Effects of selective TNFR1 inhibition or TNFR2 stimulation, compared to non-selective TNF inhibition, on (neuro)inflammation and behavior after myocardial infarction in male mice.

BACKGROUND: Myocardial infarction (MI) coinciding with depression worsens prognosis. Although Tumor Necrosis Factor alpha (TNF) is recognized to play a role in both conditions, the therapeutic potential of TNF inhibition is disappointing. TNF activates two receptors, TNFR1 and TNFR2, associated with opposite effects. Therefore, anti-inflammatory treatment with specific TNF receptor interference was compared to non-specific TNF inhibition regarding effects on heart, (neuro)inflammation, brain and behavior in mice with MI. METHODS: Male C57BL/6 mice were subjected to MI or sham surgery. One hour later, MI mice were randomized to either non-specific TNF inhibition by Enbrel, specific TNFR1 antagonist-, or specific TNFR2 agonist treatment until the end of the protocol. Control sham and MI mice received saline. Behavioral evaluation was obtained day 10-14 after surgery. Eighteen days post-surgery, cardiac function was measured and mice were sacrificed. Blood and tissue samples were collected for analyses of (neuro)inflammation. RESULTS: MI mice displayed left ventricular dysfunction, without heart failure, (neuro) inflammation or depressive-like behavior. Both receptor-specific interventions, but not Enbrel, doubled early post-MI mortality. TNFR2 agonist treatment improved left ventricular function and caused hyper-ramification of microglia, with no effect on depressive-like behavior. In contrast, TNFR1 antagonist treatment was associated with enhanced (neuro)inflammation: more plasma eosinophils and monocytes; increased plasma Lcn2 and hippocampal microglia and astrocyte activation. Moreover, increased baseline heart rate, with reduced beta-adrenergic responsiveness indicated sympathetic activation, and coincided with reduced exploratory behavior in the open field. Enbrel did not affect neuroinflammation nor behavior. CONCLUSION: Early receptor interventions, but not non-specific TNF inhibition, increased mortality. Apart from this undesired effect, the general beneficial profile after TNFR2 stimulation, rather than the unfavourable effects of TNFR1 inhibition, would render TNFR2 stimulation preferable over non-specific TNF inhibition in MI with comorbid depression. However, follow-up studies regarding optimal timing and dosing are needed.

The role of neutrophil gelatinase associated lipocalin (NGAL) as biological constituent linking depression and cardiovascular disease.

Depression is more common in patients with cardiovascular disease than in the general population. Conversely, depression is a risk factor for developing cardiovascular disease. Comorbidity of these two pathologies worsens prognosis. Several mechanisms have been indicated in the link between cardiovascular disease and depression, including inflammation. Systemic inflammation can have long-lasting effects on the central nervous system, which could be associated with depression. NGAL is an inflammatory marker and elevated plasma levels are associated with both cardiovascular disease and depression. While patients with depression show elevated NGAL levels, in patients with comorbid heart failure, NGAL levels are significantly higher and associated with depression scores. Systemic inflammation evokes NGAL expression in the brain. This is considered a proinflammatory effect as it is involved in microglia activation and reactive astrocytosis. Animal studies support a direct link between NGAL and depression/anxiety associated behavior. In this review we focus on the role of NGAL in linking depression and cardiovascular disease.

J147 affects cognition and anxiety after surgery in Zucker rats.

