Steady-State Kinetics of NADH:coenzyme Q Oxidoreductase Isolated from Bovine Heart Mitochondria

Steady-state kinetics of the bovine heart NADH:coenzyme Q oxidoreductase reaction were analyzed in the presence of various concentrations of NADH and coenzyme Q with one isoprenoid unit (Q₁). Product inhibitions by NAD⁺ and reduced coenzyme Q₁ were also determined. These results show an ordered sequential mechanism in which the order of substrate binding and product release is Q₁–NADH–NAD⁺–Q₁H₂. It has been widely accepted that the NADH binding site is likely to be on the top of a large extramembrane portion protruding to the matrix space while the Q₁ binding site is near the transmembrane moiety. The rigorous controls for substrate binding and product release are indicative of a strong, long range interaction between NADH and Q₁ binding sites.     

A redox cycle with complex II prioritizes sulfide quinone oxidoreductase-dependent H2S oxidation

The dual roles of H₂S as an endogenously synthesized respiratory substrate and as a toxin raise questions as to how it is cleared when the electron transport chain is inhibited. Sulfide quinone oxidoreductase (SQOR) catalyzes the first step in the mitochondrial H₂S oxidation pathway, using CoQ as an electron acceptor, and connects to the electron transport chain at the level of complex III. We have discovered that at high H₂S concentrations, which are known to inhibit complex IV, a new redox cycle is established between SQOR and complex II, operating in reverse. Under these conditions, the purine nucleotide cycle and the malate aspartate shuttle furnish fumarate, which supports complex II reversal and leads to succinate accumulation. Complex II knockdown in colonocytes decreases the efficiency of H₂S clearance while targeted knockout of complex II in intestinal epithelial cells significantly decreases the levels of thiosulfate, a biomarker of H₂S oxidation, to approximately one-third of the values seen in serum and urine samples from control mice. These data establish the physiological relevance of this newly discovered redox circuitry between SQOR and complex II for prioritizing H₂S oxidation and reveal the quantitatively significant contribution of intestinal epithelial cells to systemic H₂S metabolism.     

Impairment of the mitochondrial respiratory enzyme activity triggers sequential activation of apoptosis-inducing factor-dependent and caspase-dependent signaling pathways to induce apoptosis after spinal cord injury

The mitochondrion participates in caspase-independent or caspase-dependent apoptotic pathways through the release of apoptosis-inducing factor or cytochrome c. Whether both mitochondrial apoptotic cascades are triggered in the injured spinal cord remains unknown. Here, we demonstrated that neurons, astrocytes and microglia in spinal segments proximal to a complete spinal cord transection underwent two phases of apoptotic cell death. The early phase of high-molecular weight (HMW) DNA fragmentation was associated with nuclear translocation of apoptosis-inducing factor, reduction in mitochondrial respiratory chain enzyme activity and decrease in cellular ATP concentration. The delayed phase of low-molecular weight (LMW) DNA fragmentation was accompanied by cytosolic release of cytochrome c , activation of caspases 9 and 3, and resumption of mitochondrial respiratory functions and ATP contents. Microinfusion of coenzyme Q₁₀, an electron carrier in mitochondrial respiratory chain, into the epicenter of the transected spinal cord attenuated both phases of induced apoptosis, and reversed the elicited mitochondrial dysfunction, bioenergetic failure, and activation of apoptosis-inducing factor, cytochrome c , or caspases 9 and 3. We conclude that mitochondrial dysfunction after spinal cord transection represents the initiating cellular events that trigger the sequential activation of apoptosis-inducing factor-dependent and caspase-dependent signaling cascades, leading to apoptotic cell death in the injured spinal cord.     

