The effects of botulinum toxin A and papaverine on human saphenous vein and internal mammary artery grafts: an in vitro study.

Introduction: Autologous saphenous vein (SV) and internal mammary artery (IMA) are used as bypass conduits during coronary artery bypass graft surgery. Vasospasm of the arterial and venous grafts may constitute a significant clinical problem. Pretreatment with a vasodilator drug of the graft ex vivo or intraluminal injection before implantation may be used for spasm prophylaxis. This in vitro study was designed to assess the vasoactive effects and time-dependent changes of botulinum toxin A (BTX-A) and papaverine pretreatment on vasospasm of human SV and IMA grafts. Also, histomorphology of the vessels was assessed. Material and methods: SV and IMA segments were suspended in organ baths, and isometric contraction responses to 2 different concentrations of 5-hydroxytryptamine (5-HT) and endothelin-1 (ET-1) were recorded after incubation with 2 different concentrations of BTX-A and papaverine at 2 time points (0 h and 2 h). Results: The results revealed the following: 1) incubation with BTX-A and papaverine relaxes both SV and IMA rings contracted with 5-HT and ET-1; 2) the duration of the relaxant effect of BTX-A lasts longer than papaverine; and 3) no apparent histomorphological changes were observed in the grafts under light microscopy. Conclusions: This study demonstrates that in human SV and IMA grafts, pretreatment with both BTX-A and papaverine are safe and have a potent inhibitory effect depending on the vessel and vasoconstrictor agent. The long-lasting vasodilatory effect of BTX-A on vascular smooth muscle may provide promising results in the prevention of venous and arterial graft spasm.

Neuroprotective Effect of Intrastriatal Caffeic Acid Phenethyl Ester Treatment in 6-OH Dopamine Model of Parkinson's Disease in Rats.

Parkinson's disease (PD) is the second most common neurodegenerative disorder, and the main cause of PD is still not known. Until now, no cure for Parkinson's disease is yet in sight. Caffeic acid phenethyl ester (CAPE) is a polyphenolic component of the propolis, which can be derived from honeybee hive propolis. We aimed to determine the effect of intrastriatal CAPE administration as a neuroprotective agent on 6-hydroxydopamine (6-OHDA)-induced PD model. Adult male Wistar rats weighing 280-320 g were used. The PD model was induced with unilateral intrastriatal 6-OHDA injection. Treatment groups received 20 µmol/5 µL/4 day and 80 µmol/5 µL/4 day CAPE 24 h after 6-OHDA injection. Eight days after 6-OHDA application, behavioral studies (adhesive tape removal test, open-field test, cylinder test, and apomorphine-induced asymmetric rotational behavior) were performed once more to compare the effects of CAPE on behavior tests. Striatal histological verifications, immunohistochemistry, and stereological quantitation were performed. Our results for the first time showed that, besides improving the motor performance, CAPE treatment also prevents 6-OHDA-induced loss of TH-positive neurons. From our results, CAPE may be a promising clinical agent in the treatment of PD.

Relaxation matters: comparison of in-vitro vasodilatory role of botulinum toxin-A and papaverine in human radial artery grafts.

BACKGROUND: Radial artery (RA) is widely used in coronary artery bypass (CABG) surgery and the prevention of spasm is crucial for graft patency. Botulinum toxin A (BTX-A) and B are commonly used for aesthetic reasons and neuromuscular disorders. They are proven to raise blood flow and increase survival of ischemic skin flaps. In this study we evaluated and compared the vasodilator effects of BTX-A and papaverine on human RA grafts. METHODS: After resting 60 min in isolated organ baths, human RA grafts were examined. Contraction responses for different doses of serotonin (5-HT) and endothelin-1 (ET-1) were evaluated as a percent of maximum contraction response elicited by 80 mM potassium chloride (KCl). The inhibitory effects of BTX-A and papaverine on contraction responses taken at the 0th hour were compared with the 1st and 2nd hour responses. Inhibitory effects of BTX-A and papaverine against the contractile agent were evaluated by comparing the results of the first and last (0th and 2nd hour) application. RESULTS: In low concentrations, when we compared the effects of BTX-A (10- 8 M) and papaverine (10- 6 M) on 5-HT, papaverine was found to be more effective at both the 0th and 2nd hour (p < 0.05). Both BTX-A and papaverine inhibited the maximum contractile effect of ET-1 to the same extent at the 0th hour; but, the inhibitory effect of BTX-A was significantly stronger at the 2nd hour (p < 0.05). In high concentrations, when we compared the effects of BTX-A (10- 6 M) and papaverine (10- 4 M) on 5-HT, papaverine showed stronger inhibition (p < 0.05), whereas both agents had similar action of inhibition on ET-1 mediated maximum contraction responses. CONCLUSION: BTX-A inhibits both ET-1 and 5-HT induced contractions and its effectiveness does not decrease over time as observed with papaverine. This study is the first in the literature using human RA for prevention of vasospasm by BTX-A.

