小剂量氢化可的松对脓毒症大鼠脑细胞线粒体功能的影响

目的:探讨糖皮质激素对脓毒症大鼠脑细胞线粒体功能的影响.方法:将42只大鼠随机分为对照组、模型组、药物干预组.模型组和药物干预组采用盲肠结扎穿孔术致脓毒症模型.药物干预组术后即给予小剂量氢化可的松(6 mg/kg).于术后8、16、24 h测定各组大鼠脑细胞线粒体琥珀酸脱氢酶(SDH)、细胞色素C氧化酶(CCO)活性及ATP的含量.结果:与对照组比较.模型组及药物干预组术后8 h SDH、CCO活性及ATP含量均明显降低(P＜0.01);药物干预组术后8 h SDH活性和ATP含量高于模型组(P＜0.05),CCO活性差异无显著性(P＞0.05).术后16 h和24 h,药物干预组SDH、CCO活性及ATP含量均明显高于模型组(P＜0.05或P＜0.01).结论:在脓毒症早期,小剂量氢化可的松可以改善脑细胞的线粒体功能及能量代谢,对线粒体损伤具有保护功能.     

银杏叶提取物对糖尿病大鼠心肌线粒体的保护作用

目的观察银杏叶提取物(EGb)对糖尿病大鼠心肌线粒体的保护作用。方法40只SD大鼠随机分为正常对照组、糖尿病组和EGb治疗组，观察各组大鼠心肌线粒体超微结构，检测心肌线粒体琥珀酸脱氢酶(SDH)、超氧化物歧化酶(SOD)、一氧化氮合酶(NOS)的活性，以及一氧化氮(NO)、丙二醛(MDA)的含量。结果糖尿病大鼠心肌线粒体主要表现为肿胀，嵴变短，空泡化；SDH、SOD活性下降，NOS活性及MDA、NO含量增高。EGb治疗组病变较糖尿病组明显减轻，心肌线粒体SDH、SOD活性升高，NOS活性及NO、MDA含量下降。结论EGb能保护自由基和过量一氧化氮对心肌线粒体的损伤，提高SDH活性，从而对糖尿病大鼠心肌起保护作用。     

培哚普利对慢性心力衰竭大鼠心肌能量代谢及超微结构的影响

目的 研究培哚普利对异丙肾上腺素(lSO)诱导的慢性心力衰竭(CHF)大鼠心肌能量代谢和超微结构的影响。方法 55只雄性SD大鼠随机(随机数字法)分为对照组(C组)和模型组。模型制备采用皮下多点注射ISO法,4周后模型组再随机(随机数字法)分为未治疗组(M组)和培哚普利治疗组(P组)。平均治疗5周后进行二维心脏超声检查,测定心肌组织中ATP、ADP、AMP、乳酸(LA)含量、肌浆网Ca2 -ATP酶(SERCA)活性以及进行病理形态学的观察。结果 与未治疗组相比,培哚普利治疗组大鼠左室射血分数(EF)、左室短轴缩短率(FS)分别提高了3.25％和7.33％。光镜和电镜结果显示,培哚普利治疗组心肌损伤程度较模型组明显减轻。与对照组相比,未治疗组大鼠心肌ATP、AMP、TAN(总腺苷)和LA含量均显著下降(P＜0.05),但培哚普利治疗组大鼠心肌ADP、AMP、ATP/ADP和TAN(总腺苷)含量与对照组相比差异无统计学意义(P＞0.05)。与未治疗组相比,培哚普利治疗组大鼠心肌SERCA活性提高了16.41％ (P ＞0.05)。结论 培哚普利能够改善心力衰竭大鼠心肌能量代谢、病理和超微结构。     

