应用脑微透析技术观察大鼠纹状体基础环鸟苷酸来源及一氧化氮在纹状体细胞内对环鸟苷酸的转导过程

目的：应用清醒大鼠脑微透析技术以选择性可溶性鸟苷酸环化酶抑制剂1H-[1，2，4]恶二唑[4，3-A]喹喔啉-1-酮为工具药观察大鼠纹状体一氧化氮-环鸟背酸信号转导。方法：实验于2004—02／08在锦州医学院药理教研室完成，选择雄性健康SD大鼠35只，平衡麻醉液麻醉后，将大鼠固定在立体定位仪上，横跨纹状体植入一透析探头，待大鼠清醒后24h进行透析灌流实验，灌流速度为5mL／min，每20min收集1次透析液，用放射免疫方法测定环鸟苷酸含量。结果：35只大鼠均进入结果分析。①纹状体内局部灌流1H-[1，2，4]恶二唑[4，3-A]喹喔啉-1-酮10，50，100μmol／L可剂量依赖性降低细胞外基础环鸟苷酸水平，1H-[1，2，4]恶二唑[4，3-A]喹喔啉-1-酮100μmol／L作用最强，最大，可使细胞外基础环鸟苷酸水平降低约50％。②1H-[1，2．4]恶二唑[4，3-A]喹喔啉-1-酮100μmol／L可完全对抗N-甲基-D-天冬氨酸（250μmol／L）引起的细胞外环鸟苷酸水平增加。③1H-[1，2，4]恶二唑[4，3-A]喹喔啉-1-酮也可阻断一氧化氮供体S-亚硝基-N-乙酰青霉胺（1mmol／L）引起的细胞外环鸟苷酸水平增加。纹状体内局部灌流α-氨基羟甲基异恶唑丙酸（100μmol／L）不能使环鸟苷酸水平增加。结论：①在基础状态下纹状体的环鸟苷酸约50％来源于可溶性鸟苷酸环化酶的激活。②纹状体的一氧化氮-环鸟苷酸神经通路的激活是通过N-甲基-D-天冬氨酸受体完成的。③在大鼠纹状体一氧化氮也是通过激活可溶性鸟苷酸环化酶而使环鸟苷酸增加的．     

应用脑微透析技术观察大鼠纹状体基础环鸟苷酸来源及一氧化氮在纹状体细胞内对环鸟苷酸的转导过程

目的:应用清醒大鼠脑微透析技术以选择性可溶性鸟苷酸环化酶抑制剂1H-犤1,2,4犦恶二唑犤4,3-A犦喹喔啉-1-酮为工具药观察大鼠纹状体一氧化氮-环鸟苷酸信号转导。方法:实验于2004-02/08在锦州医学院药理教研室完成,选择雄性健康SD大鼠35只,平衡麻醉液麻醉后,将大鼠固定在立体定位仪上,横跨纹状体植入一透析探头,待大鼠清醒后24h进行透析灌流实验,灌流速度为5mL/min,每20min收集1次透析液,用放射免疫方法测定环鸟苷酸含量。结果:35只大鼠均进入结果分析。①纹状体内局部灌流1H-犤1,2,4犦恶二唑犤4,3-A犦喹喔啉-1-酮10,50,100μmol/L可剂量依赖性降低细胞外基础环鸟苷酸水平,1H-犤1,2,4犦恶二唑犤4,3-A犦喹喔啉-1-酮100μmol/L作用最强,最大,可使细胞外基础环鸟苷酸水平降低约50%。②1H-犤1,2,4犦恶二唑犤4,3-A犦喹喔啉-1-酮100μmol/L可完全对抗N-甲基-D-天冬氨酸(250μmol/L)引起的细胞外环鸟苷酸水平增加。③1H-犤1,2,4犦恶二唑犤4,3-A犦喹喔啉-1-酮也可阻断一氧化氮供体S-亚硝基-N-乙酰青霉胺(1mmol/L)引起的细胞外环鸟苷酸水平增加。纹状体内局部灌流a-氨基羟甲基异恶唑丙酸(100μmol/L)不能使环鸟苷酸水平增加。结论:①在基础状态下纹状体的环鸟苷酸约50%来源于可溶性鸟苷酸环化酶的激活。②纹状体的一氧化氮-环鸟苷酸神经通路的激活是通过N-甲基-D-天冬氨酸受体完成的。③在大鼠纹状体一氧化氮也是通过激活可溶性鸟苷酸环化酶而使环鸟苷酸增加的。     

