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Modulators of calcium and potassium channels: their effects on endothelin-1 binding to cardiac membranes
W G Haynes1, C J Waugh, M E Dockrell
1Department of Medicine, University of Edinburgh, Western General Hospital, Scotland.
Abstract:
Endothelin-1 (ET-1) causes long-lasting vasoconstriction associated with a prolonged elevation of intracellular free Ca2+. Because this may be mediated through an effect on membrane ion channels, we investigated the effects of the dihydropyridine calcium channel antagonist nifedipine; two structurally distinct K+ channel openers, pinacidil and levcromakalim; and the inactive stereoisomer of levcromakalim (D-cromakalim), as well as ET-1 and ET-3, on binding of 125I-labeled endothelin-1 to rat cardiac membranes. Specific binding of 125I-ET-1 was inhibited in a concentration-dependent manner by unlabeled ET-1 (IC50 = 1.56 +/- 0.78 nM; slope = -0.49 +/- 0.10) and ET-3 (IC50 = 314 +/- 54 nM; slope = -0.34 +/- 0.11). Nifedipine, in concentrations < or = 10(-5) M, did not affect 125I-ET-1 binding. However, levcromakalim significantly inhibited 125I-ET-1 binding (maximum binding 49 4/- 9%; p = 0.04), whereas the inactive isomer, D-cromakalim, had no effect. Pinacidil also inhibited 125I-ET-1 binding, although to a lesser extent than levcromakalim (maximum binding 63 +/- 7%). These findings provide evidence for a stereospecific interaction between K(+)-channel openers and ET-1 binding in rat cardiac membranes. Because the slope of the logistic fit was substantially less than unity, and the effects of pinacidil and levcromakalim were incomplete, there may be two or more receptors for ET-1 in rat heart, only one of which is sensitive to K(+)-channel openers.
Insights
Potassium channel openers, levcromakalim and pinacidil, interact with endothelin-1 (ET-1) binding sites in rat cardiac membranes. This suggests a stereospecific interaction and potential for multiple ET-1 receptors in the heart.
Area of Science:
- Cardiovascular Pharmacology
- Molecular Cardiology
- Ion Channel Modulation
Background:
- Endothelin-1 (ET-1) induces prolonged vasoconstriction and elevated intracellular calcium, potentially via ion channel mechanisms.
- Investigating the interaction between ET-1 and cardiac ion channels is crucial for understanding vasoconstriction regulation.
Purpose of the Study:
- To examine the effects of calcium channel blockers and potassium channel openers on endothelin-1 binding to rat cardiac membranes.
- To elucidate the role of specific ion channels in mediating ET-1's cardiovascular effects.
Main Methods:
- Radioligand binding assays using 125I-labeled endothelin-1 (125I-ET-1) on rat cardiac membranes.
- Competitive inhibition studies with unlabeled ET-1, ET-3, nifedipine, pinacidil, and levcromakalim (and its inactive isomer D-cromakalim).
Main Results:
- Unlabeled ET-1 and ET-3 inhibited 125I-ET-1 binding in a concentration-dependent manner.
- Levcromakalim and pinacidil, but not nifedipine or D-cromakalim, significantly inhibited 125I-ET-1 binding.
- The incomplete inhibition by K+ channel openers suggests the presence of multiple ET-1 receptor subtypes in rat cardiac membranes.
Conclusions:
- Potassium channel openers exhibit a stereospecific interaction with endothelin-1 binding sites in rat cardiac membranes.
- These findings support the existence of distinct endothelin-1 receptor populations, with at least one subtype sensitive to K+ channel openers.