Functional properties of homomeric, human alpha 7-nicotinic acetylcholine receptors heterologously expressed in the SH-EP1 human epithelial cell line.
alpha 7-Nicotinic acetylcholine receptors (alpha 7-nAChRs) are broadly distributed in the central nervous system, where they play important roles in chemical and electrical signaling, and perhaps in neurite outgrowth, synaptic plasticity, and neuronal death/survival. To help elucidate their normal and pathophysiological roles, we have heterologously expressed human alpha 7-nAChR in transfected SH-EP1 human epithelial cells. Reverse transcription-polymerase chain reaction and mRNA fluorescence in situ hybridization analyses demonstrate expression of human alpha 7 subunits as messenger RNA. Patch-clamp recordings exploiting a novel strategy to prevent functional rundown of whole-cell peak current responses to repeated acute challenges with nicotinic agonists show successful expression of functional alpha 7-nAChR that mediate inward currents characterized by rapid phases of activation and inactivation. Concentration-response curves show that nicotine, acetylcholine, and choline are efficacious agonists at human alpha 7-nAChRs. Current-voltage relationships show inward rectification for agonist-induced currents. Human alpha 7-nAChRs exhibit some sensitivity to alpha 7-nAChR antagonists alpha-bungarotoxin (Bgt) or methyllycaconitine (MLA) when applied coincidentally with agonist, but much higher affinity block occurs when cells and alpha 7-nAChRs are pre-exposed to antagonists for 2 min before challenge with agonist. Both Bgt and MLA are competitive inhibitors of alpha 7-nAChR function. Whole-cell current peak amplitudes and half-times for inactivation of alpha 7-nAChR functional responses to nicotine are dramatically reduced in the absence of extracellular Ca2+, suggestive of high Ca2+ permeability of the alpha 7-nAChR channel. Thus, heterologously expressed human alpha 7-nAChR in mammalian cells have properties of native alpha 7-nAChR or of alpha 7-nAChR heterologously expressed in other systems and serve as excellent models for studies of molecular bases of alpha 7-nAChR function.
Zhao
Lingke
L
Division of Neurology, Barrow Neurological Institute, 350 West Thomas Rd., Phoenix, AZ 85013-4496, USA.
Kuo
Yen-Ping
YP
George
Andrew A
AA
Peng
Jian-Hong
JH
Purandare
Madhuri Singh
MS
Schroeder
Katherine M
KM
Lukas
Ronald J
RJ
Wu
Jie
J
D000109:Acetylcholine / Q000494:pharmacology; D000157:Aconitine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002038:Bungarotoxins / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D009538:Nicotine / Q000494:pharmacology; D018722:Nicotinic Agonists / Q000494:pharmacology; D018733:Nicotinic Antagonists / Q000494:pharmacology; D011978:Receptors, Nicotinic / Q000187:drug effects / Q000502:physiology*; D014162:Transfection; D064569:alpha7 Nicotinic Acetylcholine Receptor
D002038:Bungarotoxins; C579167:Chrna7 protein, human; D018722:Nicotinic Agonists; D018733:Nicotinic Antagonists; D011978:Receptors, Nicotinic; D064569:alpha7 Nicotinic Acetylcholine Receptor; C054634:methyllycaconitine; D009538:Nicotine; D000109:Acetylcholine; D002118:Calcium; D000157:Aconitine