衰老对大鼠阴茎组织中一氧化氮,cGMP,cAMP的影响及与勃起功能障碍的关系

目的:观察衰老对一氧化氮NO,cAMP,cGMP含量变化,探讨其与阴()茎勃起功能障碍的关系。方法:应用亚硝酸还原酶法及放免法对不同月龄,8,16,24个月)大(2鼠阴茎组织中NO及cGMP,cAMP含量分别进行检测。结果:随月龄增加,NO含量先增高,再降低,其中月龄最高,24月龄8最低,各月龄组间差异有非常显著性意义(P<0.001);cGMP,cAMP含量均随月龄增加而逐渐下降,各月龄组间差异也有非常显著性意义(P<0.001)。结论:衰老对NO及cGMP,cAMP有显著影响,提示临床应用增加NO,cGMP,cAMP的药物对勃起功能障碍的康复有重要价值。     

学习记忆过程中磷酸化的环磷酸腺苷反应单元结合蛋白在大鼠脑纹状体内的表达

背景脑纹状体边缘区是在大鼠脑纹状体发现的一个新亚区,已证明边缘区与脑的学习记忆功能密切相关.而且,即刻早期基因c-fos,c-jun涉及边缘区内学习记忆的信号转导过程.但边缘区在参与学习记忆过程中还启动了其他哪些中间环节?环磷酸腺苷反应单元结合蛋白是长期记忆形成过程所必需的一种关键分子.检测磷酸化的环磷酸腺苷反应单元结合蛋白在纹状体是否有表达及其分布情况,有助于从分子水平探索学习记忆过程中纹状体的信号转导机制问题.目的探讨大鼠在学习记忆过程中磷酸化环磷酸腺苷反应单元结合蛋白在脑纹状体内的表达情况.设计完全随机对照.单位第一军医大学珠江医院神经科学研究所.材料实验于2003-04/08在第一军医大学珠江医院神经科学研究所完成.选择第一军医大学实验动物中心提供的健康雄性成年SD大鼠48只,经两次Y型迷宫(MG-2型,三兴声电公司)实验筛选后获得合格动物40只.方法40只合格动物随机分成4组训练组10只大鼠在Y迷宫中接受厌暗逃避反射训练;假训练灯光刺激组10只大鼠进入Y迷宫后仅接受灯光刺激,迷宫底部电网无电流通过;假训练电网刺激组10只大鼠进入Y迷宫后,则在无灯光刺激的情况下仅接受迷宫底部的电网刺激;对照组10只大鼠被单纯置于无光无电的Y迷宫环境中.动物在Y迷宫中进行学习记忆训练后,应用免疫组织化学方法检测磷酸化的环磷酸腺苷反应单元结合蛋白在脑纹状体内的表达.主要观察指标磷酸化的环磷酸腺苷反应单元结合蛋白在脑纹状体内的表达.结果40只实验大鼠均进入结果分析.①训练组大鼠经电Y迷宫训练后,脑纹状体内侧的边缘区内即有明显的磷酸化的环磷酸腺苷反应单元结合蛋白阳性表达,而假训练组或对照组的边缘区内均无明显的磷酸化的环磷酸腺苷反应单元结合蛋白阳性表达.②在海马、前额叶皮质和扣带回等处也有较多的磷酸化的环磷酸腺苷反应单元结合蛋白阳性表达.结论大鼠进行电Y迷宫厌暗学习时,脑纹状体边缘区内的转录因子磷酸化的环磷酸腺苷反应单元结合蛋白参与学习记忆的信号转导过程.     

