Neural progenitor cells are widespread throughout the adult central nervous system but only give rise to neurons in specific loci. Negative regulators of neurogenesis have therefore been postulated, but none have yet been identified as subserving a significant role in the adult brain. Here we report that nitric oxide (NO) acts as an important negative regulator of cell proliferation in the adult mammalian brain. We used two independent approaches to examine the function of NO in adult neurogenesis. In a pharmacological approach, we suppressed NO production in the rat brain by intraventricular infusion of an NO synthase inhibitor. In a genetic approach, we generated a null mutant neuronal NO synthase knockout mouse line by targeting the exon encoding active center of the enzyme. In both models, the number of new cells generated in neurogenic areas of the adult brain, the olfactory subependyma and the dentate gyrus, was strongly augmented, which indicates that division of neural stem cells in the adult brain is controlled by NO and suggests a strategy for enhancing neurogenesis in the adult central nervous system. T he vast majority of neurons in the mammalian brain are produced during embryonic development. However, remnants of the germinal zones of the developing brain continue to proliferate into adulthood, generating large numbers of neurons in the adult brain (1-3). The subventricular zone (SVZ) of the lateral ventricles (LVs), its anterior extension, the rostral migratory stream (RMS), and the subgranular cell layer (S-GCL) of the dentate gyrus (DG) of the hippocampus are the major sites of adult neurogenesis, although other regions of the adult brain retain the potential to generate new neurons (4-6). Many of the newly generated neurons undergo physiological cell death (7), but it is becoming clear that some of these new neurons become integrated into existing neuronal circuits, thus potentially contributing to a previously unanticipated form of neuroplasticity (8). Several protein growth factors have been shown to affect adult neurogenesis in vivo (5, 6, 9-11). However, the signaling systems involved in regulating cell division in the adult brain are only beginning to be understood.Increasingly diverse functions of NO, a transcellular signaling molecule (12), are continuing to be demonstrated, and there is growing evidence that NO may be involved in controlling proliferation of neuronal cells. Neuronal NO synthase (nNOS), the major NOS isoform in the mammalian brain, is transiently expressed in the developing brain in a pattern suggesting its involvement in neural development (13). Furthermore, NO has been shown to effectively and reversibly suppress cell division (14, 15); this property of NO, coupled to its ability to regulate gene expression, is exploited in a number of developmental contexts (16). Materials and MethodsA full description of the methods used in this work can be found in Supporting Materials and Methods, which is published as supporting information on the PNAS web site, www.pnas.org.All a...

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Mutations and deletions in mitochondrial DNA (mtDNA) lead to a number of human diseases characterized by neuromuscular degeneration. Accumulation of truncated mtDNA molecules (∆-mtDNA) lacking a specific 4977-bp fragment, the common deletion, leads to three related mtDNA diseases : Pearson's syndrome; Kearns-Sayre syndrome; and chronic progressive external ophthalmoplegia (CPEO). In addition, the proportion of ∆-mtDNA present increases with age in a range of tissues. Consequently, there is considerable interest in the effects of the accumulation of ∆-mtDNA on cell function. The 4977-bp deletion affects genes encoding 7 polypeptide components of the mitochondrial respiratory chain, and 5 of the 22 tRNAs necessary for mitochondrial protein synthesis. To determine how the accumulation of ∆-mtDNA affects oxidative phosphorylation we constructed a series of cybrids by fusing a human osteosarcoma cell line depleted of mtDNA (ρ 0 ) with enucleated skin fibroblasts from a CPEO patient. The ensuing cybrids contained 0Ϫ86 % ∆-mtDNA and all had volumes, protein contents, plasma-membrane potentials and mitochondrial contents similar to those of the parental cell line. The bioenergetic consequences of accumulating ∆-mtDNA were assessed by measuring the mitochondrial membrane potential, rate of ATP synthesis and ATP/ADP ratio. In cybrids containing less than 50Ϫ55% ∆-mtDNA, these bioenergetic functions were equivalent to those of cybrids with intact mtDNA. However, once the proportion of ∆-mtDNA exceeded this threshold, the mitochondrial membrane potential, rate of ATP synthesis, and cellular ATP/ADP ratio decreased. These bioenergetic deficits will contribute to the cellular pathology associated with the accumulation of ∆-mtDNA in the target tissues of patients with mtDNA diseases.

Photosynthetic microbial fuel cells (PMFCs) are an emerging technology for renewable solar energy conversion. Major efforts have been made to explore the electrogenic activity of cyanobacteria, mostly using practically unsustainable reagents. Here we report on photocurrent generation (≈8.64 μA cm(-2)) from cyanobacteria immobilized on electrodes modified with an efficient electron mediator, an Os(2+/3+) redox polymer. Upon addition of ferricyanide to the electrolyte, cyanobacteria generate the maximum current density of ≈48.2 μA cm(-2).

