scholarly journals The control of adenosine concentration in polymorphonuclear leucocytes, cultured heart cells and isolated perfused heart from the rat

1983 ◽  
Vol 214 (2) ◽  
pp. 317-323 ◽  
Author(s):  
A C Newby ◽  
C A Holmquist ◽  
J Illingworth ◽  
J D Pearson
Keyword(s):  
Adenine Nucleotide ◽  
Maximal Activity ◽  
Adenosine Kinase ◽  
Neonatal Rat ◽  
Heart Cells ◽  
Polymorphonuclear Leucocytes ◽  
Perfused Heart ◽  
Intact Cells ◽  
Adenosine Concentration ◽  
Rat Heart Cells

Rat polymorphonuclear leucocytes or neonatal-rat heart cells in culture were treated with 2′-deoxycoformycin and 5-iodotubercidin at concentrations that inhibited adenosine deaminase (EC 3.5.4.4) and adenosine kinase (EC 2.7.1.20) inside the intact cells, and the rate of adenosine accumulation was determined. The basal rate of adenosine formation was 2% (polymorphonuclear leucocytes) or 9% (heart cells) of the maximal activity of adenosine kinase also measured in intact cells. Greatly increased rates of adenosine formation were observed during adenine nucleotide catabolism. This condition also led to a decrease in adenosine kinase activity. When isolated rat hearts were perfused with 5-iodotubercidin alone at a concentration which inhibited adenosine kinase, no increase in tissue or perfusate adenosine or inosine concentration was observed. However, perfusion with hypoxic buffer or infusion of adenosine into the coronary circulation at a rate (20 nmol/min) equivalent to 40% of the activity of adenosine kinase caused large increases in effluent perfusate adenosine and inosine concentrations. These data argue unanimously against the existence of a substrate cycle controlling adenosine concentration. They suggest instead that an increase in the rate of adenosine formation is the principal cause of elevations in adenosine concentration during ATP catabolism.

Localization of the Na/Ca exchange-dependent Ca compartment in cultured neonatal rat heart cells

1995 ◽  
Vol 268 (1) ◽  
pp. C119-C126 ◽  
Author(s):  
G. A. Langer ◽  
S. Y. Wang ◽  
T. L. Rich
Keyword(s):  
Total Content ◽  
Cardiac Cells ◽  
Neonatal Rat ◽  
High Dose ◽  
Heart Cells ◽  
Cell Content ◽  
Intact Cells ◽  
Content Ratio ◽  
On Line ◽  
Rat Heart Cells

It has been previously established, in both adult and cultured neonatal cardiac cells, that there is a discrete Na/Ca exchange-dependent Ca compartment. It has been proposed that a component of junctional sarcoplasmic reticulum (JSR) Ca and Ca bound to the apposed inner sarcolemmal leaflet represent together the subcellular locus of the compartment. The present study examines this proposal. The amount of Ca in the total compartment is measured isotopically in intact functional cells (using the on-line "scintillation disk" technique) under a variety of perfusion conditions. Under identical labeling conditions, sarcolemmal membranes are rapidly (within a few hundred milliseconds) isolated from another set of intact cells by "gas dissection," and the amount of Ca bound to the membranes is measured. Probes that specifically decrease SR Ca content (thapsigargin, caffeine, low-dose ryanodine) decrease total cell content and sarcolemmal binding proportionally. High-dose ryanodine (producing closure of SR channels) markedly reduces sarcolemmal binding relative to total content of the compartment. The sarcolemmal sites saturate between 1 and 2 mM extracellular Ca ([Ca]o), whereas the total compartment saturates between 4 and 6 mM [Ca]o. Below 1 mM [Ca]o, sarcolemmal binding is maintained relative to total compartment content. Finally, the total compartment increases after reversal of the intracellular Na to extracellular Na ([Na]i/[Na]o) gradient with sarcolemmal content-to-total content ratio dependent on the method used to reverse the [Na]i/[Na]o ratio. The results are consistent with localization of the Na/Ca exchange-dependent compartment to the subsarcolemmal region ("cleft") where JSR Ca is in equilibrium with anionic inner sarcolemmal leaflet Ca binding sites.

