Functional, morphological and metabolic characteristics of isolated hearts from normotensive and spontaneously hypertensive rats before, during and after renal hypertension

It has been much discussed whether left ventricular hypertrophy (LVH) in hypertension implies improved or impaired cardiac performance, mainly because experiments in various hypertensive models have given controversial results. It has, for example, been suggested that increased collagen content may depress LV function both in renal and spontaneous hypertension (SHR) in rats. For such reasons cardiac performance was studied both in SHR and in normotensive control rats (NCR) before and during superimposed two-kidney, one clip Goldblatt hypertension, and also 1 week after reversal of hypertension by unclipping. The LV function of the isolated hearts was determined in an antegrade working heart perfusion system. Further, myocardial morphology, with regard to fibrous tissue infiltration and energy metabolic status, were evaluated in the renal hypertensive rats before and after unclipping. Compared with NCR, maximal cardiac performance was elevated in the hypertrophied SHR hearts, but depressed when renal hypertension was superimposed, even though this led to further LVH. However, one week after reversal of renal hypertension, when LVH was still considerable, cardiac function was increased well above the control level, even though the stores of high energy compounds and the content of myocardial fibrous tissue was almost the same as during renal hypertension. It is concluded that LVH generally enhances cardiac performance, but that concomitant renal hypertension exerts a cardio-depressive influence which can neither be ascribed to an increased fibrous tissue content nor to a reduced energy charge potential. It is therefore suggested that some negative inotropic agent of renal or extrarenal origin is released during two-kidney, one clip renal hypertension, which offsets the enhanced performance induced by the left ventricular hypertrophy.     

Functional and biochemical effects of a K+-ionophore on the isolated perfused rat heart

Valinomycin, a K⁺-specific ionophore, influenced function and metabolism of isolated, perfused rat hearts in a dose-dependent fashion. At a concentration of 0.05 μg ml^-1 in perfusion fluid a 50% reduction of heart rate (HR) and a 90% reduction in max dP/dt were observed. These effects were paralleled by a substantial decrease of myocardial energy charge from about 0.80 to 0.20. A 2.5 fold increase in tissue lactate concentration indicated an increased rate of glycolytic activity. Low ATP combined with high ADP and AMP levels as found in these valinomycin-treated hearts is known to promote phosphofructokinase activity and may explain the elevated lactate levels. A significant increase in the concentrations of adenosine, IMP and inosine was observed as well.     

Inducible collagenolytic activity in isolated perfused rat hearts.

There is an extensive collagen network in the heart. The precise anatomy and function of this system has not been fully elucidated. The system does appear to contribute to diastolic compliance, and evidence indicates that the system may be important in directing the stress generated by sarcomeres to the ventricular cavity. Little is known about the mechanisms controlling collagen deposition and resorption in the heart. In this paper the authors demonstrate that disulfide reagents are capable of inducing a collagenolytic reaction in the isolated perfused heart that removes all components of the collagen matrix of the heart as visualized by scanning electron microscopy. The expression of collagenolytic activity requires perfusion of the heart for 1 hour with a disulfide reagent followed by 2 hours with Krebs-Hensleit alone. These results suggest that an inducible and active collagenolytic system exists in cardiac tissue and that this system may be expressed under conditions of oxidative stress.     

Cocaine-induced small vessel spasm in isolated rat hearts.

Cocaine abuse has been associated with pathologic cardiovascular events including acute myocardial infarction (AMI) and sudden death. Although coronary vasospasm has been proposed as a possible mechanism, the ability of cocaine to induce coronary spasm has not been conclusively demonstrated. In these studies, isolated rat hearts were perfused with cocaine (100 micrograms to 500 micrograms/ml) for 1 minute, perfusion-fixed with glutaraldehyde, and histologically assessed for evidence of coronary spasm through light and electron microscopy. Light micrographs revealed that cocaine induced spasm in coronary arterioles up to 65 microns in diameter, whereas larger caliber vessels did not constrict. Ultrastructurally, vacuolation was observed in the endothelial and smooth muscle cells of constricted arterioles. Endothelial integrity was maintained and interendothelial junctions remained intact. Morphologic evidence of constriction was supported by data obtained from Langendorff-heart preparations in which cocaine reduced myocardial flow rate under constant pressure conditions and increased aortic perfusion pressure under constant flow conditions. Spasm induced by cocaine was prevented by the calcium entry blocker nitrendipine, but not by phentolamine, an alpha-adrenergic antagonist. The finding of small vessel spasm in this study may explain the significant number of clinical cases of cocaine-associated AMI in which the main coronary arteries appear angiographically normal.     

