Effects of the cardioselective KATP channel blocker HMR 1098 on cardiac function in isolated perfused working rat hearts and in anesthetized rats during ischemia and reperfusion.

It has been argued that activation of KATP channels in the sarcolemmal membrane of heart muscle cells during ischemia provides an endogenous cardioprotective mechanism. In order to test whether the novel cardioselective KATP channel blocker HMR 1098 affects cardiac function during ischemia, experiments were performed in rat hearts during ischemia and reperfusion. Isolated perfused working rat hearts were subjected to 30 min of low-flow ischemia in which the coronary flow was reduced to 10% of its control value, followed by 30-min reperfusion. In the first set of experiments the hearts were electrically paced at 5 Hz throughout the entire protocol. At the end of the 30-min ischemic period the aortic flow had fallen to 44 +/- 2% (n=8) of its nonischemic value in vehicle-treated hearts, whereas in the presence of 0.3 micromol/l and 3 micromol/l HMR 1098 it had fallen to 29 +/- 7% (n=5, not significant) and 8 +/- 2% (n=12, P<0.05), respectively. Glibenclamide (3 micromol/l) reduced the aortic flow to 9.5 +/- 7% (n=4, P<0.05). In control hearts the QT interval in the electrocardiogram shortened from 63 +/- 6 ms to 36 +/- 4 ms (n=10, P<0.05) within 4-6 min of low-flow ischemia. This shortening was completely prevented by 3 micromol/l HMR 1098 (60 +/- 5 ms before ischemia, 67 +/- 6 ms during ischemia, n=9, not significant). When rat hearts were not paced, the heart rate fell spontaneously during ischemia, and HMR 1,098 (3 micromol/l) caused only a slight, statistically non-significant reduction in aortic flow during the ischemic period. In order to investigate whether HMR 1098 shows cardiodepressant effects in a more pathophysiological model, the left descending coronary artery was occluded for 30 min followed by reperfusion for 60 min in anesthetized rats. Treatment with HMR 1098 (10 mg/kg i.v.) had no statistically significant effects on mean arterial blood pressure and heart rate during the control, ischemia and reperfusion periods. At the end of the reperfusion period, aortic blood flow was slightly reduced by HMR 1098, without reaching statistical significance (two-way analysis of ANOVA, P=0.15). Myocardial infarct size as a percentage of area at risk was not affected by HMR 1098 (vehicle: 75 +/- 3%, HMR 1098: 72 +/- 2%, n=7 in each group). In conclusion, cardiodepressant effects of HMR 1098 were observed only in isolated perfused working rat hearts which were continuously paced during global low-flow ischemia. In the model of anesthetized rats subjected to regional ischemia, HMR 1098 had no significant effect on cardiac function or infarct size.

The broad bean nodulin VfENOD18 is a member of a novel family of plant proteins with homologies to the bacterial MJ0577 superfamily.

Full-length transcript sequences were isolated from broad bean root nodules, which encode a novel nodulin designated VfENOD18. The corresponding transcripts were detected in early and in late stages of nodule development and were localized exclusively in the nitrogen-fixing zone III. The VfENOD18 sequence is not only homologous to a number of ESTs from various mono- and dicotyledonous plants, but also to the ATP-binding protein MJ0577 from Methanococcus jannaschii and to a range of bacterial proteins that belong to the MJ0577 superfamily. Hence, VfENOD18 is a member of a ubiquitous family of plant proteins that might function as ATP-binding proteins or ATPases. On the genomic level, VfENOD18 genes can be divided into two groups on the basis of differences in their 5' UTRs. One group lacks the 5' UTR region including the ATG initiation codon, whereas the second group contained the complete 5' UTR region. Further upstream of this VfENOD18 gene, a retrotransposon sequence was identified. The -14/-964 VfENOD18 promoter fragment was devoid of complete organ-specific elements known from other nodulin gene promoters. Nevertheless, this region was able to mediate full promoter activity in the central region of transgenic Vicia hirsuta root nodules.

