In:
The Journal of Membrane Biology, Springer Science and Business Media LLC, Vol. 256, No. 1 ( 2023-02), p. 63-77
Abstract:
Most blockers of both hERG (human ether-à-go-go -related gene) channels and pancreatic ß-cell ATP-sensitive K + (K ATP ) channels access their binding sites from the cytoplasmic side of the plasma membrane. It is unknown whether binding to intracellular components competes with binding of these substances to K + channels. The whole-cell configuration of the patch-clamp technique, a laser-scanning confocal microscope, and fluorescence correlation spectroscopy (FCS) were used to study hERG channels expressed in HEK (human embryonic kidney) 293 cells and K ATP channels from the clonal insulinoma cell line RINm5F. When applied via the pipette solution in the whole-cell configuration, terfenadine blocked both hERG and K ATP currents with much lower potency than after application via the bath solution, which was not due to P-glycoprotein-mediated efflux of terfenadine. Such a difference was not observed with dofetilide and tolbutamide. 37–68% of hERG/EGFP (enhanced green-fluorescent protein) fusion proteins expressed in HEK 293 cells were slowly diffusible as determined by laser-scanning microscopy in the whole-cell configuration and by FCS in intact cells. Bath application of a green-fluorescent sulphonylurea derivative (Bodipy-glibenclamide) induced a diffuse fluorescence in the cytosol of RINm5F cells under whole-cell patch-clamp conditions. These observations demonstrate the presence of intracellular binding sites for hERG and K ATP channel blockers not dialyzable by the patch-pipette solution. Intracellular binding of terfenadine was not influenced by a mutated hERG (Y652A) channel. In conclusion, substances with high lipophilicity are not freely diffusible inside the cell but steep concentration gradients might exist within the cell and in the sub-membrane space. Graphical Abstract
Type of Medium:
Online Resource
ISSN:
0022-2631
,
1432-1424
DOI:
10.1007/s00232-022-00252-y
Language:
English
Publisher:
Springer Science and Business Media LLC
Publication Date:
2023
detail.hit.zdb_id:
1459323-3
SSG:
12
Permalink