亚低温对急性缺氧缺血新生鼠脑线粒体能量代谢的作用

目的：研究亚低温治疗对缺氧缺血脑损伤（hypoxic-ischemic brain damage,HIBD)新生大鼠的脑线粒体琥珀酸脱氢酶(succinate dehydrogenase,SHD)复合酶Ⅱ的活性及ATP合成能力的影响，以探讨亚低温治疗的神经保护机制。方法:将HIBD模型鼠随机分为缺氧缺血(HI)常温恢复组和亚低温干预组，同时设对照组。各组动物在HI后不同时间点(0、2、6、24、48、72h)断头取脑，用密度离心及差速离心方法提取线粒体，生化法测定脑线粒体SDH和复合酶Ⅱ的活性，并用酶萤光法检测脑线粒体的ATP合成能力。结果：HI常温恢复组结扎侧大脑脑线粒体SDH的活性在HI后2—6h先下降后再逐渐恢复，72h最高但仍明显低于正常；脑线粒体复合酶Ⅱ的活性及脑线粒体ATP合成能力在HI后2h明显恢复，6h出现第2次下降，72h恢复正常。亚低温干预组各时间点脑线粒体SDH活性、脑线粒体复合酶Ⅱ的活性及ATP的合成能力均显著高于同一时间点常温组，SDH在72h恢复正常，复合酶Ⅱ在48h恢复正常，ATP合成能力在24h恢复正常。结论：亚低温可减轻HIBD鼠线粒体SDH及复合酶Ⅱ活性的下降，增加脑ATP的合成，缩短继发性能量衰竭的时间，从而保护脑组织。     

益肾降浊汤对高脂血症小鼠脑缺血再灌后能量代谢的影响

目的：探讨能量代谢障碍在缺血性神经元损伤中的作用及益肾降浊汤对高脂血症小鼠脑缺血再灌后能量代谢的影响。方法：在造成小鼠高脂血症基础上，进行反复脑缺血再灌注，制备智能障碍模型。采用高效液相色谱法和生物化学法分别测定脑细胞能量负荷值和脑组织琥珀酸脱氢酶(succinate dehydrogenase，SDH)，细胞色素氧化酶(cytochrome coxidase，CCO)，Na^+-K^+-ATPase活性。结果：模型组小鼠脑细胞能量负荷值，脑组织SDH，CCO，Na^+-K^+-ATPase活性均显著低于正常对照组(P&;lt;0．01)，益肾降浊汤可提高模型小鼠脑组织SDH，CCO，Na^+-K^+-ATPase活性及脑细胞能量负荷水平(P&;lt;0．01)。结论：益肾降浊汤可改善高脂血症小鼠脑缺血再灌后能量代谢障碍。     

大剂量维生素C对病毒性心肌炎心肌腺苷酸酶活性和钠钙代谢的影响

目的 :探讨病毒性心肌炎小鼠心肌腺苷酸酶活性及钠、钙含量变化 ,并用大剂量维生素 C干预治疗。方法 :雄性 Balb/ c小鼠随机分为柯萨奇 B3病毒 (CVB3)感染组、CVB3感染加大剂量维生素 C治疗组及对照组。在不同时间点分别测定心肌腺苷酸酶活性、钠和钙含量及组织病理变化。结果 :与对照组比较 ,感染组心肌Na K ATP酶、Ca2 ATP酶活性明显下降 ,钠、钙含量显著升高 (P均 <0 .0 1) ,并早于心肌结构变化 ;治疗组上述各项指标均较感染组显著改善 (P<0 .0 5或 P<0 .0 1) ,心肌炎症、坏死也明显减轻 (P<0 .0 5或 P<0 .0 1)。结论 :心肌腺苷酸酶活性下降 ,反常性钠、钙超载可能是造成病毒感染后心肌继发性损伤的重要因素 ;大剂量维生素 C能明显保护病毒性心肌炎时心肌腺苷酸酶活性 ,降低心肌钠、钙含量 ,改善心肌组织结构 ,促进心肌预后。     