学习记忆过程中磷酸化的环磷酸腺苷反应单元结合蛋白在大鼠脑纹状体内的表达

背景：脑纹状体边缘区是在大鼠脑纹状体发现的一个新亚区，已证明边缘区与脑的学习记忆功能密切相关。而且，即刻早期基因c-fos，c-jun涉及边缘区内学习记忆的信号转导过程。但边缘区在参与学习记忆过程中还启动了其他哪些中间环节？环磷酸腺苷反应单元结合蛋白是长期记忆形成过程所必需的一种关键分子。检测磷酸化的环磷酸腺苷反应单元结合蛋白在纹状体是否有表达及其分布情况，有助于从分子水平探索学习记忆过程中纹状体的信号转导机制问题。目的：探讨大鼠在学习记忆过程中磷酸化环磷酸腺苷反应单元结合蛋白在脑纹状体内的表达情况。设计：完全随机对照。单位：第一军医大学珠江医院神经科学研究所。材料：实验于2003-04/08在第一军医大学珠江医院神经科学研究所完成。选择第一军医大学实验动物中心提供的健康雄性成年SD大鼠48只，经两次Y型迷宫（MG-2型，三兴声电公司）实验筛选后获得合格动物40只。方法：40只合格动物随机分成4组：训练组10只大鼠在Y迷宫中接受厌暗逃避反射训练；假训练灯光刺激组10只大鼠进入Y迷宫后仅接受灯光刺激，迷宫底部电网无电流通过；假训练电网刺激组10只大鼠进入Y迷宫后，则在无灯光刺激的情况下仅接受迷宫底部的电网刺激；对照组10只大鼠被单纯置于无光无电的Y迷宫环境中。动物在Y迷宫中进行学习记忆训练后，应用免疫组织化学方法检测磷酸化的环磷酸腺苷反应单元结合蛋白在脑纹状体内的表达。主要观察指标：磷酸化的环磷酸腺苷反应单元结合蛋白在脑纹状体内的表达。结果：40只实验大鼠均进入结果分析。①训练组大鼠经电Y迷宫训练后，脑纹状体内侧的边缘区内即有明显的磷酸化的环磷酸腺苷反应单元结合蛋白阳性表达，而假训练组或对照组的边缘区内均无明显的磷酸化的环磷酸腺苷反应单元结合蛋白阳性表达。②在海马、前额叶皮质和扣带回等处也有较多的磷酸化的环磷酸腺苷反应单元结合蛋白阳性表达。结论：大鼠进行电Y迷宫厌暗学习时，脑纹状体边缘区内的转录因子磷酸化的环磷酸腺苷反应单元结合蛋白参与学习记忆的信号转导过程。     

黄蒲通窍胶囊对血管性痴呆大鼠血浆及脑组织环磷腺苷酸、环磷酸鸟苷和一氧化氮的影响

目的:观察中药黄蒲通窍胶囊对血管性痴呆大鼠血浆及脑组织环磷腺苷酸、环磷酸鸟苷和一氧化氮变化的影响。方法:实验于2004-05在安徽中医学院第一附属医院实验中心进行。选用雄性Wistar大鼠100只,分为空白组15只;假手术组15只;余下70只全部造模,随机选取造模成功大鼠45只,分成模型组、黄蒲通窍组、阿米三嗪3组各15只。空白组不干预,假手术组不阻断大脑中动脉,其他3组线栓法制作大脑中动脉栓塞所致的血管性痴呆大鼠模型。黄蒲通窍组和阿米三嗪组按照大鼠体质量以0.01mL/g的剂量灌胃给药,其他3组予以蒸馏水(0.01mL/g)灌胃,于造模结束第2天开始给药,1次/d,连续处理28d。观察各组大鼠血浆及脑组织环磷腺苷酸、环磷酸鸟苷和一氧化氮的变化。结果:58只大鼠进入结果分析。①环磷腺苷酸:模型组大鼠血浆和脑组织中的环磷腺苷酸浓度降低犤(0.34±0.11),(17.07±2.81)nmol/L犦,黄蒲通窍组、阿米三嗪组较模型组明显升高犤(0.96±0.11),(28.61±6.98),(1.08±0.13),(32.10±6.41)nmol/L犦。②环磷酸鸟苷:模型组大鼠血浆和脑组织中的环磷酸鸟苷浓度升高犤(0.26±0.05),(5.92±1.15)nmol/L犦,黄蒲通窍组、阿米三嗪组较模型组明显降低犤(0.14±0.05),(4.08±0.30),(0.14±0.05),(3.74±0.25)nmol/L犦。③模型组一氧化氮浓     

Characterization of subtypes of hypertension in CAPD patients by cyclic guanosine monophosphate.

