Stable expression of the neuronal BI (class A) calcium channel in baby hamster kidney cells.
Plasmids containing the BI (alpha 1) cDNA with the dihydrofolate reductase (DHFR) gene, skeletal muscle alpha 2-subunit cDNA with the neo marker gene, and beta-subunit cDNA were co-transfected into baby hamster kidney (BHK) cells. BHK cells lack endogenous calcium channel activity. Twenty percent of the methotrexate (MTX) and G418 resistant clones were found to express calcium channel activity using the patch-clamp technique. A single clone, BHKBI147, demonstrated stable electrophysiological characteristics over 20 passages. Ca2+ currents of the BI channel in BHKBI147 cells were largely blocked by a specific P-type blocker, omega-AgaIVA, with an IC50 of 150 nM. Unlike the BI channel, Ca2+ currents of cardiac L-type channels expressed in BHK cells were completely blocked by the L-type antagonist, nifedipine, with an IC50 of 56 nM.
Niidome
T
T
Tsukuba Research Laboratories, Eisai Co., Ltd., Ibaraki, Japan.
Teramoto
T
T
Murata
Y
Y
Tanaka
I
I
Seto
T
T
Sawada
K
K
Mori
Y
Y
Katayama
K
K
D000818:Animals; D001483:Base Sequence; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000096:biosynthesis* / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D002999:Clone Cells; D006224:Cricetinae; D017931:DNA Primers; D015870:Gene Expression; D005819:Genetic Markers; D005839:Gentamicins / Q000494:pharmacology; D007668:Kidney; D007700:Kinetics; D008727:Methotrexate / Q000494:pharmacology; D008969:Molecular Sequence Data; D009132:Muscles / Q000378:metabolism; D009474:Neurons / Q000378:metabolism*; D009543:Nifedipine / Q000494:pharmacology; D010957:Plasmids; D016133:Polymerase Chain Reaction; D015183:Restriction Mapping; D013111:Spider Venoms / Q000494:pharmacology; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics; D014162:Transfection / Q000379:methods; D020867:omega-Agatoxin IVA
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D017931:DNA Primers; D005819:Genetic Markers; D005839:Gentamicins; D013111:Spider Venoms; D020867:omega-Agatoxin IVA; C010680:antibiotic G 418; D013762:Tetrahydrofolate Dehydrogenase; D009543:Nifedipine; D008727:Methotrexate