Effect of the side chain structure of coenzyme Q on the steady state kinetics of bovine heart NADH: coenzyme Q oxidoreductase

Steady state kinetics of bovine heart NADH: coenzyme Q oxidoreductase using coenzyme Q with two isoprenoid unit (Q₂) or with a decyl group (DQ) show an ordered sequential mechanism in which the order of substrate binding and product release is NADH-Q₂ (DQ) -Q₂H₂ (DQH₂)-NAD⁺ in contrast to the order determined using Q₁ (Q₁-NADH-NAD⁺-Q₁H₂) (Nakashima et al., J. Bioenerg. Biomembr. 34, 11–19, 2002). The effect of the side chain structure of coenzyme Q suggests that NADH binding to the enzyme results in a conformational change, in the coenzyme Q binding site, which enables the site to accept coenzyme Q with a side chain significantly larger than one isoprenoid unit. The side chains of Q₂ and DQ bound to the enzyme induce a conformational change in the binding site to stabilize the substrate binding, while the side chain of Q₁ (one isoprenoid unit) is too short to induce the conformational change.     

东北地区落叶松人工林的根系呼吸

落叶松根系呼吸速率在6～9月期间逐渐升高,8月达到高峰,之后明显下降.幼林根系呼吸速率和根系呼吸占土壤总呼吸的比例均高于成熟林.根系呼吸速率与根生物量呈线性相关,与土温呈指数相关,与土壤含水量无明显相关关系,但温度较高时,土壤湿度的增加能促进根系呼吸.成熟林和幼林根系呼吸的Q10值分别为5.56和4.17.     

云南香格里拉地区亚高山草甸不同放牧管理方式下的碳排放

青藏高原由于高寒低温限制了有机碳的分解,大量的碳积累在土壤碳库中,对全球升温的反应很敏感.放牧可能会对该区草甸的碳排放产生显著影响.采用闭合箱动态法测定了云南香格里拉地区不同放牧管理方式下的亚高山草甸生态系统呼吸与土壤呼吸.常年放牧草甸与季节性放牧草甸的生态系统呼吸和土壤呼吸均呈现相似且明显的单峰季节变化特征,7月份达最大值,生态系统呼吸分别为9.77、8.03 μmol CO2 m-2 s-1,土壤呼吸分别为8.05、7.74 μmol CO2 m-2 s-1;1月份达最低值,生态系统呼吸分别为0.21、0.48 μmol CO2 m-2 s-1,土壤呼吸分别为0.16、0.49 μmol CO2 m-2 s-1.受一天中气温和土温的影响,常年放牧草甸的生态系统呼吸与土壤呼吸的日变化在夏季与冬季均呈现明显的单峰曲线变化,最高值都出现在14:00左右,最低值出现在凌晨.在夏季6～10月份,常年放牧草甸的呼吸显著大于季节性放牧草甸,表明较高的放牧强度增加了亚高山草甸的碳排放.土壤温度的指数模型F = aebT比土壤水分能更好地解释呼吸的变异性(R2 = 0.50～0.78,P < 0.0001).二元回归模型F=aebTWc比单因子模型的效果更好(R2=0.56～0.89,P < 0.0001).土壤呼吸在整个亚高山草甸生态系统呼吸中占主导地位,在常年放牧草甸与季节性放牧草甸分别为63.0%～92.7%和47.5%～96.4%,地上植物呼吸随生长季的变化而变化,在生长旺季占有较大的比例.生态系统呼吸和土壤呼吸的长期Q10(1a)是短期Q10(1d)的2倍左右.季节性放牧草甸的长期Q10小于常年放牧草甸,表明在温度上升的背景下,放牧压力较小的草甸碳库较为稳定,具有较好的碳截存能力.     