大鼠肥厚心肌线粒体ADP/ATP载体活性变化的实验研究

目的 探讨压力超负荷大鼠肥厚心肌线粒体内膜ADP/ATP载体(AAC)转运活性的变化.方法 将雄性SD大鼠随机分为假手术组和腹主动脉缩窄组,术后5周及15周观察大鼠血流动力学参数、心室重构指标,密度梯度离心法提取大鼠心肌线粒体,用抑制剂终止法测定线粒体AAC的转运活性,高效液相色谱法测量心肌线粒体内腺苷酸含量.结果 大鼠腹主动脉缩窄术后5周出现左心室肥厚,术后15周加重伴心功能减退;术后5周AAC活性降低,但无统计学意义,15周时AAC活性显著减低与线粒体内(ATP+ADP)含量下降相一致.结论 压力超负荷后心功能减退的肥厚心肌AAC转运活性降低,使能量产生和利用异常,提示AAC活性改变是肥厚心肌组织能量代谢障碍及心功能减退的重要机制.     

心肌康对白介素-2和6在病毒性心肌炎中的作用研究

目的 :探讨病毒性心肌炎 (VMC)免疫发病机制及心肌康对 VMC小鼠血白介素 2和 6 (IL 2、IL 6 )水平的调节作用。方法 :腹腔注射柯萨奇 B3 病毒复制小鼠 VMC模型。将昆明种小鼠随机分为正常对照组、模型组、生脉散组及心肌康大、中、小剂量治疗组 ,用放射免疫法测定各组小鼠血清 IL 2、IL 6含量。结果 :与正常对照组比较 ,模型组 IL 2、IL 6水平明显升高 ,而生脉散组及心肌康治疗组小鼠血 IL 2、IL 6水平明显低于模型组。结论 :IL 2、IL 6分泌增加在 VMC急性期发病机制中起着重要作用。心肌康能有效地改善并调节 VMC感染机体的免疫功能 ,为治疗 VMC安全、有效的手段。     

左旋精氨酸对实验性缺血/再灌注损伤心肌能量代谢的影响

目的探讨左旋精氨酸(L-Arg)对心肌缺血/再灌注损伤(MIRI)时心肌细胞能量代谢的影响及其机制。方法实验兔30只,随机分为假手术对照组(A组)、心肌缺血/再灌注组(B组)和心肌缺血/再灌注+L-Arg治疗组(C组)。在再灌注20min时,分别检测心肌组织内三磷酸腺苷(ATP)、二磷酸腺苷(ADP)、一磷酸腺苷(AMP)含量,总腺苷酸量(TAN),能荷(EC),丙二醛浓度(MDA),超氧化物歧化酶(SOD)活性,一氧化氮(NO)水平及内皮素(ET)浓度,同时观察心肌超微结构的改变。结果C组与B组比较,心肌组织内ATP、ADP、TAN、NO含量,SOD活性及EC均明显增高(P<0.05,P<0.01),MDA、ET浓度均显著减少(P<0.05);与A组比较,AMP、TAN、NO、ET、MDA差异无显著性(P>0.05);心肌超微结构的异常改变显著减轻。结论L-Arg可通过降低体内氧自由基、ET水平,提高体内NO水平、SOD活性而改善缺血/再灌注损伤心肌的能量代谢。     

