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Nicotine Activates and Up-Regulates Nicotinic Acetylcholine Receptors in Bronchial Epithelial Cells

¹ Division of Neuroscience, Oregon National Primate Research Center, Oregon Health & Science University, Beaverton, Oregon; and ² Department of Neuroscience, University of Pennsylvania, Philadelphia, Pennsylvania

Correspondence and requests for reprints should be addressed to Eliot Spindel, M.D., Ph.D., Division of Neuroscience, Oregon National Primate Research Center, 505 NW 185th Ave., Beaverton, OR 97006. E-mail: Spindele{at}ohsu.edu

Prenatal nicotine exposure impairs normal lung development and leads to diminished pulmonary function after birth. Previous work from our laboratory has demonstrated that nicotine alters lung development by affecting a nonneuronal cholinergic autocrine loop that is expressed in lung. Bronchial epithelial cells (BECs) express choline acetyltransferase, the choline high-affinity transporter and nicotinic acetylcholine (ACh) receptor (nAChR) subunits. We now demonstrate through a combination of morphological and electrophysiological techniques that nicotine affects this autocrine loop by up-regulating and activating cholinergic signaling. RT-PCR showed the expression of 3, 4, 7, 9, 10, β2, and β4 nAChR mRNAs in rhesus monkey lung and cultured BECs. The expression of 7, 4, and β2 nAChR was confirmed by immunofluorescence in the cultured BECs and lung. The electrophysiological characteristics of nAChR in BECs were determined using whole-cell patch–clamp on cultured BECs. Both ACh and nicotine evoked an inward current, with a rapid desensitizing current. Nicotine induced inward currents in a concentration-dependent manner, with an EC₅₀ of 26.7 µM. Nicotine-induced currents were reversibly blocked by the nicotinic antagonists, mecamylamine, dihydro-β-erythroidine, and methyllcaconitine. Incubation of BECs with 1 µM nicotine for 48 hours enhanced nicotine-induced currents by roughly 26%. The protein tyrosine phosphorylation inhibitor, genistein, increased nicotine-induced currents by 58% and enhanced methyllcaconitine-sensitive currents (7 nAChR activities) 2.3-fold, whereas the protein tyrosine phosphatase inhibitor, pervanadate, decreased the effects of nicotine. These results demonstrate that chronic nicotine exposure up-regulates nAChR activity in developing lung, and that nAChR activity can be further modified by tyrosine phosphorylation.

Key Words: nicotinic acetylcholine receptors • electrophysiology • bronchial epithelial cells • nicotine • lung

CLINICAL RELEVANCE This research explains the fundamental mechanism by which nicotine affects lung development and function—by activating as opposed to desensitizing nicotinic cholinergic receptors in lung. This will help to develop therapies to block the effects of nicotine on lung.

 

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