The standardized Ginkgo biloba extract Egb-761 protects vascular endothelium exposed to oxidized low density lipoproteins.

Dietary antioxidants are frequently proposed as protective agents for the vascular endothelium during the onset of atherosclerosis. This protection may occur at two distinct levels. First, they prevent oxidative modification of atherogenic lipoproteins (LDL). Second, they can provide a cellular protection against oxidized LDL-mediated endothelium dysfunction, although this mechanism remains poorly considered in many instances. To gain insight into the mechanism underlying such cellular protection against oxidized LDL, we examined the impact of a popular traditional medicine, an extract from Ginkgo biloba with well-known antioxidant properties, on two endothelial cells properties: cell adhesion and ionic homeostasis. Cellular lipoperoxides levels were also measured as a marker of cellular oxidative stress. Human umbilical-vein endothelial cells were exposed to native (nat-) or oxidized (ox-) LDL, the latter prepared to be compatible with clinically observed levels of oxidation. Although nat-LDL had little effect, ox-LDL increased endothelial adhesive properties (35%, p<0.01) and lipoperoxidation (45%, p<0.01). Na,K-ATPase activity, a key regulator of ionic homeostasis, was significantly decreased after exposure to nat-LDL (30%, p<0.01) and dramatically depressed after exposure to ox-LDL (65%, p<0.001). The standardized preparation of Ginkgo biloba EGb-761 totally protected adhesive properties and endothelial lipoperoxide levels. Moreover, it limited the decrease in Na,K-ATPase activity induced by ox-LDL to levels similar to nat-LDL. This suggests that EGb-761 protects endothelial adhesive properties and helps prevent the disruption of ionic homeostasis. The EGb-761-mediated inhibition of ox-LDL-induced lipoperoxide levels in endothelial cells appears to be an important mechanism by which Ginkgo biloba extract protects endothelial properties.

Inhibition of type 4 phosphodiesterase by rolipram and Ginkgo biloba extract (EGb 761) decreases agonist-induced rises in internal calcium in human endothelial cells.

The effects of Gingko biloba extract EGb 761 on 5 isolated, vascular, cyclic nucleotide phosphodiesterase (PDE) isoforms were evaluated. EGb 761 preferentially inhibited PDE4 (IC(50)=25.1 mg/L), the isoform that is mainly present in endothelial cells, in a competitive manner (K:(i)=12.5 mg/L). Because changes in cyclic nucleotide levels may affect intracellular calcium ([Ca(2+)](i)) levels in endothelial cells, we examined the effects of EGb 761 on both resting [Ca(2+)](i) levels and agonist-induced rises in [Ca(2+)](i) in single human umbilical vein endothelial cells (HUVECs) in culture. The effects of EGb 761 were compared with those of rolipram, a selective PDE4 inhibitor that increases cellular cAMP levels, and the cAMP analogue dibutyryl cAMP (db-cAMP). EGb 761 (20 and 100 mg/L), rolipram (50 micromol/L), and db-cAMP (100 micromol/L) significantly inhibited histamine-, ATP-, and thrombin-induced [Ca(2+)](i) increases in HUVECs without modifying resting [Ca(2+)](i) levels. Similar results were obtained by using a Ca(2+)-free bath solution. EGb 761 (100 mg/L), but not rolipram (50 micromol/L) or db-cAMP (100 micromol/L), also inhibited Ca(2+) influx into cells having thapsigargin-depleted internal Ca(2+) stores and bathed in a Ca(2+)-free external solution. Our results are consistent with an inhibition of PDE activity that causes a reduction of agonist-induced increases in [Ca(2+)](i) in HUVECs, mainly by inhibition of Ca(2+) mobilization from internal stores. It thus may be that the cardiovascular effects of EGb 761 involve inhibition of PDE4 activity and subsequent modification of Ca(2+) signaling in endothelial cells.

KATP channel modulation in working rat hearts with coronary occlusion: effects of cromakalim, cicletanine, and glibenclamide.