褪黑素对肺炎小鼠心肌的保护作用

目的：了解褪黑素对肺炎小鼠心肌细胞线粒体形态和心脏功能的影响，以探讨在肺炎时应用自由基清除剂对肺外器官的保护作用。方法：雄性NIH小鼠48只随机分为4组：生理盐水对照组(以生理盐水注入气管模拟制作肺炎模型)、肺炎组(经气管注入细菌悬液制作肺炎模型)、褪黑素+肺炎组(在肺炎模型基础上每日经腹腔注射褪黑素溶液进行干预)、溶媒+肺炎组(在肺炎模型基础上每日经腹腔注射褪黑素溶液的溶媒作为对照)。各组动物观察72h后检测血流动力学指标，取心肌组织制备光镜和电镜标本，剩余心肌组织制备匀浆，检测心肌组织中丙二醛含量、总超氧化物歧化酶(total superoxide dismutase，T-SOD)活性和铜锌一超氧化物歧化酶(cu，Zn-superoxide dismutase，Cu,Zn-SOD)活性、肿瘤坏死因子-α(TNF-α)含量，同时检测血浆中TNF-α含量；电镜观察心肌线粒体形态改变。结果：肺炎后小鼠心脏功能受损、心肌组织中丙二醛、TNF-α含量增加(F=3．15-8.03，P&;lt;0．01-0.05)；T-SOD[肺炎组、生理盐水对照组：(110．3&;#177;27．4)，(152.16&;#177;27．4)kNU／g]，Cu，Zn-SOD[肺炎组、生理盐水对照组：(22.89&;#177;4.17)，(30.50&;#177;6.96)kNU／g]活性降低(F=4．05，P&;lt;0．05；F：5．78，P&;lt;0.01)；ATP含量降低(F=2．91。P&;lt;0．05)；心肌组织线粒体损伤明显(F=8．55。P&;lt;0．01)。褪黑素干预可部分抑制上述改变(F=2．91～8．55，P&;lt;0．01～0．05)。结论：在肺炎小鼠中，褪黑素通过降低心肌脂质过氧化、保护心肌抗氧化能力、保护心肌线粒体等作用进而保护小鼠心脏功能。     

病毒性肝炎患者中柯萨奇B组病毒的感染情况

目的：了解柯萨奇B组病毒（coxackie virus B,CVB)在病毒性肝炎中的感染情况。方法：用免疫酶法对203例各型肝炎组进行CVB1至6型免疫球蛋白G（IgG)检测，并与105例病毒性心肌炎组及正常对照组进行分析。结果：肝炎组阳性率（39．9％）高于正常对照组（13．4％），P＜0．005，而低于心肌炎组（53．3％），P＜0．025，B1－6型都有一定的检出率，且以各型交叉感染为主。结论：病毒性肝炎患者对CVB各型普遍易感，其致病性应引起重视与探讨。     

Coxsackievirus B3 induces apoptosis in the early phase of murine myocarditis: a comparative analysis of cardiovirulent and noncardiovirulent strains

OBJECTIVE: To investigate the relationship between enteroviral infection and myocardial tissue apoptosis during the development of viral myocarditis in a murine model. METHODS: C3H/HeJ mice were inoculated with two strains of coxsackievirus B3, specifically CVB3 (cardiovirulent Nancy strain) and CVB3/0 (noncardiovirulent strain). Mice were sacrificed at 4, 7 and 10 days postinfection (p.i.). Hearts were removed, and plaque assays and RT-PCR were performed to detect the presence of viruses. Myocardial tissue sections were additionally evaluated by hematoxylin and eosin staining for inflammation, VP1 and Bax immunohistochemical staining for detection of virus and Bax expression, and TUNEL and Apostain for localization of apoptosis. RESULTS: CVB3 replicated to significantly higher titers than CVB3/0 at all time points. Histopathological analyses revealed significant inflammatory changes at all time points in CVB3-infected mice, in contrast to minimal changes in CVB3/0-infected mice. TUNEL and Apostain assays of myocardial tissues from mice infected with CVB3 disclosed maximum apoptotic lesions at 4 days p.i. and to a lesser extent at 7 and 10 days p.i. Moreover, CVB3-infected myocardial tissues displayed significantly enhanced Bax expression at 4 days p.i., and lesions overlapped with VP1-stained areas. CONCLUSIONS: These data indicate that (1) the cardiovirulent strain CVB3 induces more severe inflammation and apoptosis than the noncardiovirulent CVB3/0 strain, (2) viral replication is localized in inflammatory and apoptotic lesions in myocardial tissues, (3) apoptotic changes are observed in the early stages of myocarditis and (4) Bax may be associated with the apoptosis process in CVB3-induced myocarditis.     

