The biochemical activation of T-type Ca2+ channels in HEK293 cells stably expressing alpha1G and Kir2.1 subunits.
In order to investigate the currently unknown cellular signaling pathways of T-type Ca(2+) channels, we decided to construct a new cell line which would stably express alpha(1G) and Kir2.1 subunits in HEK293 cells (HEK293/alpha(1G)/Kir2.1). Compared to cells which only expressed alpha(1G) (HEK293/alpha(1G)), HEK293/alpha(1G)/Kir2.1 cells produced an enormous inward rectifying current which was blocked by external Ba(2+) and Cs(+) in a concentration-dependent manner. The expression of Kir2.1 channels contributed significantly to the shift of membrane potential from -12.2+/-2.8 to -57.3+/-3.7mV. However, biophysical and pharmacological properties of alpha(1G)-mediated Ca(2+) channels remained unaffected by the expression of Kir2.1 subunits, except for the enlarging of the window current region. Biochemical activation of alpha(1G) channels using 150mM KCl brought about an increase in [Ca(2+)](i), which was blocked by mibefradil, the T-type Ca(2+) channel blocker. These data suggest that the HEK293/alpha(1G)/Kir2.1 cell line would have potential uses in the study of T-type Ca(2)(+) channel-mediated signaling pathways and possibly useful in the development of new therapeutic drugs associated with T-type Ca(2)(+) channels.
Biomedical Research Center, Korea Institute of Science and Technology (KIST), Seoul 136-791, Republic of Korea.
Taehyun
T
Kim
Juhyun
J
Choi
Sunoh
S
Kim
Ohyeun
O
Kwon
Seung-Yeol
SY
Nah
Ye Sun
YS
Han
Hyewhon
H
Rhim
D001464:Barium / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000378:metabolism; D020747:Calcium Channels, T-Type / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002586:Cesium / Q000378:metabolism; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000196:embryology; D008564:Membrane Potentials / Q000502:physiology; D020748:Mibefradil / Q000378:metabolism; D018408:Patch-Clamp Techniques; D024661:Potassium Channels, Inwardly Rectifying / Q000235:genetics / Q000378:metabolism*; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D014162:Transfection
D002121:Calcium Channel Blockers; D020747:Calcium Channels, T-Type; C489457:KCNJ2 protein, human; D024661:Potassium Channels, Inwardly Rectifying; D021122:Protein Subunits; D002586:Cesium; D001464:Barium; D020748:Mibefradil; D002118:Calcium