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Biphasic vasodilatory responses to adenosine and 5'-N-ethylcarboxamidoadenosine (NECA) were observed in the coronary vasculature of K(+)-arrested perfused rat hearts. Dose-response data for both agonists were best represented by two-site models. For adenosine, two sites with negative log ED50 (pED50) values of 8.1 +/- 0.1 (mean +/- S.E.M) and 5.2 +/- 0.1 were obtained, mediating 31 +/- 2% and 69 +/- 2% of the total response. In the presence of 8-phenyltheophylline, the vasodilatory response to adenosine remained best fitted to a two-site model with pED50 values of 7.0 +/- 0.2 and 5.4 +/- 0.2. The relative contribution of each site to the total response remained unchanged. For NECA, pED50 values of 9.6 +/- 0.1 and 6.8 +/- 0.2 were obtained, representing 48 +/- 3% and 52 +/- 3% of the sites, respectively. In contrast, ATP produced a monophasic response with a pED50 value of 8.8 +/- 0.1. These results provide evidence of adenosine receptor and response heterogeneity in the in situ coronary vasculature.

Exogenous adenosine causes a monophasic dilation of the coronary vessels in paced, perfused rat heart preparations. Because levels of endogenous adenosine in paced hearts may mask the presence of high potency adenosine receptors, we have developed a method to measure coronary vascular responses in a potassium-arrested heart. Hearts from adult male, Wistar rats were perfused at a constant flow rate of 10 mL/min in the nonrecirculating, Langendorff mode, using Krebs-Henseleit buffer. After 30 min, coronary perfusion pressure was 44 +/- 1 mmHg (mean +/- SEM). Hearts were then perfused with a modified Krebs-Henseleit buffer containing 35 mM potassium. Coronary perfusion pressure increased by 84 +/- 3 mmHg. Adenosine-induced reductions in coronary perfusion pressure were expressed as a percentage of the maximal increase in pressure produced by modified Krebs-Henseleit buffer from the equilibration level. A concentration-response curve for adenosine (n = 6) was biphasic and best described by the presence of two adenosine receptors, with negative log EC50 values of 8.8 +/- 0.3 and 4.3 +/- 0.1, representing 29 +/- 3 and 71 +/- 3%, respectively, of the observed response. Interstitial adenosine sampled by microdialysis during potassium arrest was 25% of the concentration found in paced hearts. Endogenous adenosine in nonarrested hearts may obscure the biphasic response of the coronary vessels to adenosine.

Actively transcribed genes are more susceptible to nuclease digestion, an observation suggested to reflect an altered state of chromatin organization. It has been hypothesized that exposure or sequestration of chromatin domains is a higher order gene regulatory mechanism. In order to test whether tissue lineage is organized by mechanisms at the level or chromatin structure, three cardiac phenotype-conferring genes (atrial natriuretic factor, myosin light chain-1-ventricular and alpha-tropomyosin) have been assessed for DNase 1 sensitivity in nuclei prepared from tissues of the developing guinea pig. These data have been related to the level of tissue mRNA expression of these genes to ascertain whether the exposed state of genes can occur when transcription is low or undetectable. Although this phenomenon was evident in some cases, the data were not consistent with mechanisms at the level of chromatin structure directing tissue type.

As a prerequisite to investigating the specification and differentiation of cardiac tissue in vitro, the ontogeny of a number of putative cardiac-specific, and striated muscle-specific gene transcripts has been studied. The probes used include cDNAs of alpha-actins, myosin heavy chains, myosin light chains, alpha-tropomyosin, troponin-T and atrial natriuretic factor. The expression of these genes was monitored by Northern analysis of heart and various other tissues at three developmental ages, viz, adult, neonatal and mid-foetal. The aim of this exercise was to confirm the efficacy of a number of markers to represent a cardiac-specific subset of gene expression in our mammalian model, the guinea pig. Our results indicate predominantly cardiac expression for the mRNA transcripts of cardiac alpha-actin (c alpha-actin), cardiac myosin heavy chain-alpha (MHC alpha), cardiac myosin heavy chain-beta (MHC beta), myosin light chain-1A (MLC1A), myosin light chain-1V (MLC1V), alpha-tropomyosin (alpha TM), cardiac troponin-T (cTnT) and atrial natriuretic factor (ANF). Furthermore, cardiac-specific expression at the midfoetal time point was observed for five gene transcripts, MLC1V, MHC alpha, MHC beta, striated alpha TM and ANF. No genes were expressed exclusively in cardiac tissue; for example, expression of the genes for c alpha-actin, both cardiac MHCs, both MLCs, alpha TM and cTnT was evident in skeletal and vascular smooth muscles at some stages of development. An interesting difference between this species and those of previous studies was the minor contribution of skeletal alpha-actin to cardiac phenotype.(ABSTRACT TRUNCATED AT 250 WORDS)