OBJECTIVES: We studied the effects of ATP-sensitive potassium channel (KATP) modulation on ischemic cardiac performance and reperfusion-induced ventricular fibrillation (VF), and assessed the contribution of KATP to the cardioprotective and anti-arrhythmic effect of the anti-hypertensive drug cicletanine. METHODS: Isolated working rat hearts, subjected to a 10-min coronary occlusion followed by reperfusion, were perfused in the presence of vehicle, 0.1-60 microM cromakalim, an opener of KATP; 3-60 microM cicletanine; and 0.1-10 microM glibenclamide, a blocker of KATP, respectively. RESULTS: All concentrations of cicletanine, similarly to 0.1-10 microM cromakalim, attenuated ischemia-induced deterioration of aortic flow, left ventricular developed pressure, and left ventricular end-diastolic pressure. In contrast to cromakalim, cicletanine did not increase coronary flow. Cicletanine (60 microM) and cromakalim (10 and 60 microM) significantly reduced the incidence of reperfusion-induced VF; however, 60 microM cromakalim triggered VF during ischemia. Lower concentrations of cromakalim and cicletanine did not produce an anti-arrhythmic effect. Cardiac functional parameters were concentration dependently worsened by glibenclamide, and the drug did not change the incidence of VF. Glibenclamide (0.1 microM) did not significantly affect cardiac performance, but it did abolish the anti-ischemic effect of cromakalim (1-10 microM) and cicletanine (60 microM). Glibenclamide suppressed the anti-arrhythmic effect of 10 and 60 microM cromakalim; however, it did not affect the anti-arrhythmic effect of cicletanine. CONCLUSIONS: (i) The anti-ischemic but not the anti-arrhythmic effect of cicletanine may involve opening of KATP. (ii) opening of KATP attenuates, inhibition of the channel exacerbates functional consequences of coronary occlusion, and (iii) KATP opening attenuates reperfusion-induced VF, but it triggers ischemia-induced VF. KATP blocking does not affect VF.

Effects of SOD, catalase, and a novel antiarrhythmic drug, EGB 761, on reperfusion-induced arrhythmias in isolated rat hearts.

Effects of superoxide dismutase (SOD), catalase, EGB 761 (Tanakan), and their combination on reperfusion-induced ventricular fibrillation (VF), tachycardia (VT), and the formation of oxygen free radicals were studied after 30 min of global ischemia followed by reperfusion in isolated rat hearts. In the first series of studies, rats received a daily dose of 10(4), 2 x 10(4), or 5 x 10(4) U/kg of SOD (i.v.); 2.5 x 10(4), 5 x 10(4), or 10(5) U/kg of catalase (i.v.); and 25, 50, 100, or 200 mg/kg of EGB 761 (per os), respectively, for 10 d (chronic administration). Neither SOD nor catalase alone reduced the incidence of reperfusion arrhythmias, but EGB 761 dose-dependently reduced the incidence of such arrhythmias. The coadministration of SOD (5 x 10(4) U/kg) with catalase (5 x 10(4) U/kg) significantly reduced the incidence of VF and VT. The same reduction in the incidence of VF and VT was observed when SOD (5 x 10(4) U/kg) was given in combination with EGB 761 (50 mg/kg). In the second series of studies, hearts were isolated and perfused with 5 x 10(4) U/l of SOD plus 5 x 10(4) U/l of catalase (acute treatment), and the incidence of reperfusion-induced VF and VT was significantly reduced. The combination of SOD (5 x 10(4) U/l) with EGB 761 (50 mg/l) also reduced the incidence of VF and VT. In these experiments, we studied the time course of oxygen radical formation using 5,5-dimethyl-pyrroline-N-oxide (DMPO), a spin trap, and it was found that EGB 761 (200 mg/l) or the coadministration of EGB 761 (50 mg/l) with SOD (5 x 10(4) U/l) almost completely abolished the formation of oxygen radicals during reperfusion measured by electron spin resonance (ESR) spectroscopy. Although SOD or catalase alone significantly reduced the formation of oxygen radicals, these drugs failed to prevent the development of reperfusion arrhythmias, while their combination significantly attenuated both the formation of free radicals and the incidence of reperfusion-induced arrhythmias. Our results indicate that the combination therapy may synergistically reduce the formation of free radicals and the incidence of reperfusion-induced VF and VT.

