Calcium tolerant ventricular myocytes prepared by preincubation in a “KB medium” View Full Text


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Article Info

DATE

1982-10

AUTHORS

Gerrit Isenberg, Udo Klockner

ABSTRACT

Ventricular myocytes were isolated by means of collagenase and hyaluronidase. Hearts of rats or guinea pigs were perfused in a retrograde manner; bovine ventricular tissue was incubated in from of tissue chunks.In “low Ca medium” (aCa=1 μM), the myocytes beat spontaneously. Resting potentials of −4 mV and input resistances of 1 MΩ, associated with a fast decay of aKi, indicated “hyperpermeability” of the cell membrane. Within 2 h, the cells aged: they granulated (swelling of mitochondria), contracted, rounded up and finally disaggregated. When the superfusate was changed to Tyrode solution (3.6 mM CaCl2), 90–98% of those cells responded with the “Ca paradox”. Initially, aCai increased from 0.2 μM to >10 μM, and the sarcomere length (SL) shortened to <1.5 μm. Within the following 3 min aCai renormalized but the contracture was sustained. After another 5 min, the SL shortened to <1.2 μm, but a delay of 1–2 min passed before aCai started to increase towards aCa0. The 2–10% Ca tolerant cells beat spontaneously. Their resting potentials were between −10 and −70 mV, their input resistances between 4 and 20 MΩ. Repolarization by anodal current flow could restore the sodium spike but not the plateau of the action potential.Freshly prepared myocytes were pre-incubated in a KB medium composed of 85 mM KCl, 30 mM K2HPO4, 5 mM MgSO4, 5 mM Na2ATP, 5 mM pyruvate, 5 mM succinate, 5 mM β-OH-butyrate, 5 mM creatine, adjusted with KOH to pH 7.2 and with EGTA to pCa 7.5. The minimum time of preincubation was 1 h, the maximum 5 days when the cells were stored at 5°C. The conditioning with the KB medium improved both Ca tolerance and electrophysiology significantly. About 70% of the rod shaped myocytes were Ca tolerant. These cells had resting potentials between −75 and −90 mV, the membrane resistances were around 7 kΩ×cm2. The action potentials showed well pronounced plateaus and lasted for 125 ms (rat), 300 ms (guinea pig) or 400 ms (bovine). Their shape showed the typical species related pecularities.We discuss that both ageing and Ca intolerance of the un-conditioned cells result from Ca overload through the hyperpermeable membrane, and that Ca overload causes ATP depletion via mitochondrial disfunction. The constituents of the KB medium may act to prevent these processes. More... »

PAGES

6-18

References to SciGraph publications

  • 1973-04. The equilibration time course of [K+]0 in cat cortex in EXPERIMENTAL BRAIN RESEARCH
  • 1980-03. Glycocalyx is not required for slow inward calcium current in isolated rat heart myocytes in NATURE
  • 1980-08. Free calcium in heart muscle at rest and during contraction measured with Ca2+-sensitive microelectrodes in NATURE
  • 1982-10. Isolated bovine ventricular myocytes in PFLÜGERS ARCHIV - EUROPEAN JOURNAL OF PHYSIOLOGY
  • 1981-09. Some properties of KCl-filled microelectrodes: Correlation of potassium “leakage” with tip resistance in THE JOURNAL OF MEMBRANE BIOLOGY
  • 1979-01. Are cardiac muscle cells ‘skinned’ by EGTA or EDTA? in NATURE
  • 1974-09. Iontophoretic application of acetylcholine: Advantages of high resistance micropipettes in connection with an electronic current pump in PFLÜGERS ARCHIV - EUROPEAN JOURNAL OF PHYSIOLOGY
  • 1980-08. Removal of sialic acid from cardiac sarcolemma does not affect contractile function in electrically stimulated guinea pig left atria in NATURE
  • 1980-02. Decoupling of heart muscle cells: Correlation with increased cytoplasmic calcium activity and with changes of nexus ultrastructure in THE JOURNAL OF MEMBRANE BIOLOGY
  • 1966-08. Paradoxical Influence of Calcium Ions on the Permeability of the Cell Membranes of the Isolated Rat Heart in NATURE
  • 1979-05. Risk and advantages of using strongly beveled microelectrodes for electrophysiological studies in cardiac Purkinje fibers in PFLÜGERS ARCHIV - EUROPEAN JOURNAL OF PHYSIOLOGY
  • 1981-04. Oxygen requirements, morphology, cell coat and membrane permeability of calcium-tolerant myocytes from hearts of adult rats in CELL AND TISSUE RESEARCH
  • 1978-03. Volume changes and potential artifacts of epithelial cells of frog skin following impalement with microelectrodes filled with 3m KCl in THE JOURNAL OF MEMBRANE BIOLOGY
  • 1979-03. Sodium current in single heart muscle cells in NATURE
  • Identifiers

    URI

    http://scigraph.springernature.com/pub.10.1007/bf00584963

    DOI

    http://dx.doi.org/10.1007/bf00584963

    DIMENSIONS

    https://app.dimensions.ai/details/publication/pub.1039653041

    PUBMED

    https://www.ncbi.nlm.nih.gov/pubmed/7177773


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