Vulnerable patients are at risk for neuroinflammation-mediated post-operative complications, including depression (POD) and cognitive dysfunction (POCD). Zucker rats, expressing multiple risk factors for post-operative complications in humans, may provide a clinically relevant model to study pathophysiology and explore potential interventions. J147, a newly developed anti-dementia drug, was shown to prevent POCD in young healthy rats, and improved early post-surgical recovery in Zucker rats. Aim of the present study was to investigate POCD and the therapeutic potential of J147 in male Zucker rats. Risk factors in the Zucker rat strain were evaluated by comparison with lean littermates. Zucker rats were subjected to major abdominal surgery. Acute J147 treatment was provided by a single iv injection (10 mg/kg) at the start of surgery, while chronic J147 treatment was provided in the food (aimed at 30 mg/kg/day), starting one week before surgery and up to end of protocol. Effects on behavior were assessed, and plasma, urine and brain tissue were collected and processed for immunohistochemistry and molecular analyses. Indeed, Zucker rats displayed increased risk factors for POCD, including obesity, high plasma triglycerides, low grade systemic inflammation, impaired spatial learning and decreased neurogenesis. Surgery in Zucker rats reduced exploration and increased anxiety in the Open Field test, impaired short-term spatial memory, induced a shift in circadian rhythm and increased plasma neutrophil gelatinase-associated lipocalin (NGAL), microglia activity in the CA1 and blood brain barrier leakage. Chronic, but not acute J147 treatment reduced anxiety in the Open Field test and protected against the spatial memory decline. Moreover, chronic J147 increased glucose sensitivity. Acute J147 treatment improved long-term spatial memory and reversed the circadian rhythm shift. No anti-inflammatory effects were seen for J147. Although Zucker rats displayed risk factors, surgery did not induce extensive POCD. However, increased anxiety may indicate POD. Treatment with J147 showed positive effects on behavioral and metabolic parameters, but did not affect (neuro)inflammation. The mixed effect of acute and chronic treatment may suggest a combination for optimal treatment.

Increased cardiovascular risk in rats with primary renal dysfunction; mediating role for vascular endothelial function.

Primary chronic kidney disease is associated with high cardiovascular risk. However, the exact mechanisms behind this cardiorenal interaction remain unclear. We investigated the interaction between heart and kidneys in novel animal model for cardiorenal interaction. Normal Wistar rats and Munich Wistar Fromter rats, spontaneously developing renal dysfunction, were subjected to experimental myocardial infarction to induce cardiac dysfunction (CD) and combined cardiorenal dysfunction (CRD), respectively (N = 5-10). Twelve weeks later, cardiac- and renal parameters were evaluated. Cardiac, but not renal dysfunction was exaggerated in CRD. Accelerated cardiac dysfunction in CRD was indicated by decreased cardiac output (CD 109 ± 10 vs. CRD 79 ± 8 ml/min), diastolic dysfunction (E/e') (CD 26 ± 2 vs. CRD 50 ± 5) and left ventricular overload (LVEDP CD 10.8 ± 2.8 vs. CRD 21.6 ± 1.7 mmHg). Congestion in CRD was confirmed by increased lung and atrial weights, as well as exaggerated right ventricular hypertrophy. Absence of accelerated renal dysfunction, measured by increased proteinuria, was supported by absence of additional focal glomerulosclerosis or further decline of renal blood flow in CRD. Only advanced peripheral endothelial dysfunction, as found in CRD, appeared to correlate with both renal and cardiac dysfunction parameters. Thus, proteinuric rats with myocardial infarction showed accelerated cardiac but not renal dysfunction. As parameters mimic the cardiorenal syndrome, these rats may provide a clinically relevant model to study increased cardiovascular risk due to renal dysfunction. Peripheral endothelial dysfunction was the only parameter that correlated with both renal and cardiac dysfunction, which may indicate a mediating role in cardiorenal interaction.

Tiotropium inhibits pulmonary inflammation and remodelling in a guinea pig model of COPD.

Airway remodelling and emphysema are major structural abnormalities in chronic obstructive pulmonary disease (COPD). In addition, pulmonary vascular remodelling may occur and contribute to pulmonary hypertension, a comorbidity of COPD. Increased cholinergic activity in COPD contributes to airflow limitation and, possibly, to inflammation and airway remodelling. This study aimed to investigate the role of acetylcholine in pulmonary inflammation and remodelling using an animal model of COPD. To this aim, guinea pigs were instilled intranasally with lipopolysaccharide (LPS) twice weekly for 12 weeks and were treated, by inhalation, with the long-acting muscarinic receptor antagonist tiotropium. Repeated LPS exposure induced airway and parenchymal neutrophilia, and increased goblet cell numbers, lung hydroxyproline content, airway wall collagen and airspace size. Furthermore, LPS increased the number of muscularised microvessels in the adventitia of cartilaginous airways. Tiotropium abrogated the LPS-induced increase in neutrophils, goblet cells, collagen deposition and muscularised microvessels, but had no effect on emphysema. In conclusion, tiotropium inhibits remodelling of the airways as well as pulmonary inflammation in a guinea pig model of COPD, suggesting that endogenous acetylcholine plays a major role in the pathogenesis of this disease.

