Search

The standardized extract of Ginkgo biloba EGb 761 has been used to reduce cognitive dysfunction. The present study was designed to evaluate the effect of postischemic oral treatment with EGb 761 in a model of vascular dementia in gerbils. Daily oral posttreatment with EGb 761 led to a significant recovery of spatial memory assessed by the object location test, inhibited the decrease in plasma SOD activity and protected the hippocampal CA1 neurons, even when administered after the insult. These data provide further evidence for the therapeutic potential of EGb 761 in the treatment of vascular dementia., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Minocycline has been reported to exert neuroprotection through inhibition of inflammatory processes and of mitochondrial cell death pathway. To further characterize the neuroprotective effect of minocycline, we determined its efficacy in different neuronal damage paradigms involving inflammation or mitochondrial dysfunction. In transient global ischaemia in gerbils, minocycline reduced hippocampal neuronal damage measured by peripheral type benzodiazepine binding sites density, a marker of microglial activation. The antiinflammatory properties of minocycline were confirmed on the model of carrageenan-induced paw oedema in rats. The use of two experimental animal models involving administration of mitochondrial toxins inhibiting a different complex of the mitochondrial respiratory chain permitted the exploration of the mitochondrial impact of minocycline. Although minocycline exhibited a marked efficacy in 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP; complex I inhibitor)-induced neurotoxicity in mice, it was ineffective in malonate (complex II inhibitor)-induced striatal lesion in rats. In vitro investigations on energized mitochondria isolated from rat liver showed that minocycline (1 microM) did not inhibit the swelling induced by MPP+(1-methyl-4-phenylpyridinium). Moreover, higher concentrations of minocycline induced swelling. From these experiments, the neuroprotective activity of minocycline appears more related to its antiinflammatory activity than to a direct beneficial action on mitochondria.

The alpha 1 adrenoceptor subtypes mediating contraction of rabbit prostate and hypogastric artery were pharmacologically characterized using an isolated organ bath technique. The prostate had the same sensitivity to the contractile action of methoxamine and phenylephrine, whereas the hypogastric artery was five times less sensitive to the action of methoxamine in comparison with phenylephrine. Clonidine elicited contraction in the hypogastric artery but not in the prostate. BMY7378 was about 70-fold more potent to antagonize the phenylephrine-induced contraction in the hypogastric artery (pA2 8.14) than in the prostate (pA2 6.28), and 5-methyl-urapidil was about three-fold more potent on prostrate than on hypogastric artery. The potency of different alpha 1-adrenoceptor antagonists tested in the rabbit prostate was significantly correlated with their binding affinity for the expressed recombinant alpha 1A-, but not alpha 1B- or alpha 1D-, adrenoceptor subtype, whereas, the potency of the alpha 1-adrenoceptor antagonists tested in the rabbit hypogastric artery was better correlated with the defined alpha 1D-adrenoceptor. Chloroethylclonidine produced a 10-fold rightward shift in the phenylephrine concentration-response curve in the hypogastric artery but only had a weak effect in the prostate. The results indicate that significant heterogeneity exists among alpha1-adrenoceptor in the rabbit hypogastric artery (alpha 1D-adrenoceptor) and the prostate (alpha 1A-adrenoceptor).

The endothelin receptors in rabbit isolated rings of saphenous artery and saphenous vein have been characterized using endothelin-1, endothelin-2, endothelin-3, sarafotoxin S6c, and BQ123. Although artery rings were more sensitive than those from vein to the contractile action of phenylephrine, endothelin-1 was about three times more potent as a contractile agonist on vein than on artery. In rings precontracted with phenylephrine, carbachol was 10 times more potent in vein than in artery rings to induce endothelium-dependent relaxation. However, in rings precontracted to a similar tone by endothelin-1, the relaxation elicited by carbachol was reduced in the vein but remained unchanged in the artery. In endothelium-denuded saphenous artery, endothelin-1 and endothelin-2 elicited contraction with equal potency, whereas endothelin-3 and sarafotoxin S6c were weak agonists. In saphenous vein, the rank order of sensitivity was sarafotoxin S6c > endothelin-2 > or = endothelin-1 = endothelin-3, whereas sarafotoxin S6c and, to a lesser extent, endothelin-3 act as partial agonists. The ETA receptor antagonist BQ123 shifted, to the right, the concentration-response curves of endothelin-1 on endothelium-denuded saphenous artery (pA2 = 7.25). In the endothelium-denuded saphenous vein, 10 microM BQ123 shifted to the right only the response to high concentrations of endothelin-1. In vein but not in artery, endothelin-1 and sarafotoxin S6c induced an endothelium-dependent relaxation, which was increased, in the case of endothelin-1, in the presence of BQ123.2+.

