HPLC法测定心衰大鼠心肌中ATP、ADP与AMP 的含量

摘 要 目的： 建立大鼠心肌组织ATP、ADP、AMP 的含量测定方法,考察心衰大鼠的ATP、ADP、AMP定量变化。 方法: 色谱柱为Thermo Hypersil C₁₈柱(250 mm×4.6 mm,5 μm);流动相为6.8 g·L^-1磷酸二氢钾和1.164 g·L^-1氢氧化钠溶液;检测波长为254 nm。SD大鼠随机分为对照组和试验组,分别腹腔注射生理盐水和多柔比星,给药6周后取心脏组织制备匀浆,用HPLC法测定大鼠心脏组织中ATP、ADP和AMP的含量。结果:ATP、ADP和AMP在0.625～40μg·ml^-1呈良好的线性(r>0.99),最低检测限和最低定量限为0.625 μg·ml^-1,精密度RSD均<9.50%,回收率为93.20%～108.0%,稳定性RSD 为3.0%～14.0%。多柔比星组ATP的含量均明显低于生理盐水组,而AMP和ADP含量则明显高于生理盐水组（P<0.01）。结论: HPLC法测定心肌组织中ATP、ADP和AMP含量满足生物样本的测定要求,可用于考察蒽环类抗肿瘤药物对心衰大鼠的影响。     

红花-甘草配伍对寒凝血瘀大鼠肠系膜动脉血管舒缩功能的影响

目的:研究红花-甘草配伍对寒凝血瘀证大鼠肠系膜动脉血管舒缩功能的影响。方法:采用冰水应激大鼠30d制作寒凝血瘀证大鼠模型,应用生物机能实验系统测定大鼠肠系膜动脉血管收缩力。结果:与对照组比较,模型组苯肾上腺素(PE)收缩血管的量效曲线、β2受体拮抗剂ICI 118551预处理后肾上腺素(AD)收缩血管的量效曲线及沙丁胺醇(SAL)舒张血管的量效曲线均明显右移,最大收缩幅度(E_max)或舒张幅度(R_max)均明显降低,pD_2值均明显减小;与模型组比较,红花组和红花-甘草组此三种量效曲线明显左移,E_max或R_max明显升高,红花-甘草组的作用强于红花组。用高K^+和PE收缩血管,Ach对模型组大鼠血管的内皮依赖性舒张作用明显减弱,红花组和红花-甘草组Ach舒张血管的作用明显增强,红花-甘草组的作用强于红花组。结论:红花-甘草配伍能明显改善寒凝血瘀证大鼠肠系膜血管的收缩和舒张功能,其机制可能与调节肾上腺素受体和保护血管内皮功能有关。     

HPLC法测定术后疲劳综合征大鼠骨骼肌ATP,ADP,AMP的含量

目的:建立改良的高效液相色谱(HPLC)法测定术后疲劳综合征(POFS)大鼠骨骼肌5’-三磷酸腺苷(ATP)、5’-二磷酸腺苷(ADP)、5’-磷酸腺苷(AMP)的含量。方法:采用色谱柱Hypersil ODS2(4.6 mm×150 mm,5μm),柱温25℃;流动相为100 mmol.L-1磷酸盐缓冲液(含12 mmol.L-1的磷酸氢二钠和88 mmol.L-1的磷酸二氢钠,pH=6.5)-甲醇(99.9∶0.1,体积比);流速1.0 mL.min-1;紫外波长为254 nm;进样量20μL。结果:ATP、ADP、AMP色谱峰在10 min内得到了较好的分离,在测定范围内均呈良好的线性关系,r分别为1.000,0.9996,0.9994;精密度试验日内RSD≤4.1%,日间RSD≤3.5%;稳定性试验RSD≤4.8%;重复性试验RSD≤2.3%;回收率在97.1%～105.4%之间,RSD≤4.8%。应用改良的方法对模型组大鼠和对照组大鼠骨骼肌ATP、ADP、AMP的含量进行检测。结果显示,与对照组相比,模型组大鼠骨骼肌ATP含量在术后第1,3 d明显下降(P<0.05),ADP含量在术后第7 d明显升...     

