OBJECTIVEIn our study, the function of the third-order branches of the mesentenc artery was measured by microvascular ring technique, which can be used to detect microvascular function in some disease related to microvascular dysfunction.

METHODSIsolated, fixed, standardized and then activated the third-order branches of rat mesenteric artery. Microvascular tone was measured by systolic and diastolic drags respectively, with the help of DMT tension apparatus and PowerLab data acquisition system.

RESULTSThe third-order branches of rat mesenteric artery showed excellent response to vasoactive drugs. The contraction effect of norepinephrine (NE) reached 19 in mN. When acetylcholine (Ach) or sodium nitroprusside (SNP) of 10(9)-10(5)mol/L was added, vascular tones showed gradient drop: 80% of maximal relaxation when adding ACh, while 95% of maximal relaxation when adding SNP.

CONCLUSIONThe third-order branches of the mesenteric artery function was successfully detected by using microvascular ring technique.

Acetylcholine ; pharmacology ; Animals ; In Vitro Techniques ; Male ; Mesenteric Arteries ; drug effects ; physiology ; Nitroprusside ; pharmacology ; Norepinephrine ; pharmacology ; Rats ; Vasoconstrictor Agents ; pharmacology ; Vasodilation ; physiology ; Vasodilator Agents ; pharmacology

The study is to investigate the effect of angiotensin II (Ang II) and its receptor blockers on migration and endothelin-1 (ET-1) expression of rat vascular adventitial fibroblast subpopulations. Vascular adventitial fibroblasts were individually expanded by using cloning rings, and the effects of Ang II on the migration of adventitial fibroblast subpopulations were evaluated by Transwell. Fluorescence quantitative-PCR detected the expression of preproET-1 mRNA induced by Ang II, and its receptor antagonists losartan and PD-123319. The concentration of ET-1 was determined by ELISA. It showed that spindle shaped and epithelioid shaped cells were isolated by using cloning rings, named as spindle cells and round cells. RT-PCR showed that fibroblast subpopulations did not have leukocytes, endothelial cells and smooth muscle cells, namely pure cell lines. Compared with respective control cells, two subpopulations had transferring ability. Ang II significantly improved round cells migration in a concentration-dependent manner, and had no obvious influence on spindle cells migration. Ang II (1 x 10(-8) - 1 x 10(-6) mol x L(-1)) significantly increased the expression of preproET-1 mRNA in round cells (P < 0.01), and had no significant effect on the expression of preproET-1 mRNA in spindle cells. Losartan blocked the expression of preproET-1 mRNA induced by Ang II in round cells, and had no significant effect on the expression of preproET-1 mRNA in spindle cells. The effects of Ang II and ET-1 receptor inhibitors on the release of ET-1 were similar to the expression of preproET-1 mRNA. The results indicate that there are two cell subpopulations: round cells and spindle cells in rat vascular adventitial fibroblasts. Ang II significantly improved cells migration, and increased the expression of ET-1 in round cell subpopulation. It suggested that there may be different migratory mechanisms in two cell subpopulations, and the two subpopulations may play a different role in vascular remodeling and reparative process.
Angiotensin II ; pharmacology ; Angiotensin Receptor Antagonists ; pharmacology ; Animals ; Cell Movement ; drug effects ; Cells, Cultured ; Endothelin-1 ; genetics ; metabolism ; Fibroblasts ; cytology ; metabolism ; Imidazoles ; pharmacology ; Losartan ; pharmacology ; Male ; Pyridines ; pharmacology ; RNA, Messenger ; metabolism ; Rats ; Rats, Sprague-Dawley ; Vasoconstrictor Agents ; pharmacology