Albumin decreases hydrogen peroxide and reperfusion injury in isolated rat hearts

Perfusion with human serum albumin decreased myocardial hydrogen peroxide (H₂O₂) levels (as assessed by inactivation of myocardial catalase activities following ammotriazole pretreatment) and increased myocardial ventricular developed pressures (DP), contractility (+dP/dt) but not relaxation rate (-dP/dt) in isolated crystalloid perfused rat hearts subjected to normothermic global ischemia (20 min) and then reperfusion (40 min). Albumin also decreased H₂O₂ concentrations in vitro. The findings support the possibility that albumin may act as a protective O₂ metabolite scavenger in vivo.     

低氧及常氧灌流对离体大鼠心脏应激蛋白的诱导

目的和方法：观察低氧灌流和常氧灌流诱导离体大鼠心脏的应激蛋白合成。结果：低氧和常氧灌流均可诱导３个分子量为７０ｋＤａ的一组应激蛋白（ｓｐ６８，７０和７２）合成显著增加，５个分子量为６０ｋＤａ的一组应激蛋白（ｓｐ５４－６５）合成也有所增加。比较不同灌流方式和时程对应激蛋白合成的影响，发现７０ｋＤａ的一组应激蛋白合成量在常氧灌流的离体心脏中高于低氧灌流；并且该组应激蛋白合成量在经两种灌流方式处理１ｈ的以脏高于灌流处理２ｈ的心脏。结论：应激蛋白合成量并不与应激原的迭加或与细胞受环境损害的严重性成正比。对灌流的离体心脏而言，缺血可能是诱导应激蛋白合成的主要因素，低氧及常氧则起调节合成量的作用     

Sodium dependence of carnitine transport in isolated perfused adult rat hearts

In heart muscle, the intracellular carnitine concentration is approximately 40 times higher than the plasma carnitine concentration, suggesting the existence of an active transport process. At physiological serum carnitine concentrations (44 microM), 80% of total myocardial carnitine uptake occurs via a carrier-mediated transport system. The mechanism of this carrier-mediated transport was studied in isolated perfused rat hearts. Carnitine transport showed an absolute dependence on the extracellular sodium concentration. The rate of carnitine transport was linearly related to the perfusate sodium concentration at every perfusate carnitine concentration examined (15-100 microM). Total removal of extracellular sodium completely abolished the carrier-mediated transport. Decreasing the perfusate potassium concentration from a control of 5.9 to 0.6 mM stimulated transport by 35%, whereas increasing the extracellular potassium concentration from 5.9 to 25 mM reduced transport by 60%. The carrier-mediated transport was inversely proportional to the extracellular potassium concentration. Acetylcholine (10(-3) M), isoproterenol (10(-7) M), or ouabain (10(-3) did not alter the rate of carnitine transport. Addition of tetrodotoxin (10(-5) stimulated carnitine transport by about 40%, while gramicidin S (5 X 10(-6) M) decreased uptake by about 18% relative to control. The data provide evidence that carnitine transport by cardiac cells occurs by a Na+-dependent cotransport mechanism that is dependent on the Na+ electrochemical gradient.     

Functional biochemical changes and their pharmacologic correction by pirroksan in renal ischemia

Bilateral one-hour ischemia of the kidneys in rats causes essential metabolic shifts in the renal tissues, and diminished intensity of the energetic status as a result of which the intactness of the cell membranes and the main functions of the nephron are disturbed. Pharmacological correction of postischemic disorders by the alpha-adrenergic blocking agent pyrroxane increases the activity of oxidative processes and intensifies the glycolysis rate. Improvement of the energy provision of the nephron function occurs in parallel with restoration of the diuresis level and processes of filtration and reabsorption.     

