Tobacco cembranoids block behavioral sensitization to nicotine and inhibit neuronal acetylcholine receptor function

Abstract

Cembranoids are cyclic diterpenoids found in tobacco and in marine invertebrates. The present study established that tobacco cembranoids inhibit behavioral sensitization to nicotine in rats and block several types of nicotine acetylcholine receptors (AChRs). 1) At the behavioral level, rat locomotor activity induced by nicotine was significantly increased after seven daily nicotine injections. This sensitization to nicotine was blocked by mecamylamine (1 mg/kg) and by the cembranoids eunicin, eupalmerin acetate (EUAC), and (4R)-2,7,11-cembratriene-4-6-diol (4R), each at 6 mg/kg. None of these compounds modified locomotor activity of nonsensitized rats. 2) In cells expressing human AChRs, cembranoids blocked carbamoylcholine-induced (86)Rb(+) flux with IC(50) in the low micromolar range. The cell lines used were the SH-EP1-halpha4beta2 cell line heterologously expressing human alpha4beta2-AChR, the SH-SY5Y neuroblastoma line naturally expressing human ganglionic alpha3beta4-AChR, and the TE671/RD cell line naturally expressing embryonic muscle alpha1beta1gammadelta-AChR. The tobacco cembranoids tested were 4R and its diastereoisomer 4S, and marine cembranoids tested were EUAC and 12,13-bisepieupalmerin. 3) At the molecular level, tobacco (4R and 4S) and marine (EUAC) cembranoids blocked binding of the noncompetitive inhibitor [(3)H]tenocyclidine to AChR from Torpedo californica electric organ. IC(50) values were in the submicromolar to low-micromolar range, with 4R displaying an order of magnitude higher potency than its diastereoisomer, 4S.

Characterization of Human α4β2-Nicotinic Acetylcholine Receptors Stably and Heterologously Expressed in Native Nicotinic Receptor-Null SH-EP1 Human Epithelial Cells

“…Excellent expression of functional ␣4␤2-nAChR assayable using ion flux (current study; Ferchmin et al, 2001), electrophysiological (see Wu et al, 2001), or calcium imaging techniques (Pacheco et al, 2001) has been achieved, giving advantage over transfected cell lines expressing rat ␣4␤2-nAChR-like binding sites but not yet shown to express high levels of functional receptors (Cooper et al, 1999;Shafaee et al, 1999). The expression of h␣4␤2-nAChR from entirely wild-type subunits and for cells maintained using conventional techniques has advantages over other model systems that have employed altered incubation temperatures or have resorted to creation of chimeric subunits with transmembrane and cytoplasmic domains from non-nAChR subunits to get good surface and functional expression (Cooper et al, 1999).…”

Neuroprotective activity of (1S,2E,4R,6R,-7E,11E)-2,7,11-cembratriene-4,6-diol (4R) in vitro and in vivo in rodent models of brain ischemia

“…Although the optimal neuroprotective dose of 4R in vivo is not known, 6 mg/kg 4R did not alter exploratory activity in naïve rats but abolished behavioral sensitization to nicotine (Ferchmin et al, 2001). In vitro experiments suggest that even at lower doses, 4R could be effective (Ferchmin et al, 2005, Eterovic et al, 2013).…”

“…Our group discovered that certain cembranoids are noncompetitive inhibitors of nicotinic acetylcholine receptors (nAChRs) (Eterovic et al, 1993; Ferchmin et al, 2001; Hann et al, 1998). As one of cembranoids, 4R (Fig.…”

4R-cembranoid protects against diisopropylfluorophosphate-mediated neurodegeneration

“…The optimal neuroprotective dose of 4R in vivo is not known; however, at 6 mg/kg, 4R did not alter exploratory activity in naïve rats but abolished behavioral sensitization to nicotine (Ferchmin et al, 2001). In collaboration with SRI (Stanford Research International; Menlo Park, California), it was found that up to 98 mg/kg 4R was not toxic (manuscript in preparation).…”