Objective: To examine the effect of hydrogen peroxide on the function and ultrastructure of cardiac muscle and the coronary vasculature in an isovolumic rat heart preparation perfused at constant flow., Design: Ventricular function was monitored via a balloon placed in the left ventricle and the response of the coronary vessels to vasodilators was assessed in hearts arrested with 35 mM potassium and treated with 5 microM phenylephrine to contract the coronary resistance vessels. Changes in coronary perfusion pressure reflect changes in resistance vessel tone. SETTING/ANIMALS: This experimental study consisted of 14 heart preparations, six control and eight treated hearts., Interventions: Hydrogen peroxide was included in the perfusate at a final concentration of 250 microM for 20 mins., Main Results: Hydrogen peroxide reduced rate-pressure product by 42%, caused a fivefold increase in end-diastolic pressure and increased coronary perfusion pressure by 33%. Also, the response of the coronary vasculature to the endothelium-dependent vasodilator, histamine, and endothelium-independent vasodilator, sodium nitrite, was decreased by 55% and 53%, respectively. Electron microscopy of hydrogen peroxide-treated hearts showed damage to both capillaries and arterioles. Endothelial cells were distorted and contained pinocytotic vesicles, endothelial cell junctions were disrupted and blood vessels were detached from surrounding tissue. A comparatively small amount of injury was seen in the myocyte population., Conclusions: The greater amount of ultrastructural damage seen in blood vessels compared with cardiac muscle suggests that the smooth muscle and endothelial cells of the vasculature are more susceptible to oxidant injury than the myocytes.

Adenosine may modulate blood flow and electrical activity in heart in response to changes in myocardial energy metabolism. In the present study, 31P NMR spectroscopy was used to examine the relation between cytosolic phosphate metabolite levels and release of adenosine into the venous effluent of isovolumic heart during graded low-flow ischaemia or metabolic stimulation with isoproterenol. When coronary flow rate was varied in steps between 1.6 and 12 ml/min/g, cytosolic ATP levels did not change significantly but the phosphorylation potential exhibited a linear correlation with flow rate below approximately 7 ml/min/g. Purine release (adenosine and inosine) correlated linearly with the cytosolic phosphorylation potential and free AMP concentration. Metabolic stimulation of hearts with isoproterenol (0.4, 3.0, and 60 nM), produced a significant fall in cytosolic ATP levels and decreased the cytosolic phosphorylation potential. Purine release in these hearts increased exponentially as the cytosolic phosphorylation potential dropped, and as cytosolic free AMP increased. These results support a link between the phosphorylation potential and the mechanism of adenosine production during ischaemia and metabolic stimulation. Presumably, this link is the activity of the enzyme 5'-nucleotidase, which is responsible for converting AMP to adenosine, together with the concentration of its substrate, AMP. In low-flow ischaemia, cytosolic AMP may control adenosine formation. With isoproterenol stimulation, a more complex relationship exists, indicating possible allosteric regulation of the enzyme(s) responsible for adenosine formation, in addition to changes in AMP concentration.

Clinical proton NMR imaging uses magnetic field strengths in the range 0.1 to 0.5 T. In addition to the large static magnetic field, patients are exposed to magnetic field gradients during imaging and under extreme conditions, such as power failure or quenching, the field may collapse precipitously. A potential source of hazard to patients under these conditions is the induction of thoracic currents which may trigger ventricular fibrillation. In the present experiments, a 0.16 T resistive magnet with a time constant of 60 ms, powered by a programmable power supply, was used to examine any possible effects of static and changing magnetic field on the ECG and arterial blood pressure of anesthetized rats and guinea pigs. Animals were exposed to the following field conditions: static fields of 0.16 T; sine, triangular, and square wave modulated fields from 0.1 to 2 Hz; rapid field switches in excess of 2.0 T/s for 25 ms timed to occur at different stages of the cardiac cycle, including the vulnerable period during ventricular repolarization; and AC fields of 50 Hz. No change was observed in the blood pressure, heart rate, or ECG under any of the field conditions examined.