Heat shock pretreatment prevents cardiac mitochondrial dysfunction during sepsis

The present study was designed to investigate the effect of previous heat shock treatment on the mitochondria function of the heart during a cecal ligation and puncture (CLP)-induced sepsis model. Rats of the heated group were heated by whole-body hyperthermia 24 h before the CLP operation. Cardiac mitochondria were freshly collected 9 and 18 h after CLP, indicating early and late sepsis, respectively. The expressions of heat shock protein 72 (Hsp72), glucose-regulated protein 75 (Grp75), and mitochondrial complexes I, II, III, and IV were evaluated by Western blot and immunochemical analysis. Enzyme activities of NADH cytochrome c reductase (NCCR), succinate cytochrome c reductase (SCCR), and cytochrome c oxidase (CCO) were measured after the reduction or oxidation of cytochrome c using a spectrophotometer. The results showed that the ATP content in the heart significantly declined during late sepsis, whereas heat shock treatment reversed this declination. The enzyme activities of NCCR, SCCR, and CCO were apparently suppressed during late stage of sepsis. The protein expressions of mitochondrial complex II and complex IV and Grp75 were also down-regulated during sepsis. Previously treated by heat shock, late-sepsis rats emerged with a high preservation of mitochondrial respiratory chain enzymes, both the protein amount and enzyme activity. Aspects of morphology were observed by electron microscopy, while heat shock treatment revealed the attenuation of cardiac mitochondrial damage induced by sepsis. In conclusion, structural deformity and the decrease of respiratory chain enzyme activity in mitochondria and its leading to a decline of ATP content are highly correlated with the deterioration of cardiac function during sepsis, and heat shock can reverse adverse effects, thus achieving a protective goal.     

Enhanced ERK-1/2 activation in mice susceptible to coxsackievirus-induced myocarditis

Group B coxsackieviral (CVB) infection commonly causes viral myocarditis. Mice are protected from CVB3 myocarditis by gene-targeted knockout of p56(Lck)(Lck), the Src family kinase (Src) essential for T cell activation. Extracellular signal-regulated kinase 1 and 2 (ERK-1/2) can influence cell function downstream of Lck. Using T cell lines and neonatal cardiac myocytes we investigated the role of ERK-1/2 in CVB3 infection. In Jurkat T cells ERK-1/2 is rapidly activated by CVB3; but, this response is absent in Lck-negative JCaM T cells. Inhibition of ERK-1/2 with UO126 reduced CVB3 titers in Jurkat cells, but not in JCaM cells. In cardiac myocytes CVB3 activation of ERK-1/2 is blocked by the Src inhibitor PP2. In addition, viral production in myocytes is decreased by Src or ERK-1/2 inhibition. In vitro, in both immune and myocardial cells, ERK-1/2 is activated by CVB3 downstream of Lck and other Src's and is necessary for efficient CVB3 replication. In vivo, following CVB3 infection, ERK-1/2 activation is evident in the myocardium. ERK-1/2 activation is intense in the hearts of myocarditis-susceptible A/J mice. In contrast, significantly less ERK-1/2 activation is found in the hearts of myocarditis-resistant C57BL/6 mice. Therefore, the ERK-1/2 response to CVB3 infection may contribute to differential host susceptibility to viral myocarditis.     

Enhanced expression of superoxide dismutase messenger RNA in viral myocarditis. An SH-dependent reduction of its expression and myocardial injury.

