A new oil-gap method for internal perfusion and voltage clamp of single cardiac cells

1987 ◽  
Vol 410 (1-2) ◽  
pp. 7-14 ◽  
Author(s):  
T. Mitsuiye ◽  
A. Noma
Keyword(s):  
Voltage Clamp ◽  
Cardiac Cells ◽  
Internal Perfusion

Voltage Clamp and Internal Perfusion With Suction-Pipette Method

1985 ◽  
pp. 151-169 ◽  
Author(s):  
Arthur M. Brown ◽  
David L. Wilson ◽  
Yasuo Tsuda
Keyword(s):  
Voltage Clamp ◽  
Internal Perfusion

Is there a second external lidocaine binding site on mammalian cardiac cells?

1989 ◽  
Vol 257 (1) ◽  
pp. H79-H84 ◽  
Author(s):  
L. A. Alpert ◽  
H. A. Fozzard ◽  
D. A. Hanck ◽  
J. C. Makielski
Keyword(s):  
Binding Site ◽  
Cardiac Arrhythmias ◽  
Local Anesthetics ◽  
Sodium Current ◽  
Cardiac Cells ◽  
Na Channel ◽  
Functional Difference ◽  
Na Channels ◽  
Internal Perfusion ◽  
Cardiac Purkinje Cells

Lidocaine and its permanently charged analogue QX-314 block sodium current (INa) in nerve, and by this mechanism, lidocaine produces local anesthesia. When administered clinically, lidocaine prevents cardiac arrhythmias. Nerve and skeletal muscle are much more sensitive to local anesthetics when the drugs are applied inside the cell, indicating that the binding site for local anesthetics is located on the inside of those Na channels. Using a large suction pipette for voltage clamp and internal perfusion of single cardiac Purkinje cells, we demonstrate that a charged lidocaine analogue blocks INa not only when applied from the inside but also from the outside, unlike noncardiac tissue. This functional difference in heart predicts that a second local anesthetic binding site exists outside or near the outside of cardiac Na channels and emphasizes that the cardiac Na channel is different from that in nerve.

scholarly journals Properties of internally perfused, voltage-clamped, isolated nerve cell bodies.

1978 ◽  
Vol 71 (5) ◽  
pp. 489-507 ◽  
Author(s):  
K S Lee ◽  
N Akaike ◽  
A M Brown
Keyword(s):  
Voltage Clamp ◽  
Nerve Cell ◽  
Action Potentials ◽  
Divalent Cations ◽  
Membrane Resistance ◽  
Membrane Properties ◽  
Neuronal Membrane ◽  
Internal Perfusion ◽  
Shunt Resistance ◽  
Membrane Function

The membrane properties of isolated neurons from Helix aspersa were examined by using a new suction pipette method. The method combines internal perfusion with voltage clamp of nerve cell bodies separated from their axons. Pretreatment with enzymes such as trypsin that alter membrane function is not required. A platinized platinum wire which ruptures the soma membrane allows low resistance access directly to the cell's interior improving the time resolution under voltage clamp by two orders of magnitude. The shunt resistance of the suction pipette was 10-50 times the neuronal membrane resistance, and the series resistance of the system, which was largely due to the tip diameter, was about 10(5) omega. However, the peak clamp currents were only about 20 nA for a 60-mV voltage step so that measurements of membrane voltage were accurate to within at least 3%. Spatial control of voltage was achieved only after somal separation, and nerve cell bodies isolated in this way do not generate all-or-none action potentials. Measurements of membrane potential, membrane resistance, and membrane time constant are equivalent to those obtained using intracellular micropipettes, the customary method. With the axon attached, comparable all-or-none action potentials were also measured by either method. Complete exchange of Cs+ for K+ was accomplished by internal perfusion and allowed K+ currents to be blocked. Na+ currents could then be blocked by TTX or suppressed by Tris-substituted snail Ringer solution. Ca2+ currents could be blocked using Ni2+ and other divalent cations as well as organic Ca2+ blockers. The most favorable intracellular anion was aspartate-, and the sequence of favorability was inverted from that found in squid axon.

scholarly journals Ionic currents in single isolated bullfrog atrial cells.