甘草素对来曲唑诱导的多囊卵巢综合征大鼠的保护作用及机制研究

目的：探究甘草素对来曲唑诱导的多囊卵巢综合征（PCOS）大鼠的保护作用及机制研究。方法：将40只雌性SD大鼠随机分为4组：A组：不做任何处理，B组：造大鼠多囊卵巢综合征模型，C组：给予甘草素300 mg·kg^-1，D组：给予甘草素100 mg·kg^-1。末次给药结束12 h后，称取大鼠卵巢质量；采用HE染色的方法观察大鼠卵巢组织形态的变化；采用ELISA法检测血清中促卵泡雌激素（FSH）、促黄体生成素（LH）、雌二醇（E2）、孕酮（P）和睾酮（T）含量变化；采用Western blot法检测各组大鼠卵巢组织中P450芳香化酶（P450arom）的表达量。结果：大鼠卵巢质量测定结果表明，C组较B组大鼠卵巢质量降低（P<0.05）。HE染色结果表明，B组大鼠的卵巢组织形态学出现异常改变，C组较B组卵巢组织形态有明显改善，D组大鼠卵巢组织形态未见明显改善。ELISA结果表明，B组与A组大鼠血清中LH、T、E2含量比较，差异有显著性（P<0.05）；C组与B组大鼠血清中LH、T、E2含量比较，差异有显著性（P<0.05）。PCR结果表明，C组大鼠卵巢组织中CYP19 mRNA含量较B组降低（P<0.05）。WB结果表明，B组大鼠卵巢组织中P450arom蛋白表达水平比A组增多（P<0.05）；C组大鼠卵巢组织中P450arom蛋白表达水平比B组减少（P<0.05）；D组与B组大鼠卵巢组织中P450arom蛋白相比无明显变化（P>0.05）；C组大鼠卵巢组织中P450arom蛋白表达水平比D组降低（P<0.05）。结论：甘草素可明显改善来曲唑诱导的多卵巢综合征大鼠的卵巢组织形态，调节性激素分泌，抑制卵巢组织中P450arom的表达。     

激光散斑衬比成像技术观察补阳还五汤对大鼠脑缺血再灌注后脑微血流的影响

目的：建立脑缺血再灌注大鼠模型,观察补阳还五汤对大鼠脑缺血再灌注后脑微血流的影响。方法：将动物分为对照组、模型组、补阳还五汤组、重组人血管内皮抑制素组、补阳还五汤+重组人血管内皮抑制素组,应用激光散斑衬比成像技术观察大鼠梗死区域脑血流状态,测定脑组织的含水量,并检测血管内皮生长因子（vascular endothelial growth factor,VEGF）蛋白及基因表达水平。结果：与对照组相比,脑缺血再灌注大鼠模型大鼠脑血流速度明显降低（P<0.01）,脑组织含水量增高（P<0.01）,VEGF蛋白和基因的表达减少（P<0.01）;与模型组相比,补阳还五汤组能明显提高脑缺血再灌注大鼠模型大鼠脑血流速度（P<0.01）,降低脑组织含水量（P<0.01）,提高VEGF蛋白和基因的表达（P<0.01）。结论：补阳还五汤具有改善缺血再灌注后脑血流的作用,而调节VEGF蛋白及基因的表达水平是其参与改善脑血流的主要途径之一。     

黄芪多糖对脓毒症急性肾损伤大鼠AMPK/SIRT1信号通路介导的肾上皮细胞能量代谢的影响

目的：研究黄芪多糖（APS）对脓毒症急性肾损伤（AKI）大鼠肾上皮细胞能量代谢的影响。方法： 60只大鼠随机抽取10只设为正常组,其余随机分为AKI组,APS低、高剂量组各10只,地塞米松组11只。建模后6,12,18 h时APS低、高剂量组APS 100,200 mg·kg^-1灌胃,地塞米松组地塞米松10 mg·kg^-1灌胃,正常组及AKI组等量生理盐水灌胃。建模后24 h测磷法检测肾上皮细胞钠-钾-三磷酸腺苷酶（Na^＋-K^＋-ATP）活性;Western blot法检测肾组织沉默信息调节因子2相关酶类1（SIRT1）。结果：与AKI组比较,APS低、高剂量组,地塞米松组Na^＋-K^＋-ATP酶活性增强（t=3.894,P=0.005;t=6.564,P<0.01;t=6.565,P<0.01）;与APS低剂量组比较,APS高剂量组、地塞米松组Na^＋-K^＋-ATP酶活性增强（t=2.544,P=0.034;t=2.635,P=0.030）;与AKI组比较,APS低、高剂量组,地塞米松组SIRT1蛋白表达量升高（t=9.923,P<0.01;t=12.042,P<0.01;t=10.960,P<0.01）;与APS低剂量组比较,APS高剂量组、地塞米松组SIRT1蛋白表达量升高（t=4.985,P=0.001;t=5.000,P=0.001）。结论： APS可改善脓毒症AKI大鼠肾上皮细胞能量代谢,可能与激活AMPK/SIRT1信号通路有关。     

