Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:1.9.3.1 (cytochrome oxidase)
 8,822 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Fluorescence of NADH and vascular volume of the brain cortex of chloralose-anesthetized cats were measured by surface fluororeflectometry. A cranial window and superfusion technique was elaborated for the topical inhibition of mitochondrial electron transport in the brain cortex by amytal (inhibits at site I) and cyanide (inhibits at site III). The changes in NAD/NADH redox state and CVV evoked by these electron transport inhibitors were compared with those elicited by anoxic anoxia. Amytal (10(-3)-10(-1) M) and cyanide (10(-5)-10(-2) M) resulted in a concentration-dependent and reversible increase in cortical NAD reduction and vascular volume, but the cerebrocortical vessels were almost completely dilatated long before maximum NAD reduction was reached. Cyanide at 10(-2) M increased cortical NAD reduction and vascular volume as much as anoxic anoxia. Amytal at 10(-1) M induced approximately half of the NAD reduction evoked by 10(-2) M cyanide or anoxic anoxia, but resulted in only slightly less vasodilatation than that following cyanide and anoxic anoxia. Since amytal inhibits mitochondrial electron transport at site I--and cyanide and anoxia at site III--but induces a comparable degree of vasodilatation, it is concluded that cytochrome oxidase cannot be the single molecular oxygen sensor in the brain cortex.
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PMID:A simple cranial window technique for optical monitoring of cerebrocortical microcirculation and NAD/NADH redox state. Effect of mitochondrial electron transport inhibitors and anoxic anoxia. 668 84

The MTT assay, which is widely used to measure cell proliferation and to screen for anticancer drugs, is based on reduction of the tetrazolium salt, MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) by actively growing cells to produce a blue formazan product. Despite broad acceptance of this assay, neither the subcellular localization, nor the biochemical events involved in MTT reduction are known. Mitochondrial involvement in MTT reduction has been inferred from studies with respiratory inhibitors using succinate as a substrate, but the contribution of this activity to overall cellular MTT reduction is unknown. Using the bone marrow-derived cell line, 32D, we investigated the subcellular localization of MTT reduction using succinate, NADH, and NADPH as substrates. At optimum substrate concentrations, MTT reduction by whole cell homogenates was greatest with NADH and least with succinate, which accounted for less than 10% of the combined activities. Using succinate, 96% of recoverable MTT reducing activity was in particulate fractions of the cell and 77% in the mitochondrial and light mitochondrial/lysosomal fractions. When NADH and NADPH were used as substrates, increased amounts of MTT reducing activity were associated with soluble fractions of the cell and association with mitochondrial fractions was less pronounced. To further characterize MTT reduction by the mitochondrial fraction, respiratory chain inhibitors were used to explore involvement of electron transport in MTT reduction. Succinate-dependent mitochondrial MTT reduction was inhibited by 80% with chlorpromazine, 70% by antimycin A, and 85-90% by thenoyltrifluoracetone (TTFA), but inhibition was not observed with rotenone at < or = 2 microM, Amytal, or azide. These results suggest that when succinate is used as an electron donor, 70-80% of mitochondrial MTT reduction occurs subsequent to transfer of electrons from cytochrome c to cytochrome oxidase, but prior to the point of azide inhibition. In contrast to succinate, NADPH-dependent mitochondrial MTT reduction was not affected by any of the respiratory inhibitors tested, and NADH-dependent reduction was only inhibited by chlorpromazine (40-50% at plateau concentrations). These results suggest that most cellular MTT reduction occurs outside the mitochondrial inner membrane and involves NADH and NADPH-dependent mechanisms that are insensitive to respiratory chain inhibitors. This interpretation is supported by whole cell studies in which rotenone failed to affect basal and interleukin-3-stimulated MTT reduction at times up to 4 h but strongly inhibited DNA synthesis. We conclude that most cellular reduction of MTT occurs extramitochondrially and probably involves the pyridine nucleotide cofactors NADH and NADPH.
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PMID:Characterization of the cellular reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT): subcellular localization, substrate dependence, and involvement of mitochondrial electron transport in MTT reduction. 839 Feb 25

Cardiac ischemia damages the mitochondrial electron transport chain. Irreversible blockade of electron transport at complex I by rotenone decreases ischemic damage to cardiac mitochondria by decreasing the loss of cytochrome c and preserving respiration through cytochrome oxidase. Therapeutic intervention to protect myocardium during ischemia and reperfusion requires the use of a reversible inhibitor that allows resumption of oxidative metabolism during reperfusion. Amobarbital is a reversible inhibitor at the rotenone site of complex I. We asked whether amobarbital administered immediately before ischemia protected respiratory function. Isolated rat hearts were perfused for 15 min followed by 25-min global ischemia at 37 degrees C. Amobarbital-treated hearts received drug for 1 min before ischemia. Subsarcolemmal (SSM) and interfibrillar (IFM) populations of mitochondria were isolated after ischemia, and oxidative phosphorylation was measured. Amobarbital protected oxidative phosphorylation, including through cytochrome oxidase, in both SSM and IFM in a dose-dependent manner, with an optimal dose of 2 to 2.5 mM. Amobarbital also preserved cytochrome c content in both SSM and IFM. Thus, reversible blockade of the electron transport chain during ischemia protects mitochondrial respiration.
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PMID:Blockade of electron transport before cardiac ischemia with the reversible inhibitor amobarbital protects rat heart mitochondria. 1617 99

Ischemia damages the mitochondrial electron transport chain (ETC), mediated in part by damage generated by the mitochondria themselves. Mitochondrial damage resulting from ischemia, in turn, leads to cardiac injury during reperfusion. The goal of the present study was to localize the segment of the ETC that produces the ischemic mitochondrial damage. We tested if blockade of the proximal ETC at complex I differed from blockade distal in the chain at cytochrome oxidase. Isolated rabbit hearts were perfused for 15min followed by 30min stop-flow ischemia at 37 degrees C. Amobarbital (2.5mM) or azide (5mM) was used to block proximal (complex I) or distal (cytochrome oxidase) sites in the ETC. Time control hearts were buffer-perfused for 45min. Subsarcolemmal mitochondria (SSM) and interfibrillar mitochondria (IFM) were isolated. Ischemia decreased cytochrome c content in SSM but not in IFM compared to time control. Blockade of electron transport at complex I preserved the cytochrome c content in SSM. In contrast, blockade of electron transport at cytochrome oxidase with azide did not retain cytochrome c in SSM during ischemia. Since blockade of electron transport at complex III also prevented cytochrome c loss during ischemia, the specific site that elicits mitochondrial damage during ischemia is likely located in the segment between complex III and cytochrome oxidase.
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PMID:Isolating the segment of the mitochondrial electron transport chain responsible for mitochondrial damage during cardiac ischemia. 2052 65