Renal failure induces telomere shortening in the rat heart.

BACKGROUND: Renal failure aggravates pathological cardiac remodelling induced by myocardial infarction (MI). Cardiac remodelling is associated with telomere shortening, a marker for biological ageing. We investigated whether mild and severe renal failure shorten cardiac telomeres and excessively shorten telomeres after MI. METHODS: Rats were subjected to sham, unilateral (UNX) or 5/6th nephrectomy (5/6NX) to induce none, mild or severe renal failure. MI was induced by left coronary artery ligation. Renal function parameters and blood pressure were measured. DNA was isolated from non-infarcted cardiac tissue. Telomere length was assessed by quantitative polymerase chain reaction (PCR). RESULTS: Proteinuria was unchanged in UNX and MI compared with control, but strongly increased in 5/6NX, UNX+MI and 5/6NX+MI. Serum creatinine levels were increased fourfold in 5/6NX and tenfold in 5/6NX+MI. 5/6NX and groups with both renal failure and MI showed an approximate 20% reduction of telomere length, similar to the MI group. No excess telomere shortening was observed in hearts from rats with renal ablation after MI. CONCLUSION: Severe renal failure, but not mild renal failure, leads to shortening of cardiac telomeres to a similar extent as found after MI. Renal failure did not induce excessive telomere shortening after MI. (Neth Heart J 2009;17:190-4.).

Effects of erythropoietin on advanced pulmonary vascular remodelling.

Erythropoietin (EPO) mobilises endothelial progenitor cells and promotes neovascularisation in heart failure. The present authors studied the effects of EPO on pulmonary vascular and cardiac remodelling in a model for flow-associated pulmonary arterial hypertension (PAH). PAH was induced in adult male Wistar rats by the injection of monocrotaline combined with an abdominal aortocaval shunt 1 week later (PAH or experimental group). Immediately afterwards, rats were randomised into those who received treatment with EPO (PAH+EPO group) and controls. Pulmonary and systemic haemodynamics, and right ventricular and pulmonary vascular remodelling were evaluated 3 weeks later. Vascular occlusion of the intra-acinar pulmonary vessels (13.4+/-0.7 versus 16.7+/-1.3% in PAH+EPO and PAH, respectively) and medial wall thickness of the pre-acinar arteries (wall-to-lumen ratio 0.13+/-0.01 versus 0.17+/-0.01 in PAH+EPO and PAH, respectively) decreased after treatment with EPO. Moreover, right ventricular capillary density was increased by therapy (2,322+/-61 versus 2,100+/-63 capillaries x mm(-2) in PAH+EPO and PAH, respectively). Increased mean pulmonary arterial pressure and decreased right ventricular contractility in the model were not altered by EPO treatment. In this rat model of flow-associated pulmonary arterial hypertension, erythropoietin treatment beneficially affected pulmonary vascular and cardiac remodelling. These histopathological effects were not accompanied by significantly improved haemodynamics.

Regenerative cell therapy and pharmacotherapeutic intervention in heart failure: Part 2: Pharmacological targets, agents and intervention perspectives.

Regenerative medicine represents a promising perspective on therapeutic angiogenesis in patients with cardiovascular disease, including heart failure. However, previous or ongoing clinical trials show ambiguous outcomes with respect to the benefit of regenerative therapy by means of bone marrow stem cell infusion in myocardial infarction patients. Therefore, it is necessary to set up a rational therapeutic strategy in the treatment of congestive heart failure. Chemokines, cytokines and growth factors, as well as pharmaceutical agents, may have an impact on endothelial progenitor cell (EPC) physiology and thus can provide targets for pharmacological intervention. Indeed, EPCs and stem cell niches both in bone marrow and myocardial tissue can be treated as an integral target for recruitment of EPCs from the bone marrow to the cardiac ischaemic niche. In this article, we individually place the signalling factors in their specified context, and explain their roles in the various phases of neovascularisation (see Part 1). (Neth Heart J 2008;16:337-43.).