Detection of myocardial cell damage in isolated rat hearts with near-infrared spectroscopy

One hallmark of cell death resulting from prolonged ischemia is cell membrane disruption. We apply optical spectroscopy to gauge membrane disruption in isolated rat hearts by monitoring (1) the washout of myoglobin (Mb) and (2) the accumulation of an exogenous contrast agent in permeabilized cells. The contrast agent, a neodymium (Nd) chelate, has several absorptions in the visible and near-IR, and when present in the perfusate, it cannot penetrate cellular membranes. When membrane integrity is disrupted, however, it is expected to accumulate within the intracellular space; moreover, cellular Mb is expected to wash out. To test this hypothesis, rat hearts (n=12) are perfused with Krebs-Henseleit buffer (KHB), followed by perfusion with KHB in which a 5 mM Nd-DTPA solution is present. Membrane damage is then induced by infusion of digitonin into the Nd-KHB perfusate to provide a digitonin concentration of 2.5, 5, or 10 microg/mL. After 30 min of infusion, Mb levels fall to 46+/-14% of baseline levels and Nd-DTPA rises to 161+/-19% of predigitonin levels. No apparent dependence of total membrane disruption on digitonin concentration over the concentration range studied is found, although higher concentrations do lead to more rapid membrane disruption.     

Evaluation of a lumped parameter model for isolated working rat hearts

When a mechanical model of the rat aortic input impedance is perfused with a pulsatile pump, the computed values of the impedance components vary linearly with flow rate and are interactive. When the model is perfused by an isolated rat heart, the total load upon the left ventricle consists of the model and coronary impedances in parallel. Adenosine triphosphate induces changes in coronary impedance, and the redistribution of cardiac output from the model to the coronary circulation causes flow-related changes in the model impedance. Thus, the mechanical model does not provide a constant load for the isolated heart, because of variations in both the model and coronary impedances.     

Evaluation of a lumped parameter model for isolated working rat hearts

When a mechanical model of the rat aortic input impedance is perfused with a pulsatile pump, the computed values of the impedance components vary linearly with flow rate and are interactive. When the model is perfused by an isolated rat heart, the total load upon the left ventricle consists of the model and coronary impedances in parallel. Adenosine triphosphate induces changes in coronary impedance, and the redistribution of cardiac output from the model to the coronary circulation causes flow-related changes in the model impedance. Thus, the mechanical model does not provide a constant load for the isolated heart, because of variations in both the model and coronary impedances.     

Functional and biochemical properties of rat Kupffer cells and peritoneal macrophages

Functional and biochemical techniques were used to further characterize heterogeneity between rat Kupffer cells and peritoneal macrophages. Both macrophage cell types were found to phagocytize antibody coated sheep red blood cells in a time-dependent manner. However, Kupffer cells were two to three times more phagocytic than were peritoneal macrophages. In contrast, the peritoneal cells released significantly more superoxide anion in response to the complement cleavage product, C5a and the phorbol ester tumor promoter, 12-0-tetradecanoyl-phorbol-13-acetate, and produced more hydrogen peroxide than did the liver macrophages. Both cell types responded chemotactically to C5a. These results suggest that macrophages may develop specialized functions depending on the needs of their local environment. Using one and two dimensional SDS-polyacrylamide gel electrophoresis, we also compared the production of newly synthesized proteins by Kupffer cells and peritoneal macrophages. In general, the macrophages were found to produce similar types and numbers of proteins with some exceptions. These included proteins that were unique to peritoneal macrophages and other proteins observed only in Kupffer cells. The production of these proteins in liver macrophages did not appear to correlate with levels of functional activation, but may be more related to the tissue origin of the cells.     

