PKC-ε mediates multiple endothelin-1 actions on systolic Ca2+ and contractility in ventricular myocytes.
Endothelin-1 (ET-1) induces positive inotropy (enhanced contractility) in cardiac muscle, but establishing underlying cellular mechanisms has been controversial in part because of a growing number of signaling pathways and end effectors targeted by ET-1. Here we present evidence that ET-1 induces positive inotropism in ventricular tissue by increasing both systolic Ca2+ and myofilament Ca2+ sensitivity. To examine the roles of PKC-δ and PKC-ε in these acute responses to ET-1, kinase inactive dominant negative PKC (dn-PKC) constructs were expressed in adult rat ventricular myocytes. Yellow fluorescent protein (YFP) was fused to dn-PKC constructs to visualize expression and localization of dn-PKC in living myocytes. Due to an alanine to glutamate mutation in the pseudosubstrate site, dn-PKCs constitutively translocated to anchoring sites and were unaffected by agonist or phorbol ester treatment. Dn-PKC-δ-YFP mainly distributed at Z-lines and at intercalated disks in adult myocytes, whereas dn-PKC-ε-YFP stained the surface sarcolemma, T-tubules/Z-lines and perinuclear region. Myocytes expressing dn-PKC-δ-YFP showed normal systolic Ca2+ and contractile responses to ET-1. In contrast, the entire ensemble of ET-1 responses was blocked in myocytes expressing dn-PKC-ε-YFP including increased Ca2+ transients, enhanced myofilament Ca2+ sensitivity, and positive inotropy. This report provides direct evidence that PKC-ε is activated early and robustly following ET-1 stimulation and thus mediates multiple intracellular changes underlying the acute actions of ET-1 on myocardium.
Molecular and Cellular Pharmacology, University of Wisconsin School of Medicine and Public Health, Madison, WI, USA.
Misuk
M
Kang
Ka Young
KY
Chung
D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D019332:Endothelin-1 / Q000494:pharmacology / Q000502:physiology*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008297:Male; D009200:Myocardial Contraction / Q000187:drug effects / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000502:physiology*; D051744:Protein Kinase C-epsilon / Q000235:genetics / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013599:Systole / Q000187:drug effects / Q000502:physiology; D016276:Ventricular Function
D001426:Bacterial Proteins; D019332:Endothelin-1; D008164:Luminescent Proteins; C054562:yellow fluorescent protein, Bacteria; D051744:Protein Kinase C-epsilon; D002118:Calcium