Regenerative cell therapy and pharmacotherapeutic intervention in heart failure: Part 1: Cardiovascular progenitor cells, their functions and sources.

It has been postulated that bone marrow derived endothelial progenitor cells (BM-EPCs) are essential for neovascularisation and endothelial repair and are involved in pharmacological treatment, and even its potential targets. There is no doubt that the ultimate success of angiogenic cell therapy will be determined by an appropriate stimulation of certain angiogenic progenitor cell subpopulations. Unfortunately, the biology of EPCs is still poorly understood. In particular, the understanding of endogenous microenvironments within the progenitor cell niches is critical, and will provide us with information about the signalling systems that supply a basis to develop rational pharmacotherapy to enhance the functional activity of endogenous or transplanted progenitor cells. The final success of clinical improvement of progenitor cell-mediated vascular repair and angiogenic therapy depends on a better understanding of EPC biology and a smart therapeutic design. In the first part of this review we first briefly discuss the possible involvement of progenitor cells in chronic heart failure. In part 2 we focus on factors that beneficially affect BMEPCs, with an emphasis on pharmacological molecular pathways involved in BM-EPC-induced neovascularisation. (Neth Heart J 2008;16:305-9.).

Low-dose aspirin improves in vivo hemodynamics in conscious, chronically infarcted rats.

OBJECTIVE: The equivalent of the clinically used low (= antiplatelet)-dose aspirin, inhibited collagen deposition in the non-infarcted myocardium in rats with myocardial infarction. In the present study, the in vivo hemodynamic consequences of this daily low-dose aspirin were investigated in conscious, chronically instrumented, infarcted rats. METHODS: Rats, treated with 25 mg/kg aspirin daily from 2 days before to 3 weeks after coronary artery ligation, were chronically instrumented with an electromagnetic flow-probe and arterial and venous catheters, to record cardiac output, and arterial and venous blood pressure, respectively, in the conscious freely moving animal. In parallel, isolated hearts were studied with regard to left ventricular stiffness (pressure/volume relationships), maximal cardiac perfusion (adenosine), and in vitro heart rate and beta-adrenergic responsiveness. Plasma catecholamine levels were measured. RESULTS: Aspirin normalized the increased heart rate after infarction, at a preserved cardiac output. This was accompanied by a (non-significant) increase in stroke volume, at unchanged cardiac loading conditions. The lower heart rate after aspirin was due to reduced intrinsic heart rate rather than to lower sympathetic activation of the heart, since similar effects were observed in isolated perfused hearts, while circulating levels of catecholamines and beta-adrenergic responsiveness were not influenced. The improved stroke volume was not explained by reduced left ventricular stiffness or increased maximal perfusion after aspirin. CONCLUSION: In addition to the antithrombotic action, effects of low-dose aspirin on cardiac remodeling could be associated with favorable hemodynamic effects, as reflected by a lower heart rate for the same cardiac output. Although the underlying mechanisms are still unknown, it suggests a clinically relevant beneficial effect which deserves further investigation.

Altered cardiac collagen and associated changes in diastolic function of infarcted rat hearts.

OBJECTIVE: Anti-inflammatory drugs have been shown to modulate collagen deposition during myocardial infarction (MI) induced remodeling. Chronic effects of methylprednisolone (5 mg/kg/day) and low-dose aspirin (25 mg/kg/day) on cardiac collagen and left ventricular diastolic function were studied in rat hearts, 21 days after MI. METHODS: Left ventricular function was assessed at baseline and after beta-adrenergic stimulation with isoproterenol in isolated perfused hearts, using an intraventricular balloon. After diastolic arrest, left ventricular pressure-volume curves were obtained. Left ventricular dilation was defined as the corresponding left ventricular volume at 20 mmHg left ventricular diastolic pressure. In histological sections, perivascular and interstitial collagen content were quantified morphometrically as the Sirius Red positive area in the non-infarcted interventricular septum. RESULTS: Impaired baseline left ventricular function of MI-hearts was improved by methylprednisolone but not by low-dose aspirin. Isoprotenerol significantly enhanced systolic function in all hearts, whereas it augmented the decrease in left ventricular diastolic pressure only in methylprednisolone-treated MI-hearts. The rightward shift of the pressure-volume curve after MI was aggravated by methylprednisolone but not with low-dose aspirin treatment. Low-dose aspirin reduced perivascular but not interstitial collagen whereas methylprednisolone decreased both perivascular and interstitial collagen. CONCLUSIONS: Our findings indicate that MI-induced collagen deposition in the spared myocardium can be affected by chronic therapy with low-dose aspirin or methylprednisolone. The effects on interstitial collagen seemed reflected in an altered left ventricular diastolic function.

