[3H]dofetilide binding to cardiac myocytes: modulation by extracellular potassium

Area of Science:

• Pharmacology
• Cardiovascular Physiology
• Biochemistry

Background:

• The radioligand [3H]dofetilide is used to study delayed rectifier potassium channels and Class III drugs.
• Previous studies examined [3H]dofetilide binding at high extracellular potassium (135 mM).
• Hyperkalemia is known to affect the pharmacological response to I(Kr) channel blockers.

Purpose of the Study:

• To investigate how altered ionic conditions, specifically extracellular potassium levels, affect [3H]dofetilide binding characteristics.
• To test the hypothesis that changes in ionic milieu modify [3H]dofetilide binding to cardiac potassium channels.

Main Methods:

• Radioligand binding assays using guinea-pig ventricular myocytes and neonatal mouse myocytes.
• Measurement of [3H]dofetilide binding under varying extracellular potassium concentrations (5 mM and 20 mM).
• Analysis of binding affinity (K(d)) and maximal binding capacity (B(max)).

Main Results:

• Under physiological conditions (5 mM K+), [3H]dofetilide bound to two sites (high and low affinity) on guinea-pig myocytes.
• Increasing extracellular K+ to 20 mM resulted in a single binding site with significantly altered affinity (K(d) 120±12 nM).
• Similar significant changes in high-affinity binding were observed in neonatal mouse myocytes at elevated extracellular K+.

Conclusions:

• Extracellular potassium concentration modulates [3H]dofetilide binding to its high-affinity site on cardiac potassium channels.
• These findings highlight the importance of physiological ionic conditions when studying drug-channel interactions.
• The study provides crucial insights into the biochemical characterization of I(Kr) channel blockers.