Inhibition of endogenous nitric oxide synthase potentiates ischemia-reperfusion-induced myocardial apoptosis via a caspase-3 dependent pathway.

OBJECTIVE: Apoptosis of cardiomyocytes may contribute to ischemia-reperfusion injury. The role of nitric oxide (NO) in apoptosis is controversial. Therefore, we investigated the effect of NO synthase inhibition on apoptosis of cardiomyocytes during ischemia and reperfusion and elucidated the underlying mechanisms. METHODS AND RESULTS: Isolated perfused rat hearts (n = 6/group) were subjected to ischemia (30 min) and reperfusion (30 min) in the presence or absence of the NO synthase inhibitor NG-mono-methyl-L-arginine. Reperfusion induced cardiomyocyte apoptosis as assessed by immunohistochemistry (TUNEL-staining) and the demonstration of the typical DNA laddering. Apoptosis during reperfusion was associated with the cleavage of caspase-3, the final down-stream executioner caspase, whereas the protein levels of the anti-apoptotic protein Bcl-2 and the pro-apoptotic protein Bax were unchanged. Inhibition of the NO synthase drastically increased ischemia and reperfusion-induced apoptosis of cardiomyocytes. Moreover, the NO synthase inhibitor enhanced the activation of caspase-3, suggesting that NO interferes with the activation of caspases in ischemia-reperfusion. CONCLUSION: The results of the present study demonstrate that inhibition of endogenous NO synthesis during ischemia and reperfusion leads to an enhanced induction of apoptosis, suggesting that the endogenous NO synthesis protects against apoptotic cell death. Inhibition of NO synthesis thereby activates the caspase cascade, whereas the Bcl-2/Bax protein levels remained unchanged.

Dose-dependent reduction of myocardial infarct mass in rabbits by the NHE-1 inhibitor cariporide (HOE 642).

The aim of this study was to investigate the dose-dependent effect of pretreatment with the selective sodium-hydrogen exchange NHE-subtype 1 inhibitor cariporide on myocardial infarct mass in a rabbit model of coronary ligation and reperfusion. Furthermore, in a second part of the study, we tested the effect of cariporide in the rabbits when given prior to reperfusion. Rabbits (n=49) were randomized in 7 groups: saline vehicle, cariporide: 0.01, 0.03, 0.1 and 0.3 mg/kg, and subjected to a 30 min occlusion of a branch of the left coronary artery followed by 2 h reperfusion. Cariporide was given as a bolus intravenously 10 min before occlusion or 5 min before reperfusion. After reperfusion, myocardial infarct mass was determined by triphenyl tetrazolium chloride staining and expressed as a percent of area at risk. Cariporide given intravenously 10 min before occlusion in doses of 0.01, 0.03, 0.1, 0.3 mg/kg, led to a dose-dependent reduction in infarct mass from 58+/-6% in controls to 48+/-4% (-17%, NS), 36+/-5% (-38%, p<0.05), 26+/-6% (-55%, p<0.05), 11+/-4% (-81%, p<0.05) respectively, whereas area at risk did not differ in between the groups. The effect of the lowest dose of 0.01 mg/kg did not reach significance. Plasma levels at different doses of cariporide were correlated to the respective infarct mass. After coronary occlusion left ventricular end-diastolic pressure (LVEDP) significantly increased throughout occlusion and reperfusion. Cariporide in the doses of 0.3, 0.1 and 0.03 mg/kg normalized LVEDP when measured after 2 h reperfusion. In controls hemodynamic parameters such as mean arterial blood pressure (MAP), heart rate (HR), left ventricular pressure (LVP) and LV dP/dt(max) were not significantly changed by ischemia/reperfusion with the exception of MAP, LVP and LV dP/dt(max) which were significantly decreased after 120 min reperfusion. Cariporide at doses of 0.1, 0.03 and 0.01 mg/kg did not significantly influence these parameters, whereas the highest dose of 0.3 mg/kg prevented the decrease of MAP and LVP. Cariporide (0.3 mg/kg i.v.) administered 5 min before reperfusion significantly reduced infarct mass by 31%. Under these conditions the increase of LVEDP after coronary occlusion was not influenced by cariporide. As in the pretreatment experiments, the decrease of MAP and LVP was prevented when measured 2 h after reperfusion. The results show that pretreatment with the NHE-subtype 1 inhibitor cariporide is cardioprotective by reducing infarct mass in rabbits in a dose-dependent manner. While the cardioprotective effect of pretreatment could be demonstrated over a broad range of doses, the efficacy of the compound when given only on reperfusion was significant but more limited.