Inducible vascular nitric oxide synthase accounts for the contractile impairment observed in endotoxemia. We provide evidence that lipoteichoic acid (LTA) from Staphylococcus aureus, a micro-organism without endotoxin, also induces nitric oxide synthase. Our study demonstrates that on endothelium-free rings of rat aorta. LTA-like lipopolysaccharide induces a loss of contractility restored by Methylene blue and NG-nitro-L-arginine-methyl ester (LNAME). Moreover in cultured vascular smooth muscle cells, LTA produces a dose-dependent increase in intracellular cyclic GMP which is antagonized by LNAME and prevented by dexamethasone.

The effects of moderate cooling and of phenylarsine oxide on the contraction induced by two vasoactive peptides, angiotensin II (AII) and endothelin (ET-1), were investigated on endothelium-free rings of rat aortas. At 37 degrees C, the contraction induced by AII (0.1 microM) was transient. This decline in tension is unlikely to be due to rapid degradation of AII. In contrast, ET-1 (10 nM) induced a slowly developing and sustained contraction similar to the one observed with phorbol 12-13 dibutyrate (PDB, 22 nM). Moderate cooling (25 degrees C) significantly potentiated and prolonged the effect of AII but reduced the velocity of the ET-1 and PDB contraction, although the rate of the phenylephrine (1 microM) response remained unchanged. Phenylarsine oxide (100 microM) reduced the decline in tension in response to AII but inhibited the contraction elicited by ET-1 and PDB. In rings incubated in calcium-free medium (37 degrees C), AII induced a phasic contraction. This was followed by a second phasic contraction after calcium (2.5 mM) had been restored to the bath. The intensity of this second contraction decreased as the time between AII and calcium injection increased. This method, using regression analysis, permitted us to determine the time taken to reduce the contraction by half (4.8 min; r: 0.96), which may reflect the half-time of receptor sequestration. In calcium-free medium, the contractions induced by ET-1 and PDB were slow and sustained. Thus, rapid AII-receptor internalization leads to a short-term regulation of vascular tone whereas activation of protein kinase C by ET-1 may induce a long-term regulation.

The effects of NG-monomethyl-L-arginine (NMMA), a specific inhibitor of nitric oxide (NO) synthesis was tested on the endotoxin-induced alterations of alpha-adrenoceptor function. In isolated aorta, there was no significant difference in the tension induced by phenylephrine (PE, 10 microM) on rings removed from control and endotoxin injected rats (10 mg/kg, ip). However, a lack of tonicity of the contraction was observed in rings of shocked rats (8 +/- 2.9 and 86 +/- 4.6% relaxation at 105 min for sham and shocked rings respectively). The gradual tension decrease to PE was more potent in rings possessing endothelial cells. However, in both preparations, the loss of tonicity was significantly inhibited by NMMA (30 microM). In endothelium-free rings, L-arginine (100 microM) potentiated the loss of tonicity to PE and reversed the inhibitory effect of NMMA. NMMA, like methylene blue, was also able to restore the PE-contraction. The results indicate that the endotoxin-induced alterations of vascular reactivity may be due, in part, to NO formation from L-arginine independent of the endothelium.

The alterations of vascular reactivity in endotoxemia were investigated using the aorta from intraperitoneally endotoxin-injected rats (E. Coli 0111: B4; 10 mg/kg). After 3 hours, rings were removed and isometric contractions recorded. Using single maximal agonist concentration, similar contractions were observed in sham and shocked preparations for phenylephrine (PE), endothelin (ET-1) and phorbol 12, 13 dibutyrate (PDB) but not for KCl. Nevertheless, contraction elicited by PE lost tonicity (105 min observation). This fact was not due to endothelium, prostaglandins (since indomethacin and aspirin were ineffective) PE degradation (since medium contained, propranolol, hydrocortisone acetate, cocaine and EDTA) or excitation-contraction coupling fading (since contraction was restored by other agonists) but may involve synthesis of protein (since cycloheximide significantly antagonized this phenomenon). This loss of tonicity was also observed with ET-1 but was less pronounced with KCl and PDB. In conclusion, endotoxin induced impairment of vascular reactivity may occur through different pathways: contractile events (e.g. calcium influx as exemplified by KCl) and receptor events (loss of tonicity and/or desensitization as exemplified by PE and ET-1) from the different agonists tested, PDB, which activates the protein kinase C, was the less affected by endotoxin.