Determinants of coronary reserve in rats subjected to coronary artery ligation or aortic banding.

OBJECTIVE: We investigated whether decreased coronary reserve in hearts after coronary artery ligation or in hearts from rats after aortic banding can be related to remodeling of resistance arteries. METHODS: Maximal coronary flow (absolute flow) and cardiac perfusion (flow corrected for heart weight) were determined in isolated, perfused rat hearts after adenosine or nitroprusside, at 3 and 8 weeks after coronary artery ligation or 4-5 weeks after aortic banding. Perivascular collagen and medial thickness of resistance arteries were determined by morphometry. RESULTS: maximal coronary flow of infarcted hearts had been restored to sham values at 3 weeks. Growth of cardiac muscle mass from 3 to 8 weeks exceeded the increase in maximal coronary flow, leading to a decreased perfusion at 8 weeks. A slight, transient increase in perivascular collagen, but no medial hypertrophy, was found after infarction. After aortic banding perivascular fibrosis and medial hypertrophy led to a decreased maximal coronary flow in both the hypertrophied left and the non-hypertrophied right ventricle. Consequently, perfusion of the left ventricle was most severely reduced. CONCLUSIONS: Reduced maximal perfusion after aortic banding is determined by both cardiac hypertrophy and vascular remodeling. In contrast, during infarction-induced remodeling, reduction of perfusion is not determined by vascular remodeling, but mainly by disproportional cardiac hypertrophy relative to vascular growth.

Hypothermia extends the cardioprotection by ischaemic preconditioning to coronary artery occlusions of longer duration.

OBJECTIVE: To test the hypothesis that mild hypothermia potentiates the cardioprotection afforded by ischaemic preconditioning so that infarct size limitation can be obtained after coronary artery occlusion (CAO) durations which exceed the cardioprotective range (> 90 min) of either hypothermia or ischaemic preconditioning alone. METHODS: Four groups of anaesthetized rats were subjected to different durations of CAO: (i) normothermia (N, 36.5-37.5 degrees C, n = 29), (ii) normothermia + ischaemic preconditioning (N + IP, 15 min CAO followed by 10 min of reperfusion, n = 35), (iii) hypothermia (H, 30-31 degrees C, n = 31) and (iv) hypothermia + ischaemic preconditioning (H + IP, n = 24). Infarct size (IA/AR) was determined after 3 hours of reperfusion using trypan blue to delineate the area at risk (AR) from non-risk region and nitroblue tetrazolium to delineate infarcted area (IA) from viable myocardium. RESULTS: In N the CAO duration versus infarct size relation had a sigmoid shape with virtually no infarction occurring at 15 min CAO and 56 +/- 5% of the area at risk being infarcted at 30 min CAO reaching a plateau of 71 +/- 2% at 60 min CAO. Hypothermia produced a rightward shift of the relation resulting in an approximately 15 min delay in onset of infarction. Ischaemic preconditioning produced a similar reduction in infarct size (23 +/- 4%) at 30 min CAO compared to hypothermia (13 +/- 3%) but also limited infarct size at 45 min to 36 +/- 3% and at 60 min CAO to 50 +/- 3% suggesting a slowing of infarct progression. Neither intervention limited IA/AR produced by 120 min CAO. In H + IP, combined hypothermia and ischaemic preconditioning resulted in synergistic infarct size reduction so that at 45 min and 60 min CAO IA/AR was reduced to 17 +/- 3% and 23 +/- 3%, respectively, and even at 120 min CAO to 58 +/- 5%, which was significantly smaller than during normothermic control conditions (p < 0.05 vs. N). CONCLUSION: Mild hypothermia limited IA/AR modestly but markedly enhanced the cardioprotection afforded by ischaemic preconditioning in the in situ rat heart so that irreversible damage produced by even prolonged coronary artery occlusions was limited.