Modification of reperfusion-induced ionic imbalance by free radical scavengers in spontaneously hypertensive rat retina.

We studied the effects of free radical scavengers, superoxide dismutase (SOD), vitamin E, and EGB 761, on ion shifts (Na+, K+, and Ca2+) induced by ischemia reperfusion in rat retina obtained from spontaneously hypertensive rats. Eyes were subjected to 90 min of retinal ischemia followed by 24 h of reperfusion. Two basic protocols were used: (1) chronic application, in which rats received SOD (7500, 15,000, and 30,000 U/kg, i.v.), vitamin E (50, 100, and 200 mg/kg, i.v.), and EGB 671 (50, 100, and 200 mg/kg, orally) for 10 d, respectively; and (2) acute administration, in which 7500, 15,000, and 30,000 U/kg of SOD, 50, 100, and 200 mg/kg of vitamin E, and 50, 100, and 200 mg/kg of EGB 761 were administered after an ischemic episode, at the onset of reperfusion, respectively. In the drug-free control group, 90 min ischemia followed by 24 h of reperfusion resulted in an accumulation of retinal sodium and calcium from their nonischemic control values of 76 +/- 4 and 3.2 +/- 0.1 mumol/g dry weight to 112 +/- 6 (p < .001) and 6.2 (p < .001) mumol/g dry weight, respectively. Tissue potassium loss was also observed in this model of retinal ischemia reperfusion, and after 90 min ischemia followed by 24 h of reperfusion potassium content was significantly reduced from its nonischemic control value of 266 +/- 5 to 207 +/- 6 (p < .001) mumol/g dry weight. The chronic administration of SOD, vitamin E, and EGB 761 dose dependently reduced the reperfusion-induced ionic imbalance and improved the recovery of retinal ion contents.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of natural antioxidant ginkgo biloba extract (EGB 761) on myocardial ischemia-reperfusion injury.

Recently, it was reported that Ginkgo biloba extract (EGb 761), which is known to have antioxidant properties, also has antiarrhythmic effects on cardiac reperfusion-induced arrhythmias. In the present study, effects of EGb 761 on cardiac ischemia-reperfusion injury were investigated from the point of view of recovery of mechanical function as well as the endogenous antioxidant status of ascorbate. Isolated rat hearts were perfused using the Langendorff technique, and 40 min of global ischemia were followed by 20 min of reperfusion. EGb 761 improved cardiac mechanical recovery and suppressed the leakage of lactate dehydrogenase (LDH) during reperfusion. Furthermore, EGb 761 diminished the decrease of myocardial ascorbate content after 40 min of ischemia and 20 min of reperfusion. Interestingly, EGb 761 also suppressed the increase of dehydroascorbate. These results indicate that EGb 761 protects against cardiac ischemia-reperfusion injury and suggest that the protective effects of EGb 761 depend on its antioxidant properties.

EGb 761 protects liver mitochondria against injury induced by in vitro anoxia/reoxygenation.

The present study investigated the protective effects of Ginkgo biloba extract (EGb 761) on rat liver mitochondrial damage induced by in vitro anoxia/reoxygenation. Anoxia/reoxygenation was known to impair respiratory activities and mitochondrial oxidative phosphorylation efficiency. ADP/O (2.57 +/- 0.11) decreased after anoxia/reoxygenation (1.75 +/- 0.09, p < .01), as well as state 3 and uncoupled respiration (-20%, p < .01), but state 4 respiration increased (p < .01). EGb 761 (50-200 microg/ml) had no effect on mitochondrial functions before anoxia, but had a specific dose-dependent protective effect after anoxia/reoxygenation. When mitochondria were incubated with 200 microg/ml EGb 761, they showed an increase in ADP/O (2.09 +/- 0.14, p < .05) and a decrease in state 4 respiration (-22%) after anoxia/reoxygenation. In EPR spin-trapping measurement, EGb 761 decreased the EPR signal of superoxide anion produced during reoxygenation. In conclusion, EGb 761 specially protects mitochondrial ATP synthesis against anoxia/reoxygenation injury by scavenging the superoxide anion generated by mitochondria.