The vasoconstrictive properties of endothelin (ET-1) and the protein kinase C activator, phorbol 12-13 dibutyrate (PDB) were comparatively investigated in isolated rat aorta. ET-1 (0.3-100 nM) and PDB (10 nM-3 microM) induced a slowly developing sustained contraction in endothelium denuded aorta. Maximal contractions induced by ET-1 and PDB were unaffected by diltiazem (10 microM). Substantial contraction to ET-1 (30 nM) and PDB (0.1 microM) remained in calcium-free medium. Contractions of ET-1 and PDB in calcium-free medium were unaffected by intracellular calcium depletion induced by phenylephrine. Following the response to ET-1 and PDB in a calcium-free medium, an additional sustained contraction was observed after calcium (2.5 mM) was added to the bath. The protein kinase C inhibitor, H7 (100 microM) was more potent in inhibiting contractions induced by phenylephrine and KCl than the ones elicited by ET-1 and PDB. The other protein kinase C inhibitors i.e. staurosporine (50 nM) and phloretin (100 microM) inhibited to a similar extent all the agonists tested. These results suggest that protein kinase C may play an important role in mediating the contraction to ET-1 in rat aorta.

Spirally-cut strips of rabbit aorta were used to examine the relaxations produced by carbachol and extract of Ginkgo biloba (Gb) under isometric conditions. After precontracting the strips with phenylephrine (10(-7) M), carbachol produced a dose-related relaxation (PD2 congruent to 6.2 +/- 0.1) and this effect was antagonized competitively by atropine (PA2 congruent to 9.4 +/- 0.1). Gb (0.2 or 0.3 mg/ml) also relaxed the strips. Removal of the endothelium or a 30-min pre-treatment of the strips with a substance that has lipoxygenase-inhibitor activity (nordihydroguaiaretic acid, NDGA, 10(-5) M) abolished the relaxant effect of carbachol and partially blocked the relaxant effect of Gb. Thus, at least part of the relaxant effect of Gb is mediated by a factor(s) (e.g., EDRF) that is released from endothelial cells.

The effects of atrial natriuretic factor (ANF) on contractile events triggered by phenylephrine (PE) were investigated in rat aorta. Isometric contraction of endothelium-free rat aorta rings was recorded in Ca2+-free medium containing 1 mM EGTA. PE (1 microM) induced a phasic contraction and a sustained contraction following addition of Ca2+ (2.5 mM) to the medium. The phasic contraction was due to intracellular Ca2+ release whereas the sustained one was dependent on extracellular Ca2+ influx. ANF (1-3 nM) and prazosin (3-10 nM) both reduced the two types of contraction. However, during the sustained contraction, a rhythmic activity was observed when ANF, but not prazosin, was used as an inhibitory agent. The calcium antagonist diltiazem (0.1-1 microM) abolished this rhythmic activity, which was attributed to a Ca2+ influx through potential-dependent channels. The results indicate that ANF, unlike prazosin, may discriminate between PE-triggered contractile events in the rat aorta.

The contractile properties of endothelin (ENDO) and its interactions with some putative antagonists were investigated in endothelium free ring of rat aorta. ENDO induced a slowly developing contraction which is only partially affected by sodium nitroprusside (10(-8) M - 10(-5) M) and to a lesser extend by the calcium antagonist nifedipine (10(-8) M - 10(-5) M). Atrial natriuretic factor (ANF) (10(-9) M - 10(-8) M), cicletanine (3.10(-5) M - 3.10(-4) M) and quercetin (10(-5) M - 10(-4) M) induced a dose dependent relaxation in ENDO precontracted preparations. ANF was less effective in inhibiting ENDO- than phenylephrine precontracted aorta. In addition, the ANF vasodilating effect upon ENDO contraction is potentiated by cicletanine (10(-4) M). The protein kinase C inhibitor phloretin, induced a dose-dependent relaxation (10(-5) M - 3.10(-4) M) in both ENDO and phorbol 12, 13-dibutyrate precontracted aorta. Whereas H7 (10(-5) M - 3.10(-4) M) an other protein kinase C inhibitor was only effective in ENDO-induced contraction. These data indicate that in isolated rat aorta, the contraction induced by ENDO does not mainly occur through membrane potential-dependent calcium channels. The vasoconstrictor mechanism of ENDO, which is different from the one triggered by phenylephrine could involve activation of protein kinase C.