Renin-angiotensin system components in the interstitial fluid of the isolated perfused rat heart. Local production of angiotensin I.

We used a modification of the isolated perfused rat heart, in which coronary effluent and interstitial transudate were separately collected, to investigate the uptake and clearance of exogenous renin, angiotensinogen, and angiotensin I (Ang I) as well as the cardiac production of Ang I. The levels of these compounds in interstitial transudate were considered to be representative of the levels in the cardiac interstitial fluid. During perfusion with renin or angiotensinogen, the steady-state levels (mean +/- SD) in interstitial transudate were 64 +/- 34% (P < .05 for difference from the arterial level, n = 8) and 108 +/- 42% (n = 6) of the arterial level, respectively; the levels in coronary effluent were not significantly different from those in interstitial transudate. Ang I was not detectable in interstitial transudate during perfusion with Tyrode's buffer or angiotensinogen. It was very low in interstitial transudate during perfusion with renin and rose to much higher levels during combined renin and angiotensinogen perfusion. The total production rate of Ang I present in interstitial fluid could be largely explained by the renin-angiotensinogen reaction in the fluid phase of the interstitial compartment. In contrast, the total production rate of Ang I present in coronary effluent and the net ejection rate of Ang I via coronary effluent were, respectively, 4.6 +/- 2.2 and 2.8 +/- 1.3 (P < .01 and P < .05 for difference from 1.0, n = 6) times higher than could be explained by Ang I formation in the fluid phase of the intravascular compartment. Ang I from the interstitial fluid contributed little to the Ang I in the intravascular fluid and vice versa. These data reveal two tissue sites of Ang I production, ie, the interstitial fluid and a site closer to the blood compartment, possibly vascular surface-bound renin. There was no evidence that the release of locally produced Ang I into coronary effluent and interstitial transudate occurred independently of blood-derived renin or angiotensinogen.

Organ blood flow protection in hypertension and congestive heart failure.

The nutrients to the different organs of the body are provided through the blood supply to these organs. Since the nutrient requirements change considerably and abruptly according to the demands set by, for example, physical activity, organ blood flow has to be adjusted accordingly. This is achieved by a complex interplay between neural (parasympathetic and sympathetic nerves), humoral (vasopressin, angiotensin, etc.), and local (ions, pH, adenosine, etc.) factors. It is also well recognized that diseases, such as hypertension and heart failure, as well as the drugs to treat them, can substantially affect organ blood flow. Using tracer microspheres, several studies in animals have shown that, during the established phase of hypertension, there is some decrease in cardiac output, which is then redistributed, with the brain, small intestines, and heart receiving a higher proportion. Blood flow to and vascular conductance in the other organs, particularly the kidneys, is decreased. Similar findings have also been observed in human hypertension. During congestive heart failure, most organs show a decrease in blood flow and there is a conspicuous increase in the sympathetic nerve activity. The drug responses may differ in the disease state and, from a clinical viewpoint, changes in some circulatory beds (e.g., cerebral, cardiac, renal, skeletal muscles) as well as the partition of blood flow into nutrient (tissue) and non-nutrient (arteriovenous anastomoses or shunts) parts can be of great importance. For example, overzealous antihypertensive therapy or the use of cerebral vasodilators may adversely affect cerebral circulation in certain situations and increased arterial blood flow may only be in the arteriovenous anastomotic part of the circulation.

Cardiovascular effects of social stress in borderline hypertensive rats.