A Ginkgo biloba extract (EGb 761) prevents mitochondrial aging by protecting against oxidative stress.

The effect of aging on indices of oxidative damage in rat mitochondria and the protective effect of the Ginkgo biloba extract EGb 761 was investigated. Mitochondrial DNA from brain and liver of old rats exhibited oxidative damage that is significantly higher than that from young rats. Mitochondrial glutathione is also more oxidized in old than in young rats. Peroxide formation in mitochondria from old animals was higher than in those from young ones. According to morphological parameters (size and complexity), there are two populations of mitochondria. One is composed of large, highly complex mitochondria, and the other population is smaller and less complex. Brain and liver from old animals had a higher proportion of the large, highly complex mitochondria than seen in organs from young animals. Treatment with the Ginkgo biloba extract EGb 761 partially prevented these morphological changes as well as the indices of oxidative damage observed in brain and liver mitochondria from old animals.

Cicletanine reverses vasoconstriction induced by the endogenous sodium pump ligand, marinobufagenin, via a protein kinase C dependent mechanism.

RATIONALE: Cicletanine (CIC), an anti-hypertensive compound with direct vascular and natriuretic actions, is especially effective in salt-sensitive hypertension, in which dysregulation of the sodium pump plays an important pathogenic role, and digitalis-like cardiotonic steroids contribute to increased vascular tone. The purpose of the present study was to investigate whether, and by what mechanisms, cicletanine antagonizes the vasoconstrictor effects of cardiotonic steroids in isolated human arteries. METHODS: The effects of cicletanine on vascular tone were studied in isolated, endothelium-denuded rings of 2nd-3rd-order branches of human mesenteric arteries pre-contracted with bufodienolide marinobufagenin (MBG), an Na/K-ATPase inhibitor, or endothelin-1 (ET-1). Na/K-ATPase activity was measured in sarcolemmal membranes from the mesenteric artery. Activity of rat brain protein kinase C (PKC) was measured using the PepTag phosphorylation assay. RESULTS: MBG and ET-1 both induced sustained vasoconstriction in human mesenteric artery rings, and cicletanine relaxed rings pre-contracted with either MBG (EC50 = 11 +/- 2 micromol/l) or ET-1 (EC50 = 6.4 +/- 1.1 micromol/l). Although 8-Br-cGMP (100 micromol/l) caused complete vasorelaxation of arterial rings pre-contracted with ET-1, it did not affect the MBG-induced vasoconstriction. An activator of PKC, phorbol diacetate (PDA) (50 nmol/l), attenuated CIC-induced vasorelaxation of mesenteric artery rings pre-contracted with MBG (EC50 > 100 micromol/l), but not rings pre-contracted with ET-1 (EC50 = 6.5 +/- 1.2 micromol/l). In mesenteric artery sarcolemma, 100 nmol/l MBG inhibited the Na/K-ATPase by 68 +/- 5% and cicletanine (100 micromol/l) attenuated this Na/K-ATPase inhibition by 85 +/- 6%. In the PepTag PKC assay, cicletanine produced a concentration-dependent inhibition of rat brain PKC activity (IC50 45 +/- 11 micromol/l). In the presence of 50 nmol/l PDA, 100 micromol/l cicletanine did not antagonize the Na/K-ATPase inhibition by MBG, and did not inhibit the PKC from rat brain. CONCLUSIONS: Cicletanine antagonizes vasoconstriction induced by Na/K-ATPase inhibition via a PKC-dependent mechanism that does not involve inhibition of cyclic GMP phosphodiesterase (cGMP-PDE). This mechanism of action may be relevant to the greater potency of cicletanine in salt-sensitive hypertension in which plasma levels of endogenous digitalis-like cardiotonic steroids are elevated. Our findings also suggest that PKC is an important factor for cardiotonic steroid-Na/K-ATPase interactions on the vascular tone, and is therefore a potential target for therapeutic intervention in hypertension.