东北温带次生林与落叶松人工林的土壤呼吸

2006年5—10月,使用Li-6400-09土壤呼吸系统测定了黑龙江省帽儿山地区温带次生林转化为落叶松人工林后土壤呼吸速率(Rs)的变化.结果表明:次生林与落叶松人工林土壤呼吸速率的日变化均呈单峰型曲线,与地温的日变化趋势相似.测定期间内,次生林和落叶松人工林Rs的变化范围分别为0.43~7.26μmol CO2.m-2.s-1和0.63~4.70μmol CO2.m-2.s-1,最大值出现在7—8月,最小值出现在10月.5—8月,次生林的Rs明显高于落叶松人工林.次生林和落叶松人工林枯落物层呼吸速率的季节变化范围分别为-0.65~1.26μmol CO2.m-2.s-1和-0.43~0.47μmol CO2.m-2.s-1.两林分中的Rs与土壤温度均呈明显的指数相关,且与5 cm深地温相关最紧密.用5 cm地温估算的次生林和落叶松人工林Q10分别为3.61和3.07.次生林的Rs与10~20 cm土壤含水率相关显著,而落叶松人工林的Rs与土壤含水率无明显相关.     

植物根呼吸对升温的响应

植物根呼吸碳释放量高达18Pg／a,约为全球化石燃料燃烧碳排放量（6．5Pg／a）的2．8倍。了解根呼吸对升温的响应对于构建陆地生态系统碳动态模型、评价地下碳库碳收支具有重要作用。短期升温能明显提高根呼吸速率,但在近乎恒定的温度梯度下,根呼吸速率可能逐渐恢复到温度变化前的水平。根呼吸的温度敏感性与植物种和测定的温度范围有关,其Q10值介于1．1～10之间。在野外条件下,根呼吸的温度敏感性还会受到土壤湿度、养分状况、呼吸底物有效性、太阳辐射、光合产物的地下分配模式和天气状况等影响。通常根呼吸的温度敏感性比土壤微生物呼吸的温度敏感性高,但室内控制温度下和野外环割（girdling）实验中并未观测到类似现象。根呼吸是否具有温度适应性仍是一个尚未解决的重大科学问题。有关根呼吸对升温的适应机理仍不清楚,可能是碳循环研究存在不确定性的重要来源。今后的研究方向应集中在以下几方面：（1）深入探讨根呼吸的温度适应性;（2）扩大对成年植物种的研究;（3）扩大对环境因子交互影响和模拟研究;（4）扩大对植物根呼吸测定和升温新技术的研究。     

川西亚高山针叶林土壤呼吸速率与不同土层温度的关系

采用密闭气室红外CO2分析法（IRGA）,在野外连续定位测定了川西亚高山冷杉原始林的土壤呼吸,并对其不同土层（0、5、10、15和20cm）的温度进行了同步测定．在此基础上,分析了土壤呼吸的日、季节动态变化,及其与不同土层温度的关系和土壤呼吸Q10值变化．结果表明：冷杉原始林土壤呼吸呈现明显的日变化和季节变化．土壤呼吸的日最高值出现在12：00-14：00,最低值出现在8：00—10：00,与土壤表面温度的日变化一致;土壤呼吸的季节变化表明：7—8月的土壤呼吸高于9-11月,与不同土层温度季节变化规律一致;土壤呼吸与不同土层温度呈显著的指数增长关系,土壤呼吸速率与土壤15cm深处温度的相关性明显高于其它土层;利用Q10模型计算0～20cm各土层的Q10值分别为2．36、4．75、4．90、6．27和5．46,表明海拔高、温度低的环境条件下,土壤呼吸的Q10值偏高．     