The Vicia faba lipoxygenase gene VfLOX1 is expressed in the root nodule parenchyma.

A full-length cDNA encoding the broad bean lipoxygenase VfLOX1 was isolated from a nodule cDNA library. The VfLOX1 gene was strongly expressed in nodules, and only weakly in roots. VfLOX1 transcripts were localized in the nodule parenchyma and in the cells surrounding the root stele.

Effects of the Na+/H+-exchange inhibitor Hoe 642 on intracellular pH, calcium and sodium in isolated rat ventricular myocytes.

The inhibitors of the Na+/H+-exchange (NHE1) system Hoe 694 and Hoe 642 possess cardioprotective effects in ischaemia/reperfusion. It is assumed that these effects are due to the prevention of intracellular sodium (Nai) and calcium (Cai) overload. The purpose of the present study was to investigate the effects of Hoe 642 on intracellular pH, Na+ and Ca2+ (pHi, Nai and Cai) in isolated rat ventricular myocytes under anoxic conditions or in cells in which oxidative phosphorylation had been inhibited by 1.5 mmol/l cyanide. In cells which were dually loaded with the fluorescent dyes 2, 7-biscarboxyethyl-5,6-carboxyfluorescein (BCECF) and Fura-2, anoxia caused acidification of the cells (from pHi 7.2 to pHi 6.8) and an increase in Cai from about 50 nmol/l to about 1 micromol/l. The decrease in pHi began before the cells underwent hypoxic (rigor) contracture, whereas Cai only began to rise after rigor shortening had taken place. After reoxygenation, pHi returned to its control value and Cai oscillated and then declined to resting levels. It was during this phase that the cells rounded up (hypercontracture). When 10 micromol/l Hoe 642 was present from the beginning of the experiment, pHi and Cai were not significantly different from control experiments. At reoxygenation, pHi did not recover, but Cai oscillated and returned to its resting level. To monitor Nai, the cells were loaded with the dye SBFI. After adding 1.5 mmol/l cyanide or 100 micromol/l ouabain, Nai increased from the initial 8 mmol/l to approximately 16 mmol/l. Hoe 642 or Hoe 694 (10 micromol/l) did not prevent the increase in Nai. In contrast, the blocker of the persistent Na+ current R56865 (10 micromol/l) attenuated the CN--induced rise in Nai. The substance ethylisopropylamiloride was not used because it augmented considerably the intensity of the 380 nm wavelength of the cell's autofluorescence. In conclusion, the specific NHE1 inhibitor Hoe 642 did not attenuate anoxia-induced Cai overload, nor CN--induced Nai and Cai overload. Hoe 642 prevented the recovery of pHi from anoxic acidification. This low pHi maintained after reoxygenation may be cardioprotective. Other possible mechanisms of NHE1 inhibitors, such as prevention of Ca2+ overload in mitochondria, cannot be ruled out. The increase in Nai during anoxia is possibly due to an influx of Na+ via persistent Na+ channels.

Protective effects of HOE642, a selective sodium-hydrogen exchange subtype 1 inhibitor, on cardiac ischaemia and reperfusion.

