大鼠体内硝基精氨酸的手性代谢动力学

目的　应用毛细管电色谱 (CEC)对大鼠体内的硝基精氨酸手性转化和代谢动力学进行研究。方法　采用手性配体交换法检测大鼠血浆中D型硝基精氨酸 (D NNA)和L型硝基精氨酸(L NNA)的浓度,应用非房室模型对所获得的血药浓度-时间数据进行拟合,计算药动学参数。结果　D NNA和L NNA在大鼠体内代谢具有明显的异构体选择性,清除率分别为(0 46±0 02)ml·h-1·kg-1和(0 17±0 03)ml·h-1·kg-1 (P<0 05 );T1 /2分别为 ( 1 44±0 28 )h和(3 48±0 41)h(P<0 05)。D NNA到L NNA的单项手性转化率为(50 03±8 5)%。结论　D NNA和L NNA在大鼠体内的代谢有明显的异构体选择性 (其差异可能主要源于D NNA到L NNA的单向手性转化)。     

L-精氨酸、L-硝基精氨酸对红藻氨酸诱导大鼠癫痫及其海马结构中iNOS mRNA表达的影响

目的　观察诱导型一氧化氮合酶 (iNOS)在红藻氨酸(KA)癫痫大鼠海马内的表达及L 精氨酸 (L Arg)和L 硝基精氨酸 (L NNA)慢性干预的影响。方法　采用惊厥剂量的KA(1 0mg·kg- 1 )诱导大鼠癫痫发作 ,以NOS抑制剂L NNA(50mg·kg- 1 )和NO前体L Arg(40mg·kg- 1 )进行干预 ,对大鼠的癫痫发作行为及KA后不同时间点的海马内i NOSmRNA ,通过RT PCR观察其表达。结果　KA可使动物发生时间相关性癫痫发作 ,L NNA预处理后使KA诱导的癫痫发作明显加重 ,而L Arg预处理后使KA诱导的癫痫发作减弱。iNOSmRNA在KA处理后 3h开始有微弱的表达 ,且随着时间的延长逐渐增加 ,2 4h达到最高水平 ,2d及3d时未见表达 ,但 7d时又出现高表达 ;经L NNA预处理的动物 ,KA后 1h其海马结构中未出现iNOSmRNA ,但L Arg预处理后再给予KA后 1h ,可见微弱的iNOSmRNA表达。结论　红藻氨酸给药后一定时间 ,癫痫大鼠海马结构中可出现iNOSmRNA表达 ,L Arg慢性干预也有一定影响     

NO对红藻氨酸癫痫发作和IL-6表达的影响

目的　探讨脑内一氧化氮 (NO)介质对癫痫发作及白细胞介素 6(IL 6)表达的影响。方法　采用红藻氨酸 (KA)诱导大鼠癫痫发作 ,以一氧化氮合酶 (NOS)抑制剂L 硝基精氨酸 (L NNA)和NO前体L 精氨酸 (L Arg)进行干预 ,从行为学评估及形态学的角度 ,对KA癫痫发作和IL 6的相关变化做了观察。结果　一次惊厥剂量的KA (1 0mg·kg- 1 )可使动物发生明显的时间相关性癫痫发作 ,并伴随着海马结构、梨状区及大脑皮层等相关脑区IL 6免疫反应(IL 6ir)的快速升高与增强。而经L NNA(50mg·kg- 1 )或与其等摩尔量的L Arg(40mg·kg- 1 )预处理后 ,其癫痫行为发生了明显变化。L NNA促进和加重了癫痫发作 ,KA给药后 3h许多动物死亡 ,而L Arg可使癫痫发作行为减缓。与行为干预相对应 ,L NNA对海马结构等相关脑区内的IL 6ir有明显的上调作用 ,L Arg则显示出相反的效应。结论　内源性NO介质对KA癫痫发作具有抑制作用 ,对IL 6的快速表达具有下调作用 ,但这种下调作用与内源性NO介质抗癫痫发作的稳态关系还需进一步探讨     

毛细管电色谱手性配体交换法检测生物样品中的硝基精氨酸异构体研究

目的：建立毛细管电色谱（CEC）分离检测生物样品中硝基精氨酸异构体的方法。方法：采用CEC手性配体交换模式检测血浆样品中D-硝基精氨酸（D—NNA）和L-硝基精氨酸（L—NNA），流动相为50mmol／L醋酸缓冲液[pH5．0，含2mmol／Laspartame，1mmol／L Cu^2＋和5％（v／v）甲醇]；流速为0．02mL／min；操作压为1000psi；检测波长为UV280nm。结果：L—NNA和D—NNA在0．025～0．75mmol／L的浓度范围内峰面积／浓度的相关系数均可达0．99以上，日内变异系数均小于3．0％，日间变异系数分别为3．1％和3．4％。结论：本研究方法具有快速、高分辨、检测样品量和流动相用量少等优点。     