Ischemia and reperfusion-induced histologic changes in the rat retina. Demonstration of a free radical-mediated mechanism.

Histologic alterations of ischemia- and reperfusion-induced retinal damage are critically dependent on the duration of the period of ischemia. Male Sprague Dawley rats were anesthetized, and a suture was placed behind the globe including the central retinal artery. Because it was desirable that untreated eyes show a great histologic change due to reperfusion-induced damage (in order that maximum scope would exist for demonstration of any protective effect of a drug treatment), a preliminary series of studies established the time-induced characteristics for the retina with transient regional ischemia. Eyes (n = 6-12 in each group) were subjected to 30, 60, or 90 min of ischemia followed by 0.5, 1, 2, 4, and 24 hr of reperfusion, respectively. The 30-min ischemia followed by reperfusion did not result in any histologic changes; 60-min ischemia followed by reperfusion induced a moderate retinal edema which returned to the preischemic value after 24 hr of reperfusion. The 90-min ischemia followed by reperfusion further aggravated retinal edema and increased the migration of neutrophil leukocytes. Even after 24 hr of reperfusion, the retinal edema had not disappeared although an attenuation was observed. In this study, the rats were treated with superoxide dismutase (SOD-PEG, 15 x 10(3) U/kg) or EGB 761 (100 mg/kg) for 10 days (chronic treatment). The SOD and EGB 761 significantly reduced the development of reperfusion-induced retinal edema and significantly prevented the neutrophil leukocyte infiltration. Both also had a protective effect against reperfusion-induced injury when these agents were administered just before reperfusion ("late" administration).(ABSTRACT TRUNCATED AT 250 WORDS)

Functional protection of photoreceptors from light-induced damage by dimethylthiourea and Ginkgo biloba extract.

PURPOSE: To investigate the functional protective effect of a synthetic (dimethylthiourea, DMTU) and a natural antioxidant (Ginkgo biloba extract, EGb 761) against light-induced retinal degeneration. METHODS: Wistar rats were exposed for 24 hours to 1700-lux light after treatment with DMTU or EGb 761. Electroretinograms were recorded before and on day (D)1, D3, D8, D15, D22, and D29 after light exposure. The b-wave amplitude was plotted against log L (ganzfeld luminance), providing the b-wave sensitivity curve. The Naka-Rushton function fitted to the sensitivity curve enabled derivation of the parameters Bmax (saturated amplitude) and K (luminance-inducing Bmax/2). In addition, rats from each group were killed for retinal morphometric analyses. RESULTS: In the untreated group, light exposure caused collapse of the b-wave sensitivity curves. Bmax was reduced by 51% at D1 without subsequent recovery. K increased temporarily, reverting to normal values 8 days later. The outer nuclear layer thicknesses decreased markedly in the superior retina. In the treated groups, light exposure had a weaker effect on sensitivity curves. The values of Bmax were not significantly different from those in the unexposed-untreated group, although K increased temporarily. Retinal morphometry was preserved. CONCLUSIONS: Dimethylthiourea and EGb 761 afford functional protection against light-induced retinal damage.

Peroxyl radical scavenging activity of Ginkgo biloba extract EGb 761.