二丁酰环磷腺苷钙治疗银屑病不良反应/事件发生情况及影响因素分析

目的： 了解二丁酰环磷腺苷钙治疗银屑病不良反应的发生情况并分析其影响因素。方法： 采用回顾性病例对照研究的方法，选取接受二丁酰环磷腺苷钙治疗的922例患者作为研究队列，以发生相关不良反应患者为病例组，未发生不良反应患者作为对照组，对收集到的数据进行单因素及多因素Logistics回归分析。结果： 纳入的患者中112例被判定为二丁酰环磷腺苷钙不良反应/事件（12.15%）。单因素分析结果显示，女性不良反应/事件高于男性（P<0.01），青年、中年高于老年、少年（P<0.01），既往有药物过敏史不良反应/事件高于无药物过敏史（χ²=4.101，P<0.05），合并高血压不良反应/事件发生率明显低于无高血压并发症（χ²=10.292，P<0.01）。多因素Logistics回归分析显示，二丁酰环磷腺苷钙相关不良反应/事件的危险因素是女性、青年、中年、药物过敏史，保护性因素为合并高血压。结论： 当患者具有女性、青年、中年、既往有药物过敏史等特征时应用二丁酰环磷腺苷钙应予以重视，且患者合并高血压可降低药物不良反应/事件的发生。     

回乳抑增方抗泌乳素腺瘤的药效及作用机制研究

目的：研究中药复方回乳抑增方抗泌乳素腺瘤的药效及作用机制。方法：建立SD雌性大鼠泌乳素腺瘤模型,大鼠随机分为7组,分别为正常组、模型组各14只,手术对照组、溴隐亭阳性药组、回乳抑增方高剂量组、回乳抑增方中剂量组、回乳抑增方低剂量组各6只。除正常组和手术对照组外,其他5组模型大鼠灌胃给予相应药物30 d,每天1次。应用基因芯片筛选正常组大鼠和泌乳素腺瘤模型组大鼠垂体组织基因间的差异。运用Western Blot测定各组大鼠垂体组织中泌乳素（PRL）、胞内磷脂酰肌醇激酶（PI3K）、蛋白激酶B（AKT）和哺乳动物雷帕霉素靶蛋白（mTOR）蛋白含量。结果：大鼠泌乳素腺瘤模型建立成功。与正常组相比,模型组大鼠垂体组织的PI3K基因表达显著。与正常组相比,模型组大鼠垂体组织中PRL、PI3K、AKT和mTOR表达显著升高（^**P<0.01）,而与模型组相比,高、中和低剂量的回乳抑增方给药组大鼠垂体组织中PRL、PI3K、AKT和mTOR表达显著降低（^##P<0.01）。结论：回乳抑增方可显著抑制泌乳素腺瘤的增长和泌乳素的过度分泌,作用机制与调控PI3K/AKT/mTOR信号通路有关。     

苦参碱治疗慢性萎缩性胃炎的实验研究

目的：探讨苦参碱对大鼠慢性萎缩性胃炎（CAG）的影响及其作用机制。方法：采用多因素复合法建立CAG模型,将大鼠分为正常组、模型组、阳性对照组（维酶素300 mg·kg^-1）、苦参碱高剂量组（200 mg·kg^-1）、苦参碱低剂量组（100 mg·kg^-1）,每组10只。灌胃给药,连续治疗8周后,于光镜下观察胃黏膜组织病理学并进行病理程度分级;放射免疫法测定血清胃泌素（GAS）和血浆生长抑素（SS）含量;酶联免疫吸附法测定胃黏膜匀浆液中白细胞介素I β（IL-1β）、IL-6、肿瘤坏死因子α（TNF-α）浓度,实时定量聚合酶链反应法测定胃黏膜Toll样受体4（TLR4）、髓样分化因子88（MyD88）、核因子κB（NF-κB）基因表达。结果：各给药组大鼠胃黏膜病变有显著减轻,炎性细胞浸润程度和固有腺体萎缩程度的组间差异均有统计学意义（P<0.01）;与模型组相比,苦参碱高、低剂量组大鼠血清GAS含量显著升高（P<0.01）,血浆SS含量则出现显著降低（P<0.01）;苦参碱高、低剂量组大鼠胃黏膜IL-1β、IL-6和TNF-α水平均有显著降低,与模型组相比组间差异均有统计学意义（P<0.01）;与模型组相比,苦参碱高剂量组大鼠胃黏膜TLR4、MyD88和NF-κB mRNA相对表达量均有显著降低（P<0.05或P<0.01）,苦参碱低剂量组TLR4和NF-κB mRNA相对表达量均有显著降低（P<0.05或P<0.01）。结论：苦参碱可通过抑制TLR4/MyD88/NF-κB信号转导通路的持续活化及相关炎症细胞因子的高表达,减轻CAG炎症反应,阻止炎症细胞对胃黏膜造成的损伤,发挥对胃黏膜的保护作用。     