N-硝基-L-精氨酸抗吗啡、二氢埃托啡诱导小鼠位置偏爱作用

采用条件性位置偏爱模型对N 硝基 L 精氨酸 (L NNA)在吗啡、二氢埃托啡精神依赖中的作用进行了研究。L NNA 5mg·kg-1可使吗啡、二氢埃托啡精神依赖小鼠在条件性位置偏爱箱偏爱侧停留时间分别从 6 0 9± 2 0 2min ,7 98± 0 72min显著增加到 9 31± 1 10min (P <0 0 1) ,9 4 8± 1 2 1min (P <0 0 1)。小鼠皮下注射吗啡、二氢埃托啡形成精神依赖后 ,脑内NO含量升高至 (46 3 3± 18 6 )pmol·mg-1蛋白、(470 9± 16 4 )pmol·mg-1蛋白 ,显著高于生理盐水组的 (40 7 9± 15 6 )pmol·mg-1蛋白水平 (P <0 0 1)。而连续协同给予L NNA 5mg·kg-1三天后 ,吗啡、二氢埃托啡精神依赖小鼠脑内NO的含量分别降低至 (40 3 5± 17 2 )pmol·mg-1蛋白 (P <0 0 1) ,(412 2± 2 3 8)pmol·mg-1蛋白 (P <0 0 1) ,显著低于给予L NNA前的水平。与生理盐水组小鼠水平无显著性差异。我们的结果表明 :L NNA可逆转吗啡、二氢埃托啡所致的精神依赖 ,L NNA抑制小鼠脑内NO合成的作用可能参与了该过程     

免疫荧光法检测L-NNA对脑缺血再灌流后海马区GFAP合成的影响

目的　探讨NG 硝基 L 精氨酸 (L NNA)对沙土鼠海马区胶质纤维酸性蛋白 (GFAP)合成的影响。方法　钳夹沙土鼠的双侧颈总动脉制造脑缺血模型 ,应用免疫荧光法染色。结果　脑缺血再灌流后海马区GFAP合成增加 ,GFAP阳性细胞主要分布在放射层及分子层 ,L NNA能抑制海马区GFAP的合成。结论　L NNA是一氧化氮合酶强的抑制剂 ,L NNA可能通过抑制NO的产生抑制了GFAP的合成     

吗啡依赖及戒断大鼠脊髓和脑干中一氧化氮合酶基因的表达

目的　观察吗啡依赖或吗啡戒断大鼠脊髓和脑干中一氧化氮合酶 (NOS)基因表达的变化。方法　以 β actin为内参照 ,用逆转录聚合酶链反应 (RT PCR)测定NOSmRNA的表达水平。结果　吗啡依赖大鼠脊髓和脑干NOS表达水平较正常对照大鼠降低 ,纳洛酮 ( 4mg·kg-1,ip)激发大鼠吗啡戒断症状 1h后脊髓和脑干中NOS表达水平明显升高 ,戒断 2h和 4h后NOS基因表达较 1h组减少。NOS抑制剂L N 硝基精氨酸甲酯 (L NAME ,10mg·kg-1)处理后大鼠吗啡戒断症状减少 ,同时脊髓和脑干的NOS基因表达水平较戒断 1h组明显降低。甲基东莨菪碱 ( 0 5mg·kg-1)处理组脊髓和脑干中NOS表达水平较戒断 1h组明显降低 ;选择性毒蕈碱受体M1拮抗剂 pirenzepine( 10mg·kg-1)处理组动物脊髓中NOS表达水平较戒断 1h组降低 ,而脑干中NOS表达水平没有改变 ;NMDA受体拮抗剂MK 80 1( 0 12 5mg·kg-1)处理后脊髓和脑干中NOS表达水平较戒断 1h组没有差异。结论　吗啡慢性处理后脊髓和脑干中NOSmR NA水平降低 ,抑制内源性NO生成和阻断毒蕈碱受体可以减少吗啡戒断所引起脊髓和脑干中NOS基因的表达     

Nω-硝基-L-精氨酸对大鼠八臂迷宫工作记忆的作用(英文)

目的:研究一氧化氮合成酶抑制剂N_ω-硝基-L-精氨酸(NNA)对大鼠空间工作记忆的作用.方法:采用八臂迷宫延迟插板的程序.结果:腹腔注射NNA 100 mg kg~(-1)对大鼠八臂迷宫选择的准确性没有显著影响,只能增加反应的潜伏期.东莨菪碱0.25 mg kg~(-1)使大鼠延迟后的错误选择显著增加.脑室内注射NNA(10,50,100 nmo1/1 μL)没有影响准确性.结论:急性NNA给予对大鼠空间工作记忆的形成和使用没有显著影响.     

