Efficient expression of rat brain type IIA Na+ channel alpha subunits in a somatic cell line.
Type IIA rat brain Na+ channel alpha subunits were expressed in CHO cells by nuclear microinjection or by transfection using a vector containing both metallothionein and bacteriophage SP6 promoters. Stable cell lines expressing Na+ channels were isolated, and whole-cell Na+ currents of 0.9-14 nA were recorded. The mean level of whole-cell Na+ current (4.5 nA) corresponds to a cell surface density of approximately 2 channels active at the peak of the Na+ current per microns 2, a density comparable to that observed in the cell bodies of central neurons. The expressed Na+ channels had the voltage dependence, rapid activation and inactivation, and rapid recovery from inactivation characteristic of Na+ channels in brain neurons, bound toxins at neurotoxin receptor sites 1 and 3 with normal properties, and were posttranslationally processed to a normal mature size of 260 kd. Expression of Na+ channel cDNA in CHO cells driven by the metallothionein promoter accurately and efficiently reproduces native Na+ channel properties and provides a method for combined biochemical and physiological analysis of Na+ channel structure and function.
West
J W
JW
Department of Pharmacology, University of Washington, Seattle 98195.
Scheuer
T
T
Maechler
L
L
Catterall
W A
WA
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D001923:Brain Chemistry; D016466:CHO Cells; D002467:Cell Nucleus / Q000502:physiology; D006224:Cricetinae; D004247:DNA / Q000737:chemistry / Q000235:genetics; D004553:Electric Conductivity; D015870:Gene Expression; D046911:Macromolecular Substances; D008845:Microinjections; D008969:Molecular Sequence Data; D010957:Plasmids; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011950:Receptors, Cholinergic / Q000378:metabolism; D012530:Saxitoxin / Q000378:metabolism / Q000494:pharmacology; D015222:Sodium Channels / Q000235:genetics* / Q000502:physiology; D013779:Tetrodotoxin / Q000378:metabolism / Q000494:pharmacology; D014162:Transfection
D046911:Macromolecular Substances; D012333:RNA, Messenger; D011950:Receptors, Cholinergic; D015222:Sodium Channels; D012530:Saxitoxin; D013779:Tetrodotoxin; D004247:DNA