K+, Na+, and Cl- activities in ventricular myocytes isolated from rabbit heart

1986 ◽  
Vol 251 (2) ◽  
pp. C197-C208 ◽  
Author(s):  
M. Desilets ◽  
C. M. Baumgarten
Keyword(s):  
Ventricular Myocytes ◽  
Reference Electrode ◽  
Rabbit Heart ◽  
Cardiac Cells ◽  
Adult Rabbit ◽  
Intracellular Ion Activities ◽  
Ion Activities ◽  
Tip Potential ◽  
Isolated Cardiac Cells ◽  
Intracellular Milieu

Intracellular K+, Na+, and Cl- activities (aiK, aiNa and aiCl) were measured in ventricular myocytes enzymatically isolated from adult rabbit heart. The activities in normal Tyrode solution containing 2.5 mM Ca2+ were the following (in mM): aiK = 100.0 +/- 3.5 (n = 9); aiNa = 8.4 +/- 1.5 (n = 6); and aiCl = 17.9 +/- 1.5 (n = 11) (mean +/- SE). Membrane potential was -81.6 +/- 0.7 mV (n = 26). These values were determined after correction for changes of junction and tip potential at the reference electrode, estimated to be 4.9 +/- 0.6 mV (n = 7) for 0.15 M KCl-filled electrodes; and intracellular interference detected by the Cl- ion-selective electrode, 11.2 +/- 0.6 mM (n = 4). Extended-tip shunting was avoided by fabricating Na+ ion-selective microelectrodes from aluminosilicate rather than borosilicate glass. These results show that isolated cardiac cells can maintain normal intracellular ion activities. Diffusion of electrolyte from the reference electrode can rapidly alter the intracellular milieu, however. After 10 min of impalement with 0.15 M KCl-filled microelectrodes (resistance approximately equal to 25 M omega), aiK increased by 8.7 +/- 2.0 mM and aiCl by 10.3 +/- 3.1 mM. In contrast, aiNa did not significantly change during the double impalement.

Effect of partial Na pump and Na-H exchange inhibition on [Ca]i during acidosis in cardiac cells

1991 ◽  
Vol 261 (5) ◽  
pp. C758-C766 ◽  
Author(s):  
T. Nakanishi ◽  
M. Seguchi ◽  
T. Tsuchiya ◽  
E. J. Cragoe ◽  
A. Takao ◽  
...  
Keyword(s):  
Intracellular Ph ◽  
Fluorescent Dyes ◽  
Contractile Function ◽  
Cardiac Cells ◽  
Adult Rabbit ◽  
Mechanical Function ◽  
Intracellular Acidosis ◽  
Na Pump ◽  
Intracellular Ca ◽  
Isolated Cardiac Cells

This study investigated the effects of partial Na pump inhibition and Na-H exchange inhibition on contractile function, intracellular pH (pHi), and intracellular Ca concentration ([Ca]i) during intracellular acidosis, using the fluorescent dyes 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein and fura-2 in isolated cardiac cells of adult rabbits. Intracellular acidosis with normal extracellular pH was induced by an NH4Cl (10 mM) prepulse technique. A nontoxic concentration (0.5 microM) of ouabain was used to inhibit the Na pump. 5-(N-ethyl-N-isopropyl)amiloride (EIPA) was used to inhibit Na-H exchange. In both the absence and presence of ouabain, pHi fell transiently and then recovered after removal of NH4Cl. Ouabain did not alter the pHi changes observed after removal of NH4Cl. Diastolic and systolic [Ca]i increased during acidosis after NH4Cl removal. In the presence of ouabain, the increase in [Ca]i during acidosis was greater than that in the absence of this drug. Ouabain enhanced the recovery of contractile function during acidosis. In both the absence and presence of ouabain, Na-H exchange inhibition by EIPA reduced the recovery of pHi and mechanical function and the increase in [Ca]i, which were normally observed after NH4Cl removal. These data suggest that in adult rabbit myocytes the Na pump inhibition enhances the increase in [Ca]i during acidosis, and the Na-H exchange inhibition reduces it. The [Ca]i increase during acidosis may be in part due to the altered Na-Ca exchange, which in turn results from the increased Na-H exchange.

