Role of nicotinic acetylcholine receptor subtypes α4β2 and α7 in nicotine-ethanol interaction and cross-tolerance: functional correlation with cerebellar nitric oxide

Abstract

The prevalent co-abuse of alcohol and tobacco products results in higher morbidity and mortality rates than those caused by either drug alone, thus posing a serious health problem. Understanding the interactive relationship between nicotine and ethanol might lead to effective strategies for the treatment of co-addiction. This dissertation project was designed to investigate the role of two nicotinic-acetylcholine-receptor (nAChR) subtypes (i.e. α4β2 and α7) on nicotine’s attenuation of ethanol ataxia and its functional correlation with alterations in cerebellar nitric oxide concentration. Expression levels of cerebellar α4β2 and α7 subtypes were also determined immunohistochemically.

Ethanol (2g/kg;i.p.)-induced ataxia was assessed by Rotorod in stereotaxically cannulated CD-1 male mice following the acute or repeated intracerebellar (ICB) microinfusions of α4β2- and α7-selective agonists, RJR-2403 and PNU-282987. Acute RJR-2403 (31, 62, 125ng), and PNU-282987 (25ng, 250ng, 2.5μg), dose-dependently reduced ethanol ataxia. Pretreatment with potent α4β2 [Dihydro-β-erythroidine (DHβE)] and α7 [Methyllycaconitine (MLA)]-subtype–selective antagonists prevented RJR-2403 and PNU-282987’s attenuation of ethanol ataxia, respectively. Both antagonists also offset nicotine’s reduction of ethanol ataxia, confirming the contribution of α4β2 and α7 subtype in ethanol ataxia. There was no tonic role of either subtype in ethanol ataxia. Additionally, ICB α4β2 and α7 subtype antisense treatment data correspond with agonist-induced behavioral responses. Animals repeatedly microinfused with RJR- 2403 or PNU-282987 became tolerant to ethanol ataxia. The observed cross-tolerance was faster in onset and longer in duration with PNU-282987 than RJR-2403. Pretreatment with DHβE and MLA, prevented the development of cross-tolerance.

The cerebellar nitrite+nitrate (NOx) levels were significantly enhanced and reduced following acute/repeated RJR- 2403/PNU-282987 microinfusion and acute ethanol

injection, respectively. Pretreatment with RJR- 2403 or PNU-282987 followed by ethanol prevented the ethanol-induced decrease in NOx concentration in both acute and repeated treatment paradigms, thus correlating the decrease in NOx concentration with ataxia and elevation with attenuation of ataxia. Both α4β2 and α7 subtypes exhibited high immunoreactivity in Purkinje, however, expression in molecular and granular cell layers was sparse. Overall, the results of the project support the role of α4β2 and α7 subtypes in the functional interaction between nicotine and acute ethanol ataxia, with NO-cGMP signaling as a participating factor.