上海典型城市草坪土壤呼吸特征

采用CFX-2开放式呼吸测定系统测定了上海城区百慕大、黑麦草-百慕大混播、结缕草和狗牙根4种典型草坪的土壤呼吸速率。结果表明：4种草坪的土壤呼吸速率均呈明显季节变化,最大值出现在7—8月,最小值出现在12月—翌年1月;4种草坪土壤呼吸平均速率依次为百慕大草坪<黑麦草-百慕大混播草坪<结缕草草坪<狗牙根草坪,其中百慕大草坪的土壤呼吸速率变化范围为0.13～2.25 μmol·m^-2·s^-1,黑麦草-百慕大混播草坪为1.16～5.95 μmol·m^-2·s^-1,结缕草草坪为0.93～8.27 μmol·m^-2·s^-1,狗牙根草坪为1.21～9.27 μmol·m^-2·s^-1;4种草坪的土壤呼吸速率与气温、5 cm地温和10 cm地温均呈极显著指数相关;百慕大草坪和黑麦草-百慕大混播草坪的日变化均呈单峰曲线,与气温、5 cm地温和10 cm地温的日变化趋势一致;4种草坪土壤呼吸对温度的敏感性指数即Q₁₀值为1.60～2.66;除结缕草外,其他草坪的土壤呼吸速率与土壤含水率相关性不显著;草坪的呼吸特征与其生长习性直接相关,而冷暖季混播草坪Q₁₀值小,对提高城市生态景观和环境质量有积极作用。     

西双版纳三叶橡胶林树干呼吸特征

采用红外气体分析法(IRGA)为期1a原位监测西双版纳三叶橡胶(Hevea brasiliensis)4个年龄段(7、15、27、40a)的树干呼吸情况,同时对每个年龄段树干监测2种高度(1.3 m-割胶部位、2.0 m-不割胶部位)和2个方向(南、北面)以及林内空气和树干1cm深温度.结果表明,4个年龄的树干呼吸有相同的季节规律,都是在雨季大于干季.林龄是影响橡胶树树干呼吸的一个重要因素,15、27a树干呼吸速率最大,分别为(4.989±0.278), (4.678±0.268) μmol·m-2·s-1,显著高于40a和7a树,40a树((3.753±0.205) μmol·m-2·s-1)显著大于7a树((2.299±0.129) μmol·m-2·s-1).所研究的高度和方向上树干呼吸速率无差异,割胶对树干表层破坏愈合后并不影响树干呼吸.树干呼吸与树干温度呈显著相关性,有良好的自然指数回归关系,Q10值为1.966～3.127,南北面Q10差异不明显,4个年龄段树干呼吸Q10值平均为2.452,大于已监测的热带树种.各年龄段橡胶林的主干(一级分枝以下部分树干)呼吸初步估算表明, 7、15、27a和40a橡胶树主干呼吸分别为1.74, 5.54, 7.53, 7.59 t C·hm-2·a-1.     

林木非同化器官与土壤呼吸的温度系数Q10值的特征分析

温度系数(Q₁₀,温度每变化10 ℃,呼吸速率的相对变化)不仅可以用来描述不同森林非同化器官(根系和树干)和土壤对温度升高的敏感性,并由此断定它们在全球变暖进程中的不同表现,而且是其呼吸总量定量估计中必不可少的参数。虽然目前已经进行了大量的研究,但不同研究者结论并不一致,影响我们对问题的整体把握。因此,有必要综合以往文献进行统计分析。该文综合大量文献,评述了林木非同化器官和土壤的Q₁₀值频率分布、不同研究方法对Q₁₀值的可能影响并探讨了它们对温度升高的敏感性。结果表明,不同非同化器官和土壤的Q₁₀值差异较大,但具有相对稳定的分布中心范围。其中,土壤呼吸Q₁₀值中,频率分布最集中的区域是2.0~2.5,占23%,其中超过80%的测定结果在1.0~4.0之间,中位数为2.74。 根系呼吸的Q₁₀值,频率分布最集中的区域2.5~3.0,占33%,而大部分(>80%)的研究结果在1.5~3.0之间,中位数为2.40。树干呼吸的Q₁₀值中,频率分布最集中的区域是1.5~2.0,占42%,而90%以上的测定结果在1.0~3.0之间,中位数为1.91。通过对比,发现不同非同化器官Q₁₀值不同(树干<根系<根系与土壤共同体<去除根系土壤)。其中树干和根系的Q₁₀值显著低于去除根系土壤的Q₁₀值(p<0.05),表明土壤微生物活动对于未来全球变暖的反应要比木质化器官更敏感。此外,常绿植物的根系和树干呼吸的Q₁₀值与落叶树木对应值差异不显著,说明同化器官叶片的着生时间长短对非同化器官Q₁₀的影响不大。不同的CO₂分析方法(碱吸收法,红外线测定技术和气相色谱方法)对土壤呼吸Q₁₀值测定结果的影响不显著(p>0.10),根系分离方法(断根测定和壕沟隔断测定)也对根系呼吸的Q₁₀值影响也不显著(p>0.10)。但是,与活体测定相比,离体测定树干呼吸显著提高了其Q₁₀值。总体来看,不同林分相同非同化器官以及不同非同化器官呼吸的Q₁₀值相对稳定但仍具有较大的差异性,研究方法也对结果产生一定影响,在进行呼吸总量的定量估计中应该注意这一点。今后研究的重点是进一步把影响森林非同化器官呼吸的外在因素和内在因素综合考虑于Q₁₀值相关模型中,以便准确定量估计其呼吸总量,而研究难点是深入研究Q₁₀值具有较大变异性的原因(如温度适应性)和内在机理以便更好的表征不同器官和生态系统组分对全球变暖的敏感性。     