Behavioural effects of basal ganglia rho-kinase inhibition in the unilateral 6-hydroxydopamine rat model of Parkinson's disease.

Parkinson's disease (PD) is one of the most common neurodegenerative disorders, which affects more than six million people in the world. While current available pharmacological therapies for PD in the early stages of the disease usually improve motor symptoms, they cause side effects, such as fluctuations and dyskinesias in the later stages. In this later stage, high frequency deep brain stimulation of the subthalamic nucleus (STN-DBS) is a treatment option which is most successful to treat drug resistant advanced PD. It has previously been demonstrated that activation of Rho/Rho-kinase pathway is involved in the dopaminergic cell degeneration which is one of the main characteristics of PD pathology. In addition, the involvement of this pathway has been suggested in diverse cellular events in the central nervous system; such as epilepsy, anxiety-related behaviors, regulation of dendritic and axonal morphology, antinociception, subarachnoid haemorrhage, spinal cord injury and amyotrophic lateral sclerosis. However, up to date, to our knowledge there are no previous reports showing the beneficial effects of the potent Rho-kinase inhibitor Y-27632 in the 6-hydroxydopamine (6-OHDA) rat model of PD. Therefore, in the present study, we investigated the behavioural effects of basal ganglia Y-27632 microinjections in this PD model. Our results indicated that basal ganglia Y-27632 microinjections significantly decreased the number of contralateral rotations-induced by apomorphine, significantly increased line crossings in the open-field test, contralateral forelimb use in the limb-use asymmetry test and contralateral tape playing time in the somatosensory asymmetry test, which may suggest that Y-27632 could be a potentially active antiparkinsonian agent.

Protective effects of Hawthorn (Crataegus oxyacantha) extract against digoxin-induced arrhythmias in rats.

OBJECTIVE: Digitalis preparations are commonly used by children and adults with heart diseases worldwide, although excessive doses may cause cardiac effects. The aim of the study is to evaluate the antiarrhythmic effect of Crataegus oxyacantha extract on digoxin-induced arrhythmias in anesthetized Wistar rats. METHODS: Control and experimental groups were evaluated for arrhythmias induced by digoxin. Fifteen rats (7 as controls and 8 as the experimental group) were included in the study. The dry fruits of 100 mg Crataegus oxyacantha were extracted by percolation method. Digoxin, at a dose of 40 µg/kg/min, was infused to form the arrhythmias in all rats. Simultaneously, the extract was infused into the experimental group, while 0.9% NaCl was infused into control group. Electrocardiographic QRS prolongation and arterial blood pressure changes were analyzed. RESULTS: The experimental group lived longer (62.13±2.20 min) than the controls (p=0.002). On the other hand, the time to beginning of QRS prolongation did not differ between the two groups (p=0.812). Bradycardia was significant in the control group (288.01±10.54 beat/min and p=0.01). The maximum QRS duration was observed in the control group during the digoxin and 0.9% NaCl infusion period (53.29±3.99 ms and p=0.001). Also, the durations of atrial and ventricular arrhythmias were shorter in the experimental group. However, arterial blood pressure dipping was significant in the experimental group (23.67±10.89 mm Hg and p<0.001). CONCLUSION: Crataegus oxyacantha alcoholic extract produced an antiarrhythmic effect that was induced by digoxin in Wistar rats. However, in the clinical use of this extract, the hypotensive effect should be considered. Also, the alcoholic extract of Crataegus oxyacantha may be an alternative treatment medication for arrhythmias induced by digoxin toxicity in humans.