OBJECTIVE: The aim was to characterise the new compound HOE642 as a selective and cardioprotective Na+/H+ exchange inhibitor in various models. METHODS: The effect of HOE642 was tested in the osmotically activated Na+/H+ exchange of rabbit erythrocytes and in propionate induced swelling of human thrombocytes. Recovery of pH after an NH4Cl prepulse and effects on other ion transport systems by patch clamp technique were investigated in rat cardiomyocytes. NHE subtype specifity of the compound was determined by 22Na+ uptake inhibition in a fibroblast cell line separately expressing subtype isoforms 1-3. Protective effects of HOE642 in cardiac ischaemia and reperfusion by ligation of coronary artery were investigated in isolated working rat hearts and in anaesthetised rats. RESULTS: HOE642 concentration dependently inhibited the amiloride sensitive sodium influx in rabbit erythrocytes, reduced the swelling of human platelets induced by intracellular acidification, and delayed pH recovery in rat cardiomyocytes. In the isolated working rat heart subjected to ischaemia and reperfusion HOE642 dose dependently reduced the incidence and the duration of reperfusion arrhythmias. It also reduced the the release of lactate dehydrogenase and creatine kinase, and preserved the tissue content of glycogen, ATP, and creatine phosphate. In anaesthetised rats undergoing coronary artery ligation intravenous and oral pretreatment with HOE642 caused a dose dependent reduction or a complete prevention of ventricular premature beats, ventricular tachycardia, and ventricular fibrillation. The compound was well tolerated and neutral to circulatory variables. Other cardiovascular agents tested in this model were not, or were only partly, effective at doses showing marked cardiodepressive effects. CONCLUSIONS: HOE642 is a very selective NHE subtype 1 inhibitor showing cardioprotective and antiarrhythmic effects in ischaemic and reperfused hearts. Further development of well tolerated compounds like HOE642 could lead to a new therapeutic approach in clinical indications related to cardiac ischaemia and reperfusion.

Na+/H+ exchange and its inhibition in cardiac ischemia and reperfusion.

The characterization of various ion transport systems has led to a better understanding of the effects, which seem to take part in the impairment of ischemic and reperfused cardiac tissue. This review discusses the role of the Na+/H+ exchange system in the pathophysiology of ischemia and reperfusion and the beneficial effects of its inhibition. At the onset of ischemia intracellular pH (pHi) decreases due to anaerobic metabolism and ATP hydrolysis, leading to an activation of Na+/H+ exchange. This in turn increases intracellular Na+ (Na+i) and activates Na+/K+ ATPase, with a consecutive increase of energy consumption. Since cellular Na+ and Ca++ transport are coupled by the Na+/Ca++ exchange system, which depends on the Na+ gradient, the high Na+i leads to increased intracellular Ca++ (Ca++i). After a certain period, Na+/H+ exchange is inactivated by a decrease of extracellular pH. In case of reperfusion the acid extracellular fluid is washed out, which reactivates Na+/H+ exchange, leading to an unfavourably fast restoration of pHi and a second time to Na+ and Ca++i overflow. High Ca++i is assumed to be one of the main reasons for ischemic and reperfusion injury, like arrhythmias, myocardial contracture, stunning and necrosis. It seems that the inhibition of Na+/H+ exchange can interrupt this process at an early phase and prevent or delay the consequences of ischemia and reperfusion as demonstrated by numerous investigators.

Hoe 694, a new Na+/H+ exchange inhibitor and its effects in cardiac ischaemia.

