A real-time screening assay for GIRK1/4 channel blockers

Area of Science:

• Cardiovascular Pharmacology
• Ion Channel Research
• Drug Discovery

Background:

• The cardiac acetylcholine-activated potassium channel (I(K,Ach)), a GIRK4/1 channel subtype, is a potential therapeutic target for atrial fibrillation (AF).
• Developing selective blockers for this channel is crucial for novel AF drug development.

Purpose of the Study:

• To establish a robust cell-based screening assay for identifying novel GIRK1/4 channel blockers.
• To validate the assay's utility in identifying potential therapeutic agents for AF.

Main Methods:

• Utilized the mouse atrial HL-1 cell line expressing GIRK1/4 channels in a 96-well plate format.
• Employed a fluorescent membrane potential-sensitive dye (DiBAC(4)(3)) and a fluorescent imaging plate reader (FLIPR) for signal detection.
• Assessed channel activity by measuring fluorescence changes upon carbachol stimulation and validated with known blockers (BaCl2) and modulators.

Main Results:

• A significant decrease in fluorescence upon carbachol application indicated K+ efflux through GIRK1/4 channels, with a Z' factor of 0.5-0.6.
• The assay demonstrated sensitivity to known blockers (BaCl2) and confirmed channel function.
• Screening a small compound library identified amiloride and propafenone analogues as GIRK1/4 channel blockers at sub-micromolar concentrations.

Conclusions:

• The developed cell-based assay is effective for screening GIRK1/4 channel blockers.
• This assay provides a valuable tool for the discovery of new, selective agents for atrial fibrillation treatment.