Infralimbic cortex Rho-kinase inhibition causes antidepressant-like activity in rats.

Depression is one of the most common psychiatric disorders in the world; however, its mechanisms remain unclear. Recently, a new signal-transduction pathway, namely Rho/Rho-kinase signalling, has been suggested to be involved in diverse cellular events in the central nervous system; such as epilepsy, anxiety-related behaviors, regulation of dendritic and axonal morphology, antinociception, subarachnoid haemorrhage, spinal cord injury and amyotrophic lateral sclerosis. However there is no evidence showing the involvement of Rho-kinase pathway in depression. In addition, the infralimbic cortex, rodent equivalent to subgenual cingulate cortex has been shown to be responsible for emotional responses. Thus, in the present study, intracranial guide cannulae were stereotaxically implanted bilaterally into the infralimbic cortex, and the effects of repeated microinjections of a Rho-kinase (ROCK) inhibitor Y-27632 (10 nmol) were investigated in rats. Y-27632 significantly decreased immobility time and increased swimming and climbing behaviors when compared to fluoxetine (10 µg) and saline groups in the forced swim test. In addition, Y-27632 treatment did not affect spontaneous locomotor activity and forelimb use in the open-field and cylinder tests respectively; but it enhanced limb placing accuracy in the ladder rung walking test. Our results suggest that Y-27632 could be a potentially active antidepressant agent.

Cardiovascular effects of resveratrol and atorvastatin treatments in an H2O2-induced stress model.

Oxidative stress has been implicated in the pathophysiology of several types of cardiovascular disease (CVD). Statins are widely used to inhibit the progression of atherosclerosis and reduce the incidence of CVD. Certain over-the-counter products, including resveratrol, show similar effects to statins and may thus be used in conjunction with statins for the treatment of the majority of patients with CVD. The aim of the present study was to evaluate the effects of atorvastatin, resveratrol and resveratrol + atorvastatin (R+A) pretreatment on myocardial contractions and vascular endothelial functions in the presence of H2O2 as an experimental model of oxidative stress in rats. Four groups were established and referred to as the control, atorvastatin, resveratrol and R+A groups. Atorvastatin (40 mg/kg, per oral) and/or resveratrol (30 mg/kg, intraperitoneal) treatments were administered for 14 days. On the 15th day, the thoracic aortas and hearts of the rats were dissected and placed into isolated organ baths. Vascular responses to cumulative doses of H2O2 (1×10-8-1×10-4 M H2O2) with and without N (G)-nitro-L-arginine methyl ester (L-NAME) incubation were measured. In addition, myocardial electrical stimulation (ES) responses to various H2O2 concentrations (1×10-7-1×10-5 M H2O2) were evaluated. In the control and atorvastatin groups, H2O2 application caused a significant dose-dependent decrease in the ES-induced contractions in the myocardial tissue of rats. In the resveratrol and R+A groups, H2O2 application did not significantly affect myocardial contraction at any dose. In all groups, incubation with L-NAME caused a significant augmentation in the H2O2 response, revealing that this effect was mediated via the vascular endothelium. In conclusion, pretreatment with R+A for CVD appears to be superior to pretreatment with either agent alone.

The effect of supplementation of grape seed proanthocyanidin extract on vascular dysfunction in experimental diabetes.