scholarly journals Effect of 5′-deoxy-5′-isobutylthioadenosine on formation and release of adenosine from neonatal and adult rat ventricular myocytes

1993 ◽  
Vol 291 (3) ◽  
pp. 833-839 ◽  
Author(s):  
P Meghji ◽  
A C Skladanowski ◽  
A C Newby ◽  
L L Slakey ◽  
J D Pearson
Keyword(s):  
Neonatal Rat ◽  
Nucleoside Transport ◽  
Minor Role ◽  
Heart Cells ◽  
Polymorphonuclear Leucocytes ◽  
Intact Cells ◽  
Rat Cardiomyocytes ◽  
Adult Rat ◽  
Atp Concentration ◽  
Rat Myocytes

1. Studies in rat polymorphonuclear leucocytes have suggested that 5′-deoxy-5′-isobutylthioadenosine (IBTA), an inhibitor of the IMP-selective cytosolic 5′-nucleotidase, may be used to test its role in adenosine formation in intact cells. We investigated adenosine formation in neonatal and adult rat cardiomyocytes. 2. 2-Deoxyglucose (30 mM) with oligomycin (2 micrograms/ml) induced a 90-100% fall in ATP concentration in 10 min in neonatal and 60 min in adult heart cells. Adenosine accumulation was substantially increased, accounting for 13% of the fall in ATP concentration in neonatal cells and 56% in adult cells. 3. Anti-(rat liver ecto-5′-nucleotidase) serum did not inhibit adenosine accumulation. Furthermore, dipyridamole (10 microM), a nucleoside-transport blocker, inhibited by 80% the appearance of the newly formed adenosine in the medium, showing that adenosine is produced intracellularly by both adult and neonatal-rat myocytes in response to inhibition of oxidative metabolism. 4. IBTA (3 mM) inhibited by 80% the appearance of adenosine in the medium, but did not inhibit total adenosine accumulation by neonatal-rat myocytes and only modestly inhibited total adenosine accumulation by adult myocytes. 5. IBTA, like dipyridamole, inhibited incorporation of extracellular adenosine (10 microM) into neonatal and adult ventricular myocyte nucleotides by 60-70%. Transport of IBTA (100 microM) into the cells did not appear to be inhibited by dipyridamole (30 microM). 6. We conclude that IBTA acted primarily to inhibit adenosine release from myocytes. The small effect on adenosine formation rates implies that the IMP-selective cytosolic 5′-nucleotidase plays a minor role in this tissue.

Myosin heavy chain turnover in cultured neonatal rat heart cells: effects of [Ca2+]i and contractile activity

1996 ◽  
Vol 271 (5) ◽  
pp. C1447-C1456 ◽  
Author(s):  
K. L. Byron ◽  
J. L. Puglisi ◽  
J. R. Holda ◽  
D. Eble ◽  
A. M. Samarel
Keyword(s):  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Protein Turnover ◽  
Ventricular Myocytes ◽  
Contractile Activity ◽  
Myofibrillar Protein ◽  
Neonatal Rat ◽  
Heart Cells ◽  
Cell Shortening ◽  
Rat Heart Cells

Blockade of L-type Ca2+ channels in spontaneously contracting cultured neonatal rat ventricular myocytes causes contractile arrest, myofibrillar disassembly, and accelerated myofibrillar protein turnover. To determine whether myofibrillar protein turnover. To determine whether myofibrillar atrophy results indirectly from loss of mechanical signals or directly from alterations in intracellular Ca2+ concentration ([Ca2+]i), contractile activity was inhibited with verapamil (10 microM) or 2,3-butanedione monoxime (BDM), and their effects on cell shortening, [Ca2+]i, and myosin heavy chain (MHC) turnover were assessed. Control cells demonstrated spontaneous [Ca2+]i transients (peak amplitude 232 +/- 15 nM, 1-2 Hz) and vigorous contractile activity. Verapamil inhibited shortening by eliminating spontaneous [Ca2+]i transients. Low concentrations of BDM (5.0-7.5 mM) had no effect on basal or peak [Ca2+]i transient amplitude but reduced cell shortening, whereas 10 mM BDM reduced both [Ca2+]i transient amplitude and shortening. Both agents inhibited MHC synthesis, but only verapamil accelerated MHC degradation. Thus MHC half-life does not change in parallel with contractile activity but rather more closely follows changes in [Ca2+]i. [Ca2+]i transients appear critical in maintaining myofibrillar assembly and preventing accelerated MHC proteolysis.