1. The benzoylguanidine derivative Hoe 694 ((3-methylsulphonyl-4- piperidino-benzoyl) guanidine methanesulphonate) was characterized as an inhibitor of Na+/H+ exchange in rabbit erythrocytes, rat platelets and bovine endothelial cells. The potency of the compound was slightly lower or comparable to ethylisopropyl amiloride (EIPA). 2. To investigate a possible cardioprotective role of the Na+/H+ exchange inhibitor Hoe 694, rat isolated working hearts were subjected to ischaemia and reperfusion. In these experiments all untreated hearts suffered ventricular fibrillation on reperfusion. Addition of 10(-7) M Hoe 694 to the perfusate almost abolished reperfusion arrhythmias in the rat isolated working hearts. 3. Hoe 694 reduced the release of lactate dehydrogenase (LDH) and creatine kinase (CK), which are indicators of cellular damage during ischaemia, into the venous effluent of the hearts by 60% and 54%, respectively. 4. The tissue content of glycogen at the end of the experiments was increased by 60% and the high energy phosphates ATP and creatine phosphate were increased by 240% and 270% respectively in the treated hearts as compared to control hearts. 5. Antiischaemic effects of the Na+/H+ exchange inhibitor, Hoe 694, were investigated in a second experiment in anaesthetized rats undergoing coronary artery ligation. In these animals, pretreatment with Hoe 694 caused a dose-dependent reduction of ventricular premature beats and ventricular tachycardia as well as a complete suppression of ventricular fibrillation down to doses of 0.1 mg kg-1, i.v. Blood pressure and heart rate remained unchanged. 6. We conclude that the new Na+/H+ exchange inhibitor, Hoe 694, shows cardioprotective and antiarrhythmic effects in ischaemia and reperfusion in rat isolated hearts and in anaesthetized rats. In view of the role which Na+/H+ exchange seems to play in the pathophysiology of cardiac ischaemia these effects could probably be attributed to Na+/H+ exchange inhibition.

Ramipril prevents left ventricular hypertrophy with myocardial fibrosis without blood pressure reduction: a one year study in rats.

1. Angiotensin converting enzyme (ACE)-inhibitors have been demonstrated to be effective in the treatment of cardiac hypertrophy when used in antihypertensive doses. The aim of our one year study with an ACE-inhibitor in rats was to separate local cardiac effects produced by a non-antihypertensive dose from those on systemic blood pressure when an antihypertensive dose was used. 2. Rats made hypertensive by aortic banding were subjected to chronic oral treatment for one year with an antihypertensive dose of the ACE inhibitor, ramipril 1 mg kg-1 daily, (RA 1 mg) or received a low dose of 10 micrograms kg-1 daily (RA 10 micrograms) which did not affect high blood pressure. 3. Chronic treatment with the ACE-inhibitor prevented left ventricular hypertrophy in the antihypertensive rats as did the low dose which had no effects on blood pressure. Similar effects were observed on myocardial fibrosis. Plasma ACE activity was inhibited in the RA 1 mg but not in the RA 10 micrograms group although conversion of angiotensin (Ang) I to Ang II in isolated aortic strips was suppressed in both treated groups. Plasma catecholamines were increased in the untreated control group, but treatment with either dose of ramipril normalized the values. The myocardial phosphocreatine to ATP ratio (an indicator of the energy state in the heart) was reduced in the vehicle control group whereas the hearts from treated animals showed a normal ratio comparable to hearts from sham-operated animals. 4. After one year, five animals were separated from each group, treatment withdrawn, and housed for additional six months. In the RA 1 mg group, blood pressure did not reach the value of the control vehicle group and surprisingly, left ventricular hypertrophy and myocardial fibrosis did not recur in animals during withdrawal of treatment.5. These data show that long term ACE inhibitor treatment with ramipril in antihypertensive and non-antihypertensive doses prevented cardiac hypertrophy and myocardial fibrosis. This protective effect was still present after 6 months treatment withdrawal.

Cardiovascular effects of the novel potassium channel opener (3S,4R)-3-hydroxy-2,2-dimethyl-4-(2-oxo- 1-pyrrolidinyl)-6-phenylsulfonylchromane hemihydrate.