This study is to investigate protective effect of safflor yellow B (SYB) against vascular endothelial cells (VECs) injury induced by angiotensin-II (Ang-II). VECs were cultured and divided into six groups: control group, Ang-II group, Ang-II + SYB (1 micromolL(-1)) group, Ang-II + SYB (10 micromolL(-1)) group, Ang-II + SYB (100 micromolL(-1)) group and Ang- II + verapamil (10 micromolL(-1)) group. Except control group, all of VECs in other groups were treated with Ang- II at the final concentration of 0.1 micromolL(-1). Mitochondria membrane potential (MMP) and free calcium concentration ([Ca2+]i) were measured by laser scanning confocal microscopy, and mitochondria complex IV activity was detected by BCA method. The levels of reactive oxygen species (ROS) in VECs were analyzed by fluorescence detector and apoptosis of VECs was observed by flow cytometer. Caspase 3 was determined by Western blotting method. Comparing with control group, Ang-II was able to increase [Ca2+]i and ROS level, decrease MMP level, inhibit complex IV activity and enhance caspase 3 activity in VECs, as a result, enhance apoptosis of VECs. But SYB could significantly reduce the result induced by Ang- II relying on different dosages (P < 0.05 or P < 0.01). SYB was able to eliminate the effect of Ang-II on VECs via regulating [Ca2+]i, mitochondrial structure and function and inhibiting apoptosis.
Angiotensin II ; adverse effects ; Antioxidants ; isolation & purification ; pharmacology ; Apoptosis ; drug effects ; Calcium ; metabolism ; Carthamus tinctorius ; chemistry ; Caspase 3 ; metabolism ; Cells, Cultured ; Chalcone ; analogs & derivatives ; isolation & purification ; pharmacology ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Electron Transport Complex IV ; metabolism ; Endothelial Cells ; cytology ; metabolism ; Humans ; Membrane Potential, Mitochondrial ; drug effects ; Mitochondrial Proton-Translocating ATPases ; metabolism ; Plants, Medicinal ; chemistry ; Reactive Oxygen Species ; metabolism ; Vasoconstrictor Agents ; adverse effects

The Chinese herbal medicine Tianma (Gastrodia elata) has been used for treating and preventing primary headache over thousands of years, but the exact pharmacological mechanism of the main bioactive ingredient gastrodin remains unclear. In present study, the effects of gastrodin on calcitonin gene-related peptide (CGRP) and phosphorylated extracellular signal-regulated kinase1/2 (pERK1/2) expression were observed in rat trigeminal ganglion (TG) after in vitro organ culture to explore the underlying intracellular mechanism of gastrodin on primary vascular-associated headache. CGRP-immunoreactivity (CGRP-ir) positive neurons count, positive area, mean optical density and integrated optical density by means of immunohistochemistry stain were compared at different concentrations of gastrodin, which was separately co-incubated with DMEM in SD rat TG for 24 hours. Only at 5 or 10 mmol L(-1) concentration, gastrodin demonstrated significantly concentration-dependent reduction of CGRP-ir (+) expression and its action closed to 1.2 mmol L(-1) sumatriptan succinate. While at 2.5, 20, and 40 mmol L(-1) concentration, gastrodin did not show remarkable effects on CGRP-ir (+) expression. The optimal concentration of gastrodin (5 and 10 mmol L(-1)) similarly inhibited CGRP-mRNA expression level separately compared with 1.2 mmol L(-1) sumatriptan succinate and 10 micromol L(-1) flunarizine hydrochloride, which was quantitatively analyzed by real-time PCR (RT-PCR). pERK1/2 level was examined by Western blotting after co-cultured with optimal concentration of gastrodin and effective specific ERK1/2 pathway inhibitors PD98059, U0126. The result indicated that gastrodin significantly reduced pERK1/2 protein actions similarly to ERK1/2 pathway specific blockade. It suggests ERK1/2 signaling transduction pathway may be involved in gastrodin intracellular mechanism. This study indicates gastrodin (5 and 10 mmol L(-1)) can remarkably reduce CGRP-ir (+) neuron, CGRP-mRNA and pERK1/2 expression level in cultured rat TG, with its actions similar to the effective concentration of sumatriptan succinate, flunarizine hydrochloride and specific ERK1/2 pathway blocker. The intracellular signaling transduction ERK1/2 pathway may be involved in the gastrodin reducing CGRP up-regulation in rat TG after organ culture.
Animals ; Benzyl Alcohols ; administration & dosage ; isolation & purification ; pharmacology ; Butadienes ; pharmacology ; Calcitonin Gene-Related Peptide ; genetics ; metabolism ; Dose-Response Relationship, Drug ; Flavonoids ; pharmacology ; Flunarizine ; pharmacology ; Gastrodia ; chemistry ; Glucosides ; administration & dosage ; isolation & purification ; pharmacology ; MAP Kinase Signaling System ; drug effects ; Male ; Mitogen-Activated Protein Kinase 1 ; antagonists & inhibitors ; metabolism ; Mitogen-Activated Protein Kinase 3 ; antagonists & inhibitors ; metabolism ; Nitriles ; pharmacology ; Organ Culture Techniques ; Plants, Medicinal ; chemistry ; RNA, Messenger ; Rats ; Rats, Sprague-Dawley ; Sumatriptan ; pharmacology ; Trigeminal Ganglion ; metabolism ; Vasoconstrictor Agents ; pharmacology ; Vasodilator Agents ; pharmacology