The oxygen free radical system has been reported to be activated by influenza virus infection in the lungs. However, the involvement of oxygen radicals in viral myocarditis is still unknown. Captopril, an angiotensin-converting enzyme (ACE) inhibitor and potent free radical scavenger with a sulfhydryl group, was effective for the treatment of viral myocarditis, while enalapril, an ACE inhibitor without a sulfhydryl group, was not effective against acute myocarditis. In this study, we investigated the role of oxygen radicals in the pathogenesis of viral myocarditis and the therapeutic effects of agents with a sulfhydryl group. 4-wk-old BALB/c mice were inoculated with the encephalomyocarditis virus, and treated with captopril or N,2-mercapto-propionyl glycine (MPG), a sulfhydryl-containing amino acid derivative without ACE inhibiting property, from days 4 to 14. On day 14, captopril and MPG significantly improved survival of mice and myocardial injury (necrosis, cellular infiltration, and calcification) in a dose-dependent manner compared with the infected control group. Thus, captopril and MPG were effective for the treatment of virus-induced myocarditis. Furthermore, a striking induction of manganese superoxide dismutase (Mn-SOD) and copper/zinc SOD (Cu/Zn-SOD) mRNAs in infected hearts was found (8-13-fold for Mn-SOD and 4-11-fold for Cu/Zn-SOD) when compared with age-matched uninfected mice hearts. MPG completely inhibited the increase of both mRNAs, even when treatment was started on day 4. Thus, oxygen radicals may play an important role in the pathogenesis of viral myocarditis, and a therapeutic approach by eliminating oxygen radicals seems possible.     

黄芪甲苷对病毒性心肌炎小鼠TLR4、NF-κB及TNF-α表达的影响

目的探讨黄芪甲苷对病毒性心肌炎小鼠Toll样受体4(TLR4) mRNA、核因子-κB(NF-κB) mRNA表达及血清肿瘤坏死因子(TNF-α)的影响。方法将80只雄性BALB/c小鼠随机分为柯萨奇病毒感染组(感染组,30只)、黄芪甲苷治疗组(治疗组,30只)和正常对照组(对照组,20只)。感染组和治疗组小鼠腹腔注射柯萨奇病毒B3(CVB3),建立病毒性心肌炎模型。治疗组于注射病毒后30min开始灌服黄芪甲苷,每日1次×7d;感染组以生理盐水灌胃;对照组用不含病毒的Eagle液腹腔注射、生理盐水灌胃。各组分别在7d、14d各随机抽取8只小鼠处死,HE染色后检测心肌病理积分、逆转录-聚合酶链反应(RT-PCR)检测心肌中TLR4 mRNA和NF-κB mRNA的表达,ELLSA法检测血清TNF-α的含量。结果感染组心肌病理积分、TLR4 mRNA、NF-κB mRNA表达及血清TNF-α的含量较对照组明显增加(P<0.01),治疗组心肌病理积分、TLR4 mRNA、NF-κB mRNA表达及血清TNF-α的含量较感染组明显降低(P<0.05)。结论黄芪甲苷可以通过调节TLR4、NF-κB及TNF-α在病毒性心肌炎中的表达,减轻心肌损伤。     

Expression of intercellular adhesion molecule-1 in murine hearts with acute myocarditis caused by coxsackievirus B3.

