Electrophysiological properties of neonatal mouse cardiac myocytes in primary culture

H. B. Nuss, E. Marban

Research output: Contribution to journalArticle

Abstract

The increasing utility of transgenic mice in molecular studies of the cardiovascular system has motivated us to characterize the ionic currents in neonatal mouse ventricular myocytes. Cell capacitance measurements (30 ± 1 pF, n = 73) confirmed visual impressions that neonatal mouse ventricular myocytes in primary culture are considerably smaller than freshly isolated adult ventricular myocytes. With the use of electron microscopy, mitochondria and sarcoplasmic reticulum were found in close association with myofibrils, but transverse tubules were not observed. Action potential durations, measured at 50 and 90% repolarization, were 23 ± 1 and 42 ± 2 ms respectively (n = 46). Application of 4-aminopyridine (4-AP; 5 mM) prolonged action potential duration at 50% repolarization by 26 ± 5% (n = 3). The brevity of the action potential is explained by the rapid activation of a transient outward K+ current upon voltage-clamp depolarization to plateau potentials. Potassium currents identified include an inward rectifier, a large 4-AP-sensitive transient outward, a slowly inactivating 4-AP-insensitive outward, a slowly activating delayed rectifier and a small rapidly activating E-4031 (10 μM)-sensitive delayed rectifier K+ current. Sodium currents (-305 ± 50 pA pF-1, n = 21) were recorded in 40 mM Na+ with Ni2+ (1 mM) to block Ca2+ currents and with K+ replaced by Cs+. The relative insensitivity of the Na+ current to block by tetrodotoxin (IC50 = 2.2 ± 0.3 μM, n = 4) is distinctive of the cardiac Na+ channel isoform. Nitrendipine-insensitive (10 μM) Ba2+ currents elicited during steps from -90 to -30 mV measured -25 ± 5 pA pF-1 (n = 7, 30 mM Ba(2+)). Decay of these currents was complete during 180 ms depolarizations, even with Ba2+ as the charge carrier. These currents were not present when the holding potential was set at -50 mV. These data support the presence of a low threshold, T-type Ca2+ current. The maximal nitrendipine-sensitive L-type Ca2+ current density was -10 ± 2 pA pF-1 (n = 8) in 2 mM Ca2+ and -38 ± 5 pA pF-1 (n = 9) in 30 mM Ba2+. Exposure to isoprenaline (1 μM) resulted in an 82% increase (n = 3) in the amplitude of the Ba2+ currents elicited at 0 mV. Neonatal mouse cardiac myocytes in primary culture possess surprisingly large inward currents given the brevity of their action potentials. Their small cell size and apparent absence of transverse tubules makes them technically favourable for whole-cell patch clamp studies. For these reasons, this preparation promises to be useful for the phenotypic characterization of transgenic mice.

Original languageEnglish (US)
Pages (from-to)265-279
Number of pages15
JournalJournal of Physiology
Volume479
Issue number2
StatePublished - 1994

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Cardiac Myocytes
Action Potentials
Muscle Cells
Nitrendipine
Transgenic Mice
4-Aminopyridine
Myofibrils
Tetrodotoxin
Sarcoplasmic Reticulum
Cardiovascular System
Cell Size
Isoproterenol
Inhibitory Concentration 50
Electron Microscopy
Potassium
Mitochondria
Protein Isoforms
Sodium

ASJC Scopus subject areas

  • Physiology

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Electrophysiological properties of neonatal mouse cardiac myocytes in primary culture. / Nuss, H. B.; Marban, E.

In: Journal of Physiology, Vol. 479, No. 2, 1994, p. 265-279.

Research output: Contribution to journalArticle

Nuss, H. B. ; Marban, E. / Electrophysiological properties of neonatal mouse cardiac myocytes in primary culture. In: Journal of Physiology. 1994 ; Vol. 479, No. 2. pp. 265-279.
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abstract = "The increasing utility of transgenic mice in molecular studies of the cardiovascular system has motivated us to characterize the ionic currents in neonatal mouse ventricular myocytes. Cell capacitance measurements (30 ± 1 pF, n = 73) confirmed visual impressions that neonatal mouse ventricular myocytes in primary culture are considerably smaller than freshly isolated adult ventricular myocytes. With the use of electron microscopy, mitochondria and sarcoplasmic reticulum were found in close association with myofibrils, but transverse tubules were not observed. Action potential durations, measured at 50 and 90{\%} repolarization, were 23 ± 1 and 42 ± 2 ms respectively (n = 46). Application of 4-aminopyridine (4-AP; 5 mM) prolonged action potential duration at 50{\%} repolarization by 26 ± 5{\%} (n = 3). The brevity of the action potential is explained by the rapid activation of a transient outward K+ current upon voltage-clamp depolarization to plateau potentials. Potassium currents identified include an inward rectifier, a large 4-AP-sensitive transient outward, a slowly inactivating 4-AP-insensitive outward, a slowly activating delayed rectifier and a small rapidly activating E-4031 (10 μM)-sensitive delayed rectifier K+ current. Sodium currents (-305 ± 50 pA pF-1, n = 21) were recorded in 40 mM Na+ with Ni2+ (1 mM) to block Ca2+ currents and with K+ replaced by Cs+. The relative insensitivity of the Na+ current to block by tetrodotoxin (IC50 = 2.2 ± 0.3 μM, n = 4) is distinctive of the cardiac Na+ channel isoform. Nitrendipine-insensitive (10 μM) Ba2+ currents elicited during steps from -90 to -30 mV measured -25 ± 5 pA pF-1 (n = 7, 30 mM Ba(2+)). Decay of these currents was complete during 180 ms depolarizations, even with Ba2+ as the charge carrier. These currents were not present when the holding potential was set at -50 mV. These data support the presence of a low threshold, T-type Ca2+ current. The maximal nitrendipine-sensitive L-type Ca2+ current density was -10 ± 2 pA pF-1 (n = 8) in 2 mM Ca2+ and -38 ± 5 pA pF-1 (n = 9) in 30 mM Ba2+. Exposure to isoprenaline (1 μM) resulted in an 82{\%} increase (n = 3) in the amplitude of the Ba2+ currents elicited at 0 mV. Neonatal mouse cardiac myocytes in primary culture possess surprisingly large inward currents given the brevity of their action potentials. Their small cell size and apparent absence of transverse tubules makes them technically favourable for whole-cell patch clamp studies. For these reasons, this preparation promises to be useful for the phenotypic characterization of transgenic mice.",
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