OBJECTIVE: To observe the effect of soft tissue crush injury on the tensions of thoracic aortic rings (TARs) in rats and to investigate the potential roles of nitric oxide in the change of the tensions. METHODS: Thirty adult SD rats were randomly divided into control group and crush injury (8 h and 16 h after injury) groups. Two kinds of TARs (one with endothelium and the other without endothelium) in vitro were prepared. In the TARs with endothelium, the tensions induced by phenylephrine (PE), acetylcholine (Ach), calcium ionophore A23187 and angiotensin II (AngI) were measured by the vascular tension detective technique. Then the TARs with endothelium were preincubated with nitric oxide synthase inhibitor N-nitro-L-arginine (L-NNA) for 20 minutes, the tensions induced by PE and Ang II were measured again. In the TARs without endothelium, the tensions induced by PE and Ang II were measured by the same method. RESULTS: In the TARs with endothelium, the tension of relaxation induced by cumulative doses of Ach and A23187 decreased significantly in 8 h and 16h groups. The tension of contraction induced by cumulative doses of PE and Ang II also decreased significantly (P<0.05). The tension of contraction increased after the preincubation with L-NNA. In the TARs without endothelium, the tension of contraction induced by PE and Ang II increased comparing to that of TARs with endothelium. CONCLUSION The soft tissue crush injury can influence the tensions of TARs in rats and the vascular-derived NO can mediate the effects.
Animals ; Aorta, Thoracic/physiopathology* ; Disease Models, Animal ; Endothelium, Vascular/metabolism* ; Female ; Hindlimb/injuries* ; Male ; Muscle, Smooth, Vascular/metabolism* ; NG-Nitroarginine Methyl Ester/pharmacology* ; Nitric Oxide/biosynthesis* ; Nitric Oxide Synthase/metabolism* ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Soft Tissue Injuries/physiopathology* ; Vasoconstriction/drug effects* ; Vasoconstrictor Agents/pharmacology* ; Vasodilation/drug effects* ; Vasodilator Agents/pharmacology*

OBJECTIVETo study the relaxant effects of glycyrrhizinate and salvianolic acid B on rat portal vein in vitro.

METHODSHealthy female Wistar rats were canalized from hepatic artery, portal vein and hepatic vein in vitro. Remained blood in liver was eliminated with heparinized Krebs-Henseleit solution through hepatic artery, then the liver was isolated under infusing manner. Being constricted with phenylephrine and relaxed with acetylcholine, and infused with glycyrrhizinate or salvianolic acid B, the portal pressures of infused rat livers were consistently monitored by BL-420S physiological experiment system. The median effective concentration (EC50) of the two agents were analyzed with non-linear various slope regression using Prism-4 software.

RESULTSEC50 of glycyrrhizinate in relaxing the phenylephrine-contracted portal was 1.5556 x 10(-9) mol/L, suggesting one of the mechanism of action of diammonium glycyrhizinate for the treatment of portal hypertension was direct relaxation. Salvianolic acid B showed constrictive action on the phenylephrine-retracted portal vein, the EC50 was 1.4639 x 10(-9) mol/L, indicating that its indirect control action was took part in the portal hypertension therapy synergistically.

CONCLUSIONUnder the mode with both controlled-velocity and monitored pressure, glycyrrhizinate showed relaxation and salvianolic acid B showed constriction on portal pressure in vitro.

Animals ; Benzofurans ; pharmacology ; Blood Pressure ; drug effects ; Female ; Glycyrrhizic Acid ; pharmacology ; Hypertension, Portal ; physiopathology ; Phenylephrine ; pharmacology ; Portal Vein ; physiology ; Rats ; Rats, Wistar ; Vasoconstrictor Agents ; pharmacology ; Vasodilator Agents ; pharmacology ; Vasomotor System ; drug effects

OBJECTIVETo investigate the effect of endothelin receptors in chronic venous insufficiency (CVI) in lower extremities.

METHODSTen cases of varicose veins from CVI patients (as case group) and ten cases of non-varicose veins (as control group) were investigated in this study. The two groups were divided into two groups respectively: endothelium-intact group and de-endothelium groups. The vasoconstriction mediated by endothelin A (ETA) and endothelin B (ETB) receptors was recorded with myography. The distribution of ETA and ETB receptors was detected by immunohistochemistry method.