人参皂苷Rd对乳腺增生及伴发疼痛的治疗作用及机制研究

目的：研究人参皂苷Rd（ginsenoside Rd， GSRd）抗乳腺增生和镇痛的作用，并探讨其相关作用机制。方法：采用雌二醇（E₂）和黄体酮诱导雌性未孕大鼠形成乳腺增生模型，给予GSRd干预后观察大鼠乳腺组织病理学的变化；酶联免疫法（ELISA）检测血清中丙二醛（MDA）、谷胱甘肽过氧化物酶（GSH-px）、超氧化物歧化酶（SOD）的含量；免疫印迹法检测大鼠乳腺组织中Bcl-2与P53蛋白的表达；采用扭体和福尔马林舔足法观察GSRd的镇痛作用。结果：与模型组比较，GSRd可明显抑制乳腺组织的增生，升高血清中SOD的含量，增加GSH-px活力，降低MDA的含量（P<0.05，P<0.01）；免疫印迹的实验结果表明GSRd可增加BCL-2的蛋白表达（P<0.05，P<0.01），抑制P53蛋白蛋的表达（P<0.05，P<0.01）；镇痛实验结果显示GSRd对冰醋酸致小鼠扭体反应和福尔马林所致的Ⅰ、Ⅱ相反应均有抑制作用（P<0.05，P<0.01）。结论：GSRd对乳腺增生的治疗作用可能与增强体内抗氧化能力调节Bcl-2与P53蛋白的表达有关，并对乳腺增生所伴随的疼痛有明显的缓解作用。     

磷酸肌酸钠对体外循环下房室间隔缺损修补术患儿心肌能量代谢和心肺功能的影响

目的:探讨磷酸肌酸钠液冷晶体停跳液对体外循环下房室间隔缺损修补术患儿心肌能量代谢和心肺功能的影响。方法:采用前瞻性随机对照研究。选取在南方医科大学附属佛山医院接受体外循环下行手术治疗的房室间隔缺损患儿48例,采用随机数字表法分为试验组和对照组,每组24例。对照组患儿男14例、女10例,年龄（7.2±3.4）岁,体质量（...     

Antagonism of tetrodotoxin- and procaine-induced axonal blockade by adenine nucleotides in the frog sciatic nerve

The effects of adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP), and adenosine on compound action potentials were investigated in de-sheathed frog-sciatic nerve preparations. ATP and ADP but not adenosine antagonized the inhibitory action of tetrodotoxin (TTX) on nerve conduction. AMP had little or no antagonistic effect on TTX-induced axonal block. ATP was more effective than ADP. The effects of the nucleotides were related to the degree of the TTX-induced inhibition and were more evident where the blockade was more intense. ATP and ADP but not adenosine antagonized the procaine-induced axonal blockade which, in some experiments, was completely reversed by these nucleotides. ATP and ADP were of similar potency. The axonal blockade induced by pentobarbitone was not antagonized by ATP, ADP, AMP or adenosine. The possibility that ATP stimulates a TTX-sensitive sodium channel is discussed.     

