In normal (5mM KCI) HEPES buffer solution, BHT-920 and rauwloscine did not produce any contractile responses in dog mesenteric artery strips. However, when the preparation was bathed in 20mM KC1 HEPES buffer solution, BHT-920 and rauwloscine evoked significant contractile responses. These effects were markedly inhibited by parazosin which caused a parallel shift to the right of the concentration-response curve to BHT-920. In 45Ca uptake experiments carried out in the 20mM KC1 HEPES buffer solution BHT-920 and rauwolscine significantly increased 45Ca influxes which were reduced by prazosin. These results suggest that postsynaptic al-adrenoceptors in dog mesenteric artery mediate the contractile responses and the 45Ca influxes induced by BHT-920 and rauwloscine after partial depolarization by 20mM KC1.

We determined the antiarrhythmic effect of Rbl (from Panax notoginsengl on different ar rhythmia models and its effect on cytoplasmic Ca2+ concentrations ([Ca2+]i) in rat heart cells with fura-2 fluorescence. At doses of 30 and 50mg/Kg, Rb1 produced an antirrhythmic effect on BaClz-induced ven tricular tachyarrhythmia in rats, atria (fibrillation induced by CaCl2-ACh, and ventricular fibrillation induced by chloroform in mice. The rest [Ca22]i was 108+10.7nM in the freshly isolated rat heart cells. Sixty mM KCl caused an increase in [Ca2+]i to 379.4±77nM. Rb1 from 0.1 to 0.8mM significantly reduced this increase in a concentration-dependent manner, mM Rb1 (0.4mM) also completely inhibited the increase in [Ca2+]i induced by 1 μM isoprenaline. The radioligand binding study in rat heart cells showed that Rb1 did not change the Bmax. and Kd values of [3H]-dihydroalprenolol binding. These data suggest Rb1 can inhibit Ca2+ entry through voltage-dependent and receptor-linked Ca2+ channels, and that this is related to its antiarrhythmic effect, Drug Dev. Res. 39: 179-183.

Newborn male Wistar rats were treated with nerve growth factor daily by subcutaneous inJection for 2 weeks, and control rats were treated with either cytochrome c or buffered saline. Average body weight of the treated animals was lower than that of the controls during the 2 weeks of treatment, but became simdar to that of the controls thereafter. Tissue levers of norepinephrine were elevated in the brain, adrenal glands, nlesernteric arteries, and vas deferens of the treated animals immediately after the treatment, but became simliar in the three grcalps 2 weeks after tbe terminndon of tbe treatment. Blood pressure and heart tale were measured beginning at 4 weeks of age until 28 becks, when the rats were sacrificed and the mesenteric arteries sampled for morphometric measurements of vessel wall dimensions. Pretreatrnent with nerve growth fac tor did not affect blood pressure, nor heart rate. Stroctural atteration of the three types of mesenteric arteries was also absent in the treated animals. We conclude that even though neonatal treatment of normal Wistar rats with nerve growth factor for 2 weeks induced an elevation of the norepinephrine levels in severat tissues at the end of the treatment period, it was not sufficient to produce hypertension and structural alterations in the blood vessels.

1. The effects of chebulinic acid, which has been shown to elicit blood pressure lowering effect in rats, on aortic vascular contraction as well as cardiac contraction were studied in rats. 2. Chebulinic acid had no effect on KCI-induced aortic contraction, but irreversibly inhibited the contractile responses to phenylephrine in an apparently non-competitive manner. Chebulinic acid also inhibited contractile responses of rat aorta to 5-hydroxytryptamine and angiot ensin It. 3. Chebulinic acid inhibited the binding of pH]-prazosin to dog aortic microsomal membranes in a concentration-depen-dent manner with an IC50 value of 0.3dmmol/L Results of saturation binding experiments suggest a mixed mode of inhibition by chebulinic acid (i.e. a decrease in both the maximal number of binding sites and the affinity for prazosin). 4. Chebulinic acid concentration-dependently and reversihly inhibited the maximal left ventricuiar pressure of rat heart in a Langendorff preparation with 50% inhibition occurring at a concentration of 0.3nmol/L. 5. We conclude that chebulinic acid exerts non-specific inhibitory actions in vascular preparations. Its inhibitory effect on cardiac contraction was reversible and three orders of magnitude more potent than that on vascular contraction. We suggest that the hypotensive effect of chehulinic acid is probably mediated via the decrease in cardiac output resulting from reduced left ventricular contraction.