Cardiovascular effects of the novel potassium channel opener (3S,4R)-3-hydroxy-2,2-dimethyl-4-(2-oxo-1-pyrrolidinyl)-6- phenylsulfonylchromane hemihydrate (Hoe 234, CAS 132014-21-2) were investigated in rats, dogs and monkeys. In all species and independent of the route of administration Hoe 234 lowered systemic blood pressure accompanied with increases in heart rate. In rats after intravenous (i.v.) application Hoe 234 was 3 times more potent than cromakalim and its effects were reduced by pretreatment with the potassium channel blocker glibenclamide. Following intraduodenal application again Hoe 234 was more potent but mean arterial blood pressure (MAP) decreased more slowly and maximal effects were obtained later than after cromakalim. Oral administration of either single or repeated doses, however, revealed a somewhat higher potency for cromakalim. In anesthetized dogs Hoe 234 i.v. reduced MAP more potently than cromakalim whereas changes in heart rate were less pronounced. Cardiac output was increased and total peripheral resistance decreased for either agent. These results show that Hoe 234 is a novel potassium channel opener lowering blood pressure in animals due to peripheral vasodilation. It compares favourable with known potassium channel openers except for oral administration.

Effects of Na+/H+ exchange inhibitors in cardiac ischemia.

To investigate a possible protective role of Na+/H+ exchange inhibition under ischemic conditions isolated rat hearts were subjected to regional ischemia and reperfusion. In these experiments all 6 untreated hearts suffered ventricular fibrillation on reperfusion. Addition of 1 x 10(-5) mol/l amiloride or 3 x 10(-7) mol/l 5-(N-ethyl-N-isopropyl)amiloride (EIPA) markedly decreased the incidence and duration of ventricular fibrillation or even suppressed fibrillation completely as in the case of 1 x 10(-6) mol/l EIPA. Both compounds diminished the activities of lactate dehydrogenase and creatine kinase in the venous effluent of the hearts during ischemia. At the end of the experiments tissue contents of glycogen, ATP and creatine phosphate were increased in the treated hearts as compared to control hearts. In an additional experiment the beneficial effects of Na+/H+ exchange inhibition during ischemia was confirmed in vivo with anaesthetized rats undergoing coronary artery ligation. In these animals amiloride or EIPA pretreatment caused a marked reduction of ventricular premature beats and ventricular tachycardia as well as a complete suppression of ventricular fibrillation. The concentration dependent inhibition of Na+ influx via Na+/H+ exchange by amiloride and EIPA was investigated in erythrocytes from hypercholesterolemic rabbits with Na+/H+ exchange activated by exposure to hyperosmotic medium. Furthermore the inhibition of Na+ influx by EIPA after intracellular acidification was studied in cardiac myocytes of neonatal rats. Both agents were effective in the same order of potency in the ischemic isolated working rat heart as in the erythrocyte model in which they inhibited Na+/H+ exchange.(ABSTRACT TRUNCATED AT 250 WORDS)

T-kininogen, processing and functions.

Studies are presented which indicate that T-kininogen, the acute phase kininogen of the rat, could be a healing protein because of its properties as a cysteine protease inhibitor. Evidence is also presented that mRNA of T-kininogen synthesis may be a function of interleukin 6 production. A regulatory mechanism is postulated by which SH cofactors could determine if T-kinin is released or whether the T-kininogen molecule would remain intact. Evidence is also presented that T-kinin acts through kinin B2 receptors. No specific binding of bradykinin or T-kinin could be detected in rat heart preparations.

Lack of specific binding of bradykinin and D-Arg [Hyp3, Thi5, D-Tic, Oic8] bradykinin in membranes from rat heart.

Recent data demonstrated effects of bradykinin (BK) in the isolated perfused rat heart which could be blocked by BK antagonists. However, so far BK receptors in heart tissue have not been characterized. To search for BK receptors in rat hearts iodinated D-Arg[Hyp3,Thi5,D-Tic,Oic8]BK (Hoe 140) was used as a ligand for binding due to its high affinity and to its resistance to degradation. The labeled antagonist bound well to membranes prepared from rat ileum and could be displaced by the unlabeled antagonist as well as by BK and T-kinin in a concentration dependent manner. However, there was no specific binding detectable when membranes from rat left cardiac ventricle were used. To assure integrity of at least one other receptor in these membranes, binding of 3H-methylscopolamine and its displacement by the respective cold ligand was demonstrated. To rule out an occupation of the BK receptors by endogenous formed BK, the tissue was treated with an acidic buffer with high ionic strength to remove surface bound BK. However, this treatment did not unmask any specific binding for the BK antagonist. In view of the possibility that the structure of the labeled antagonist prevented its binding, experiments were carried out using tritiated BK itself. These experiments also failed to demonstrate specific binding sites which indicate that the structural aspects of Hoe 140 are probably not interfering in binding of the antagonist, especially since it binds to ileum. It is concluded, that there are too few binding sites for BK in the rat heart homogenate for ordinary binding studies.(ABSTRACT TRUNCATED AT 250 WORDS)