Antioxidant mechanisms have been proposed to underlie the beneficial pharmacological effects of EGb 761, an extract from Ginkgo biloba leaves used for treating peripheral vascular diseases and cerebrovascular insufficiency in the elderly. In vitro evidence has been reported that EGb 761 scavenges various reactive oxygen species, i.e. nitric oxide, and the superoxide, hydroxyl, and oxoferryl radicals. However, the ability of EGb 761 to scavenge peroxyl radicals (reactive species mainly involved in the propagation step of lipid peroxidation) has not been investigated. To characterize further the antioxidant action of EGb 761, we measured the protective effects of EGb 761 during: (1) the oxidation of B-phycoerythrin by peroxyl radicals generated in aqueous solution by 2,2'-azobis (2-amidinopropane) hydrochloride (AAPH); and (2) the reaction of luminol or cis-parinaric acid with peroxyl radicals generated from 2,2'-azobis (2,4-dimethylvaleronitrile) (AMVN) in liposomes or in human low density lipoprotein (LDL), respectively. To evaluate the peroxyl radical scavenging activity of EGb 761 in a more physiologically relevant model of damage to lipid-containing systems, we also analyzed the effect of the extract on the oxidation of human LDL exposed to the azo-initiators in terms of: (1) accumulation of cholesterol linoleate ester hydroperoxides, (2) depletion of alpha-tocopherol and beta-carotene, and (3) changes in intrinsic tryptophan fluorescence. EGb 761 afforded protection against oxidative damage in all the systems we analyzed; thus, it is an efficient scavenger of peroxyl radicals. This result extends the oxygen radical scavenging properties of the extract and supports the hypothesis of an antioxidant therapeutic action of EGb 761.

Ginkgo biloba extract (EGb 761): inhibitory effect on nitric oxide production in the macrophage cell line RAW 264.7.

The present study was conducted to evaluate the effect of Ginkgo biloba extract (EGb 761) on the synthesis of nitric oxide (NO) induced by lipopolysaccharide (LPS) plus interferon-gamma (IFN-gamma) in the mouse macrophage cell line RAW 264.7. EGb 761 inhibited nitrite and nitrate production, taken as an index for NO, in a concentration-dependent fashion. The IC50 for inhibition of nitrite production by activated macrophages was about 100 micrograms/mL EGb 761. The inducible NO synthase (iNOS) enzyme activity of cytosolic preparations from activated RAW 264.7 cells was inhibited by treatment with EGb 761. In addition, reverse transcription-polymerase chain reaction (RT-PCR) analysis revealed that the expression of iNOS mRNA in activated macrophages was suppressed by high concentrations of EGb 761. However, NF-kappa B DNA binding activity induced by activation with LPS/IFN-gamma was not inhibited by EGb 761. These findings indicate that not only does EGb 761 directly act as an NO scavenger but also that it inhibits NO production in LPS/IFN-gamma-activated macrophages by concomitant inhibition of induction of iNOS mRNA and the enzyme activity of iNOS. Thus, EGb 761 may act as a potent inhibitor of NO production under tissue-damaging inflammatory conditions.

Oxidative stress in diabetic retina.

The authors describe the alterations usually associated with diabetic retinopathy. They concern the classical thickening of the basal membrane of retinal capillaries and the associated modification of retinal vessel permeability. These alterations correspond to the blood-retinal barrier disruption. The authors then discuss the participation of oxygenated free radicals in the pathogenesis of diabetic retinopathy. They report several experimental studies establishing such a participation and finally describe their own results obtained on a model of retinas isolated from alloxan-induced diabetic rats. After one month of evolution, the electroretinograms (ERG) recorded on isolated retinas from diabetic rats had an amplitude about 20% lower than the controls, whereas after two months of diabetes, this decrease was about 60%. Under these conditions, the authors tested the protective properties of Ginkgo biloba extract (EGb 761) on their model. They observed that in EGb-treated animals (100 mg/kg/day), the ERG had a significantly (p less than 0.001) greater amplitude than untreated animals after two months of diabetes evolution. In conclusion, the authors discuss the possible utilization of a free radical scavenger, such as EGb 761, in the prevention of the retinal impairment in diabetes.

Protective action of diosmectite treatment on digestive disturbances induced by intestinal anaphylaxis in the guinea-pig.