We examined the endothelium-dependent relaxation response to acetylcholine (Ach) in treptozotocin-induced diabetic rat aorta at the stages of 2- and 6-wks' duration in vitro, and compared with another two groups which were treated with dietary supplement of 0.1% Aminoquanidine (AG) and 0.5% Erigeron breviscapus(EB) from l-week of diabetes induction. At the stage of 2-wks' duration of diabetes, relaxation responses to lower concentrations of Ach in 0.3uM phenylepherineprecontracted aortas were diminished significantly (Pc0.05) compared with agematched control, but the maximal relaxation of Ach remained unchanged. At the stage of 6-wks' duration, diabetes caused an approximately 60% (P<0.001) deficit in maximum relaxation, and this was significantly (P<0.001) prevented in AG and EB treated groups. There was an approximately 40% enhancement in the maximum contractile response to phenylepherine with diabetes (P<0.5), which was unaffected significantly by AG and EB treatments. The data suggest that the defective endothelium-dependent relaxation in diabetic rat aorta occurred as early as 2-wks' duration of diabetes, and the treatments of AG and EB could protect vascular endothelium although the deficits in vascular smooth muscle contractile responses were not protected.

We studied the Ca2+ movement induced by activation of α1A-, α1B- and α1D-adrenoceptor subtypes in transfected HEK-293 cells with the fura- probe. All these α1-AR subtypes induced both Ca2+ release and Ca2+ entry. The effect on Ca2+ release in α1b transfected HEK-293 cells was bigger than that in α1a and α1b transfected HEK-293 cells, and the effects on Ca2+ entry were the same in ala, alb and aid transfected HEK-293 cells. The Ca2+ entry was inhibited by 1mM NiSO4, but not by nifedipine. Cyclopiazonic acid (CPA) produced a biphasic Ca2+ signal response in Ca2+ medium, and only induced a transient response in Ca2+-free medium. After depletion of CPA-sensitive Ca2+ pool by 10uM CPA in Ca2+-free medium, 10uM adrenaline (Adr) still transiently increased [Ca2+]i in three different α1-adrenoceptor subtype transfected HEK-293 cells. However, after depletion of drenaline-sensitive Ca2+ pool by 10uM Adr, CPA transiently elevated [Ca2+], only in ala and aid transfected HEK-293 cells, not in a,, transfected HEK-293 cells. U73122, a phospholipase C (PLC) inhibitor, inhibited both Ca2+ release and Ca2+ entry induced by activation of α1B-AR,α1B and α1D subtypes in transfected HEK-293 cells. These results suggest that HEK-293 cell line contains two functionally separate intracellular Ca2+ pools, CPA-sensitive and Adr-sensitive pools. Activation of a,,-AR stimulates Ca2+ release from both CPA-sensitive and Adr-sensitive Ca2+ pools. α1A and α1D subtypes induce Ca2+release only from Adr-sensitive Ca2+ pool.

Patch-clamp whole-cell recording techniques were used to study ATP-induced Cl- current and the effects of Cl- channel blockers on NO release in bovine aortic endothelial cells. ATP evoked an outward rectified Cl- current with a reversal potential of 2973mV. This outward rectified Cl- current was dependent on Ca2+ influx, but not Ca2+ release. In Ca2+-free medium, ATP did not produce the Cl- current; however, subsequent addition of Ca2+ to the medium evoked an ATP-induced outward rectified Cl- current. Furosemide, glibenclamide, and DIDS inhibited ATP-activated Cl- current in a concentrationdependent manner with maximal inhibition of 8874%, 9371%, and 7973%, respectively. The IC50 values of furosemide, glibenclamide, and DIDS were 6.272.6, 29.6712.3, and 21.0713.4mM, respectively. These effects of Cl- channel blockers matched those on NO release from endothelial cells. Our data suggest that ATP-induced Ca2+ entry followed by increased [Ca2+]i activates Ca2+-activated Cl- channels which mediate NO release from endothelial cells. Drug Dev. Res. 57: 000-000, 2003

