Direct Evidence of Nitric Oxide Presence within Mitochondria

López-Figueroa, Manuel O.; Caamaño, Claudio A.; Morano, M. Inés; Rønn, Lars Christian Biilman; Akil, Huda; Watson, Stanley J.

doi:10.1006/bbrc.2000.2748

Cited by 115 publications

(70 citation statements)

References 36 publications

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“…of NO-related DAF-2 fluorescence previously attributed to NO formation within this particular cell organelle [26].…”

Section: Is Daf-2 Compartmentalized In Tissues?mentioning

confidence: 69%

“…A number of studies have reported on the nitrosation of DAF-2 for varying concentrations of NO donors, either in buffer solutions [9,10,20,21,30] using fluorometry or in cells/tissues [26,29] using fluorescence microscopy. …”

Section: Comparative Nitrosation Of Daf-2 In Buffer Solutions and Tismentioning

confidence: 99%

“…Furthermore, when used in conjunction with other probes, diamino fluorophores theoretically provide the option to test for co-localization of NO formation with other signaling events, such as a change in intracellular calcium concentration using Fura-2 [12,16,38], or its association with certain cell organelles by e.g. the use of MitoTracker to specifically stain for mitochrondria [26,50].…”

Section: Introductionmentioning

confidence: 99%

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Performance of diamino fluorophores for the localization of sources and targets of nitric oxide

Rodríguez

Specian

Maloney

et al. 2005

Free Radical Biology and Medicine

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An emergent approach to the detection of nitric oxide (NO) in tissues relies on the use of fluorescence probes that are activated by products of NO autoxidation. Here we explore the performance of the widely-used NO probe 4,5-diaminofluorescein diacetate (DAF-2 DA) for the localization of sources of NO in rat aortic tissue, either from endogenous NO synthesis or from chemically or photolytically-released NO from targets of nitrosation/nitrosylation. Of importance toward understanding the performance of this probe in tissues is the finding that, with incubation conditions commonly used in the literature (10 µM DAF-2 DA), intracellular DAF-2 accumulates to concentrations that approach the millimolar range. Whereas such high probe concentrations do not interfere with NO release or signaling, they help to clarify why DAF-2 nitrosation is possible in the presence of endogenous nitrosation scavengers (e.g. ascorbate and glutathione). The gain attained with such elevated concentrations is, however, mitigated by associated high levels of background autofluorescence from the probe. This, together with tissue autofluorescence, limits the sensitivity of the probe to low-micromolar levels of accumulated DAF-2 triazole (DAF-2 T), the activated form of the probe, which is higher than the concentrations of endogenous nitrosation/nitrosylation products found in tissues. We further show that the compartmentalization of DAF-2 around elastic fibers further limits its potential to characterize the site of NO production at the subcellular level. Moreover, we find that reaction of DAF-2 with HgCl 2 and other commonly employed reagents are associated with spectral changes that may be misinterpreted as NO signals. Finally, UV illumination can lead to high levels of nitrosating species that interfere with NO detection from enzymatic sources. These findings indicate that while DAF-2 may still represent an important tool for the localization of NO synthesis, provided important pitfalls and limitations are taken into consideration, it is not suited for the detection of basally-generated nitrosation/nitrosylation products.Performance of DAF-2 for NO Imaging 3

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“…of NO-related DAF-2 fluorescence previously attributed to NO formation within this particular cell organelle [26].…”

Section: Is Daf-2 Compartmentalized In Tissues?mentioning

confidence: 69%

Section: Comparative Nitrosation Of Daf-2 In Buffer Solutions and Tismentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Performance of diamino fluorophores for the localization of sources and targets of nitric oxide

Rodríguez

Specian

Maloney

et al. 2005

Free Radical Biology and Medicine

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“…Intriguingly, NO was reported to be locally produced by mitochondria (Ghafourifar et al, 1999;Lopez-Figueroa et al, 2000). Future investigation will clarify whether this locally produced NO (and its derivative peroxynitrite) induce any apoptosismodulatory e ects in addition to its regulatory e ect on oxygen consumption by inhibition of cytochrome c oxidase (Clementi et al, 1999).…”

Section: Discussionmentioning

confidence: 96%

The adenine nucleotide translocator: a target of nitric oxide, peroxynitrite, and 4-hydroxynonenal

et al. 2001

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“…All three NOS isoforms (eNOS, iNOS, and nNOS), as a key element in production of PN, have been shown to be up-regulated in brain tissue following TBI (Cobbs et al, 1997); (Wada et al, 1998) (Gahm et al, 2000); (Orihara et al, 2001). In addition, a novel isoform of Ca ++ -sensitive NOS (mtNOS), discovered within mitochondria (Bates et al, 1995); (Lopez-Figueroa et al, 2000); (Giulivi, 2003), has been shown to contribute to mitochondrial production of •NO and PN (Radi et al, 2002). The PN-derived free radicals (•NO 2 , •OH, •CO 3 − ) can induce extensive oxidative damage to cellular membranes, proteins and DNA (Beckman, 1996) (Murphy et al, 1998); (Radi, 1998).…”

Section: Introductionmentioning

confidence: 99%

Temporal relationship of peroxynitrite-induced oxidative damage, calpain-mediated cytoskeletal degradation and neurodegeneration after traumatic brain injury

Deng

Thompson

Gao

et al. 2007

Experimental Neurology

131

168

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We assessed the temporal and spatial characteristics of PN-induced oxidative damage and its relationship to calpain-mediated cytoskeletal degradation and neurodegeneration in a severe unilateral controlled cortical impact (CCI) traumatic brain injury (TBI) model. Quantitative temporal time-course studies were performed to measure two oxidative damage markers: 3-nitrotyrosine (3NT) and 4-hydroxynonenal (4HNE) at 30 min, 1, 3, 6, 12, 24, 48, 72 hrs, 7 days after injury in ipsilateral cortex of young adult male CF-1 mice. Secondly, the time course of Ca ++ -activated, calpain-mediated proteolysis was also analyzed using quantitative western-blot measurement of breakdown products of the cytoskeletal protein α-spectrin. Finally, the time course of neurodegeneration was examined using de Olmos silver staining. Both oxidative damage markers increased in cortical tissue immediately after injury (30 min) and elevated for the first 3-6 hrs before returning to baseline. In the immunostaining study, the PN-selective marker, 3NT, and the lipid peroxidation marker, 4HNE, were intense and overlapping in the injured cortical tissue. α-Spectrin breakdown products, which was used as biomarker for calpain-mediated cytoskeletal degradation, were also increased after injury, but the time course lagged behind the peak of oxidative damage and did not reach its maximum until 24 hrs post-injury. In turn, cytoskeletal degradation preceded the peak of neurodegeneration which occurred at 48 hrs post-injury. These studies have led us to the hypothesis that PN-mediated oxidative damage is an early event that contributes to a compromise of Ca ++ homeostatic mechanisms which causes a massive Ca ++ overload and calpain activation which is a final common pathway that results in post-traumatic neurodegeneration.

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Direct Evidence of Nitric Oxide Presence within Mitochondria

Cited by 115 publications

References 36 publications

Performance of diamino fluorophores for the localization of sources and targets of nitric oxide

Performance of diamino fluorophores for the localization of sources and targets of nitric oxide

The adenine nucleotide translocator: a target of nitric oxide, peroxynitrite, and 4-hydroxynonenal

Temporal relationship of peroxynitrite-induced oxidative damage, calpain-mediated cytoskeletal degradation and neurodegeneration after traumatic brain injury

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