METHODS: Colonic transit time, faecal moisture and intestinal permeability were assessed in guinea-pigs sensitized intraperitoneally with cow's milk and challenged with an oral administration of beta-lactoglobulin. One group of animals was treated for 1 week with diosmectite (500 mg.kg/day) and another with placebo. A control group was not sensitized but treated with diosmectite. RESULTS: In sensitized animals receiving placebo, challenge with beta-lactoglobulin induced a significant (P < 0.05) decrease in colonic transit time, and increases in faecal moisture and intestinal permeability. These changes were not observed in animals treated with diosmectite. CONCLUSION: Diosmectite pre-treatment protects against allergic digestive disturbances induced by antigen administration in guinea-pigs sensitized to cow's milk.

Vasorelaxant effects of cicletanine and its (+)- and (-)-enantiomers in isolated human pulmonary arteries.

The purpose of the study was to investigate, in isolated human pulmonary artery, the ability of cicletanine and its (-) and (+)-enantiomers to attenuate the endothelin-1 (Et-1) induced vasoconstriction, and to potentiate vasorelaxation (relative to plateau of the effect of Et-1) by sodium nitroprusside (SNP) and human atrial natriuretic peptide (ANP). In pulmonary artery rings, Et-1 induced a concentration-dependent vasoconstriction with median effective concentration (EC50 = 26+/-2.8 nmol/L. Pretreatment of the vessels with 100 micromol/L (+/-)-cicletanine reduced the effect of Et-1 (EC50 = 36+/-3.5 nmol/L; P < .01). (-)-enantiomer displayed greater capacity to antagonize the vasoconstrictor action of Et-1 (EC50 = 47+/-4.2 nmol/L) v (+)-enantiomer (EC50 = 29.9+/-6.5 nmol/L; P < .01). In arterial rings, precontracted with 10 nmol/L Et-1, ANP caused vasorelaxation (EC50 = 9.7+/-1.9 nmol/ L). The relaxant effect of ANP was potentiated by 100 micromol/L of (-)-(EC50 = 4.2+/-0.6 nmol/L; P < .01), but not (+)-cicletanine (EC50 = 7.6+/-0.7 nmol/L). Sodium nitroprusside relaxed pulmonary artery rings precontracted with 10 nmol/L Et-1 (EC50 = 41+/-11 nmol/L). The effect of SNP was potentiated by 10 micromol/L (+/-)-cicletanine (EC50 = 9.0+/-0.7 nmol/L; P < .05). The potentiating effect of 10 micromol/L (+)-cicletanine was weaker (EC50 = 7.9+/-1.8 nmol/L) than that of (-)-enantiomer (EC50 = 3.3+/-0.54 nmol/L; P < .05). The relaxant effect of SNP was not further potentiated by 100 micromol/L (+/-)-cicletanine. The present results demonstrate that, cicletanine antagonizes Et-1 induced vasoconstriction in an isolated human pulmonary artery and potentiates vasorelaxation by two guanylate cyclase activators, ANP and SNP. (-)-Cicletanine displays greater vasorelaxant activity v (+)-enantiomer.

Inhibition by (-)-cicletanine of the vascular reactivity to angiotensin II and vasopressin in isolated rat vessels.

In pithed rats, the levorotatory (-)-enantiomer of cicletanine reduces the pressor responses to angiotensin II (AII) and also, to a lesser extent, those to arginine-vasopressin (AVP). Here we have attempted to characterize further these inhibitory effects by studies of isolated perfused rat kidney and mesenteric vascular beds. In the isolated rat kidney, (-)-cicletanine behaves as a noncompetitive antagonist of AII- and AVP-receptor stimulation, with Ki values of 9.6 and 208 micromol/L respectively. In the isolated mesenteric vascular bed, (-)-cicletanine antagonized both AII dependent contractions with an inhibitory concentration (IC50) of 54.0 +/- 20.5 micromol/L (n = 6), and AVP dependent contractions with an IC50 of 31.6 +/- 5.0 micromol/L (n = 8). In conclusion, (-)-cicletanine antagonizes AII more effectively in rat kidney than in mesenteric vascular beds. Moreover, in rat kidney vascular beds (-)-cicletanine is more potent in blocking the pressor responses to AII than in blocking those to AVP. A selective blockade of AII induced contractions in kidney vascular beds can be one factor explaining both the greater antagonistic potency of (-)-cicletanine against AII compared with AVP in pithed rats, and the renal protective properties of cicletanine in both hypertensive and aged rats.

