In situ measurement of neuronal nitric oxide synthase activity in the spinal cord by NADPH-diaphorase histochemistry

Xu, Li; Matsumura, Shinji; Mabuchi, Tamaki; Takagi, Kunio; Abe, Tsuneyuki; Ito, Seiji

doi:10.1016/j.jneumeth.2005.06.017

Cited by 15 publications

(12 citation statements)

References 45 publications

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“…Our results showed that NADPH-d activity was not altered in the pericentral region. In accordance with these localizations, it is well accepted that NO is involved in nociceptive processing and persistent pain as both intra-and intercellular messenger in the spinal cord (Xu et al, 2006). In both the RA group and the control group, no NADPH-dstained neurons were found in the ventral horn at P1 or P21.…”

Section: Discussionsupporting

confidence: 56%

“…NO as a gaseous neurotransmitter can infl uence synaptic transmission, plasticity, neurotoxicity, and develop ment in the CNS (Li et al, 2006). In laminae I and II in mice, there was no or little expression of nNOS by P10, while NADPH-d activity in the superfi cial layer of the spinal cord increased gradually with age from P10 to P30 (Xu et al, 2006). It is well known that superfi cial dorsal horn neurons are the last spinal neurons to mature (Fitzgerald, 2005).…”

Section: Discussionmentioning

confidence: 99%

“…Through production of NO, the enzyme is involved in the promotion of axonal elongation and in the regulation of the advancement of the growth cone ( Van-Wagenen and Rehder, 1999) as well as in triggering the cell death in the nervous system during development (Iadecola, 1997) ing to nitrosative stress (Freire et al, 2009). In the spinal cord, NADPH-d activity has been detected in the dorsal horn, around the central canal, and at the intermediolateral cell column of the spinal cord (Xu et al, 2006).…”

Section: Introductionmentioning

confidence: 99%

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Postnatal Development of Spinal Cord and Liver Antioxidant Status in the Young of Retinol-Overdosed Female Rats

Patlevič

Vašková

Vaško

et al. 2013

Zeitschrift Für Naturforschung C

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The metabolic form of vitamin A, retinol, has a pivotal role in the nervous system development and neuronal differentiation, both during embryogenesis through maternal-fetal support and in the early postnatal life. Retinoic acid was administered orally at a dose of 10 mg/kg body weight to pregnant female rats through days 8 -10 of gestation. Spinal cord sections were processed for histochemical visualization one day after birth and on day 21, when weaning is expected. NADPH-diaphorase (NADPH-d)-positive neurons were found in the dorsal horn, around the central canal, and at the intermediolateral cell column on postnatal days 1 and 21 in both control and experimental groups. There were no NADPHd-positive structures in the ventral horn. The results suggest that prenatal administration of high doses of retinoic acid is not associated with postnatal morphological changes in NADPH-d-positive neurons in the rat spinal cord. Levels of antioxidants and related enzymes in retinoid storage organs were measured to estimate possible side effects. The activities of enzymes detoxifying superoxide radicals and peroxides were supressed after birth. A decrease in the level of reduced glutathione was observed on postnatal day 21, indicating an unbalanced redox environment.

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Section: Discussionsupporting

confidence: 56%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Postnatal Development of Spinal Cord and Liver Antioxidant Status in the Young of Retinol-Overdosed Female Rats

Patlevič

Vašková

Vaško

et al. 2013

Zeitschrift Für Naturforschung C

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“…Based on previous reports indicating that paraformaldehyde-resistant NADPH-diaphorase (NADPH-d) activity is identical to neuronal NOS (nNOS) in the central nervous system [25,26], we demonstrated earlier that phosphorylation of NMDA receptor subunit NR2B at its Tyr 1472 and subsequent nNOS activation are involved in the maintenance of neuropathic pain [27-29]. Furthermore, to elucidate biochemical and molecular mechanisms for nNOS activation in the spinal cord, we established an ex vivo system for NADPH-d histochemistry [30], which enabled us to clarify whether NO itself regulated nNOS activity in situ by use of NO donors, NOR1, NOR3 and SNAP, with different half-lives of 1.8 min, 30 min, and 6 h, respectively [31]. Both NOR1 and NOR3 (100 μM), but not SNAP, enhanced the cGMP level 10- to 15-fold in isolated spinal cords in the presence of IBMX.…”

Section: Discussionmentioning

confidence: 99%

Rapid S-Nitrosylation of Actin by NO-Generating Donors and in Inflammatory Pain Model Mice