RESULTSEndothelin-1 (ET-1) and sarafotoxin 6c (S6c) induced concentration-dependent contraction in the veins. In endothelium-intact veins, the E(max) and pD(2) of contraction curve induced by ET-1 were 132.30% +/- 43.42% and 6.03 +/- 0.35, respectively in control group;and were 19.24% +/- 12.94% and 6.78 +/- 0.46, respectively in case group. The E(max) and pD(2) in case group were much lower than in control group (P < 0.05). The E(max) and pD(2) induced by S6c were 30.10% +/- 12.90% and 6.54 +/- 0.36, respectively in control group, and were 9.61% +/- 1.32% and 6.75 +/- 0.29, respectively in case group; The E(max) in case group was lower than in control group (P < 0.05). In de-endothelium veins, E(max) and pD(2) of S6c were 146.18% +/- 32.33% and 6.50 +/- 0.17 in control group, and 32.93% +/- 3.00% and 6.69 +/- 0.39 in case group; The E(max) in case group was significantly lower than in control group (P < 0.05). ETA receptors was located in endothelium mainly, and ETB receptors in smooth muscle cells mainly. The sites of both ETA and ETB receptors were decreased in case group obviously.

CONCLUSIONSThe contraction mediated by ETA receptor and ETB receptor was decreased with a decrease of ETA receptor and ETB receptor sites in varicose veins of CVI. The contraction insufficiency and down-expression of ETA receptor and ETB receptor are correlated with CVI.

Adult ; Endothelin-1 ; pharmacology ; Humans ; In Vitro Techniques ; Lower Extremity ; blood supply ; Male ; Middle Aged ; Receptors, Endothelin ; drug effects ; physiology ; Vasoconstriction ; drug effects ; physiology ; Vasoconstrictor Agents ; pharmacology ; Venous Insufficiency ; physiopathology ; Viper Venoms ; pharmacology

OBJECTIVETo investigate the vasoconstriction effect of Ixeris sonchifolia in rat thoracic aortic rings and the underlying mechanisms.

METHODI. sonchifolia 10-160 g x L(-1) was cumulatively added into organ bath to observe the isometric tension of thoracic aortic rings with intact endothelium or denuded endothelium in basal tension, preconstricted by phenylephrine (PE) or potassium chloride (KCl), and thoracic aortic rings with intact endothelium preincubated frist with captopril, phosphoramidon and indomethacin, respectively, then preconstricted by PE and KCl. The response was recorded and expressed by "relative contraction".

RESULTCumulative administration of I. sonchifolia 10-160 g x L(-1) did not affect the vasomotion of aortic rings with endothelium or without endothelium in basal tension. Exposure of intact endothelium rings preconstricted by PE or KCl to I. sonchifolia at concentration (20-160 g x L(-1) induced a significant constriction, which was inhibited by preincubation with captopril, but was not inhibited by preincubation with phosphoramidon or indomethacin. Exposure of endothelium-denuded rings preconstricted by PE or KCl to I. sonchifolia at concentration (10 to approximately 160 g x L(-1) did not effect the vasoconstriction.

CONCLUSIONThe results indicate that I. sonchifolia (20 to approximately 160 g x L(-1) can contract the rat thoracic aortic rings with endothelium. The effect of contraction may enhance angiotensin converting enzyme activity and promote endothelium to synthesize angiotensin II. It has no relationship to endothelin or thromboxane A2.

Angiotensin-Converting Enzyme Inhibitors ; pharmacology ; Animals ; Aorta, Thoracic ; drug effects ; physiology ; Asteraceae ; chemistry ; Captopril ; pharmacology ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Endothelium, Vascular ; physiology ; In Vitro Techniques ; Male ; Phenylephrine ; pharmacology ; Plants, Medicinal ; chemistry ; Rats ; Rats, Sprague-Dawley ; Vasoconstriction ; drug effects ; Vasoconstrictor Agents ; pharmacology

AIMTo investigate the relaxation mechanisms of neferine (Nef) on the rabbit corpus cavernosum tissue in vitro.