The effect of cicletanine on cerebrovascular injury in stroke-prone spontaneously hypertensive rats.

The stroke-prone spontaneously hypertensive rat (SHR-SP) is one of the most suitable models for stroke study. The present trial work was undertaken so as to obtain further information concerning the action of a new furopyridine, cicletanine. Forty-six males--SHR-SP/Iffa Credo rats--aged 7 weeks, were divided into three groups. Group 1 was a control group, groups 2 and 3 were orally treated with cicletanine at 30 and 100 mg/kg. Their drinking water contained 1% NaCl. Systolic blood pressure, body weight, and survival were recorded. After 6 weeks, all the rats were sacrificed. Samples of heart, brain, and kidney were fixed for light and ultrastructural examination. We found that cicletanine treatment (30 and 100 mg/kg) had significantly inhibited the incidence of hypertensive cerebral damages as characterized by cerebral infarction and vascular alterations with fibrinoid necrosis. Compared with the control group, the rats treated with the cicletanine had a significantly increased survival rate (P less than .001); the cicletanine also had an important protective effect on tissue. Cicletanine administration prevented the development of hypertensive cerebral vascular damage, probably through direct action on the vascular walls.

Relaxation of vascular smooth muscle by cicletanine in aged wistar aorta under stress conditions: importance of nitric oxide.

The vascular mechanism of action of cicletanine, an antihypertensive agent, was studied on isolated Wistar rat aortas (24-months-old) in presence and in absence of endothelium in two different stress conditions, normoxic and hypoxic, in presence of norepinephrine (NE). Under normoxic conditions, in presence of endothelium, cicletanine (10(-9)-10(-5)M) induced a concentration-dependent relaxation, whereas in absence of endothelium, cicletanine (10(-9)-10(-5)M) was ineffective although it relaxed the smooth muscle at higher concentrations (10(-4)M). At pharmacologic concentrations (below or equal 10(-5)M), relaxation induced by cicletanine, in presence of endothelium, was prevented by N(omega)-nitro-L-arginine (L-NNA) (P <.005) and relaxation induced by the highest concentration (10(-4)M) was reversed by BaCl2 (P <.005). Under hypoxic conditions, in presence of NE and endothelium, the aorta displayed an increased developed tension that was significantly (P <.05) attenuated by cicletanine (10(-5)M) and insensitive to indomethacine (10(-7)M). When the two compounds were added together, the relaxation induced by cicletanine was significantly improved (P <.005). These results indicated that cicletanine, under stress conditions, relaxes vascular smooth muscle through an endothelium-dependent action mediated by the nitric oxide (NO) synthase pathway. We proposed that the observed vascular effects could be associated with the counter-regulation mechanisms linked to the antihypertensive action of cicletanine.

Ginkgo biloba extract (EGb 761) protects Na,K-ATPase activity during cerebral ischemia in mice.

Neuroprotective drugs such as Ginkgo biloba extract (EGb 761) could prevent the ischemia-induced impairment of the Na,K-ATPase activity. In this study, Na,K-ATPase activity and expression, contents in fatty acids and malondialdehyde, an index of lipoperoxidation, were compared in the ipsilateral (ischemic) and the contralateral (unlesioned) cortices after 1 h of unilateral focal cortices cerebral ischemia in the mouse. EGb 761 (110 mg/kg) was administered daily to half of the animals for 10 days before ischemia. Ischemia significantly reduced Na,K-ATPase activity by about 40% and increased malondialdehyde content; EGb 761 pretreatment abolished these effects. The free radical scavenger properties of EGb 761 are a potential mechanism by which Na,K-ATPase injury and lipoperoxidation are prevented.