OBJECTIVE: To test the hypothesis that chronic exposure to psychosocial stress, alone or in combination with elevated levels of dietary salt, leads to hypertension and cardiac pathology in a susceptible strain of rats. DESIGN AND METHODS: In four experiments, borderline hypertensive rats, maintained on normal or high-salt diets, were exposed to 14-16 weeks aggregation in a colony housing or in larger breeder cages. Pulsatile blood pressure was measured once a week in unrestrained male rats by pressure telemetry. Direct carotid pressures of the aggregated rats and of control rats were measured before they were killed; at necropsy cardiac and adrenal weights and ventricular design were determined. RESULTS: Despite continuous fighting, their weekly measured blood pressures remained stable; no differences in final carotid pressures between experimental and control rats were found. Rats from three aggregations showed significant increases in left and right ventricular and adrenal weights. CONCLUSION: No hypertension developed in any aggregation, although most of the rats showed signs of perceived stress (significantly reduced weight gain, enlarged adrenals and a large number of body wounds). Cardiac hypertrophy did ensue, possibly reflecting increased physical activity or intermittent increases in sympathetic activity, or both.

Bradykinin-induced release of nitric oxide by the isolated perfused rat heart: importance of preformed pools of nitric oxide-containing factors.

OBJECTIVE: To study whether the vasorelaxant effect of bradykinin in the coronary vascular bed depends on the release of NO from preformed pools and/or de-novo synthesis of NO resulting from bradykinin-induced stimulation of NO synthase. DESIGN AND METHODS: Rat hearts were perfused according to Langendorff's method. Coronary flow was measured continuously. We constructed concentration-response curves for bradykinin and L-arginine under control conditions, after downregulation of NO synthase by exposing the heart to high concentrations (10 mmol/l) of NO and during chronic inhibition of NO synthase, obtained by perfusing the heart for 30 min with 0.1 mmol/l N(omega)-nitro-L-arginine methyl ester. The effect of acute inhibition of NO synthase was studied by infusing single submaximal doses of bradykinin and of L-arginine in the absence and presence of 0.1 mmol/l N(omega)-nitro-L-arginine methyl ester. RESULTS: Coronary flow [baseline 9 +/- 2 ml/min (mean +/- SD)] increased to maximally 23 +/- 6 ml/min with bradykinin and to 16 +/- 4 ml/min with L-arginine. Maximal coronary flow, established as the maximal effect in response to NO, was 22 +/- 4 ml/min. Chronic inhibition of NO synthase reduced coronary flow to 4 +/- 1 ml/min. Coronary flow did not change after downregulation of NO synthase by NO. Neither downregulation nor acute inhibition of NO synthase affected the response to bradykinin, whereas chronic inhibition of NO synthase blocked the bradykinin-induced increase in coronary flow by > 90%. Administration of L-arginine no longer increased coronary flow under all tested conditions. CONCLUSIONS: Preformed pools of NO-containing factors exist within the isolated perfused heart and bradykinin exerts its vasorelaxant effects at least in part by the mobilization of these preformed pools. These data may reconcile previous discrepancies about the (lack of) effect of NO synthase inhibitors on bradykinin-induced coronary vasodilatation.

Cardioprotection by ischemic and nonischemic myocardial stress and ischemia in remote organs. Implications for the concept of ischemic preconditioning.

