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Guanosine Neuroprotective Action in Hippocampal Slices Subjected to Oxygen and Glucose Deprivation Restores ATP Levels, Lactate Release and Glutamate Uptake Impairment: Involvement of Nitric Oxide

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Abstract

Stroke is a major cause of disability and death worldwide. Oxygen and glucose deprivation (OGD) in brain tissue preparations can reproduce several pathological features induced by stroke providing a valuable ex vivo protocol for studying the mechanism of action of neuroprotective agents. Guanosine, an endogenous guanine nucleoside, promotes neuroprotection in vivo and in vitro models of neurotoxicity. We previously showed that guanosine protective effect was mimicked by inhibition of nitric oxide synthases (NOS) activity. This study was designed to investigate the involvement of nitric oxide (NO) in the mechanisms related to the protective role of guanosine in rat hippocampal slices subjected to OGD followed by reoxygenation (OGD/R). Guanosine (100 μM) and the pan-NOS inhibitor, l-NAME (1 mM) afforded protection to hippocampal slices subjected to OGD/R. The presence of NO donors, DETA-NO (800 μM) or SNP (5 μM) increased reactive species production, and abolished the protective effect of guanosine or l-NAME against OGD/R. Guanosine or l-NAME treatment prevented the impaired ATP production, lactate release, and glutamate uptake following OGD/R. The presence of a NO donor also abolished the beneficial effects of guanosine or l-NAME on bioenergetics and glutamate uptake. These results showed, for the first time, that guanosine may regulate cellular bioenergetics in hippocampal slices subjected to OGD/R injury by a mechanism that involves the modulation of NO levels.

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Abbreviations

ATP:

Adenosine-5′-triphosphate

DCFH:

2′,7′-Dichlorodihydrofluorescein

DETA-NO:

(Z)-1-[N-(2-Aminoethyl)-N-(2-ammonioethyl) amino] diazen-1-ium-1,2-diolate

OGD:

Oxygen and glucose deprivation

OGD/R:

Oxygen and glucose deprivation followed by reoxygenation

iNOS:

Inducible nitric oxide synthase

I/R:

Ischemia and reperfusion

KRB:

Krebs–Ringer bicarbonate buffer

L-NAME:

l-NG-nitro-l-arginine methyl ester

MTT:

3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

nNOS:

Neuronal nitric oxide synthase

NO:

Nitric oxide

NOS:

Nitric oxide synthase

SNP:

Sodium nitroprusside

14C-2-DG:

[U-14C]-2-deoxy-d-glucose

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Acknowledgements

Research supported by grants from the Brazilian funding agencies to C.I.T: CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico) –INCT for Excitotoxicity and Neuroprotection; CNPq Productivity Fellowship. D.T.T. is recipient of PhD fellowship from CAPES (Coordenação de Aperfeiçoamento de Pessoal de Ensino Superior). The authors would like to express their acknowledgments to Dr. A. Latini for her critical analysis.

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Authors and Affiliations

  1. Departamento de Bioquímica, Centro de Ciências Biológicas, Universidade Federal de Santa Catarina, Trindade, Florianópolis, SC, 88040-900, Brazil

    Daniel Tonial Thomaz, Rafaela Rafognatto Andreguetti, Luisa Bandeira Binder, Débora da Luz Scheffer, Alisson Willms Corrêa, Fátima Regina Mena Barreto Silva & Carla Inês Tasca

  2. Programa de Pós-Graduação em Bioquímica, Centro de Ciências Biológicas, Universidade Federal de Santa Catarina, Florianópolis, SC, Brazil

    Daniel Tonial Thomaz, Débora da Luz Scheffer, Fátima Regina Mena Barreto Silva & Carla Inês Tasca

  3. Programa de Pós-Graduação em Neurociências, Centro de Ciências Biológicas, Universidade Federal de Santa Catarina, Florianópolis, SC, Brazil

    Luisa Bandeira Binder & Carla Inês Tasca

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Thomaz, D.T., Andreguetti, R.R., Binder, L.B. et al. Guanosine Neuroprotective Action in Hippocampal Slices Subjected to Oxygen and Glucose Deprivation Restores ATP Levels, Lactate Release and Glutamate Uptake Impairment: Involvement of Nitric Oxide. Neurochem Res 45, 2217–2229 (2020). https://doi.org/10.1007/s11064-020-03083-2

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