高尿酸血症动物模型研究进展

高尿酸血症发生的机制为尿酸生成增多或肾脏尿酸排泄减少。目前国内外高尿酸血症模型的复制方法主要有3种 :①给予模型动物饲喂、注射次黄嘌呤 6 0 0～ 1 0 0 0mg·kg-1 、黄嘌呤 6 0 0mg·kg-1 、腺嘌呤 1 5 0～ 30 0mg·kg-1 、酵母 1 5～ 30g·kg-1 、尿酸 2 5 0或 35 0～ 70 0mg·kg-1 ,或同时给抑制尿酸分泌的药物乙胺丁醇 2 5 0mg·kg-1 、烟酸 1 0 0mg·kg-1 均可引起体内血尿酸含量增高导致高尿酸血症。②用氧嗪酸抑制大鼠或小鼠尿酸酶活性 ,氧嗪酸钾盐 30 0mg·kg-1 一次性腹腔注射 ,可致小鼠血尿酸升高 ;大鼠饲喂氧嗪酸 0 4g·d-1和尿酸 0 6g·d-1 ,3～ 4wk后血尿酸持续性升高。③通过胚胎干细胞同源性重组 ,破坏小鼠尿酸酶基因 (EC 1 7 3 3) ,再用基因重组法获得尿酸酶缺乏的突变小鼠 ,制成高尿酸血症模型小鼠。小鼠和大鼠体内存在尿酸酶 ,可以将体内尿酸进一步分解为尿囊素 ,而禽类 (鸡、鹌鹑等 )体内缺乏尿酸酶     

芝麻素的抗氧化作用及其对代谢综合征大鼠肾病的影响

目的探讨芝麻素(sesamin,Ses)对代谢综合征大鼠肾病的影响。方法高脂、高糖诱导大鼠代谢综合征24wk,第9周口服含药芝麻素(120、60、30mg·kg-1.d-1)和辛伐他汀(5mg·kg-1.d-1)16wk,称体重和左肾湿重;测血糖、血脂、血压、肾功能、肾皮质氧化和抗氧化指标;HE和Masson染色观察肾脏形态及胶原沉积;免疫组化法表达诱导型一氧化氮合酶和硝基酪氨酸。结果芝麻素高中剂量组能明显降低代谢综合征大鼠血糖、血脂和血压,提高肾皮质总超氧化物歧化酶、过氧化氢酶、谷胱甘肽过氧化酶活性,减少丙二醛、NO2-/NO3-和羟自由基含量,下调诱导型一氧化氮合酶和硝基酪氨酸,减轻肾小球与肾间质胶原沉积,逆转肾小球硬化和肾间质纤维化,改善肾功能。结论芝麻素具有抗氧化应激和改善代谢综合征肾病的作用。     

Chiral pharmacokinetics and inversion of N^G-nitro-arginine in the rat

AIM: To explore pharmacokinetics of N^G-nitro-D-arginine (D-NNA) and N^G-nitro-L-arginine (L-NNA) in conscious rats.METHODS: The plasma concentration of D-NNA and L-NNA were determined by chiral ligand exchange method with capillary electrochromatography (CEC). Pharmacokinetic parameters were estimated using non-compartment model and were fitted using a computer program DAS. Chiral inversion rate of D-NNA to L-     

Bilateral kidney ligation abolishes pressor response to N(G)-nitro-D-arginine

We have shown that N(G)-nitro-D-arginine (D-NNA) is 50% as potent as N(G)-nitro-L-arginine (L-NNA) in causing pressor response and 2-3% as potent as L-NNA in inhibiting endothelium-dependent relaxation in vitro. These results suggest in vivo activation of D-NNA. Furthermore, the potency of D-NNA was markedly increased after it had been incubated with homogenate of the kidney, but not plasma or homogenate of the aorta, lungs or liver. This study examined if bilateral ligation of the kidneys attenuated the biological action of D-NNA. I.v. bolus of D-NNA (16 mg/kg), L-NNA (3 mg/kg) and norepinephrine (0.25-16 microg/kg) increased arterial pressure in sham-operated rats. Bilateral ligation of the kidneys abolished pressor response to D-NNA, but not L-NNA and norepinephrine. I.v. bolus D-NNA in sham-operated rats, but not kidney-ligated rats, inhibited relaxation response to acetylcholine in pre-constricted aortic rings ex vivo. These results indicate that the kidney is the primary organ which activates D-NNA.     