Electrophysiology of the Sodium-Potassium-ATPase in Cardiac Cells

2001 ◽  
Vol 81 (4) ◽  
pp. 1791-1826 ◽  
Author(s):  
Helfried Günther Glitsch
Keyword(s):  
Cell Membrane ◽  
Potential Gradient ◽  
Outward Current ◽  
Pump Current ◽  
Cardiac Cells ◽  
Animal Cells ◽  
Excitable Cells ◽  
Extracellular Medium ◽  
Enzymatic Isolation ◽  
Isolated Cardiac Cells

Like several other ion transporters, the Na+-K+ pump of animal cells is electrogenic. The pump generates the pump current I p. Under physiological conditions, I p is an outward current. It can be measured by electrophysiological methods. These methods permit the study of characteristics of the Na+-K+ pump in its physiological environment, i.e., in the cell membrane. The cell membrane, across which a potential gradient exists, separates the cytosol and extracellular medium, which have distinctly different ionic compositions. The introduction of the patch-clamp techniques and the enzymatic isolation of cells have facilitated the investigation of I p in single cardiac myocytes. This review summarizes and discusses the results obtained from I p measurements in isolated cardiac cells. These results offer new exciting insights into the voltage and ionic dependence of the Na+-K+ pump activity, its effect on membrane potential, and its modulation by hormones, transmitters, and drugs. They are fundamental for our current understanding of Na+-K+ pumping in electrically excitable cells.

Neurofilament proteins are co-expressed with desmin in heart conduction system myocytes

1990 ◽  
Vol 97 (1) ◽  
pp. 11-21
Author(s):  
M. Vitadello ◽  
M. Matteoli ◽  
L. Gorza
Keyword(s):  
Subcellular Distribution ◽  
Ultrastructural Level ◽  
Rabbit Heart ◽  
Cardiac Cells ◽  
Structural Components ◽  
Minor Population ◽  
Neuronal Proteins ◽  
Tissue Homogenates ◽  
A Minor

We have recently shown that specialized myocytes of the rabbit heart express a cytoskeletal protein similar to the M subunit of neurofilaments (NF). Since this result was obtained using a single anti-NF-M monoclonal antibody, we tested on conduction myocytes a panel of five anti-NF antibodies, specific for each of the three NF subunits and for phosphorylated and non-phosphorylated epitopes. Two antibodies, one specific for the L subunit and one for phosphorylated M subunit of NF, reacted with specialized myocytes in immunohistochemistry. In immunoblots on conduction tissue homogenates the two antibodies recognized two polypeptides with electrophoretic mobility and solubility properties identical to those of NF-L and NF-M in the sciatic nerve. The subcellular distribution of NF immunoreactivity in specialized myocytes was very similar to desmin localization; namely, it was distributed on large filamentous bundles and on fine filaments localized transversely at the level of the Z line. At the ultrastructural level, immunoreactive filaments were localized in the intermyofibrillar space and connected myofibrils with mitochondria. Co-expression of NF proteins and desmin was also observed in vitro in a minor population of cardiac myocytes cultured from embryonic rabbit heart. In most cases NF immunoreactivity co-localized with desmin, especially where filaments were well organized, but in some cells anti-NF and anti-desmin antibodies labelled different filamentous structures. These results indicate that NF proteins are structural components of the cytoskeleton of specialized myocytes and show a subcellular distribution very similar to desmin. Such a composition of intermediate filaments indicates that in these cardiac cells muscle differentiation is compatible with the expression of neuronal proteins.

Hyperthyroid adult rat cardiomyocytes. I. Nucleotide content, beta- and alpha-adrenoreceptors, and cAMP production

1989 ◽  
Vol 257 (5) ◽  
pp. C948-C956 ◽  
Author(s):  
C. M. Hohl ◽  
S. Wetzel ◽  
R. H. Fertel ◽  
D. K. Wimsatt ◽  
G. P. Brierley ◽  
...  
Keyword(s):  
Ventricular Myocytes ◽  
Adenine Nucleotide ◽  
Phosphodiesterase Inhibitor ◽  
Cardiac Cells ◽  
Beta Agonist ◽  
Camp Production ◽  
Adult Rats ◽  
Rat Cardiomyocytes ◽  
Adult Rat ◽  
Half Maximal Effective Concentration

Ventricular myocytes isolated from the hypertrophied hearts of thyrotoxic adult rats have an increase in mean protein content per myocyte (6.3 +/- 0.2 vs. 4.4 +/- 0.2 ng) compared with euthyroid cells. Viability and adenine nucleotide profiles are similar in both populations, but NAD content of the hyperthyroid myocytes is depressed (4.9 +/- 0.2 vs. 5.5 +/- 0.2 nmol/mg for controls) and UTP is higher (1.2 +/- 0.09 vs. 0.9 +/- 0.04 nmol/mg). Binding of (-)-[125I]iodocyanopindolol to intact hyperthyroid myocytes is increased by 42% compared with controls, with no change in the dissociation constant (Kd). This elevation in beta-receptor number is correlated to enhanced beta-agonist-induced adenosine 3',5'-cyclic monophosphate (cAMP) production. The half-maximal effective concentration (EC50) for the euthyroid isoproterenol dose-response curve is 2.14 x 10(-7) M but is decreased to 2.51 x 10(-8) M in hyperthyroid cardiac cells. Basal adenylate cyclase activity is apparently not affected by thyroid hormones, since basal cAMP levels for both groups are identical (5 pmol/mg) and both rise roughly twofold in the presence of a phosphodiesterase inhibitor. Forskolin-induced cAMP production and cAMP-specific phosphodiesterase activity are similar as well. In contrast to beta-adrenergic response, there are no significant differences in alpha 1-antagonist [3H]prazosin binding parameters between hyperthyroid and euthyroid cardiomyocytes.