Airway pharmacology of the potassium channel opener, HOE 234, in guinea pigs: in vitro and in vivo studies.

The smooth muscle relaxant effects of the novel potassium channel opener, HOE 234, were investigated in guinea pig airways and compared with those of lemakalim (BRL 38227). Both agents evoked concentration-related reduction in spontaneous tracheal tone or in the tone induced by histamine, prostaglandin E2 or carbachol. HOE 234 was more potent, particularly against carbachol, and was considerably longer acting than lemakalim in a wash-out experiment. On testing for preventive efficacy against histamine-induced bronchoconstriction in anaesthetized animals a dose-related decrease of pulmonary resistance (RL) was observed. HOE 234 given either intravenously (i.v.) or by inhalation was longer acting and 3 and 6 times more potent than lemakalim. Administration of 30 micrograms/kg i.v. HOE 234 during continuous bronchoconstriction maintained by infusion of histamine decreased RL for more than 20 min whereas the effect of 100 micrograms/kg i.v. lemakalin disappeared within 4 min. These results show that HOE 234 is effective against contractile response induced by asthma mediators in guinea pig airways and compares favourably with lemakalim. Moreover it acts on acute existing bronchospasm and therefore has the potential to act against asthma attacks.

Hoe 140 a new potent and long acting bradykinin-antagonist: in vitro studies.

1. Hoe 140 (D-Arg-[Hyp3, Thi5, D-Tic7, Oic8]bradykinin) is a new bradykinin (BK)-antagonist. It was tested in several in vitro assays and compared with D-Arg-[Hyp2,Thi5,8,D-Phe7]BK. 2. In receptor binding studies in guinea-pig ileum preparations, Hoe 140 showed an IC50 of 1.07 x 10(-9) mol l-1 and a KI value of 7.98 x 10(-10) mol l-1. 3. In isolated organ preparations Hoe 140 and D-Arg-[Hyp2,Thi5,8, D-Phe7]BK inhibited bradykinin-induced contractions concentration dependently, with IC50-values in the guinea-pig ileum preparation of 1.1 x 10(-8) mol l-1 and 3 x 10(-5) mol l-1, respectively. pA2 values in this tissue were 8.42 and 6.18, respectively. In the rat uterus preparation the IC50 value was 4.9 x 10(-9) mol l-1 for Hoe 140. D-Arg-[Hyp2, Thi5,8, D-Phe7]BK showed an IC50 of 4.0 x 10(-6) mol l-1. The IC50 values in the guinea-pig isolated pulmonary artery were 5.4 x 10(-9) mol l-1 and 6.4 x 10(-6) mol l-1, respectively. In the rabbit aorta no inhibitory effects on Des-Arg9-BK induced contractions were observed. 4. In cultured bovine endothelial cells, Hoe 140 antagonized (IC50 = 10(-8) mol l-1) bradykinin-induced endothelium-derived relaxing factor (EDRF) release and the bradykinin-induced increase in cytosolic free calcium (IC50 = 10(-9) mol l-1). 5. Hoe 140 (10 -7mol I1) totally suppressed the bradykinin-induced (10 8 to 10- mol I') prostacyclin (PGI2) release from cultured endothelial cells of bovine aorta. D-Arg-[Hyp2, Thi5'8, D-Phe7]BK (10- 7 mol I1- ) showed a weaker antagonism. 6. Taken together these results show that Hoe 140 is a highly potent bradykinin antagonist. It was two to three orders of magnitude more potent than D-Arg-[Hyp2, Thi5 8, D-Phe7]BK.