Co-release of endogenous ATP and noradrenaline from guinea-pig mesenteric veins exceeds co-release from mesenteric arteries

1. The present study was designed to compare the overflow of sympathetic neurotransmitters of guinea-pig inferior mesenteric artery and mesenteric vein evoked by electrical field stimulation (EFS) with special emphasis on the simultaneous release of ATP and noradrenaline (NA). The stimulation-evoked overflow of ADP, AMP and adenosine was also evaluated. 2. Endothelium-denuded segments of inferior mesenteric arteries or veins were superfused in a small volume (200 microL)-chamber for EFS and subsequent detection of NA (HPLC- electrochemical detection) and adenine nucleotides and adenosine (HPLC-fluorescence detection) in samples of the superfusate. 3. Both arteries and veins responded to EFS (15 V, 4-16 Hz, 0.3 msec for 60 s) with overflow of ATP and NA in a tetrodotoxin (1 micromol/L)- and guanethidine (10 micromol/L)-sensitive manner. The EFS-evoked overflow of NA in veins exceeded the overflow of NA in arteries at all frequencies of stimulation, whereas the EFS-evoked overflow of ATP, ADP and AMP in veins exceeded the overflow of adenine nucleotides in arteries at 4 and 8 Hz but not at 16 Hz stimulation. The EFS-evoked overflow of adenosine was similar in arteries and veins. 4. Activation of alpha1-adrenoceptors with methoxamine (10 micromol/L) did not produce overflow of ATP. 5. Blockade of alpha1/alpha2-adrenoceptors with phentolamine (1 micromol/L) did not affect EFS-evoked overflow of ATP, ADP, AMP and adenosine. 6. It is concluded that overflow of ATP and NA from sympathetic nerves may constitute an effective mechanism in the complex balance between capacitance and resistance in splanchnic circulation.     

A study of the purinoceptors mediating contraction in the rat colon.

1. The effects of a number of purine analogues were examined on the rat isolated colon muscularis mucosae. Adenosine, adenosine 5'-monophosphate (AMP), adenosine 5'-diphosphate (ADP), adenosine 5'-triphosphate (ATP), 2-methylthioATP (MeSATP), adenosine 5'-(2-fluorodiphosphate) (ADP beta F), adenosine 5'-(beta, gamma-methylene)triphosphonate (AMPPCP) and adenosine 5'-(alpha, beta-methylene)triphosphonate (AMPCPP) each contracted the muscularis mucosae in the concentration range 1-100 microM. 2. MeSATP was the most potent purine agonist, with a threshold concentration for contraction of 0.05 microM and an EC50 of approximately 0.3 microM, and AMPCPP was less potent than ATP. The enantiomer of AMPPCP, L-AMPPCP, was inactive at concentrations up to 100 microM. 3. The adenosine receptor antagonist 8-(p-sulphophenyl)theophylline (8-SPT, 50 microM) produced approximately 50 fold shifts of the dose-response curves to adenosine, AMP and AMPPCP, whereas those to ATP, MeSATP and substance P (SP) were unaffected. Intermediate shifts were observed for the dose-response curves to ADP, ADP beta F and AMPCPP. With a lower concentration of 8-SPT (10 microM) a dose ratio of approximately 11 was observed for the inhibition of the effects of both adenosine and AMPPCP. 4. ATP was rapidly degraded by the tissue to ADP, AMP and adenosine, ADP beta F was more slowly degraded to AMP and adenosine, and no significant degradation of AMPPCP was detected during 20 min incubation. 5. The results are consistent with the existence in the rat colon muscularis mucosae of a mixed population of purine receptors of P2Y and P1 types. The colon thus contains the first documented incidence of a P2Y-receptor mediating contraction.(ABSTRACT TRUNCATED AT 250 WORDS)     

川芎嗪联用维生素C对实验性缺血-再灌注损伤肝脏能量代谢的影响

目的:探讨川芎嗪(LGT)、维生素C(VitC)联合使用对肝缺血-再灌注损伤(HIRI)时肝细胞能量代谢的影响及其机制。方法:实验兔40只,随机分为肝缺血-再灌注组(A组)和肝缺血-再灌注+ LGT治疗组(B组)、肝缺血-再灌注+VitC治疗组(C组)和肝缺血-再灌注+LGT+VitC治疗组(D组)。在再灌注45min时,分别检测肝组织内三磷酸腺苷(ATP)、二磷酸腺苷(ADP)、一磷酸腺苷(AMP)含量、总腺苷酸量(TAN)、能荷(EC)、丙二醛(MDA)含量、超氧化物歧化酶(SOD)活性及一氧化氮代谢产物(NO_2~-/NO_3~-)水平。结果:与A组比较,D组肝组织内ATP、TAN、EC、NO_2~-/NO_3~-含量及SOD活性均明显增高(P＜0．05,P＜0．01),MDA含量显著减少(P＜0．01)。结论:LGT联用VitC可通过降低体内氧自由基水平、提高一氯化氮水平,而改善缺血-再灌注损伤肝脏的能量代谢。     