武夷山不同海拔高度土壤氮矿化对温度变化的响应

采集了武夷山4个不同海拔的植物群落（常绿阔叶林、针叶林、亚高山矮林和高山草甸）的土壤样品,在实验室条件下, 将含水量调节为田间持水量60%,置于5 ℃、15 ℃、25 ℃和35 ℃人工气候箱中培养30 d,以测定土壤净氮矿化对温度的敏感性。结果表明：相同海拔植物群落的土壤净氮矿化量和氮矿化速率均随温度的升高显著增加;不同海拔间土壤氮矿化量和氮矿化速率大小均表现为：亚高山矮林>常绿阔叶林>高山草甸>针叶林。土壤氮矿化的Q₁₀在1.03～1.54,并且15 ℃升高到25 ℃时的Q₁₀比5 ℃升高到15 ℃和25 ℃升高到35 ℃时的Q₁₀高,表明土壤氮矿化对温度的敏感性在15 ℃～25 ℃较高。     

冻融交替对科尔沁沙地不同土地利用方式土壤呼吸的影响

在中高纬度和高海拔地区,冻融作用影响土壤的理化性质和微生物性状,进而影响土壤呼吸过程。研究冻融作用下土壤呼吸的变化,对准确估算全球碳循环具有重要意义。以科尔沁沙地沙质草地、樟子松疏林草地和农田为研究对象,通过冻融实验比较不同土地利用方式和冻融循环对土壤呼吸的影响。结果表明:土地利用方式对土壤呼吸有显著影响,在未发生冻融作用时沙质草地土壤呼吸速率显著大于樟子松疏林草地和农田(P0.05),3种土地利用方式的土壤呼吸平均速率分别为0.339、0.258和0.234μmolCO2.m-2.s-1;不同冻融循环对沙质草地和樟子松疏林草地土壤呼吸影响显著(P0.05)。其中,一次冻融循环条件下沙质草地、樟子松疏林草地和农田土壤呼吸平均速率分别为0.276、0.243和0.233μmolCO2.m-2.s-1,多次冻融循环条件下分别为0.314、0.274和0.259μmolCO2.m-2.s-1;沙质草地、樟子松疏林草地和农田的Q10值分别为116.0、26.2和16.4,表明冬季低温条件下土地利用方式强烈影响土壤呼吸对温度的敏感性。     