Superoxide reacts with nitric oxide to form peroxynitrite, a potent oxidising agent which may contribute to tissue damage in pathological situations such as inflammation and ischaemia/reperfusion. One mechanism by which oxidative stress damages tissues is the induction of a specitic Cyclosporin A-sensitive mitochondrial calcium eflhrx pathway. Here we show that peroxynitrite induces calcium efflux from mammalian mitochondria and that this efflux is blocked by Cyclosporin A. These dam suggest that disruption of mitochondrial calcium eBhtx may contribute to tissue damage when superoxide and nitric oxide are present together in vivo.Key words: Peroxynitrite; Mitochondrial oxidative damage; Calcium efi%rx; Cyclosporin A; Mitochondrial permeability transition IIltroductloIlNitric oxide ('NO) is formed in many mammalian tissues where it acts as a chemical messenger [l] and superoxide (O,'-) is a by product of normal metabolism [2]. The reaction of 02'-with 'NO produces the potent oxidant peroxynitrite (ONOO-) [3,4] which contributes to tissue damage in inflammation and ischaemia/reperfusion where significant amounts of 'NO and Or*-are produced [4,5]. At 37"C, the pK, of ONOO-is 6.8 and the half life of its protonated form, peroxynitrous acid, is less than 1 second [6]. Peroxynitrous acid spontaneously cleaves to produce the powerful oxidants nitrogen dioxide and the hydroxyl radical [4,7]. The ONOO-anion itself oxidises thiols to distides [7] and therefore formation of ONOO-from 'NO and 02'-may lead to oxidative damage in vivo by a number of mechanisms.Mitochondria are an important target for oxidative damage by ONOO-since they produce 02'- [2]. Therefore exposure to 'NO can lead to the formation of ONOO-within mitochondria [6]. Furthermore, it has been shown that 'NO and ONOO-disrupt mitochondrial function [6,8]. In addition to directly inactivating mitochondrial enzymes [9, lo], many oxidants disrupt mitochondrial calcium metabolism by inducing a specific mitochondrial calcium efflux pathway [ 111. This calcium efflux is associated with, or leads to, the opening of a non-specific pore in the mitochondrial inner membrane which depolarises the mitochondrion pathway [l 1,121. While the role of calcium efflux and pore opening is unclear, it may contribute to cell damage following oxidative stress as CsA protects cells from oxidants [13].We have investigated whether or not oxidative damage by ONOO-disrupts mitochondrial calcium transport. To do this we measured calcium efflux and swelling in isolated rat liver mitochondria exposed to ONOO-and found that ONOO-induces mitochondrial calcium efflux and opening of the inner membrane pore, both of which can be prevented by CsA. Materials and methods MaterialsArsenaxo III, superoxide dismutase and catalase were from Sigma. "CaClz was from Amersham. Cyclosporin A was a kind gift from Sandoz Pharma Ltd., Basel, Switzerland. Preparation of mitochondriaLiver mitochondria were prepared from fed female Wistar rats (150-200 g) by homogenisation and differential centrifugatio...

Studies on biological photovoltaics based on intact organisms are challenging and in most cases include diffusing mediators to facilitate electrochemical communication with electrodes. However, using such mediators is impractical. Instead, surface confined Os‐polymers have been successfully used in electrochemical studies including oxidoreductases and bacterial cells but not with algae. Photoelectrogenic activity of a green alga, Paulschulzia pseudovolvox, immobilized on graphite or Os‐polymer modified graphite is demonstrated. Direct electron transfer is revealed, when no mediator is added, between algae and electrodes with electrons emerging from photolysis of water via the cells to the electrode exhibiting a photocurrent density of 0.02 μA cm−2. Os‐polymers with different redox potentials and structures are used to optimize the energy gap between the photosynthetic complexes of the cells and the Os‐polymers and those of greater solubility, better accessibility with membranes, and relatively higher potentials yielded a photocurrent density of 0.44 μA cm−2. When benzoquinone is included to the electrolyte, the photocurrent density reaches 6.97 μA cm−2. The photocurrent density is improved to 11.50 μA cm−2, when the cells are protected from reactive oxygen species when either superoxide dismutase or catalase is added. When adding an inhibitor specific for photosystem II, diuron, the photocurrent is decreased by 50%.

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Over the last decades, several studies have reported emissions of nitrous oxide (N O) from microalgal cultures and aquatic ecosystems characterized by a high level of algal activity (e.g. eutrophic lakes). As N O is a potent greenhouse gas and an ozone-depleting pollutant, these findings suggest that large-scale cultivation of microalgae (and possibly, natural eutrophic ecosystems) could have a significant environmental impact. Using the model unicellular microalga Chlamydomonas reinhardtii, this study was conducted to investigate the molecular basis of microalgal N O synthesis. We report that C. reinhardtii supplied with nitrite (NO ) under aerobic conditions can reduce NO into nitric oxide (NO) using either a mitochondrial cytochrome c oxidase (COX) or a dual enzymatic system of nitrate reductase (NR) and amidoxime-reducing component, and that NO is subsequently reduced into N O by the enzyme NO reductase (NOR). Based on experimental evidence and published literature, we hypothesize that when nitrate (NO ) is the main Nitrogen source and the intracellular concentration of NO is low (i.e. under physiological conditions), microalgal N O synthesis involves the reduction of NO to NO by NR followed by the reduction of NO to NO by the dual system involving NR. This microalgal N O pathway has broad implications for environmental science and algal biology because the pathway of NO assimilation is conserved among microalgae, and because its regulation may involve NO.