Pressor effects of L and D enantiomers of NG-nitro-arginine in conscious rats are antagonized by L- but not D-arginine.

The effects of NG-nitro-L-arginine (L-NNA) and NG-nitro-D-arginine (D-NNA) on mean arterial pressure (MAP) were studied in conscious, unrestrained rats. I.v. bolus of either L-NNA (1-64 mg/kg) or D-NNA (2-64 mg/kg) dose dependently increased MAP to similar maximum values of 55 +/- 7 and 52 +/- 4 mm Hg and with ED50 values of 4.0 +/- 0.9 and 8.9 +/- 1.2 mg/kg (P less than 0.05), respectively. The time course of the MAP response to a single dose (32 mg/kg i.v. bolus) of L-NNA and D-NNA were also obtained. The pressor effects of L-NNA and D-NNA each lasted greater than 2 h with the rise phase t 1/2 of 5 and 27 min (P less than 0.05), respectively. I.v. infusions (10 mg/kg per min) of L-arginine (L-Arg) and D-arginine (D-Arg) did not alter the pressor response to noradrenaline nor angiotensin II. L-Arg but not D-Arg attenuated the pressor responses to both L-NNA and D-NNA. Therefore, both L-NNA and D-NNA are efficacious and long-lasting pressor agents; the pressor effects of both can be antagonized by L-Arg but not D-Arg. Our results suggest that the pressor effects of both L-NNA and D-NNA involve the L-Arg/nitric oxide pathway.     

D-硝基精氨酸对小鼠肾损伤及氧化应激作用

目的研究D-硝基精氨酸(D-NNA)对小鼠的肾损伤及其氧化应激机制。方法 ICR小鼠ig给予D-NNA150,50和15 mg·kg-1,连续30 d。测定并计算肾系数;血液生化分析仪检测血清中肌酐(Crea)和尿素氮(BUN);分光光度法测定肾组织一氧化氮(NO),硫代巴比妥酸法测丙二醛(MDA)含量,比色法测定谷胱甘肽过氧化酶(GSH-Px)和超氧化物歧化酶(SOD)活性;观察肾病理组织学变化。结果与5%葡萄糖对照组相比,D-NNA 150,50和15 mg·kg-1组血清中BUN分别明显升高了83.6%,36.2%和27.4%(P<0.05),D-NNA150和50 mg·kg-1组血清中Crea分别明显升高了281.6%和10.6%(P<0.05);D-NNA150 mg·kg-1组肾系数和NO水平分别明显降低了5.6%和25.5%(P<0.05);D-NNA150和50 mg·kg-1组肾组织中MDA水平分别明显升高了69.0%和36.9%(P<0.01),SOD活性和GSH-Px活性分别明显下降了17.4%和17.7%,7.3%和13.7%(P<0.05);D-NNA150 mg·kg-1组病理检查可见肾小管损伤,嗜碱性变,萎缩或囊性扩张和间质炎性浸润,D-NNA50和15 mg·kg-1组出现炎症细胞浸润。结论 D-NNA对小鼠肾有一定的损伤作用,其作用机制可能与D-NNA的手性转化产物L-NNA导致NO合成减少,产生ROS有关。     

Inhibitory effects of benzoate on chiral inversion and clearance of N(G)-nitro-arginine in conscious rats.

N(G)-nitro-arginine (NNA) is known to exhibit stereoselective pharmacokinetics in which N(G)-nitro-d-arginine (d-NNA) has a faster clearance rate than N(G)-nitro-l-arginine (l-NNA) in anesthetized rats, and d-NNA undergoes unidirectional chiral inversion. It was postulated that chiral inversion of d-NNA was performed in a two-step pathway by d-amino acid oxidase (DAAO) followed by an unidentified transaminase. Such chiral inversion contributes (at least partially) to the pharmacokinetic stereoselectivity of NNA. This study used the selective inhibitor of DAAO, sodium benzoate, to test the above hypothesis. An i.v. bolus injection of d-NNA (32 mg/kg) and l-NNA (16 mg/kg) in conscious rats exhibited biphasic disposition with different pharmacokinetic parameters in a stereospecific manner (approximately 5-10-fold differences). Unidirectional chiral inversion of d-NNA but not l-NNA was found from these animals. In addition to its similar inhibitory effects on the d-NNA conversion and DAAO activity in kidney homogenates, sodium benzoate completely blocked chiral inversion of d-NNA and led to a smaller stereospecific difference, reflected by a nearly 50% reduction of d-NNA clearance and a 2-fold increase in t(1/2) and area under the curve of d-NNA in benzoate-pretreated rats. The results suggest that DAAO plays an essential role in chiral inversion of d-NNA and chiral inversion contributes mostly to the pharmacokinetic stereospecificity of NNA.     

