Am J Physiol Cell Physiol AJP: Regulatory, Integrative and Comparative Physiology
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Am J Physiol Cell Physiol 285: C567-C574, 2003. First published May 14, 2003; doi:10.1152/ajpcell.00422.2002
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RECEPTORS AND SIGNAL TRANSDUCTION

Nicotinic acetylcholine receptor {alpha}7 regulates cAMP signal within lipid rafts

Jin Oshikawa,1 Yoshiyuki Toya,1 Takayuki Fujita,1 Masato Egawa,2 Junichi Kawabe,3 Satoshi Umemura,1 and Yoshihiro Ishikawa1,3

1Departments of Physiology and Medicine, Yokohama City University School of Medicine, and 2Department of Medical Pathophysiology, Yokohama City University College of Nursing, Yokohama 236-0004, Japan; and 3Cardiovascular Research Institute, Departments of Medicine and Cell Biology and Molecular Medicine, University of Medicine and Dentistry of New Jersey, Newark, New Jersey 07103

Submitted 16 September 2002 ; accepted in final form 7 May 2003

Neuronal nicotinic acetylcholine receptors (nAChRs) are made of multiple subunits with diversified functions. The nAChR {alpha}7-subunit has a property of high Ca2+ permeability and may have specific functions and localization within the plasma membrane as a signal transduction molecule. In PC-12 cells, fractionation by sucrose gradient centrifugation revealed that nAChR{alpha}7 existed in low-density, cholesterol-enriched plasma membrane microdomains known as lipid rafts where flotillin also exists. In contrast, nAChR {alpha}5- and {beta}2-subunits were located in high-density fractions, out of the lipid rafts. Type 6 adenylyl cyclase (AC6), a calcium-inhibitable isoform, was also found in lipid rafts and was coimmunoprecipitated with nAChR{alpha}7. Cholesterol depletion from plasma membranes with methyl-{beta}-cyclodextrin redistributed nAChR{alpha}7 and AC6 diffusely within plasma membranes. Nicotine stimulation reduced forskolin-stimulated AC activity by 35%, and this inhibition was negated by either treatment with {alpha}-bungarotoxin, a specific antagonist of nAChR{alpha}7, or cholesterol depletion from plasma membranes. The effect of cholesterol depletion was negated by the addition of cholesterol. These data suggest that nAChR{alpha}7 has a specific membrane localization relative to other nAChR subunits and that lipid rafts are necessary to localize nAChR{alpha}7 with AC within plasma membranes. In addition, nAChR{alpha}7 may regulate the AC activity via Ca2+ within lipid rafts.

cholesterol; PC-12 cells


Address for reprint requests and other correspondence: Y. Ishikawa, Cardiovascular Research Institute, Depts. of Medicine and Cell Biology & Molecular Medicine, Univ. of Medicine and Dentistry of New Jersey, 185 South Orange Ave. Newark, NJ 07103 (E-mail: ishikayo{at}umdnj.edu).




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