Expression of native alpha3beta4* neuronal nicotinic receptors: binding and functional studies investigating turnover of surface and intracellular receptor populations.
Several pathological conditions involve alterations in expression of neuronal nicotinic acetylcholine receptors (nAChRs). Although some studies have addressed processes involved with muscle nAChR expression, knowledge of the regulation of neuronal nAChRs is particularly sparse. The following studies were designed to investigate cellular mechanisms involved with expression of neuronal alpha3beta4* nAChRs. Catecholamine secretion assays and receptor binding studies coupled with receptor alkylation were used to study the nAChR regulation and turnover. Alkylation of adrenal nAChRs results in a rapid and complete loss of receptor-mediated neurosecretion and surface [(3)H]epibatidine binding sites. After alkylation, both neurosecretory function and nAChR binding slowly (24-48 h) return to prealkylation levels. When cells are treated with the protein synthesis inhibitor puromycin, after alkylation, receptor-mediated neurosecretion does not recover. Long-term treatment (24-48-h) with puromycin, in the absence of alkylation, results in a slow, time-dependent shift to the right, followed by a downward shift, in the nicotine concentration-response curve, documenting a disappearance of surface nAChRs. Puromycin treatment alone also results in a loss to both surface and intracellular [(3)H]epibatidine binding sites. nAChR beta4 subunit levels are significantly decreased after treatment with puromycin. These data support a constitutive turnover of adrenal alpha3beta4* nAChRs, requiring continual de novo synthesis of new receptor protein.
Free
R Benjamin
RB
Division of Pharmacology, College of Pharmacy, The Ohio State University, 500 West 12th Avenue, Columbus, OH 43210, USA.
McKay
Susan B
SB
Gottlieb
Paul D
PD
Boyd
R Thomas
RT
McKay
Dennis B
DB
D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D005786:Gene Expression Regulation / Q000502:physiology; D007425:Intracellular Membranes / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D011978:Receptors, Nicotinic / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism
D008565:Membrane Proteins; D011978:Receptors, Nicotinic; C403530:nicotinic receptor alpha3beta4