METHODSStrips of rabbit corpus cavernosum were mounted in organ chambers. The effects of Nef were examined on isolated muscle strips precontracted with phenylephrine (PE) alone, in the presence of N(W)-nitro-L-arginine (LNNA, a nitric oxide synthase inhibitor), 1-H-[1,2,4]oxadiazolo[4,3-alpha]quinoxalin-1-one (ODQ, a guanylyl cyclase inhibitor), indomethacin (cyclooxygenase inhibitor), tetraethylammonium (Ca(2+)-activated K(+) channel blocker), 4-aminopiridine (4-AP, voltage dependent K(+) channel blocker) and glibenclamide (ATP sensitive K(+) channel blocker). The effects of Nef on KCl-induced contraction of isolated muscle strips were also investigated. The procedure of calcium absence-calcium addition was designed to observe the effect of Nef on two components of the contractile responses to PE based on the source of Ca(2+) (extracellular vs. intracellular).

RESULTSCorpus cavernosum strips relaxed in response to Nef (10(-9)-10(-4) mol/L) in a concentration-dependent manner with an IC(50) of 4.60 X 10(-6) mol/L. However, they were not affected by LNNA, ODQ, indomethacin or K(+)-channel blockers. Nef (10(-6) mol/L, 10(-5) mol/L) concentration dependently reduced the maximal contraction response of isolated strips induced by KCl to 79.3%+/-5.5% and 61.5%+/-3.2%, respectively (P < 0.01). In the calcium absence-calcium addition procedure, Nef 10(-5) mol/L inhibited both intracellular calcium-dependent and extracellular calcium-dependent contraction induced by PE (2 X 10(-5) mol/L) (P < 0.05). The inhibition ratios were 26.2%+/-5.4% and 48.3%+/-7.6%, respectively.

CONCLUSIONThe results of the present study suggest that Nef possesses a relaxant effect on rabbit corpus cavernosum tissues, which is attributable to the inhibition of extracellular Ca2+ influx and the inhibition of release of intracellular stored Ca(2+), but not mediated by the release of nitric oxide, prostaglandins or by the activation of potassium channels.

Animals ; Benzylisoquinolines ; pharmacology ; Calcium ; pharmacology ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; pharmacology ; Male ; Muscle Contraction ; drug effects ; Muscle Relaxation ; drug effects ; Penis ; drug effects ; physiology ; Phenylephrine ; pharmacology ; Potassium Chloride ; pharmacology ; Rabbits ; Vasoconstrictor Agents ; pharmacology

AIMTo determine the involvement of extracellular signal-regulated kinase 1/2 (ERK1/2) pathway in the expression of endothelin receptor type B (ETB) during culture.

METHODSSB386023, a specific inhibitor for ERK1/2 pathway, was used to define the intracellular signaling pathway for the upregulation of ETB receptors and sarafotoxin 6c (S6c), a selective agonist for ETB receptors, induced contraction in isolated rat superior mesenteric arteries. The contraction was recorded by a sensitive in vitro myograph and the receptor mRNA was quantified by a real-time PCR. The phosphorylated ERK1/2 proteins were analyzed by phosphoELISA assay.

RESULTSS6c induced strong contractile responses of the artery after culture for 24 h, while there was no response to S6c in fresh vessel segments. The enhanced contractile response to S6c paralleled with an increase of mRNA for ETB receptors. The phosphorylated ERK1/2 proteins significantly increased after culture for 3 h. After co-culture with SB386023 for 24 h, S6c-induced contractions significantly decreased with reduction of Emax from (217 +/- 14) % to (127 +/- 23) % (P <0.01). This response paralleled with a decreased level of ETB receptor mRNA.

CONCLUSIONERK1/2 pathway was involved in the up-regulation of ETB receptors on smooth muscle cells isolated from rat mesenteric arteries during culture.

Animals ; Cells, Cultured ; Male ; Mesenteric Arteries ; cytology ; Mitogen-Activated Protein Kinase 1 ; antagonists & inhibitors ; metabolism ; Mitogen-Activated Protein Kinase 3 ; antagonists & inhibitors ; metabolism ; Muscle Contraction ; drug effects ; Muscle, Smooth, Vascular ; cytology ; metabolism ; Organ Culture Techniques ; Phosphorylation ; RNA, Messenger ; biosynthesis ; genetics ; Rats ; Rats, Sprague-Dawley ; Receptor, Endothelin B ; biosynthesis ; genetics ; Signal Transduction ; Up-Regulation ; Vasoconstrictor Agents ; pharmacology ; Viper Venoms ; pharmacology