MODULATORY EFFECTS OF TYPE-C NATRIURETIC PEPTIDE ON SYMPATHETIC COTRANSMISSION IN THE RAT ISOLATED TAIL ARTERY

1. Rat type-C natriuretic peptide (CNP) has been studied for its effects on the neurogenically induced overflow of adenosine 5′-triphosphate (ATP), adenosine 5′-diphosphate (ADP), adenosine 5-monophosphate (AMP), adenosine (ADO) and noradrenaline (NA) in endothelium-free segments of rat isolated tail artery. The overflow of each was evoked by electrical field stimulation (EFS) of 0.5 ms pulses at 8 Hz for 3 min and the amount of ATP, ADP, AMP and ADO was quantified by high-performance liquid chromatography (HPLC)-fluorescent detection, while the amount of NA was quantified by HPLC-electrochemical detection. 2. Type-C natriuretic peptide (100 nmol/L) was found to cause a significant reduction of the overflow of all adenine purines and NA. However, at lower concentrations (1 and 10 nmol/L), CNP caused a significant reduction of the overflow of NA but did not change ATP overflow. 3. The overflow of ADP, AMP and ADO was significantly reduced by either concentration of CNP, so that the ratio ATP:ADP was diminished from 1:2 in controls to 1:1 after 1 nmol/L CNP and to 1:1.2 after 10 nmol/L CNP. 4. The production of inorganic phosphate (P_i) in response to the exogenous application of ATP was significantly reduced by 1,10 or 100 nmol/L CNP. 5. Type-C natriuretic peptide exerts neuromodulatory effects on the neurogenically induced release of the cotransmitters ATP and NA in rat tail artery, consisting of an inhibition of the release of both ATP and NA. This effect is accompanied by inhibition of the breakdown of ATP by ecto-ATPases. Either effect results in apparent CNP-induced differential modulation of the overflow of the cotransmitters ATP and NA.     

Modulation of Energy Metabolism in C6 Glioma Cells as Possible Mechanism Contributing to Zinc Neurotoxicity

Elevated tissue levels of zinc (Zn) have been associated with neurodegenerative diseases such as global ischemia, seizure, and Alzheimer's. The mechanism of action of Zn in causing neuronal injury is not clear. One of the possible mechanisms is the ability of Zn to alter cellular energy metabolism. Using the C6 glioma cell as a model, the present study aimed to determined the effects of increasing concentrations of Zn on cellular energy states, as defined by the levels of adenosine 5'-triphosphate (ATP), adenosine 5'-diphosphate (ADP), and adenosine 5'-monophosphate (AMP), the total adenosine nucleotides (TAN) (TAN = ATP + ADP + AMP), and the energy charge potential (ECP = [ATP + 0.5 ADP]/TAN). Uptake of Zn was visualized by the appearance of N-(6-methoxy-8-quinolyl)-p-toluene sulfonamide (TSQ)-stained fluorescent granules after a 3-h exposure to Zn in the medium. At [Zn] = 1 mM, cells appeared apoptotic. Levels of ATP and TAN decreased as the level of Zn increased. The change mirrors the increase in cell death as determined by the trypan blue exclusion test. However, when the ratio of ATP:ADP:AMP within the TAN was calculated, the percentage of ATP in the TAN increased significantly, while that of AMP decreased. The change in the relative AMP level mirrored the change in cell viability as measured by the MTT assay, which indicated a decreased in mitochondrial activity. Cellular ECP increased significantly from 0.85 ± 0.007 to 0.92 ± 0.04. The elevated ECP and relative ATP level, together with a significant decrease in the relative AMP level, are all indicators of inhibition of cellular metabolism. These results support the notion that acute exposure of C6 glioma cells to a high concentration of Zn might initially result in a decrease in relative AMP and an inhibition of mitochondrial activity. However, the ultimate toxic action of Zn on the C6 glioma cells appears to be due to a gradual inhibition of energy utilization, leading to cell shrinkage and apoptosis.     

Potentiation of adenosine triphosphate-induced contractile responses of the guinea-pig isolated vas deferens by adenosine monophosphate and adenosine 5''-monophosphorothioate.