Ischemic preconditioning studies employ one or more brief total coronary artery occlusions separated by complete reperfusion to limit infarct size during a subsequent prolonged coronary artery occlusion. We now present evidence that in anesthetized pigs a partial coronary artery occlusion without intervening reperfusion between the partial and prolonged total occlusions can also precondition the myocardium provided that the reduction in coronary blood flow is sufficiently severe. Thus infarct size was reduced after a 60 min total coronary artery occlusion when the total occlusion was preceded by a partial coronary occlusion that reduced coronary blood flow by 70% but not when the flow reduction was only 30%. In this two-stage coronary occlusion model the degree of protection appears greater in the epicardial than in the endocardial half. In view of evidence that brief occlusions of a coronary artery also protect myocardium outside its perfusion territory, we subsequently investigated whether ischemia in remote organs can protect myocardium. Because of reports that development of infarct size may be temperature dependent, we also investigated whether the cardioprotection by remote organ ischemia was temperature dependent. In anesthetized rats a 15 min coronary artery occlusion was more effective in reducing infarct size produced by a subsequent 60 min total coronary artery occlusion when the experiments were performed at a body core temperature of 30-31 degrees C than at 36-37 degrees C, while infarct size of animals which were subjected to only the 60 min total coronary artery occlusion was the same for the two body core temperatures. In rats with a body core temperature of 36-37 degrees C a 15 min mesenteric artery occlusion, but not a 15 min renal artery occlusion, reduced infarct size produced by a subsequent 60 min coronary artery occlusion. When the experiments were performed at 30-31 degrees C both the mesenteric and renal artery occlusions were protective. These observations indicate the local myocardial ischemia is not required to protect the myocardium during a prolonged coronary occlusion. We further investigated whether myocardium could also be protected by a cardiac stimulus which does not produce ischemia at all. For this purpose we electrically paced the left ventricle of anesthetized pigs to produce heart rates of 200 bpm (which did not lead to ischemia as assessed by a number of functional and biochemical variables) and found that 30 min of ventricular pacing reduced myocardial infarct size produced by a subsequent 60 min coronary artery occlusion. The protection by ventricular pacing involved activation of K+ATP channels as pretreatment with glibenclamide abolished the protection by ventricular pacing. We conclude that a number of distinctly different stimuli can protect the myocardium suggesting that ischemic myocardial preconditioning could be just one feature of a more general protection phenomenon.

Sensitivity to ischemia of chronically infarcted rat hearts; effects of long-term captopril treatment.

Myocardial infarction induced hypertrophy of non-infarcted myocardium, in parallel with interstitial and perivascular fibrosis and a decreased capillary density, could increase sensitivity to ischemia. The structural cardiac changes can be reversed by long-term captopril treatment. In the present study, ischemic sensitivity in relation to cardiac perfusion was studied in isolated, perfused hearts of untreated and captopril-treated infarcted rats. In chronically (8 weeks) infarcted hearts, maximal vasodilation in response to administered adenosine and nitroprusside, as well as to endogenously released vasodilators during reperfusion, was decreased, suggesting impaired cardiac perfusion. Ischemic release of purines and lactate was reduced in infarcted hearts, indicating decreased sensitivity to ischemia of the remodeled myocardium. Captopril treatment (3-8 weeks post myocardial infarction), which reversed hypertrophy without affecting the flow capacity of the coronary vascular bed, restored maximal cardiac perfusion. Ischemic ATP breakdown was not affected by captopril, whereas lactate release was even further reduced, suggesting alterations towards a more aerobic ATP production. These data indicate that despite the reduced maximal cardiac perfusion, the remodeled myocardium of infarcted hearts is less sensitive to ischemia. Reversal of hypertrophy by chronic captopril restored maximal cardiac perfusion and led to a better preservation of aerobic ATP production during ischemia.

'Quality of life' after therapy in rats with myocardial infarction: dissociation between hemodynamic and behavioral improvement.

Rats with myocardial infarction provide a clinically relevant model for hemodynamic and survival studies. Moreover, behavioral changes in this model, i.e. increased anxiety and reduced interest in environment and social interactions, mimic aspects of the reduced quality of life of patients. In the present study, we investigated whether pharmacological treatment that is known to improve hemodynamics and prognosis could also affect the behavioral changes associated with quality of life. Rats with 3-week-old infarcts were treated with intermittent dobutamine (1 mg/kg i.p., twice daily) or captopril (2 milligrams in drinking water). After 2 weeks of treatment, when from previous studies hemodynamics are expected to be restored, behavioral tests were performed. In the free exploration test, which primarily evaluates exploratory behavior, dobutamine normalized the reduced interest in the environment. In the standard open field and social interaction tests, which also include an anxiety component, the beneficial effects of dobutamine were not observed. In contrast, captopril normalized all behavioral changes that indicated increased anxiety. In conclusion, the expected similar hemodynamic improvement with dobutamine and captopril treatment resulted in improvement of different aspects of the changed behavior of rats with myocardial infarction, indicating that there is no direct relationship between hemodynamics and quality of life. The behavioral tests used, in combination with our previously described functional hemodynamic measurements, could provide a new basis for evaluating the effects of therapy on hemodynamic function as well as the quality of life.