A cell-mediated autoimmune mechanism has been strongly implicated in the pathogenesis of viral myocarditis. Using a murine model of myocarditis caused by coxsackievirus B3 (CVB3), we previously reported that the heart is infiltrated first by natural killer cells, which express a cytolytic factor, perforin, and then by activated T cells. This action may play an important role in the pathogenesis of the observed myocardial cell damage. Cell-cell contact and adhesion is required in immune responses, and intercellular adhesion molecule-1 (ICAM-1), which is a ligand for lymphocyte function-associated antigen-1 (LFA-1), plays an important role in this process. To investigate the essential role of the ICAM-1/LFA-1 pathway in the cell-mediated cytotoxicity involved in viral myocarditis, we examined by immunofluorescence the expression of ICAM-1 in murine hearts with acute myocarditis caused by CVB3. We also evaluated the induction of ICAM-1 in cultured cardiac myocytes treated with cytokines by immunofluorescence and Northern blot hybridization. Furthermore, we analyzed the effects of in vivo administration of anti-ICAM-1 mAbs on the inflammation associated with acute viral myocarditis. We found that CVB3-induced murine acute myocarditis resulted in enhanced expression of ICAM-1 in myocardial cells. The expression of ICAM-1 in myocardial cells could be induced in vitro by IFN-gamma and TNF-alpha, which were shown to be synthesized by the infiltrating cells. In vivo treatment with F(ab')2 fragments of an anti-ICAM-1 mAb significantly reduced the myocardial inflammation induced by CVB3. These data strongly suggest that the expression of ICAM-1 in myocardial cells plays a critical role in the cell-mediated cytotoxicity involved in acute viral myocarditis.     

Quantitative analysis of viral RNA in the murine heart and pancreas with different concentration of coxsackievirus B3

OBJECTIVES: We investigated the clinical features, pathologic changes, and viral RNA kinetics in the course of acute and subacute experimental coxsackievirus B3 (CVB3) infection in a murine model. METHODS: Five-week-old A/J inbred male mice were divided into 5 groups. Four of those groups were inoculated intraperitoneally with 5 x 10(4) (group 1), 1 x 10(5) (group 2), 5 x 10(5) (group 3), or 1 x 10(6) (group 4) PFU of CVB3. Control mice were inoculated with uninfected Vero cell lysate in DMEM. Mice from each group were sacrificed on days 7 or 14 after inoculation. RESULTS: Bloody diarrhea, earlier weight loss, perianal swelling, and death were correlated with higher viral load. One of ten mice in group 3 and 5 of 10 mice in group 4 died spontaneously between days 4 and 12 after inoculation. All of the remaining 34 mice of infected groups demonstrated extensive pancreatic inflammation. Focal myocarditis developed in only 4 (11.8%) of those 34 subjects. Amylase and creatine kinase activities in the serum were increased in the mice of infected groups. CVB3 RNA was detected in the heart and pancreatic tissue in all subjects. The CVB3 RNA copy number in pancreatic tissue was not correlated with the severity of inflammation. CONCLUSIONS: In the murine model, viral loading dose determines the clinical features of CVB3-induced infection, and the severity of pancreatitis is not correlated with the viral loading dose or tissue level of viral RNA. .     

Sulfonylureas inhibit metabolic flux through rat liver pyruvate carboxylase reaction

The effect of oral hypoglycemic sulfonylureas, tolbutamide and glyburide, on metabolic flux through the pyruvate carboxylase reaction was evaluated in liver mitochondria isolated from 24-hr fasted rats. Both these sulfonylureas inhibited the metabolic flux through the pyruvate carboxylase reaction in a concentration dependent manner. Half-maximal inhibition was achieved at tolbutamide and glyburide concentrations of 0.85 mM and 63.3 microM, respectively. Neither sulfonylurea altered the activity of pyruvate carboxylase or the Km of the enzyme for ATP and pyruvate. However, glyburide and tolbutamide decreased mitochondrial ATP content and elevated mitochondrial ADP and AMP levels. The decrease in mitochondrial ATP was greater with 400 microM glyburide compared with 2.0 mM tolbutamide. Glyburide also decreased mitochondrial acetyl-coenzyme A/CoASH ratio. Additionally, glyburide and tolbutamide stimulated pyruvate (5 mM) supported mitochondrial respiration in the absence of ADP. These data indicate that these sulfonylureas inhibit the metabolic flux through the pyruvate carboxylase reaction by decreasing mitochondrial ATP/ADP and acetyl-coenzyme A/CoASH ratios. Decreased mitochondrial nucleotide content and increased mitochondrial respiration caused by sulfonylureas suggest that these compounds may uncouple oxidative phosphorylation.