Hoe 140 a new potent and long acting bradykinin-antagonist: in vivo studies.

1. The potency, duration of action and tolerability of Hoe 140, a novel and highly potent bradykinin (BK) antagonist in vitro, has been tested in different in vivo models and compared with the well-known BK antagonist D-Arg-[Hyp2, Thi5,8, D-Phe7]BK. 2. Hoe 140 is highly potent and long acting in inhibiting BK-induced hypotensive responses in the rat. Four hours after s.c. administration of 20 nmol kg-1, inhibition still amounted to 60% whereas the effect of 200 nmol kg-1 of D-Arg-[Hyp2, Thi5,8, D-Phe7]BK was not significant. 3. BK-induced bronchoconstriction in guinea-pigs was strongly inhibited by Hoe 140. The magnitude and duration of inhibition confirmed the findings obtained in the blood pressure experiments in the rat. 4. Carrageenin-induced inflammatory oedema of the rat paw was considerably inhibited at i.v. doses between 0.1 and 1 mg kg-1. 5. In conscious dogs, intravenous doses of 0.01 and 0.1 mg kg-1 of Hoe 140 and D-Arg-[Hyp2, Thi5,8, D-Phe7]BK were well tolerated. At doses of 1 mg kg-1 adverse effects occurred that were attributed to the residual BK agonistic activity of both compounds. 6. Hoe 140 has been shown to be a highly potent and long acting BK antagonist in vivo in different animal species and models. This makes it appropriate to investigate further the physiological and pathophysiological role of BK.

Cardiac arrhythmias are ameliorated by local inhibition of angiotensin formation and bradykinin degradation with the converting-enzyme inhibitor ramipril.

We investigated the influence of the angiotensin-converting enzyme (ACE) inhibitor ramipril on cardiac arrhythmias in guinea pigs and rats. Ramiprilat, the active moiety of ramipril, did not influence action potentials of isolated guinea-pig papillary muscle or rabbit sinus node, thereby excluding cellular electrophysiological evidence of anti-arrhythmic properties. Ramipril protected against cardiac arrhythmias induced by digoxin infusion in guinea pigs. This effect was comparable with that of lidocaine. In isolated perfused ischemic working rat hearts, angiotensin (ANG) I (3 x 10(-9) M/l) and ANG II (1 x 10(-9) M/l) aggravated reperfusion arrhythmias, accompanied by deterioration of cardiodynamic and metabolic events. Bradykinin (BK) (1 x 10(-10)-1 x 10(-8) M/l), in contrast, protected against reperfusion arrhythmias, which corresponded to an increase in energy-rich phosphates and glycogen stores and a decrease in lactate levels in myocardial tissue. Identical changes were seen in hearts from rats pretreated with ramipril (1 mg/kg PO) or perfused with ramiprilat (2.58 x 10(-7)-2.58 x 10(-5) M/l). Local ACE inhibition in these ischemic hearts antagonized ANG I but not ANG II effects and enhanced BK effects. The BK antagonist D-Arg-(Hyp2, Thi5,8, D-Phe7)BK abolished the beneficial effects of BK, ramipril, and ramiprilat. Increased concentrations of BK or ramiprilat were able to reverse the antagonism. The antiarrhythmic agent nicainoprol, a fast-sodium-channel blocking drug (class Ib), also protected isolated rat hearts against reperfusion arrhythmias, but was without beneficial effects on cardiac hemodynamics and biochemical parameters, in contrast to the ACE inhibitor. These results suggest that the beneficial effects of the ACE inhibitor ramipril on digoxin and reperfusion arrhythmias are not mediated by their direct actions on ionic channels in the cell membrane. It seems that other factors are responsible for its beneficial effects on reperfusion arrhythmias, cardiac function, and metabolism, which are associated with a reduction in ANG II generation and BK degradation by local ACE inhibition in the heart.