Cicletanine (CIC) (pA2: 7.0) was found to be as potent as diphenhydramine (DIPH) (pA2: 7.2) in competitively inhibiting histamine-induced contraction of isolated rabbit aorta. Both CIC (pA2: 7.3) and DIPH (pA2: 7.5) similarly shifted histamine-induced endothelium-dependent relaxation of isolated rat aorta to the right. The similarity of results (in terms of pA2 values) indicates that in spite of their opposite responses on vascular tone, the H1 receptors on rabbit and rat aortae may be similar. DIPH was found to be as potent as cocaine in potentiating the chronotropic action of noradrenaline on isolated right atria, whereas CIC was without effect. The ability of DIPH to inhibit the noradrenaline uptake at the neuronal membrane seems to be independent of its H1-receptor antagonism potency. Finally, the furopyridine framework of CIC does not elicit the cocaine-like activity of the oxyethylamine residue of DIPH.

1. Dose-response effects of captopril in an acute model of renovascular hypertension in the anaesthetized rat and in normotensive anaesthetized rats were compared. 2. Intravenous infusions of captopril lowered the blood pressure of both hypertensive and normotensive rats, its minimal active dose being about 0.05 mg/kg. 3. The 0.1 mg/kg dose of captopril normalized the blood pressure of hypertensive rats. 4. The model described herein might be useful for screening inhibitors of the angiotensin-converting enzyme.

The mechanism of vasoconstriction induced by endothelin was investigated in rat isolated aorta in comparison with the Ca++ agonist, Bay K 8644. Endothelin (EC50 = 4 nM) induced a slow and sustained contraction in control medium whereas the one elicited by Bay K 8644 (EC50 = 14 nM) necessitating a partly K+ depolarized medium was fast with superimposed rhythmic contraction. By opposition with Bay K 8644, endothelin contraction was not inhibited by the calcium antagonists (1 microM), nifedipine, diltiazem and D 600, and substantially persisted in Ca++ free medium or after depletion of intracellular Ca++ by phenylephrine (1 microM). These data show that endothelin does not act as an activator of potential dependent Ca++ channels but probably through specific receptor(s) as suggested by its mode of vasoconstriction.

Oral treatment for 2 weeks with cicletanine [1,3-dihydro-6-methyl-7-hydroxy-3-(4-chloro-phenyl)furo(3,4-c)pyridine] at 30 mg/kg/day delayed the onset of hypertension in spontaneously hypertensive rats (SHR) (15.7 mmHg, p less than 0.001). This antihypertensive effect was increased when the animals were maintained on a high-salt diet (40.8 mmHg, p less than 0.001). The ability of cicletanine to alter calcium movements in phenylephrine (PE)- and angiotensin II (ANGIO)-triggered contraction was tested on isolated SHR aorta. PE (1 microM) and ANGIO (0.1 microM) induced a phasic contraction in calcium-free medium due to intracellular calcium release. Upon addition of calcium (2.5 mM) a second sustained (PE) or biphasis (ANGIO) contraction was observed. This second contraction was dependent on extracellular calcium influx. Cicletanine (0.1-0.3 mM) both reduced the phasic (77%, p less than 0.001 for PE; 68%, p less than 0.05 for ANGIO with cicletanine 0.3 mM) and the second contraction elicited by the two agonists (27%, p less than 0.001 for PE; 84%, p less than 0.001 for ANGIO with cicletanine 0.3 mM). These results suggest that in addition to its stimulatory effect on prostaglandin, a direct vascular action of cicletanine could also be involved in the antihypertensive properties of the drug.

The interaction between cicletanine (CIC) and atrial natriuretic factor (ANF) on vessels was studied using the isolated rat aorta as model. Contractions induced by endothelin (ET1), a vasoconstrictor peptide from vascular endothelial cells, and by phenylephrine (PE), an alpha 1-adrenoceptor agonist, were recorded isometrically on aortic rings the endothelium of which had deliberately been damaged. Both agonists produced a concentration-dependent contraction (ET1:EC50 1.8 nM, Emax 2.2 g; PE:EC50 30 nM, Emax 2.4 g) which was antagonized by a 30 min pretreatment with either CIC (100 microM) or ANF (1 nM). However, PE was more sensitive than ET1 to the inhibitory action of CIC or ANF. Moreover, during inhibition by ANF spasmodic contractions were observed with PE but not with ET1; this was not observed when the antagonist was CIC. The inhibitory effect produced by the combination of CIC and ANF was additive with ET1 whereas a potentiation was observed with PE. Thus, on the isolated rat aorta model the contraction induced by PE was more sensitive to the action of ANF than that induced by ET1, which indicates that the two antagonists have a different mechanism of action. A direct action of CIC on the alpha-adrenoceptor might account for the potentiation observed between ANF and CIC against PE.