OBJECTIVE: While most hypertensive patients with end-stage renal disease normalize high blood pressure with fluid removal by continuous ambulatory peritoneal dialysis (CAPD), there is a significant proportion of CAPD patients whose blood pressure can be controlled only by antihypertensive drugs. METHOD AND PATIENTS: To study the hypothesis that such patients are still volume overloaded, we used plasma cyclic guanosine monophosphate (cGMP) as a marker for hydration status. Thirty-two CAPD patients were divided into 3 groups: group 1, normotensive patients (n = 12); group 2, hypertensive patients who normalized their blood pressure with fluid removal (n = 12); group 3, hypertensive patients whose blood pressure was refractory to intensified fluid removal (n = 8). RESULTS: Mean cGMP levels were significantly higher in dialysis-sensitive hypertension (27 +/- 5 pmol/mL) than in dialysis-refractory hypertension (15 +/- 2 pmol/mL), or in normotensive patients (13 +/- 4 pmol/mL). Reduction of excess fluid in volume overloaded hypertensive CAPD patients resulted in a normalization of cGMP levels (14 +/- 8 pmol/mL), but did not affect this volume marker in patients with dialysis-resistant hypertension (10 +/- 4 pmol/mL). CONCLUSION: Plasma cGMP levels are elevated in volume overload-induced hypertension complicating CAPD. Hypertensive CAPD patients whose plasma cGMP levels are within normal limits have raised blood pressure refractory to volume removal. Our findings are consistent with the hypothesis that inadequate removal of excess volume plays a major role in a subset of patients with CAPD hypertension.     

Effects of atrial natriuretic factor on cyclic guanosine monophosphate and cyclic adenosine monophosphate accumulation in microdissected nephron segments from rats.

Atrial natriuretic factor (ANF) (1 microM) markedly increased cyclic guanosine monophosphate (cGMP) content in microdissected glomeruli (35-fold) and in microdissected inner medullary collecting ducts (IMCD) (20-fold). ANF caused little or no increase in cGMP content in other nephron segments. The threshold concentration for increased cGMP accumulation by ANF was 0.1-1 nM in IMCD, which is in the range reported for rat plasma. Sodium nitroprusside (1 mM), which selectively stimulates soluble guanylate cyclase, increased cGMP content in glomeruli but not in IMCD. ANF did not alter cAMP accumulation in the absence or presence of vasopressin (AVP) or parathyroid hormone (PTH) in outer and inner medullary tubule suspensions, or in microdissected proximal convoluted tubules (PCT), medullary thick ascending limbs (MAL) or IMCD. These data are compatible with the hypothesis that cGMP is a second messenger for a physiologic action of ANF in the inner medullary collecting duct. ANF apparently activates membrane-bound guanylate cyclase in this segment.     

针刺对肥胖大鼠纹状体组织一氧化氮含量和一氧化氮合酶活性的作用

背景已知针灸减肥机制涉及神经及神经体液调节.考虑到纹状体参与了对下丘脑摄食中枢、植物神经中枢、体温调节中枢等的调控.针刺对肥胖大鼠纹状体组织一氧化氮含量和一氧化氮合酶(nitric oxide syn-thase,NOS)活性的调整作用如何?目的探讨针刺对肥胖大鼠纹状体组织一氧化氮和NOS含量的影响,进一步深化针刺减肥的中枢神经作用机制.设计以自身对照、相互对照为主的实验研究.单位南京中医药大学第二临床医学院针灸研究所、南京人口管理学院生殖医学教研室.材料本实验在南京中医药大学第二临床医学院针灸研究所完成.1月龄刚断乳SD雄性大鼠,体质量50～70 g,由南京军区总医院实验动物中心提供,动物级别为清洁级.干预以普通全价鼠饲料喂养的大鼠作为正常组,将造模成功的实验性肥胖大鼠随机分为对照组和针刺组,每组6只.针刺组大鼠给予针刺治疗14 d,正常组和对照组大鼠分别每天放入大鼠固定器中适应15 min,持续14 d.采用神经生化技术观察针刺治疗前后肥胖大鼠体质量、体脂及纹状体组织一氧化氮和NOS含量的变化.主要观察指标①针刺对实验性肥胖大鼠肥胖指标及对心包、肾周和附睾脂肪量的影响.②针刺对各组大鼠纹状体组织一氧化氮含量和一氧化氮合酶活性的影响.③大鼠纹状体组织一氧化氮含量和NOS活性与体质量和Lee's指数的相关性.结果肥胖大鼠体质量和体脂均显著高于正常大鼠水平,差异均有显著性意义(t=8.86,9.99,8.31,P均＜0.01);纹状体组织一氧化氮和NOS水平均显著低于正常大鼠水平,差异均有显著性意义(t=5.48,4.37,P均＜0.01);大鼠纹状体组织一氧化氮和NOS水平与体质量和Lee's指数呈负相关(r=-0.78,-0.71;P＜0.01).针刺治疗能使肥胖大鼠纹状体组织一氧化氮和NOS水平明显回升,与针刺前比较,差异均有显著性意义(t=3.74,3.48,P均＜0.01).结论纹状体组织一氧化氮和NOS水平异常可能是肥胖发病的因素;针刺对肥胖机体纹状体组织一氧化氮和NOS水平的良性调整作用可能是实现减肥效应的中枢作用机制之一.     