1983 ◽  
Vol 81 (2) ◽  
pp. 153-194 ◽  
Author(s):  
J R Hume ◽  
W Giles
Keyword(s):  
Voltage Clamp ◽  
Single Cells ◽  
Ionic Current ◽  
Technical Difficulty ◽  
Ionic Currents ◽  
Cardiac Cells ◽  
Small Cells ◽  
Inward Current ◽  
Electrophysiological Properties ◽  
Atrial Cells

Enzymatic dispersion has been used to yield single cells from segments of bullfrog atrium. Previous data (Hume and Giles, 1981) have shown that these individual cells are quiescent and have normal resting potentials and action potentials. The minimum DC space constant is approximately 920 microns. The major goals of the present study were: (a) to develop and refine techniques for making quantitative measurements of the transmembrane ionic currents, and (b) to identify the individual components of ionic current which generate different phases of the action potential. Initial voltage-clamp experiments made using a conventional two-microelectrode technique revealed a small tetrodotoxin (TTX)-insensitive inward current. The small size of this current (2.5-3.0 X 10(-10)A) and the technical difficulty of the two-microelectrode experiments prompted the development of a one-microelectrode voltage-clamp technique which requires impalements using a low-resistance (0.5-2 M omega) micropipette. Voltage-clamp experiments using this new technique in isolated single atrial cells reveal five distinct ionic currents: (a) a conventional transient Na+ current, (b) a TTX-resistant transient inward current, carried mainly by Ca++, (c) a component of persistent inward current, (d) a slowly developing outward K+ current, and (e) an inwardly rectifying time-independent background current. The single suction micropipette technique appears well-suited for use in the quantitative study of ionic currents in these cardiac cells, and in other small cells having similar electrophysiological properties.

Effects of endothelin 1 on calcium and sodium currents in isolated human cardiac myocytes

1995 ◽  
Vol 73 (12) ◽  
pp. 1774-1783 ◽  
Author(s):  
Tzu-Hurng Cheng ◽  
Chung-Yi Chang ◽  
Jeng Wei ◽  
Cheng-I Lin
Keyword(s):  
Voltage Clamp ◽  
Cardiac Myocytes ◽  
Ventricular Myocytes ◽  
Cell Voltage ◽  
Cardiac Cells ◽  
Sodium Currents ◽  
Atrial Myocytes ◽  
Whole Cell ◽  
Voltage Clamp Technique ◽  
Endothelin 1

We have used the whole-cell voltage-clamp technique to study the effects of endothelin 1 (ET-1, 10 nM) on L-type Ca2+ currents and voltage-dependent Na+ inward currents in human cardiac cells. Myocytes were enzymatically isolated from atrial specimens obtained during open-heart surgery and from human ventricular tissues of explanted hearts. Extracellular application of ET-1 decreased the peak amplitude of Ca2+ currents by 26 ± 6% (n = 13) in atrial myocytes and by 19 ± 3% (n = 8) in ventricular myocytes. In three atrial cells, treatment with 1 μM BQ123 prevented the decrease in Ca2+ currents induced by ET-1. When GTP (0.2 mM) was added to the dialyzing pipette solution, ET-1 still caused a small decline by 12 ± 5% (n = 16), in peak Ca2+ currents, in atrial myocytes. When Ca2+ currents were increased (+210 ± 19%) by a β-adrenoceptor agonist (0.1 μM isoproterenol) or by the phosphodiesterase inhibitor isobutylmethylxanthine (10 μM), ET-1 reduced Ca2+ currents by 35 ± 6% (n = 4) and 30 ± 4% (n = 5), respectively. In human ventricular myocytes in the presence of 1 μM isoproterenol, which increased the peak Ca2+ currents by 150 ± 30%, ET-1 also induced a drastic reduction in Ca2+ currents, by 40 ± 11% (n = 5). The tetrodotoxin-sensitive Na+ currents measured in the presence of 5 mM [Na]o were significantly enhanced (+28 ± 7%) by ET-1 in five atrial myocytes. The stimulatory effect of ET-1 on Na+ currents was partially reversible. The present findings in human cardiac cells show that ET-1 did not enhance the Ca2+ currents in the absence or presence of internal GTP. The positive inotropic actions induced by ET-1 in human heart may be mediated mainly by signal-transduction pathways other than the G-protein – adenylyl cyclase – cAMP system.Key words: endothelin 1, human cardiac myocytes, whole-cell voltage-clamp technique, calcium currents, sodium currents.

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