Lu¹,

Katano²,

Uta

et al. 2011

Mol Pain

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BackgroundS-Nitrosylation, the reversible post-translational modification of reactive cysteine residues in proteins, has emerged as an important mechanism by which NO acts as a signaling molecule. We recently demonstrated that actin is a major S-nitrosylated protein in the spinal cord and suggested that NO directly attenuates dopamine release from PC12 cells by causing the breakdown of F-actin. However, the occurrence of S-nitrosylation of actin remained unclarified in animal pain model. Kinetic analysis of S-nitrosylation of actin in the present study was made by using NO-generating donors. The biotin-switch assay and purification on streptavidin-agarose were employed for identification of S-nitrosylated actin.ResultsDopamine release from PC12 cells was markedly attenuated by NOR1 (t1/2 = 1.8 min) and much less by NOR3 (t1/2 = 30 min), but not by S-nitroso-glutathione, an endogenous NO donor. A membrane-permeable cGMP analogue could not substitute for NOR1 as a suppressor nor could inhibitors of soluble guanylate cyclase and cGMP-dependent protein kinase attenuate the suppression. S-Nitrosylated actin was detected by the biotin-switch assay at 5 min after the addition of NOR1. Consistent with the kinetic analysis, actin in the spinal cord was rapidly and maximally S-nitrosylated in an inflammatory pain model at 5 min after the injection of 2% formalin into the hind paws. In vivo patch-clamp recordings of the spinal dorsal horn, NOR3 showed an inhibitory action on inhibitory synaptic transmission in interneurons of the substantia gelatinosa.ConclusionsThe present study demonstrates that rapid S-nitrosylation of actin occurred in vitro in the presence of exogenous NO-generating donors and in vivo in inflammatory pain model mice. Our data suggest that, in addition to the well-known cGMP-dependent protein kinase pathway, S-nitrosylation is involved in pain transmission via disinhibition of inhibitory neurons.

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“…Procedures for histochemical examinations followed essentially those described in our previous studies. 4,10) Mice were anesthetized with pentobarbital sodium and perfused transcardially with phosphate-buffered saline (PBS) followed by 4% paraformaldehyde (Nacalai Tesque) in phosphate buffer. Then the brain was isolated and post-fixed overnight with 4% paraformaldehyde, then immersed in 15% sucrose.…”

Section: Histochemical Examinationsmentioning

confidence: 99%

Endogenous Nitric Oxide Inhibits, Whereas Awakening Stimuli Increase, the Activity of a Subset of Orexin Neurons

Yamakawa

Kurauchi

Hisatsune

et al. 2018

Biological & Pharmaceutical Bulletin

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The lateral hypothalamic area contains neurons expressing neuronal nitric oxide synthase (nNOS), in addition to orexin neurons. Here we examined whether the activity of orexin neurons was regulated by endogenous nitric oxide (NO) in male C57BL/6 mice. Caffeine (30 mg/kg, intraperitoneally (i.p.)) increased the number of orexin neurons positive for c-Fos, a marker of neuronal activity, and also increased the number of NOS/c-Fos-positive cells as identified by reduced nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase histochemistry and c-Fos immunohistochemistry. Diphenhydramine hydrochloride (10 mg/kg. i.p.) decreased c-Fos-positive orexin neurons but had no significant effect on the number of c-Fos-positive NOS neurons. nNOS inhibitor 7-nitroindazole (25 mg/kg, i.p.) alone increased c-Fos-positive orexin neurons, and combined treatment with caffeine and 7-nitroindazole did not show additive effect in the number of c-Fospositive orexin neurons. In contrast, 7-nitroindazole decreased c-Fos-positive NOS neurons and attenuated caffeine-induced increase in c-Fos-positive NOS neurons. Sleep deprivation increased c-Fos-positive cells in both orexin neurons and NOS neurons, and 7-nitroindazole did not show additive effect with sleep deprivation in the activation of orexin neurons. Together, these results suggest that endogenous NO negatively regulates the activity of a subset of orexin neurons, and this subset of orexin neurons overlaps with that activated by awakening stimuli.

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In situ measurement of neuronal nitric oxide synthase activity in the spinal cord by NADPH-diaphorase histochemistry

Cited by 15 publications

References 45 publications

Postnatal Development of Spinal Cord and Liver Antioxidant Status in the Young of Retinol-Overdosed Female Rats

Postnatal Development of Spinal Cord and Liver Antioxidant Status in the Young of Retinol-Overdosed Female Rats

Rapid S-Nitrosylation of Actin by NO-Generating Donors and in Inflammatory Pain Model Mice

Endogenous Nitric Oxide Inhibits, Whereas Awakening Stimuli Increase, the Activity of a Subset of Orexin Neurons

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