Interleukin 1 induces prolonged L-arginine-dependent cyclic guanosine monophosphate and nitrite production in rat vascular smooth muscle cells.

The cytokine interleukin 1 (IL-1) inhibits contractile responses in rat aorta by causing endothelium-independent and prolonged activation of soluble guanylate cyclase. The present study tested whether IL-1 activates guanylate cyclase by inducing prolonged production of nitric oxide in cultured rat aortic vascular smooth muscle cells (VSMC). IL-1 induced a marked time-dependent increase in cyclic guanosine monophosphate (cGMP) in VSMC which was significant at 6 h, and increased progressively for up to 36 h. This effect of IL-1 was abolished when protein synthesis was inhibited with cycloheximide or actinomycin D, suggesting that the effect of IL-1 involves new protein synthesis. IL-1-induced cGMP accumulation was inhibited by the soluble guanylate cyclase inhibitors, methylene blue, LY83583, and hemoglobin and by the L-arginine analogue NGmonomethyl-L-arginine (L-NMMA). The inhibitory effect of L-NMMA was reversed by a 10-fold excess of L-arginine, but not by D-arginine. Nitrite, an oxidation product of nitric oxide, accumulated in the media of VSMC incubated with IL-1 for 24 h in the presence of L-arginine, whereas both IL-1-induced cGMP accumulation and nitrite production were attenuated in VSMC incubated in L-arginine-deficient medium. In L-arginine-depleted VSMC, IL-1-induced cGMP accumulation was restored to control levels by a 15-min incubation with L-arginine. These results demonstrate that IL-1 activates guanylate cyclase in rat VSMC by inducing production of nitric oxide via a pathway dependent on extracellular L-arginine.     

Relationship between cyclic guanosine monophosphate accumulation and relaxation of canine trachealis induced by nitrovasodilators.

The role of cyclic GMP (cGMP) in mediating relaxation of canine trachealis produced by nitrovasodilators (NVDs), compounds that activate guanylate cyclase, was examined. Sodium nitroprusside (SNP) produced a concentration-dependent relaxation of the canine trachealis that was accompanied by a concentration-related increase in cGMP content. In time course studies, relaxation of isolated trachealis strips induced by 30 microM SNP was paralleled by an increase in cGMP that reached a maximum of 18-fold above basal levels within 2 min. Zaprinast, an inhibitor of the cGMP-specific phosphodiesterase, potentiated both SNP-induced relaxation and cGMP accumulation. A cell-permeable analog of cGMP, 8-bromo-cGMP, mimicked the relaxant effects of SNP. Also assessed were the effects of methylene blue, an agent that inhibits soluble guanylate cyclase activity, and hemoglobin, an agent that competitively binds NO-containing compounds. In these experiments, tissues were pretreated with the above agents for 10 min, contracted with 1 or 3 microM methacholine, and then relaxed by the cumulative addition of SNP or two other NVDs, S-nitroso-N-acetyl-penicillamine (SNAP) and glyceryl trinitrate (GTN). Tissues were flash-frozen after adding the final concentration of the various NVDs and assayed for cGMP. Methylene blue and hemoglobin suppressed both cGMP accumulation and relaxation in response to SNAP and GTN. in contrast, methylene blue and hemoglobin inhibited SNP-induced cGMP accumulation but, paradoxically, potentiated SNP-induced relaxation. The results of this study generally support a role for cGMP in NVD-induced relaxation of airway smooth muscle.(ABSTRACT TRUNCATED AT 250 WORDS)