Recent growing evidence suggests that chloride (Cl-) channels are critical to the cell cycle. In cultured rat aortic vascular smooth muscle cells (VSMCs), we have previously found that Cl- channel blockers inhibit endothelin-1 (ET-1)-induced cell proliferation. The present study was designed to further identify the specific Cl- channels responsible for VSMC proliferation. Due to the lack of a specific blocker or opener of any known Cl- channels, we used the antisense strategy to investigate the potential role of ClC-3, a member of the voltage-gated Cl- channel gene family, in cell proliferation of cultured rat aortic VSMCs. With [3H]-thymidine incorporation and immunoblots, we found that ET-1-induced cell proliferation was parallel to a significant increase in the endogenous expression of ClC-3 protein. Transient transfection of rat aortic VSMCs with antisense oligonucleotide specific to ClC-3 caused an inhibition in ET-1-induced expression of ClC-3 protein and cell proliferation of VSMCs in the same concentration- and time-dependent pattern, whereas sense and missense oligonucleotides resulted in no effects on ClC-3 protein xpression and cell proliferation. These results strongly suggest that ClC-3 may be the Cl-channel involved in VSMC proliferation and thus provide compelling molecular evidence linking a specific Cl- channel to cell proliferation. The full text of this article is available at http://www.circresaha.org. (Circ Res. 2002; 91: e28-e32.)

AIM: To investigate the effects of 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) on cell proliferation and apoptosis in ECV304 endothelial cells and related molecular mechanism. METHODS: MTT, Hoechst33258, TUNEL, Flow cytometry, DNA ladder, RT-PCR, Western blot, and electrophoretic mobility shift assay (EMSA) analysis were employed. RESULTS: The 15d-PGJ2 induced apoptosis in ECV304 endothelial cells in a dose-dependent manner (the percentage of apoptosis was enhanced from 10.0%±1.3% to 32.8%±1.6%), which was accompanied by inhibition of NF-κB and AP-1 DNA binding activity, down-regulation of c-myc, upregulation of Gadd45 and p53, and activation of p38 kinase. However, the expression of p21 was found no significant change. CONCLUSION: peroxisome proliferator-activated receptor gamma ligand, 15d-PGJ2, can inhibit proliferation and induce apoptosis in ECV304 endothelial cells through different mechanisms.

目的：研究随着糖尿病的发生发展，大鼠主动脉平滑肌对苯肾上腺素等激动剂收缩反应的变化及其可能机制。方法：用链尿菌素诱导糖尿病后，在第2、6、12周，取主动脉环进行实验观察。结果：苯肾上腺素的浓度依赖性收缩反应曲线，与对照相比：在第2周，低浓度时（0.01-0.03μmol·L-1）明显增加（P<0.01），最大反应无明显变化；在第6周，各浓度点均显著增加，且最大收缩反应增加约40%；然而，在第12周1）苯肾上腺素10μmol·L-1引起的最大收缩反应趋向降低（P<0.05），2）在无Ca2+液，也较对照明显减小（P<0.05），3）在无Ca2+液，在尼非地平1μmol·-L-1和苯肾上腺素10μmol·L-1存在下，复Ca2+引起的收缩在两组问的差异未见显著性，4）在正常Krebs‘液，环匹阿尼酸10μmol·L-1引起的收缩反应较对照也显著减小（P<0.001)。结论：（1）在糖尿病的第2周，平滑肌α-1肾上腺素能受体的敏感性增加。（2）糖尿病大鼠主动脉平滑肌对苯肾上腺素收缩反应的异常变化，与通过电压依赖性钙通道的Ca+内流大小、胞内功能性Ca2+池大小及其胞内Ca2+池耗竭后所引起的充电性内流变化密切相关。