Biological activation of NG-nitro-D-arginine by kidney homogenate

N^G-nitro-L-arginine (L-NNA) and D-NNA have been shown to inhibit endothelium-dependent relaxation. This study examined if the inhibitory effect of L-NNA or D-NNA on relaxation is increased following incubation of the drug with the supernatant of tissue homogenates. Acetylcholine (ACh) caused concentration-dependent relaxation of pre-constricted rat aortic rings with maximum relaxation of 95%. Maximum relaxations to ACh were reduced to 71 and 37% in the presence of D-NNA (40 μM) and L-NNA (1 μM), respectively. Relaxation to ACh was further reduced to 18% in the presence of D-NNA that was incubated for 1 h with the supernatant of kidney homogenate, but unaffected by D-NNA incubated with the supernatant of trichloroacetic acid-denatured kidney homogenate. Incubation of L-NNA (1 μM) with either kidney supernatant or denatured kidney supernatant for 1 h did not affect its inhibitory effect on ACh-induced relaxation. Neither 1 h’s incubation with plasma, or supernatants of liver, lungs or aorta homogenates affected the inhibitory action of D-NNA (40 or 120 μM) on ACh-induced relaxation. After D-NNA was incubated in kidney supernatant, its inhibitory effect on ACh-induced relaxation of the aorta was abolished by pretreatment of the aorta with L-arginine (L-Arg) but not D-Arg suggesting involvement of the L-Arg pathway. The results suggest that D-NNA is converted by the kidney to a compound that acts similar to L-NNA. There appears to be little conversion of L-NNA to D-NNA. Received: 21 April 1997 / Accepted: 20 June 1997     

Possible dependence of pressor and heart rate effects of NG-nitro-L-arginine on autonomic nerve activity.

1. The effects of NG-nitro-L-arginine (L-NNA) on mean arterial pressure (MAP) and heart rate (HR) were investigated in conscious rats. 2. Intravenous bolus cumulative doses of L-NNA (1-32 mg kg-1) dose-dependently increased MAP. Both mecamylamine and phentolamine increased MAP responses to L-NNA, angiotensin II and methoxamine. Propranolol, reserpine, atropine and captopril did not affect MAP response to L-NNA. 3. A significant negative correlation of HR and MAP responses to L-NNA was obtained in control rats but not in rats pretreated with reserpine or mecamylamine. Significant negative correlations also occurred in the presence of atropine, propranolol, phentolamine or captopril. 4. A single i.v. bolus dose of L-NNA (32 mg kg-1) raised MAP to a peak value of 53 +/- 3 mmHg and the effect lasted more than 2 h; the rise and recovery of MAP were accompanied by significant decrease and increase in HR, respectively. While both phentolamine and mecamylamine increased peak MAP response to L-NNA, mecamylamine abolished the biphasic HR response and phentolamine potentiated the bradycardiac component of HR. 5. Blockade of the autonomic nervous and renin-angiotensin systems did not attenuate the pressor effects of L-NNA. However, the biphasic HR response to L-NNA is mediated via modulation of autonomic nerve activities.     

磷酸喹哌抗实验性心律失常作用

磷酸喹哌(PQP)9mg·kg~(-1)iv明显降低小鼠室颤的死亡率;PQP 18mg·kg~(-1)ip对氯仿诱发小鼠室颤具有保护作用;PQP 6.3mg·kg~(-1)ip显著增加恒速(10mg·L~(-1)·min~(-1)滴注乌头碱引起麻醉大鼠室性早搏(VE)、室性心动过速(VT)、室性纤颤(VF)所需的乌头碱用量;PQP5.4 mg·kg~(-1)iv显著增加恒速(50mg·L~(-1)·min~(-1))滴注哇巴因引起麻醉豚鼠VE、VT和VF所需哇巴因用量;PQP3.36mg·kg~(-1)iv明显缩短肾上腺素诱发家兔室性心律失常的持续时间.结果表明PQP具有抗心律失常作用。小鼠PQPLD_(50)iv为93.33 mg·kg~(-1)。