The effects of incubating the guinea-pig isolated vas deferens in the presence of adenine nucleotides (adenosine triphosphate, ATP; adenosine diphosphate, ADP; and adenosine monophosphate, AMP), or in the presence of their phosphorothioate analogues (adenosine 5'-O-(3-thiotriphosphate), ATP gamma S; adenosine 5'-O-(2-thiodiphosphate), ADP beta S; and adenosine 5'-monophosphorothioate, AMP alpha S), on contractile responses to ATP were compared. After challenge with a low (1 microM) or high (300 microM) concentration of ATP to obtain control responses, one vas deferens of a pair was incubated for 5 min with one of the adenine nucleotides, while the contralateral preparation was incubated with the corresponding phosphorothioate analogue. At the conclusion of the incubation the preparations were challenged again with ATP. Incubation with AMP or AMP alpha S resulted in a transient potentiation of responses to 1 microM and 300 microM ATP. The potentiation following incubation with AMP alpha S was larger than that produced by AMP. After incubation with ADP, ADP beta S, ATP and ATP gamma S, responses to 1 microM ATP were decreased, while those to 300 microM ATP were unaffected. Thus, incubation with AMP and AMP alpha S results in potentiation, rather than inhibition, of ATP-induced responses. On the other hand, 5'-diphosphate, 5'-triphosphate, 5'-O-(2-thiodiphosphate) and 5'-O-(3-thiotriphosphate) moieties on adenosine have no effect or cause autoinhibition. These results indicate that AMP exerts a potentiating effect on reactivity to exogenous ATP. AMP arising from the enzymatic degradation of ATP might modulate the level of response to ATP released endogenously as a cotransmitter.     

P1-purinoceptors in the cerebrovascular bed of the goat in vivo

The possible existence and function of specific P1-purinoceptors in the cerebrovascular bed of the unanesthetized goat have been investigated. Blood flow to one cerebral hemisphere (cerebral blood flow) was measured by means of an electromagnetic flow probe previously implanted around the ipsilateral internal maxillary artery. The injection of adenosine, AMP, ADP and ATP (3-30 micrograms) directly into the internal maxillary artery increased cerebral blood flow and decreased cerebrovascular resistance in a dose-dependent manner. Continuous infusion of 8-phenyltheophylline (8-PT), 100 micrograms/min, into the internal maxillary artery did not alter the resting cerebral blood flow or the cerebrovascular resistance, but significantly inhibited the cerebral vasodilation induced by adenosine, AMP, ADP and, to a lesser degree, ATP. The acetylcholine- and histamine-induced cerebral vasodilation was unaffected by 8-PT. These results indicate that adenosine, AMP, ADP and, at least in part, ATP increase cerebral blood flow by acting on specific P1-purinoceptors located in the cerebrovascular wall. These P1-purinoceptors do not appear to be tonically activated under physiological conditions.     

Extracellular metabolism of adenine nucleotides and adenosine in the innervated skeletal muscle of the frog.

The effects of coformycin, alpha,beta-methylene ADP, dipyridamole in the absence and presence of erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), nitrobenzylthioinosine (NBTI), mioflazine and ouabain on the metabolic pathways of exogenously applied ATP and its metabolites in the frog innervated sartorius muscle were investigated. ATP catabolism yielded ADP, AMP, IMP, adenosine and inosine; the ecto-ATPase in situ was shown to be Ca(2+)- or Mg(2+)-activated with a Kmapp for ATP of 767 +/- 48 microM. AMP catabolism yielded IMP, adenosine and inosine; inosine was formed from either exogenous IMP or exogenous adenosine. Catabolism of AMP into IMP was blocked by coformycin, which enhanced adenosine and inosine formation from AMP. alpha,beta-Methylene ADP blocked adenosine formation from AMP and inosine formation from IMP; formation of IMP from AMP was enhanced by alpha,beta-methylene ADP. Complete blockade of AMP degradation was achieved with the simultaneous use of coformycin and alpha,beta-methylene ADP. Dipyridamole attenuated but did not completely block extracellular adenosine removal and inosine appearance in the bath. EHNA, applied in the presence of dipyridamole, did not cause any further attenuation of extracellular adenosine removal. Mioflazine, NBTI and ouabain did not affect adenosine disappearance from the bath. The results suggest that, in the frog innervated sartorius muscle, ATP can be sequentially catabolized into AMP which is then catabolized either into IMP or into adenosine. This extracellular degradation of AMP into IMP might then constitute a shunt-like mechanism to control the levels of adenosine formed from adenine nucleotides.