The purpose of the present investigation was to study the relation between energy metabolism and contractile function in the isovolumic guinea pig heart. 31P nuclear magnetic resonance spectroscopy was used to measure changes in the intracellular levels of creatine phosphate, ATP, inorganic phosphate, and pH during 2.43 min total global ischemia and 2.43 min reperfusion, with a time resolution of 9.7 s. From these data, cytosolic changes in the phosphorylation potential, [ATP]-to-[ADP] ratio, free-energy change of ATP hydrolysis, and concentration of free ADP were estimated. The simultaneous monitoring of functional and biochemical parameters allowed them to be directly correlated with respect to time and with respect to each other. No significant changes in ATP were detected at any time, but changes in all other biochemical data were highly correlated with changes in contractile function. Kinetic analysis, using a nonlinear least-squares fit of the experimental points, revealed that the changes in most parameters fitted monoexponential functions. Each parameter was ranked according to its half time, which revealed that the phosphorylation potential was the only metabolic parameter to change at a rate faster than loss of contractile function during ischemia, and all metabolic changes, with the exception of pH, led the recovery of contractile function during reperfusion, the most rapid change occurring in the free ADP concentration. It is concluded that the cytosolic phosphorylation potential controls the contractile function of the heart and that cytosolic free ADP is important in the control of mitochondrial oxidative phosphorylation.

Injury is sustained by isolated hearts on repletion with calcium after a short period of perfusion with calcium-free medium at 37 degrees. A possible mechanism for the 'calcium paradox' is that exposure to a calcium-fre medium removes extracellular calcium rendering the sarcolemma more permeable to calcium. On calcium repletion, cell injury is triggered by calcium influx. Since lanthanum is known to displace calcium from extracellular pools in heart, it was used in an attempt to modulate the injury of the calcium paradox. The presence of 10 microM lanthanum in the calcium-free perfusion fluid was found to inhibit totally protein release normally produced by subsequent calcium exposure. When lanthanum was added after the calcium-free period and before calcium repletion, protein release was only partly prevented. This shows that a change in membrane properties occurred during the calcium-free perfusion period which could be prevented, but not reversed, by the addition of lanthanum.

The extent to which endogenous, extracellular adenosine mediates increased coronary flow in crystalloid-perfused, isovolumic rat hearts stimulated with either norepinephrine or isoproterenol was examined. When infused into the coronary circulation, norepinephrine (1 x 10(-7) M) rapidly increased left ventricular developed pressure (LVDP) from 81 +/- 6 to 235 +/- 13 mmHg (1 mmHg = 133.3 Pa) and coronary flow from 12.7 +/- 0.8 to 18.4 +/- 0.7 mL.min-1.g-1. The presence of either adenosine deaminase (2 U.mL-1) or the adenosine receptor antagonist, 8-phenyltheophylline (5 x 10(-6) M) in the perfusate of norepinephrine-stimulated hearts augmented the increase in LVDP and +/- dP/dtmax by 10-20% but reduced the increase in coronary flow by 34%. Doubling the rate of adenosine deaminase infusion, or infusing the enzyme and 8-phenyltheophylline together did not alter their inhibitory effectiveness. Similar results were observed with hearts stimulated with isoproterenol (5 x 10(-8) M). These data show that about a third of the vasodilation that results from the metabolic stimulation of rat heart by catecholamines is due to the receptor-mediated action of extracellular adenosine.

This study examines the possibility that endogenous adenosine modulates efficiency in isovolumic perfused rat hearts stimulated with isoproterenol or norepinephrine. Efficiency in these hearts is calculated as the rate of pressure work divided by the myocardial oxygen consumption. Within 2 min of infusion of isoproterenol (50 nM), heart rate increased by 35%, the rate pressure product by 290%, oxygen consumption by 142%, and efficiency by 67%. Infusion of adenosine deaminase (2-4 IU/ml), or 8-phenyltheophylline (5 microM), into stimulated hearts augmented the increase in heart rate by 40-45%, rate-pressure product by 18-20%, and oxygen consumption by 50-55%. Efficiency was reduced by 30-35%. Adenosine release into the coronary venous effluent increased from 195 +/- 20 pmol/min/g to 2400 +/- 180 pmol/min/g after 5 min. A similar pattern of results was observed when norepinephrine (0.1 mM) was used. The results indicate that extracellular adenosine, released by catecholamine treatment, inhibits the effects of the catecholamines on rate and contractility. Consequently, adenosine reduces cardiac work (rate-pressure product), but in so doing, improves efficiency.