Cation exchange and glycoside binding in cultured rat heart cells

1979 ◽  
Vol 236 (1) ◽  
pp. C87-C95 ◽  
Author(s):  
D. McCall
Keyword(s):  
Neonatal Rat ◽  
Heart Cells ◽  
Turnover Rates ◽  
Ouabain Binding ◽  
Na Pump ◽  
Mammalian Tissues ◽  
Flux Measurements ◽  
The Mean ◽  
Rat Heart Cells ◽  
K Transport

The Na/K-exchange characteristics, ouabain-binding kinetics, and Na pump turnover rates of synchronously contracting monolayers of neonatal rat myocardial cells were studied. The cells exchange Na rapidly (T1/2 = 35 s) with a mean Na flux of approximately 25 (pmol/cm2)/s. The half time (T1/2) of K exchange is much longer (12 min); the mean K flux is 13 (pmol/cm2)/s. Active Na/K transport, as measured by K influx, is relatively ouabain sensitive, and 10(-6) M ouabain produces half-maximal inhibition. Ouabain (10(-2)M) inhibits 60% of the Na efflux and 75% of the K influx. The cells bind [3H]ouabain rapidly (T1/2 = 8 min), but release it very slowly (T1/2 = 11 h), and both the amount bound and the rate of binding were inversely proportional to extracellular K. Specific [3H]ouabain binding demonstrates saturation reaching a maximum of 1.6 x 10(6) molecules per cell at 2 x 10(-7) M [3H]ouabain. From cell surface area and ouabain-sensitive flux measurements, the Na pump density was calculated at 720/micrometer2 with an individual pump turnover rate of 50/s. Thus the studies indicate that despite their neonatal origin, the behavior of the Na pump in these cells is very similar to that in other mammalian tissues.

Chronotropic effect of histamine on cultured neonatal rat heart cells

1979 ◽  
Vol 58 (2) ◽  
pp. 117-123 ◽  
Author(s):  
Klara Csete ◽  
Marie-Claude Auclair ◽  
Paul Lechat
Keyword(s):  
Rat Heart ◽  
Neonatal Rat ◽  
Heart Cells ◽  
Chronotropic Effect ◽  
Neonatal Rat Heart ◽  
Rat Heart Cells

Arachidonic acid and lipoxygenase metabolites uncouple neonatal rat cardiac myocyte pairs

1992 ◽  
Vol 263 (2) ◽  
pp. C494-C501 ◽  
Author(s):  
K. D. Massey ◽  
B. N. Minnich ◽  
J. M. Burt
Keyword(s):  
Arachidonic Acid ◽  
Gap Junctions ◽  
Cardiac Myocyte ◽  
Underlying Mechanism ◽  
Neonatal Rat ◽  
Heart Cells ◽  
Membrane Phospholipids ◽  
Lipoxygenase Pathway ◽  
Rat Heart Cells ◽  
The Right

The effects of arachidonic acid (AA) and its metabolites on the conductance (gj) of the gap junctions between neonatal rat myocardial cells was investigated. AA reduced gj in a dose- (2, 5, and 20 microM) and time-dependent fashion. Pretreatment of the cells with an inhibitor of the 5-lipoxygenase pathway, U-70344A, shifted the dose-response curve to the right; pretreatment with indomethacin, an inhibitor of the cyclooxygenase pathway, had no effect. The mean time to uncoupling was 3.7 +/- 0.3, 3.8 +/- 0.9, and 4.6 +/- 0.6 min (means +/- SE, P less than 0.05) for 5 microM AA, 5 microM AA + indomethacin, and 5 microM AA + U-70344A, respectively. Incorporation of AA into membrane phospholipids was not affected by the inhibitor. These studies suggest that complete uncoupling of the cells occurred at membrane concentrations of 3-4 mol%. The data indicate that AA and a 5-lipoxygenase metabolite uncouple neonatal rat heart cells. The data are discussed with respect to the possible underlying mechanism of uncoupling and the potential role of gap junctions in arrhythmia formation in ischemic heart disease.

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