Expression and regulation of connexins in cultured ventricular myocytes isolated from adult rat hearts

Gap junctions were assayed during re-differentiation of adult rat cardiomyocytes in long-term culture to gain insight into the processes of remodeling. Double immunostaining allowed the localization of connexins Cx40, Cx43, and Cx45 between myocytes and demonstrated co-expression and co-localization in individual cells and gap junction plaques, respectively. Immunoblots showed differential time-dependent changes in connexin expression and phosphorylation. The total amount of connexins and the ratio of phosphorylated/non-phosphorylated isoforms gradually increased during the re-establishment of intercellular communication. Dual voltage-clamp studies showed the involvement of several types of gap junction channels. Multichannel currents yielded diverse spectra of g(j,inst)=f( V(j)) and g(j,ss)=f( V(j)) relationships ( g(j,inst): instantaneous gap junction conductance; g(j,ss): conductance at steady state; V(j): transjunctional voltage), indicative of homotypic and heterotypic channels. Single-channel currents revealed two prominent conductances reflecting gamma(j,main) and gamma(j,residual). The histograms of gamma(j,main) showed four discrete peaks (41-44, 59-61, 70-76, and 100-107 pS) attributable to a combination of Cx45-Cx45, Cx40-Cx45 and Cx43-Cx45 channels (1st peak), Cx43-Cx43 and Cx40-Cx43 channels (2nd peak), Cx43-Cx43 channels (3rd peak) and Cx40-Cx40 and Cx40-Cx43 channels (4th peak). However, the presence of heteromeric channels cannot be excluded. The data are consistent with an up-regulation of Cx45 and Cx43 during re-differentiation.     

A microcomputer system for haemodynamic measurements in isolated, working rat hearts

This study describes the application of an Apple IIe microcomputer in combination with a pre-processor in the on-line calculation of haemodynamic variables of the isolated working rat heart and of relative rapid changes in these variables, induced by variations in left atrial filling pressure (preload) and end-diastolic aortic pressure (afterload). Variables such as heart rate, systolic and diastolic left ventricular pressure, the maximal positive and negative first derivative of the left ventricular pressure, systolic and diastolic aortic pressure and aortic flow were continuously calculated and printed at minimal intervals of 6 s. A newly designed procedure to detect the activation of the electrogram, which was necessary to start the detection of a new cardiac cycle, is described.     

Erythropoietin changes contractility, cAMP, and nitrite levels of isolated rat hearts

There is enough evidence that erythropoietin (EPO) may be involved in cardiovascular function. Therefore we have investigated the possible effects of EPO on left ventricular developed pressure, +dP/dt(max), heart rate, tissue cAMP, and nitrite levels. Isolated rat hearts were perfused under constant flow (10 ml/min) conditions with modified Krebs-Henseleit solution and recombinant human erythropoietin at doses of 100, 200, 500, and 1,000 IU/kg was administered as bolus injections. EPO at 100 IU/kg decreased, but higher doses (500 and 1,000 IU/kg) raised the developed pressure and +dP/dt(max). However, it did not affect heart rate or coronary perfusion pressure when all the respective doses were applied. EPO at 100 IU/kg increased nitrite, and at 1,000 IU/kg it raised cAMP. Our results suggest that EPO may produce dose-dependently negative and positive inotropic effects on myocardial contractility in isolated rat hearts. NO and cAMP may be involved in negative and positive inotropic effects of EPO, respectively.     

川芎嗪对缺血再灌注所致大鼠离体心脏损伤的保护作用

目的:观察川芎嗪对缺血再灌注所致大鼠离体心脏损伤的保护作用。方法:心脏缺血再灌注模型:大鼠离心脏Langen-dorff灌流,全心停灌20分钟后复灌40分钟造成缺血--再灌注损伤。将充水乳胶球囊通过左心房插入左心室,Medlab-u/81生物信号采集处理系统记录分析心功能指标LVP,±dp/dtmax,HR和CF,收集再灌5分钟时,冠脉流出液。用分光光度法测定肌酸激酶(CK)活性。结果:缺血再灌注能降低冠脉流量,LVP,±dp/dtmax值,减慢心率、增加CK释放量,两个浓度的川芎嗪(40与80mg·L-1)对离体大鼠心脏缺血再灌注损伤有保护作用,表现为改善心功能,增加冠脉流量,减少心肌细胞内酶CK的释放,川芎嗪与维拉帕米有相似的心肌保护作用。结论:川芎嗪对心脏缺血再灌注心功能损伤具有保护作用。