BACKGROUND AND PURPOSENicotinic acetylcholine receptors (nAChRs) containing a6b2 subunits expressed by dopamine neurons regulate nicotine-evoked dopamine release. Previous results show that the a6b2* nAChR antagonist, N,NЈ-dodecane-1,12-diyl-bis-3-picolinium dibromide (bPiDDB) inhibits nicotine-evoked dopamine release from dorsal striatum and decreases nicotine self-administration in rats. However, overt toxicity emerged with repeated bPiDDB treatment. The current study evaluated the preclinical pharmacology of a bPiDDB analogue. EXPERIMENTAL APPROACHThe C10 analogue of bPiDDB, N, , was evaluated preclinically for nAChR antagonist activity. KEY RESULTSbPiDI inhibits nicotine-evoked [ 3 H]dopamine overflow (IC50 = 150 nM, Imax = 58%) from rat striatal slices. Schild analysis revealed a rightward shift in the nicotine concentration-response curve and surmountability with increasing nicotine concentration; however, the Schild regression slope differed significantly from 1.0, indicating surmountable allosteric inhibition. Co-exposure of maximally inhibitory concentrations of bPiDI (1 mM) and the a6b2* nAChR antagonist a-conotoxin MII (1 nM) produced inhibition not different from either antagonist alone, indicating that bPiDI acts at a6b2* nAChRs. Nicotine treatment (0.4 mg·kg -1 ·day -1 , 10 days) increased more than 100-fold the potency of bPiDI (IC50 = 1.45 nM) to inhibit nicotine-evoked dopamine release. Acute treatment with bPiDI (1.94-5.83 mmol·kg -1 , s.c.) specifically reduced nicotine self-administration relative to responding for food. Across seven daily treatments, bPiDI decreased nicotine self-administration; however, tolerance developed to the acute decrease in food-maintained responding. No observable body weight loss or lethargy was observed with repeated bPiDI. CONCLUSIONS AND IMPLICATIONSThese results are consistent with the hypothesis that a6b2* nAChR antagonists have potential for development as pharmacotherapies for tobacco smoking cessation.Abbreviations bPIDDB, N, bPiDI, N,picolinium diiodide; DHbE, dihydro-b-erythroidine; FR5, fixed ratio 5; NAcc, nucleus accumbens; nAChR, nicotinic acetylcholine receptor; s.c., subcutaneous; TO180, 180 min timeout; TO20, 20 min timeout; VTA, ventral tegmental area; a-CtxMII, a-conotoxin MII BJP British Journal of Pharmacology DOI:10.1111DOI:10. /j.1476DOI:10. -5381.2011 346 British Journal of Pharmacology (2011) IntroductionDrugs of abuse activate the dopaminergic reward circuitry, leading to dopamine release in nucleus accumbens (NAcc) and striatum, which is associated with primary reward and habit formation respectively (Di Chiara et al., 2004;Koob and Volkow, 2010). The transition from reward seeking to compulsive behaviour associated with drug abuse appears to result from a shift from NAcc to striatal control (Koob and Volkow, 2010). Nicotine activates nicotinic acetylcholine receptors (nAChRs; receptor nomenclature follows Alexander et al., 2009) that modulate dopamine release. Identifying nAChRs regulating dopamine release is important because dopami...

The novel nicotinic receptor (nAChR) antagonist, N,N’-dodecane-1,12-diyl-bis-3-picolinium dibromide (bPiDDB), and its chemically-reduced analog, r-bPiDDB, potently inhibit nicotine-evoked dopamine (DA) release from rat striatal slices. Since tobacco smokers self-administer nicotine repeatedly, animal models incorporating repeated nicotine treatment allow for mechanistic evaluation of therapeutic candidates following neuroadaptive changes. The current study determined the ability of bPiDDB, r-bPiDDB and α-conotoxin MII (α-CtxMII), a peptide antagonist selective for α6β2-containing nAChRs, to inhibit nicotine-evoked [3H]DA release from striatal slices from rats repeatedly administered nicotine (0.4 mg/kg for 10 days) or saline (control). Concomitant exposure to maximally effective concentrations of r-bPiDDB (1 nM) and α-CtxMII (1 nM) resulted in inhibition of nicotine-evoked [3H]DA release no greater than that produced by either antagonist alone, suggesting that r-bPiDDB inhibits α6β2-containing nAChRs. Repeated nicotine treatment increased locomotor activity, demonstrating behavioral sensitization. Concentration-response curves for nicotine-evoked [3H]DA release were not different between nicotine-treated and control groups. Maximal inhibition for α-CtxMII was greater following repeated nicotine compared to control (Imax=90% vs. 62%), with no change in potency. bPiDDB was 3-orders of magnitude more potent in inhibiting nicotine-evoked [3H]DA release in nicotine-treated rats compared to control rats (IC50=5 pM vs 6 nM), with no change in maximal inhibition. Neither a shift to left in the concentration response nor a change in maximal inhibition was observed for r-bPiDDB following repeated nicotine. Thus, repeated nicotine treatment may differentially regulate the stoichiometry, conformation and/or composition of α6β2-containing nAChRs mediating nicotine-evoked striatal DA release. Therefore, bPiDDB and r-bPiDDB appear to target different α6β2-containing nAChR subtypes.

A series of tetrakis-azaaromatic quaternary ammonium salts was synthesized in order to identify compounds with higher affinity and selectivity as antagonists at neuronal nicotinic receptor subtypes that mediate nicotine-evoked DA release. A high hit rate was achieved in identifying potent analogues that inhibit these nAChRs. Three tetrakis analogues, 11j, 11f and 11g, were identified as potent (IC50 = 3, 28 and 56 nM, respectively) antagonists at these receptors. These compounds represent a novel structural class of nicotinic receptor antagonists.

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Although several therapeutic agents are available to aid in tobacco smoking cessation, relapse rates continue to be high, warranting the development of alternative pharmacotherapies. Nicotine-evoked dopamine release from its presynaptic terminals in the central nervous system leads to reward which maintains continued tobacco use. The ability of indolizidine (−)-235B′ and a sub-library of structurally-related analogs to inhibit nicotine-evoked [3H]dopamine release from rat striatal slices was determined in the current study. Indolizidine (−)-235B′ inhibited nicotine-evoked [3H]dopamine release in a concentration-dependent manner (IC50 = 42 nM, Imax = 55%). Compound (−)-237D, the double bond-reduced analog, afforded the greatest inhibitory potency (IC50 = 0.18 nM, Imax = 76%), and was 233-fold more potent than indolizidine (−)-235B′. The des-8-methyl aza-analog of indolizidine (−)-235B′, ZZ-272, also inhibited nicotine-evoked [3H]dopamine release ((IC50 = 413 nM, Imax = 59%). Concomitant exposure to maximally effective concentrations of indolizidine (−)-235B′, ZZ-272 or (−)-237D with a maximally effective concentration of α-conotoxin MII, a selective antagonist for α6β2-containing nicotinic receptors, resulted in inhibition of nicotine-evoked [3H]dopamine release no greater than that produced by each compound alone. The latter results suggest that indolizidine (−)-235B′, (−)-237D, ZZ-272 and α-conotoxin MII inhibit the same α-conotoxin MII-sensitive nicotinic receptor subtypes. Thus, indolizidine (−)-235B′ and its analogs act as antagonists of α6β2-nicotinic receptors and constitute a novel structural scaffold for the discovery of pharmacotherapies for smoking cessation.