土壤水热条件对东北森林土壤表面CO2通量的影响

东北地区森林生态系统因其面积大,碳贮量高而在本地区和我国碳平衡中占有重要的地位.土壤表面CO2通量(RS)作为陆地生态系统向大气圈释放的主要CO2源,其时空变化直接影响到区域碳循环.该研究采用红外气体分析法比较测定我国东北东部次生林区6个典型的森林生态系统的RS及其相关的土壤水热因子,并深入分析土壤水热因子对RS的影响.研究结果表明影响RS的主要环境因子是土壤温度、土壤含水量及其交互作用,但其影响程度因生态系统类型和土壤深度而异.包括这些环境因子的综合RS模型解释了67.5%～90.6%的RS变异.在整个生长季中,不同生态系统类型的土壤温度差异不显著,而土壤湿度的差异显著(α=0.05).蒙古栎(Quercus mongolica)林、红松(Pinus koraiensis)林、落叶松(Larix gmelinii)林、硬阔叶林、杂木林和杨桦(Populus davidiana-Betula platyphylla)林的RS变化范围依次为1.89～5.23 μmol CO2·m-2·s-1,1.09～4.66μmol CO2·m-2·s-1,0.95～3.52 μmol CO2·m-2·s-1,1.13～5.97μmol CO2·m-2·s-1,1.05～6.58 μmol CO2·m-2·s-1和1.11～5.76μmol CO2·m-2·s-1.RS的季节动态主要受土壤水热条件的驱动而呈现单峰曲线,其变化趋势大致与土壤温度的变化相吻合.Q10从小到大依次为蒙古栎林2.32,落叶松林2.57,红松林2.76,硬阔叶林2.94,杨桦林3.54和杂木林3.55.Q10随土壤湿度的升高而增大;但超过一定的阈值后,土壤湿度对Q10起抑制作用.该研究结果强调对该地区生态系统土壤表面CO2通量的估测应同时考虑土壤水热条件的综合效应.     

Inhibition of oxidative stress by coenzyme Q10 increases mitochondrial mass and improves bioenergetic function in optic nerve head astrocytes

Oxidative stress contributes to dysfunction of glial cells in the optic nerve head (ONH). However, the biological basis of the precise functional role of mitochondria in this dysfunction is not fully understood. Coenzyme Q10 (CoQ₁₀), an essential cofactor of the electron transport chain and a potent antioxidant, acts by scavenging reactive oxygen species (ROS) for protecting neuronal cells against oxidative stress in many neurodegenerative diseases. Here, we tested whether hydrogen peroxide (100 μM H₂O₂)-induced oxidative stress alters the mitochondrial network, oxidative phosphorylation (OXPHOS) complex (Cx) expression and bioenergetics, as well as whether CoQ₁₀ can ameliorate oxidative stress-mediated alterations in mitochondria of the ONH astrocytes in vitro. Oxidative stress triggered the activation of ONH astrocytes and the upregulation of superoxide dismutase 2 (SOD2) and heme oxygenase-1 (HO-1) protein expression in the ONH astrocytes. In contrast, CoQ₁₀ not only prevented activation of ONH astrocytes but also significantly decreased SOD2 and HO-1 protein expression in the ONH astrocytes against oxidative stress. Further, CoQ₁₀ prevented a significant loss of mitochondrial mass by increasing mitochondrial number and volume density and by preserving mitochondrial cristae structure, as well as promoted mitofilin and peroxisome-proliferator-activated receptor-γ coactivator-1 protein expression in the ONH astrocyte, suggesting an induction of mitochondrial biogenesis. Finally, oxidative stress triggered the upregulation of OXPHOS Cx protein expression, as well as reduction of cellular adeonsine triphosphate (ATP) production and increase of ROS generation in the ONH astocytes. However, CoQ₁₀ preserved OXPHOS protein expression and cellular ATP production, as well as decreased ROS generation in the ONH astrocytes. On the basis of these observations, we suggest that oxidative stress-mediated mitochondrial dysfunction or alteration may be an important pathophysiological mechanism in the dysfunction of ONH astrocytes. CoQ₁₀ may provide new therapeutic potentials and strategies for protecting ONH astrocytes against oxidative stress-mediated mitochondrial dysfunction or alteration in glaucoma and other optic neuropathies.     

