A cardiac dihydropyridine receptor II-III loop peptide inhibits resting Ca(2+) sparks in ferret ventricular myocytes.
1. We studied the effect of a peptide (Ac-10C) on cardiac ryanodine receptor (RyR) opening. This decapeptide (KKERKLARTA) is a fragment of the cardiac dihydropyridine receptor (DHPR) from the cytosolic loop between the second and third transmembrane domains (II-III loop). Studies were carried out in ferret ventricular myocytes by simultaneously applying ruptured-patch voltage clamp and line-scan confocal microscopy with fluo-3 to measure intracellular [Ca(2+)] ([Ca(2+)](i)) and Ca(2+) sparks. 2. Inclusion of Ac-10C in the dialysing pipette solution inhibited resting Ca(2+) spark frequency (due to diastolic RyR openings) by > 50 %. This occurred without changing sarcoplasmic reticulum (SR) Ca(2+) content, which was measured via the caffeine-induced Ca(2+) transient amplitude and the caffeine-induced Na(+)-Ca(2+) exchange current (I(NCX)) integral. Ac-10C also reduced slightly the size of Ca(2+) sparks. 3. Ac-10C did not alter either resting [Ca(2+)](i) (assessed by indo-1 fluorescence) or DHPR gating (measured as L-type Ca(2+) current). 4. The SR Ca(2+) fractional release was depressed by Ac-10C at relatively low SR Ca(2+) content, but not at higher SR Ca(2+) content. 5. A control scrambled peptide (Ac-10CS) did not alter any of the measured parameters (notably Ca(2+) spark frequency or SR Ca(2+) fractional release). Thus, the Ac-10C effects may be sequence or charge distribution specific. 6. Our results suggest an inhibitory regulation of RyRs at rest via the cardiac DHPR II-III loop N-terminus region. The mechanism of the effect and whether this interaction is important in cardiac excitation-contraction coupling (E-C coupling) per se, requires further investigation.
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Department of Physiology, Stritch School of Medicine, Loyola University Chicago, 2160 South First Avenue, Maywood, IL 60153, USA.
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D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D002118:Calcium / Q000502:physiology*; D020746:Calcium Channels, L-Type / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D005289:Ferrets; D007425:Intracellular Membranes / Q000378:metabolism; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D009994:Osmolar Concentration; D018408:Patch-Clamp Techniques; D010446:Peptide Fragments / Q000235:genetics / Q000502:physiology; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D016276:Ventricular Function / Q000502:physiology*
D020746:Calcium Channels, L-Type; D010446:Peptide Fragments; D002118:Calcium