will activate a unique set of genes that contribute to cognitive deficits, or altered affective behaviors, some of which will be good candidates for medication screening or development. Brain regions identified in our circuitry mapping and/or in Project 2 will be macrodissected. RNA analysis in conjunction with chromatin immunoprecipitation will be performed using a CREB specific antibody. DNA elements occupied by the transcription factor CREB will be identified using Whole-genome Chromatin IP Sequencing (ChIP-Seq). All data will be analyzed, annotated, and made available as high quality reference data to the community using the tools available in the Biospecimen and Genotyping Core.
The successful use of our behavioral models in mice will allow us to screen other novel compounds in these tests with clinically relevant established behavioral criteria which ultimately would inform Project 2 as to classes of medications that could be tested in humans following early abstinence in imaging studies and ultimately in a clinical population in Project 3. Mice are a tractable model system with which to investigate underlying molecular mechanisms for nicotine dependence. The action of varenicline as a prototypical smoking cessation compound will enable us to compare this compound to nicotine in Aims 2 and 3 of this project with respect to neural activation and gene profiling. Results obtained in our gene profiling experiments in Aim 3 of this project will inform Projects 2 and 3 as to candidate genes to examine in smokers for associations with withdrawal-induced cognitive deficits. Principal Investigators: Julie Blendy, Ph. D. & Thomas Gould, Ph.D.
Neural Substrates of Smoking Cessation and Medication Response (Project 2) - Lerman
This project seeks to identify the neural and behavioral mechanisms underlying medication effects on cognitive and other symptoms of nicotine withdrawal, using blood oxygen level dependent (BOLD) functional MRI (fMRI). Varenicline, an α4β2 nicotinic acetylcholine receptor partial agonist, will be used as the model medication based on its documented clinical efficacy. The primary study is a within-subject double-blind cross-over neuroimaging study of short-term (13 days) varenicline (vs. placebo) treatment. Treatment-seeking smokers will undergo BOLD fMRI during 2 medication periods: (a) after 3 days of monitored abstinence while on varenicline, and (b) after 3 days of monitored abstinence while on placebo (medication order counterbalanced with a 2-week washout). BOLD fMRI data will be acquired while subjects perform working memory and emotional processing tasks. The primary outcome is medication effects (within subject) on: task-related BOLD activation (BOLD fMRI) after 3 days of abstinence. Changes in behavioral performance and subjective symptoms will be explored in relation to brain activity changes.
A second experiment seeks to characterize the effects of early nicotine withdrawal on brain and behavior, including measures of cerebral blood flow, functional connectivity, resting BOLD response, and task-induced BOLD response. Participants are followed prospectively to assess quitting success.
These experiments will contribute to pharmacotherapy development for nicotine dependence by: (a) providing a profile of varenicline effects on brain activity against which future novel compounds can be compared, and (b) establishing neuroimaging as an early “surrogate marker” for smoking relapse and medication response. Principal Investigator: Caryn Lerman, Ph.D.
Project 2 - Related Publications
Patterson F, Jepson C, Strasser AA, Loughead J, Perkins KA, Gur RC, Frey JM, Siegel S, Lerman C. Varenicline improves mood and cognition during smoking abstinence. Biological Psychiatry, 2009; 65(2):144-149.
Loughead J, Ray R, Wileyto EP, Ruparel K, Sanborn P, Siegel S, Gur RC, Lerman, C. Effects of the a4b2 partial agonist varenicline on brain activity and working memory in abstinent smokers. Biological Psychiatry, 2010; 67(8):715-721.
Patterson F, Jepson C, Loughead J, Perkins K, Strasser AA, Siegel S, Frey J, Gur R, Lerman C. Working memory deficits predict short-term smoking resumption following brief abstinence. Drug and Alcohol Dependence, 2010; 106:61-64.
Loughead J, Ray R, Wileyto EP, Ruparel K, O’Donnell GP, Senecal N, Siegel S, Gur RC, Lerman C. Brain activity and emotional processing in smokers treated with varenicline. Addiction Biology, 2011, Epub ahead of print.
Early Human Screening for Cessation Medications (Project 3) - Perkins
As part of the CIRNA P50 Center at The University of Pennsylvania, we are developing a short-term procedure to improve the efficiency of early screening of potential smoking cessation medications in humans. The aim is to optimally combine laboratory and clinical trial methods to take advantage of the experimental control of the former and the clinical validity of the latter. This new approach is expected to identify drugs that should, or should not, be subjected to long and expensive clinical trials of efficacy, so that resources are better directed toward more promising medications. Our first study, with nicotine patch, indicated that medications enhance short-term abstinence as a function of intrinsic quit motivation (i. e. intention to quit soon) but not extrinsic quit motivation (monetary reinforcement of abstinence). A subsequent cross-validation study, with varenicline, shows that these findings are applicable broadly to other medications, confirming the sensitivity of our approach. Ongoing work will determine the sensitivity and specificity of the procedure prior to the testing of novel medications. When final, this screening procedure may greatly enhance the efficiency of medication screening, decreasing costs and increasing the speed of development. Principal Investigator: Kenneth Perkins, Ph.D.
Project 3 - Related Publications
Perkins KA, Mercincavage M, Fonte C, Lerman C. Varenicline’s effects on acute smoking behavior and reward and their association with subsequent abstinence. Psychopharmacology, 2010; 210(1):45-51.Perkins KA, Lerman C, Fonte CA, Mercincavage M, Stitzer ML, Chengappa KNR, Jain A. Cross-validation of a new procedure for early screening of smoking cessation medications in humans. Clinical Pharmacology and Therapeutics, 2010; 88(1):109-114.
Perkins KA, Lerman C. Early human screening of medications to treat drug addiction: novel paradigms and the relevance of pharmacogenetics. Clinical Pharmacology and Therapeutics, 2011; 89(3):460-463.