Increased oxidative stress and impaired endothelium-dependent relaxation could underlie many of the vascular complications associated with diabetes. We aimed to investigate the effect of supplementation with grape seed proanthocyanidin extract (GSPE), a natural antioxidant, on vascular responses and oxidative stress in streptozotocin-induced diabetic rats. Male Sprague-Dawley rats were divided into three groups: control rats, untreated diabetic rats, and GSPE (100 mg/kg, for 6 weeks)-supplemented diabetic rats. Thoracic aorta rings of the rats were mounted in organ baths, and relaxant responses to acetylcholine (ACh), A23187, and sodium nitroprusside (SNP) were assayed in tissues precontracted with 60 mM KCl. Plasma samples used for the measurement of malondialdehyde (MDA) level and superoxide dismutase (SOD) activity. The endothelium-dependent relaxations in response to ACh and A23187 were impaired, but endothelium-independent relaxation in response to SNP did not change in diabetic rats. Supplementation with GSPE significantly improved the relaxant responses to ACh and A23187. The MDA level was significantly elevated and the plasma SOD activity was decreased in diabetic rats, but supplementation with GSPE attenuated the elevated MDA levels and increased plasma SOD activity. Thus supplementation of GSPE may attenuate oxidative stress through the inhibition of lipid peroxidation and may restore endothelial function and reduce the risk of vascular disease in diabetes.

Preconditioning with levosimendan prevents contractile dysfunction due to H2O2-induced oxidative stress in human myocardium.

We studied the inotropic and possible antioxidant effects of levosimendan in human atrial strips, before and after induction of oxidative stress induced by H2O2. Levosimendan (10(-9) to 10(-6) M) increased contractions induced by electrical stimulation (ES) in human atrial strips. The maximal positive inotropic effect of levosimendan was 145.6 +/- 4.6% of predrug values. H2O2 (10(-6) to 10(-3) M) significantly reduced contractions induced by ES. The maximum inhibition by H2O2 on the ES induced contraction was 47.2 +/- 3.5%. Levosimendan significantly increased the isometric contractions induced by ES when compared with the values obtained in the presence of 10 M H2O2 by 89.0 +/- 4.7%, 98.9 +/- 3.4%, and 111.2 +/- 3.7% at 10(-8), 10(-7), and 10(-6) M concentrations, respectively. In concentrations of 10(-7) and 10(-6) M levosimendan, the maximum responses to ES increased when compared with the values obtained in the presence of 10(-3) M H2O2 by 87.1 +/- 3.6% and 95.1 +/- 5.3%, respectively. The cumulatively applied H2O2 (10(-6)-10(-3) M) did not change the positive inotropic response to levosimendan. In conclusion, levosimendan reverses the myocardial dysfunction induced by oxidative stress in human right atrial strips. Levosimendan prevents myocardial dysfunction if administered before oxidative stress.

The mechanisms of vasorelaxant effect of leptin on isolated rabbit aorta.

In the present study, we analysed the effects of leptin on rabbit aorta and the mechanisms underlying these effects. Leptin (10(-12)-10(-9) m) induced concentration-dependent relaxation in intact rabbit aorta rings precontracted with phenylephrine (10(-6) m). Removal of endothelium abolished the effects of leptin. Pretreatment of rings with N(omega)-nitro-l-arginine methyl ester (10(-4) m 20 min) or catalase (1200 U/mL 20 min) significantly reduced the relaxant response to leptin when compared with the control group. The incubation of brefeldin A (3.5 x 10(-5) m 90 min), indomethacin (10(-5) m 20 min), tetraethylammonium (10(-4) m 20 min), and glibenclamide (10(-5) m, 20 min) did not affect the leptin-induced vasodilation. These results suggest that leptin relaxes the rabbit aorta. The relaxation is mediated by endothelium-derived nitric oxide and hydrogen peroxide.

Effect of cooling on the responses of human saphenous vein to fentanyl, remifentanil and sufentanil.

We studied the vasodilatory effects of fentanyl, remifentanil and sufentanil on the human saphenous vein strips at 37, 32 and 28 degrees C. Fentanyl produced concentration-dependent relaxation of human saphenous vein strips precontracted with 5-hydroxytryptamine (5-HT) at every temperature studied. Compared with vein strips at 37 degrees C, relaxant responses to each one concentration of fentanyl were significantly reduced at 32 and 28 degrees C. Remifentanil relaxed vein strips in a concentration-dependent way and the relaxation for all concentrations were significantly greater at 32 and 28 degrees C compared with 37 degrees C. Sufentanil produced concentration-dependent relaxation in saphenous vein strips precontracted with 5-HT. These relaxant responses were similar at 32 degrees C compared with 37 degrees C. When bath temperature was lowered from 37 to 28 degrees C, the relaxant responses to sufentanil were significantly reduced. In summary, the present study suggests that cooling reduces the relaxation caused by fentanyl and sufentanil on human saphenous veins but augments the relaxation with remifentanil. The augmented vasodilatory effect of remifentanil with cooling may be useful on systemic vascular resistance and organ preservation under hypothermic conditions like cardiopulmonary bypass surgery.