The actions of four Ca2+-antagonists (verapamil, diltiazem, nifedipine and flunarizine) were tested on serotonin- (5-HT-) induced contraction of rabbit isolated aorta. Verapamil produced a dose-dependent, parallel shift to the right of the concentration-response curve for 5-HT (pA2 approximately equal to 7.13; Schild slope approximately equal to 1.01), indicative of competitive antagonism. The effects of diltiazem, nifedipine and flunarizine were much less pronounced and best characterized as non-competitive interactions. The effect of verapamil, which likely involves its antagonism of smooth muscle 5-HT2-receptors, might be useful in explaining its therapeutic actions and/or its side-effects.

The vasoconstrictive properties of the endothelium-derived peptide, endothelin-1 (ET-1), were investigated on rat isolated aorta and on cultured rat aortic smooth muscle cells. In rat isolated aorta, endothelin-1 induced a slow and sustained contraction in a Ca2+-free medium; after calcium readmission, an additional sustained contraction was elicited. In vascular smooth muscle cells, endothelin-1 provoked a dose-dependent Ca2+ influx that was not inhibited by calcium entry blockers (nifedipine, D 600, or diltiazem). In these cells, [125I]-endothelin-1 bound to a specific, saturable, and high affinity recognition site (Kd about 10(-9) M and Bmax = 52 +/- 2 fmol/10(6) cells). The binding was not reversible and not affected by calcium antagonists. These data do not support the hypothesis that endothelin-1 acts as an endogenous agonist of the voltage-dependent Ca2+ channels. The action of endothelin-1 can be separated into two components: one dependent on Ca2+ influx but insensitive to calcium antagonists and another independent of extracellular Ca2+. The irreversible binding of endothelin-1 may reflect an internalization of the ligand inside the cell membrane, leading to multiple contractile events.

The preferential tissue irrigatory effect of Ginkgo biloba extract in ischaemic areas is largely explained by the direct impact of this product on both arteries and veins. The adrenergic vasoregulatory system and the vascular endothelium are the preferential targets for arterial impact. Ginkgo biloba extract reinforces the physiological vasoregulation of the sympathetic nervous system directly, by acting on neuromediator release, and indirectly, by inhibiting their extraneuronal degradation by catechol-orthomethyltransferase (C.O.M.T.) In the arterial endothelium Ginkgo biloba extract stimulates the release of endogenous relaxing factors, such as endothelium-derived relaxing factor, (EDRF) and prostacyclin. The action of Ginkgo biloba extract on the venous system has been shown to have a venoconstrictor component that maintains the degree of parietal tonus essential to the dynamic clearing of toxic metabolites accumulated during tissue ischaemia. The originality of the vascular impact mechanisms of Ginkgo biloba extract is due to the fact that the product can at the same time combat the phenomena resulting from vascular spasm and with the same efficiency restore circulation in areas subject to vasomotor paralysis.

The ability of endothelium to alter contractile events in phenylephrine (PE)-triggered contraction has been tested on ring segments of the thoracic aorta removed from obese Zucker rats (plasma cholesterol 3.63 mM; n = 8) and from age matched lean rats (plasma cholesterol 2.38 mM; n = 8). In normal medium, PE (1 microM) elicited similar contractions in endothelium-denuded arteries of both strains. However, the presence of endothelium reduced these contractile events and the endothelium-dependent relaxation induced by carbachol (10 microM) was higher in obese rats. In rings incubated in Ca2+ free medium containing EGTA (1 mM), PE (1 microM) induced a phasic contraction and a sustained contraction following addition of Ca2+ (2.5 mM) to the medium. The phasic contraction was due to intracellular Ca2+ release, whereas the sustained response was dependent on extracellular Ca2+ influx. In endothelium-free preparations, the size of both the phasic and sustained contraction was similar for the two strains. The Ca2+ antagonist gallopamil (1 microM) reduced the sustained contraction of lean (24%) and obese (34%) rats without affecting the phasic contraction. In preparations possessing endothelium, the sustained, but not the phasic contraction, of both strains was inhibited. This inhibitory effect of endothelium on the sustained contraction was significantly higher in obese than in lean rats. Thus, it can be concluded that phenylephrine elicited quantitatively and qualitatively similar contractions in obese and lean rats. In both strains, the endothelium diminished the contraction induced by PE, however, this effect was more pronounced in obese rats than in lean ones. These results may explain, in part, the described absence of atherosclerotic lesions in the obese strain.