Shunt resistance and ion permeabilities in normal and cystic fibrosis airway epithelia

1989 ◽  
Vol 256 (5) ◽  
pp. C1054-C1063 ◽  
Author(s):  
N. J. Willumsen ◽  
R. C. Boucher
Keyword(s):  
Cystic Fibrosis ◽  
Steady State ◽  
Apical Membrane ◽  
Short Circuit ◽  
Strongly Correlated ◽  
Shunt Resistance ◽  
Human Nasal Epithelium ◽  
Cystic Fibrosis Airway ◽  
Intracellular Ion Activities ◽  
Ion Activities

A method for determination of shunt resistance (Rs) and absolute conductive ion permeabilities of the apical membrane in epithelia from steady-state data is described. The method assumes that the currents are satisfactorily described by the Goldman-Hodgkin-Katz regime. Its application requires measurements of standard transepithelial electrophysiological parameters and of one or more intracellular ion activities. It is applicable under both open- and short-circuit conditions. The method was tested in an electrophysiological analysis of cultured normal and cystic fibrosis (CF) human nasal epithelium. In 15 normal and 10 CF preparations with mean transepithelial resistances of 338 and 427 omega.cm2, Rs was 412 and 623 omega.cm2, respectively. The Rs values determined with the present method were strongly correlated (r = 0.94) with those obtained with another method available in the electrophysiological literature but were as a mean 20% lower. Amiloride increased Rs by 25% in CF and by 8% in normal preparations. In normal preparations, the apical Cl permeability (PCla) was 3.6 x 10(-6) cm/s, and the apical Na permeability (PNaa) was 1.6 x 10(-6) cm/s. In CF preparations, PCla was reduced to a maximum of 2.3 x 10(-7) cm/s, whereas PNaa was increased to 6.2 x 10(-6) cm/s. The apical membrane electromotive force was -1 mV in normal and 43 mV in CF preparations. It is concluded that the method can be used to calculate Rs, apical membrane ion permeabilities, and electromotive forces from steady-state electrophysiological data.

Effects of ouabain on intracellular ion activities of sensory neurons of the leech central nervous system

1991 ◽  
Vol 65 (3) ◽  
pp. 736-746 ◽  
Author(s):  
W. R. Schlue
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Input Resistance ◽  
Membrane Resistance ◽  
Membrane Potential Change ◽  
Intracellular Na Activity ◽  
Order Of Magnitude ◽  
Intracellular Ion Activities ◽  
Ion Activities ◽  
K Concentration

1. The intracellular K+, Na+, and Ca2+ of mechanosensory neurons in the central nervous system of the leech Hirudo medicinalis was measured using double-barreled ion-sensitive microelectrodes. 2. After inhibition of the Na(+)-K+ pump with 5 x 10(-4) M ouabain, the intracellular K+ activity (aKi) decreased, while the intracellular Na+ activity (aNai) increased. The input resistance decreased in the presence of ouabain. The intracellular Ca2+ increased more than one order of magnitude after ouabain addition. All changes in intracellular ion activities and membrane resistance were fully reversible. 3. When extracellular Na+ concentration ([Na+]o) was removed [replaced by tris(hydroxymethyl)aminomethane (Tris)], aNai decreased. In the absence of [Na+]o, aKi and aNai remained unchanged after inhibition of the Na(+)-K+ pump by reducing the extracellular K+ concentration ([K+]o) to 0.2 mM. The membrane resistance increased under these conditions. 4. The intracellular Ca2+ decreased or remained constant after removal of [Na+]o. Addition of ouabain in the absence of [Na+]o did not change intracellular Ca2+, which only increased after readdition of [Na+]o. 5. The relative K+ permeability (PK) measured as membrane potential change during a brief increase of the [K+]o from 4 to 10 mM, increased manyfold after addition of ouabain but only little if [Na+]o had been removed before adding ouabain. 6. The results suggest that the intracellular Na+ increase after inhibition of the Na(+)-K+ pump affects the intracellular Ca2+ level by stimulating a Nai(+)-Ca2+ exchange mechanism. The subsequent intracellular Ca2+ activity (aCai) rise may result in an increase of the membrane permeability to K+ ions.

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