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In situ and perfused rat livers showed a spon- .) The existence of such light emission, which should be termed "low-level chemiluminescence" to differentiate it from the more effective photoemission of the luciferin/luciferase systems (5, 6), was soon related to oxygendependent chain reactions involving biological lipids (3-5). This early work lay fallow for years, notwithstanding the reports by Stauff and Ostrowski on the chemiluminescence of mitochondria (7) and Howes and Steele on the chemiluminescence of microsomes (8, 9), both isolated from rat liver. The more recent reports by Nakano et al. (10) and Sugioka and Nakano (11) of light emission during lipid peroxidation and other oxidative reactions (12) in microsomes revived interest in the phenomenon and suggested chemiluminescence as a tool for the investigation of the radical reactions of lipid peroxidation under physiological conditions. We have recently reported that maximal light emission in isolated mitochondria and microsomes (13) and in submitochondrial particles (14) requires an electron transfer system, hydroperoxide, and oxygen, and that hydroperoxide-supplemented cytochrome c provides a chemiluminescent model system suitable for the elucidation of some of the molecular mechanisms responsible for light emission (15). On the other hand, isolated cells such as amoebae (16) and phagocytizing leukocytes (17) also have been found to be effective chemiluminescent sources.The most important aspect of the organ chemiluminescence is that it gives readily detectable, continuously monitorable, noninvasive signals of oxidative metabolism. This article explores the possibility of continuously monitoring the metabolism of exposed or fiberoptic probed organs in vivo by the chemiluminescent technique. In this paper we report the spontaneous and hydroperoxide-induced chemiluminescence of the in situ and perfused rat liver, as well as a partial spectral analysis of the chemiluminescence of the perfused liver. Light emission seems to indicate the generation of shkrt-lived free radicals and excited states derived from the side reactions of the free radical process of lipid peroxidation. A preliminary report on light emission has been published elsewhere (13). MATERIALS AND METHODSPhoton Counting. A single-photon-counting apparatus was used (Fig. 1). Both an EMI 9658 photomultiplier, responsive in the range 300-900 nm, with an applied potential of -1.2 kV (dark current: 20-30 counts per second), and an RCA 8850 photomultiplier, responsive in the range 300-650 nm, with an applied potential of -1.8 kV (dark current: 300-400 counts per second) were used. Phototube output was connected to an amplifier-discriminator (model 1121; Princeton Applied Res., Princeton, NJ) adjusted for single photon counting and connected to both a frequency counter (Heathkit IB 1100, Heath, Benton Harbor, MI) and a recorder. The EMI phototube, cooled .down to -400C by a thermoelectric cooler (EMI-Gencom, Plainview, NY) and the RCA phototube were placed in an Ortec housing, sealed and su...

The calory percentages of the ingredients in th:final regimen, i.e., basal diet plus ethanol solution, consumed by the animals of the alcohol group were as follows:-ethanol, 41.5%; carbohydrate> 42.6%; lipid, 6.6% and protein, 9.3%. The composition of the diet of the control group was the same except that the ethanol-derived calories were replaced by sucrose.The animals in the ethanol group \lere kept witbhout ethanol for 18 h prior to the experiment but were allowed access to the basal diet and to drinking water. Male

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The glutathione peroxidase-glutathione reductase system, an alternative pathway for metabolic utilization of H2O2 [Chance, Sies & Boveris (1979) Physiol. Rev. 59, 527-605], was investigated in Trypanosoma cruzi, an organism lacking catalase and deficient in peroxidase [Boveris & Stoppani (1977) Experientia 33, 1306-1308]. The presence of glutathione (4.9 +/- 0.7 nmol of reduced glutathione/10(8) cells) and NADPH-dependent glutathione reductase (5.3 +/- 0.4 munit/10(8) cells) was demonstrated in the cytosolic fraction of the parasite, but with H2O2 as substrate glutathione peroxidase activity could not be demonstrated in the same extracts. With t-butyl hydroperoxide or cumene hydroperoxide as substrate, a very low NADPH-dependent glutathione peroxidase activity was detected (equivalent to 0.3-0.5 munit of peroxidase/10(8) cells, or about 10% of glutathione reductase activity). Blank reactions of the glutathione peroxidase assay (non-enzymic oxidation of glutathione by hydroperoxides and enzymic oxidation of NADPH) hampered accurate measurement of peroxidase activity. The presence of superoxide dismutase and ascorbate peroxidase activity in, as well as the absence of catalase from, epimastigote extracts was confirmed. Ascorbate peroxidase activity was cyanide-sensitive and heat-labile, but no activity could be demonstrated with diaminobenzidine, pyrogallol or guaiacol as electron donor. The summarized results support the view that T. cruzi epimastigotes lack an adequate enzyme defence against H2O2 and H2O2-related free radicals.

A BSTRACT : Although the regulation of mitochondrial respiration and energy production in mammalian tissues has been exhaustively studied and extensively reviewed, a clear understanding of the regulation of cellular respiration has not yet been achieved. In particular, the role of tissue pO 2 as a factor regulating cellular respiration remains controversial. The concept of a complex and multisite regulation of cellular respiration and energy production signaled by cellular and intercellular messengers has evolved in the last few years and is still being researched. A recent concept that regulation of cellular respiration is regulated by ADP, O 2 and NO preserves the notion that energy demands drive respiration but places the kinetic control of both respiration and energy supply in the availability of ADP to F 1 -ATPase and of O 2 and NO to cytochrome oxidase. In addition, recent research indicates that NO participates in redox reactions in the mitochondrial matrix that regulate the intramitochondrial steady state concentration of NO itself and other reactive species such as superoxide radical (O 2 − ) and peroxynitrite (ONOO − ). In this way, NO acquires an essential role as a mitochondrial regulatory metabolite. NO exhibits a rich biochemistry and a high reactivity and plays an important role as intercellular messenger in diverse physiological processes, such as regulation of blood flow, neurotransmission, platelet aggregation and immune cytotoxic response.

Addition of beta-lapachone, an o-naphthoquinone endowed with trypanocidal properties to respiring Trypanosoma cruzi epimastigotes induced the release of O2- and H2O2 from the whole cells to the suspending medium. The same beta-lapachone concentration (4 micron) that released H2O2 at maximal rate completely inhibited T. cruzi growth in a liquid medium. The position isomer, alpha-lapachone, did not stimulate O2- and H2O2 release, and did not inhibit epimastigote growth. beta-Lapachone was able to stimulate H2O2 production by the epimastigote homogenate in the presence of NADH as reductant. The same effect was observed with the mitochondrial fraction supplemented with NADH, where beta-lapachone enhanced the generation of O2- and H2O2 4.5- and 2.5-fold respectively. beta-Lapachone also increased O2- and H2O2 production (2.5 and 2-fold respectively) by the microsomal fraction with NADPH as reductant. Cyanide-insensitive NADH and NADPH oxidation by the mitochondrial and microsomal fractions (quinone reductase activity) was stimulated to about the same extent by beta-lapachone. alpha-Lapachone was unable to increase O2- and H2O2 production and quinone reductase activity of the mitochondrial and microsomal fractions.

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