Climate variability and child height in rural Mexico

We examine the impacts of weather shocks, defined as rainfall or growing degree days, a cumulative measure of temperature, more than a standard deviation from their respective long run mean, on the stature of children between 12 and 47 months of age in Mexico. We find that after a positive rainfall shock children are shorter regardless of their region or altitude. Negative temperature shocks have a negative impact on height in the central and southern parts of the country as well as in higher altitudes. Although on average there are no statistically significant impacts from positive temperature shocks, certain sub-populations - namely boys, children between 12 and 23 months at the time of measurement, and children of less educated mothers - in some of the regions are negatively impacted. The results also suggest that potentially both agricultural income and communicable disease prevalence contribute to the effects.     

中国汉族人群ENPP1基因K121Q多态与2型糖尿病的关联及Meta分析

多个欧洲白人的Meta分析表明核苷酸焦磷酸酶1(Ectonucleotide pyrophosphatase/phosphodiesterase 1, ENPP1)基因K121Q多态与2型糖尿病相关联, 但在日本人、韩国人和中国台湾人的研究中没有发现相关性, 而在中国大陆人群中二者的关联研究结果不尽一致。文章调查了湖北地区539例2型糖尿病患者和404名正常人ENPP1基因K121Q多态性。基因型及等位基因频率在病例组和对照组间没有显著差异(P＞0.05), 但经性别、年龄和体重指数调整后的Logistic回归分析揭示XQ基因型与2型糖尿病显著相关(OR=1.5, 95%CI: 1.39~1.62, P<0.001)。对性别进行的亚组分析显示, 女性病例组Q等位基因和XQ基因型的频率显著高于对照组(Q: 12.4% vs. 6.1%, P=0.001; XQ: 23.7% vs. 11.7%, P=0.001)。结果表明ENPP1基因K121Q多态与湖北汉族人2型糖尿病的关联存在性别差异, 在女性中更明显。文章是对中国大陆人群进行的第一个Meta分析, 结果显示Q等位基因增加2型糖尿病的发病风险(OR=1.42, P=0.042)。     

Inhibition of mitochondrial electron transport by hydrophilic metal chelators. Determination of dehydrogenase topography

The topography of the inner mitochondrial membrane was investigated using inhibitors of electron transport on preparations of beef heart mitochondria and electron transport particles of opposite orientation. Reductions of juglone, ferricyanide, indophenol, coenzyme Q, duroquinone, and cytochrome c by NADH are inhibited to different extents on both sides of the membrane by the impermeant hydrophilic chelators bathophenanthroline sulfonate and orthophenanthroline. The extent of inhibition for each acceptor increased in the order given. At least two chelator-sensitive sites are present on each membrane face between the flavoprotein and coenzyme Q and a chelator-sensitive site is present on the matrix face between the sites of coenzyme Q and duroquinone interaction. Duroquinol oxidation in mitochondria only is stimulated by bathophenanthroline sulfonate. Juglone reduction is stimulated in electron transport particles (only) by p-hydroxymercuribenzenesulfonate, but after mercurial treatment, juglone reduction in both particles and mitochondria is more sensitive to bathophenanthroline sulfonate.Succinate dehydrogenase components are inhibited by hydrophilic orthophenanthroline or bathophenanthroline sulfonate in mitochondria only. Electron flow between the dehydrogenases of succinate and NADH occurs via a chelator-sensitive site located on the matrix face of the membrane. Inter-complex electron flow is prevented by rotenone or thenoyltrifluoroacetone. The lack of succinate-indophenol reductase inhibition by bathophenanthroline sulfonate in the presence of rotenone or thenoyltrifluoroacetone indicates that the rotenone-sensitive site may be located on the matrix face and demonstrates that electrons flow between the NADH and succinate dehydrogenases via a hydrophilic chelator and rotenone-thenoyltrifluoroacetone-sensitive site on the matrix face of the membrane. Inhibition by hydrophilic chelators only in mitochondria indicates that succinate dehydrogenase as well as NADH dehydrogenase has a transmembranous orientation.