1. David Perry Professor Dept. of Pharmacology & Physiology 994-3541 phmdcp@gwumc.edu

3. Why study nicotine? Tobacco use is a major health problem Contributes to up to 3 million deaths annually worldwide Smoking rates increasing in some U.S. populations (women, teens) Smoking rates increasing in developing countries The tobacco habit is difficult to quit Although the toxic effects of tobacco result from other components, nicotine sustains habitual use: NICOTINE is ADDICTIVE!

4. Nicotine’s effects are mediated by nicotinic cholinergic receptors 2 major classes of nicotinic receptors Muscle Neuronal (=nAChR) Ganglionic; CNS Pentameric ligand-gated cation channels Permeable to Na +, K + ; some to Ca 2+ Multiple subtypes in CNS At least 12 subunits expressed “major” subtypes:   ; also 

5. Effects of Chronic Nicotine Exposure on Neuronal Nicotinic Receptors Overall Goal: Determine changes in numbers and function of nAChR receptor subtypes after chronic exposure to nicotine Characterize neurobiological changes w/ nicotine addiction Role of different subtypes in addiction & withdrawal Test for unique vulnerability of adolescents for nicotine addiction Strategy and Methods Animal model: osmotic minipumps / rats (adults &/or adolescents) Quantify changes in numbers of receptor subtypes Quantitative autoradiography; homogenate binding; immunoprecipitation Quantify changes in receptor subtype functional activity Neurotransmitter release; rubidium efflux Behavioral responses: conditioned place preference Gene expression: microarray analysis

6. Neuronal Nicotinic Receptors are up-regulated dramatically in brains from human smokers Receptor up-regulation also occurs in rats treated w/ nicotine for 2 weeks

7. Effect of chronic nicotine exposure on functional activity of neuronal nicotinic receptors, as measured by 86 Rb efflux Change in functional activity correlates with change in receptor numbers Both changes correlate w/ presence of  4  2 subtype

8. The  6  subtype is labeled by  -conotoxin MII Quantitative autoradiography and immunoprecipitation reveal that, unlike other subtypes of nAChR, the  6 subtype is down- regulated by chronic nicotine exposure Down-regulation of  6* receptors in striatum is accompanied by a decrease in conotoxin-sensitive nicotine-stimulated dopamine release

9. Current & Future Directions Other subtypes :  7;  3  4; Adolescents: Those who initiate smoking as young adolescents are less able to quit later Model in rats (chronic nicotine in adolescents, adults) Assess differential outcomes: receptor function & number long-term behavioral changes in adult rats after adolescent exposure (CPP; motor activity) long-term changes in gene expression in adult rats after adolescent exposure (microarray analysis) Smoking & Pregnancy : w/ SPHHS & DC-STEP NRT for pregnant women ETS exposure in infants Animal studies: effects of prenatal nicotine exposure on receptor numbers and functional status