The mechanisms of the direct vascular effects of fentanyl on isolated human saphenous veins in vitro.

OBJECTIVE: The purpose of this study was to determine the mechanism of the direct effects of fentanyl on human veins in vitro. DESIGN: In vitro, prospective with repeated measures. SETTING: University research laboratory. INTERVENTIONS: Dose-response curves were obtained for cumulative doses of fentanyl (10(-9)-10(-5) mol/L) on saphenous vein strips precontracted with (10(-6) mol/L) 5-hydroxytryptamine incubated with either naloxone (10(-4) mol/L), Nomega-nitroL-arginine-methyl ester (L-NAME) (10(-4) mol/L), indomethacin (10(-5) mol/L), glibenclamide (10(-4) mol/L), tetraethylammonium (10(-4) mol/L), or ouabain (10(-5) mol/L). Vein strips were also exposed to a Ca++-free solution and 0.1 mmol/L of ethylene glycol-bis-(b-aminoethylether) N,N'-tetraacetic acid; 5-hydroxytryptamine (10(-6) mol/L) was added to the bath before cumulative Ca++ (10(-4)-10(-2) mol/L). The same procedure was repeated in the presence of fentanyl (10(-6) , 3 x 10(-6) , or 10(-5) mol/L) (p < 0.05 = significant). MEASUREMENTS AND MAIN RESULTS: Preincubation of vein strips with naloxone, L-NAME, or indomethacin did not influence the relaxant responses to fentanyl (p > 0.05). Tetraethylammonium, glibenclamide, and ouabain reduced the relaxation response to fentanyl (p < 0.05). A stepwise increase in tension was recorded with cumulative doses of Ca++ (p < 0.05). CONCLUSIONS: The present results show that fentanyl causes vasodilatation via both endothelium- and opioid receptor-independent mechanisms in the human saphenous vein. The relaxant effects of fentanyl are probably via activation of K+ channel and Na+K+-adenosine trisphosphatase and inhibition of Ca++ channel.

Cooling and response to hydrogen peroxide in human saphenous vein: role of the endothelium.

In the present work we studied the responses of human saphenous vein to H2O2 and effects of moderate cooling on these responses with analysis of the role of endothelium. H2O2 (10(-7)-10(-2) M) induced concentration-dependent contraction in the intact human saphenous vein strips at both temperatures. At 28 degrees C, the maximal contraction induced by H2O2 was significantly lower than that at 37 degrees C. Compared with intact strips, the sensitivity and the maximal contraction to H2O2 were significantly enhanced in endothelium-denuded strips at 37 and 28 degrees C. However, pD2 values and maximal contractions were not significantly different in endothelium-denuded strips at different temperatures. Pretreatment with N(G)-nitro-L-arginine methyl ester (L-NAME) increased significantly the maximal contraction and sensitivity to H2O2 at 37 and 28 degrees C. The contractions increased by L-NAME were restored by the pre-incubation of l-arginine (10(-3) M) at every temperature studied. The contractile responses of intact human saphenous veins to H2O2 were reduced significantly by 10(-5) M indomethacin at both temperatures. Our results suggest that H2O2-induced contraction of human saphenous vein are mediated by its direct effect on the smooth muscle and by the generation of products of the cyclooxygenase pathway from the endothelium. Signalling pathways of these contractile effects are the same at 37 and 28 degrees C. Under normal temperature conditions, the contraction to H2O2 is possibly modulated by endothelial nitric oxide. Cooling reduces the contraction to H2O2 by increasing release of nitric oxide.