Skip to main content
SpringerLink
Log in

Modulation of Cellular Respiration by Endogenously Produced Nitric Oxide in Rat Hippocampal Slices

  • Protocol
  • First Online:
Mitochondrial Bioenergetics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1782))

  • 3494 Accesses

Abstract

Nitric oxide (•NO) is an ubiquitous signaling molecule that participates in molecular processes associated with several neural phenomena ranging from memory formation to excitotoxicity. In the hippocampus, neuronal •NO production is coupled to the activation of NMDA type glutamate receptors. Cytochrome c oxidase has emerged as a novel target for •NO, which competes with O2 for binding to this mitochondrial complex. This reaction establishes •NO as a regulator of cellular metabolism and, possibly, mitochondrial production of reactive oxygen species which participate in cellular signaling. A major gap in the understanding of •NO bioactivity, namely, in the hippocampus, has been the lack of knowledge of its concentration dynamics. Here, we present a detailed description of the simultaneous recording of •NO and O2 concentration dynamics in rat hippocampal slices. Carbon fiber microelectrodes are fabricated and applied for real-time measurements of both gases in a system close to in vivo models. This approach allows for a better understanding of the current paradigm by which an intricate interplay between •NO and O2 regulates cellular respiration.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 109.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 199.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 199.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Laranjinha J, Santos RM, Lourenço CF, Ledo A, Barbosa RM (2012) Nitric oxide signaling in the brain: translation of dynamics into respiration control and neurovascular coupling. Ann N Y Acad Sci 1259(1):10–18

    Article  CAS  PubMed  Google Scholar 

  2. Santos RM, Lourenco CF, Gerhardt GA, Cadenas E, Laranjinha J, Barbosa RM (2011) Evidence for a pathway that facilitates nitric oxide diffusion in the brain. Neurochem Int 59(1):90–96. https://doi.org/10.1016/j.neuint.2011.05.016

    Article  PubMed  CAS  Google Scholar 

  3. Moncada S, Higgs A (1993) The L-arginine-nitric oxide pathway. N Engl J Med 329:2002–2012

    Article  CAS  PubMed  Google Scholar 

  4. Poulos TL (2006) Soluble guanylate cyclase. Curr Opin Struct Biol 16(6):736–743. https://doi.org/10.1016/j.sbi.2006.09.006

    Article  PubMed  CAS  Google Scholar 

  5. Kleppisch T, Feil R (2009) cGMP signalling in the mammalian brain: role in synaptic plasticity and behaviour. Handb Exp Pharmacol 191:549–579. https://doi.org/10.1007/978-3-540-68964-5_24

    Article  CAS  Google Scholar 

  6. Moncada S, Higgs EA (2006) Nitric oxide and the vascular endothelium. Handb Exp Pharmacol 176(Pt 1):213–254

    Article  Google Scholar 

  7. Lourenco CF, Ledo A, Barbosa RM, Laranjinha J (2017) Neurovascular-neuroenergetic coupling axis in the brain: master regulation by nitric oxide and consequences in aging and neurodegeneration. Free Radic Biol Med 108:668–682. https://doi.org/10.1016/j.freeradbiomed.2017.04.026

    Article  PubMed  CAS  Google Scholar 

  8. Poderoso JJ, Carreras MC, Lisdero C, Riobo N, Schopfer F, Boveris A (1996) Nitric oxide inhibits electron transfer and increases superoxide radical production in rat heart mitochondria and submitochondrial particles. Arch Biochem Biophys 328(1):85–92

    Article  CAS  PubMed  Google Scholar 

  9. Sarti P, Arese M, Forte E, Giuffre A, Mastronicola D (2012) Mitochondria and nitric oxide: chemistry and pathophysiology. Adv Exp Med Biol 942:75–92. https://doi.org/10.1007/978-94-007-2869-1_4

    Article  PubMed  CAS  Google Scholar 

  10. Bolanos JP, Peuchen S, Heales SJ, Land JM, Clark JB (1994) Nitric oxide-mediated inhibition of the mitochondrial respiratory chain in cultured astrocytes. J Neurochem 63:910–916

    Article  CAS  PubMed  Google Scholar 

  11. Brown GC, Cooper CE (1994) Nanomolar concentrations of nitric oxide reversibly inhibit synaptosomal respiration by competing with oxygen at cytochrome oxidase. FEBS Lett 356:295–298

    Article  CAS  PubMed  Google Scholar 

  12. Cleeter MW, Cooper JM, Darley-Usmar VM, Moncada S, Schapira AH (1994) Reversible inhibition of cytochrome c oxidase, the terminal enzyme of the mitochondrial respiratory chain, by nitric oxide: implications for neurodegenerative diseases. FEBS Lett 345:50–54

    Article  CAS  PubMed  Google Scholar 

  13. Schweizer M, Richter C (1994) Nitric oxide potently and reversibly deenergizes mitochondria at low oxygen tension. Biochem Biophys Res Commun 204:169–175

    Article  CAS  PubMed  Google Scholar 

  14. Mason MG, Nicholls P, Wilson MT, Cooper CE (2006) Nitric oxide inhibition of respiration involves both competitive (heme) and noncompetitive (copper) binding to cytochrome c oxidase. Proc Natl Acad Sci U S A 103(3):708–713. https://doi.org/10.1073/pnas.0506562103

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  15. Antunes F, Boveris A, Cadenas E (2004) On the mechanism and biology of cytochrome oxidase inhibition by nitric oxide. Proc Natl Acad Sci U S A 101:16774–16779

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Antunes F, Cadenas E (2007) The mechanism of cytochrome C oxidase inhibition by nitric oxide. Front Biosci 12:975–985

    Article  CAS  PubMed  Google Scholar 

  17. Cooper CE, Mason MG, Nicholls P (2008) A dynamic model of nitric oxide inhibition of mitochondrial cytochrome c oxidase. Biochim Biophys Acta 1777:867–876

    Article  CAS  PubMed  Google Scholar 

  18. Ledo A, Barbosa R, Cadenas E, Laranjinha J (2010) Dynamic and interacting profiles of •NO and O2 in rat hippocampal slices. Free Radic Biol Med 48(8):1044–1050. https://doi.org/10.1016/j.freeradbiomed.2010.01.024

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  19. Jarrard LE (1995) What does the hippocampus really do? Behav Brain Res 71(1–2):1–10

    Article  CAS  PubMed  Google Scholar 

  20. Garthwaite J, Garthwaite G, Palmer RM, Moncada S (1989) NMDA receptor activation induces nitric oxide synthesis from arginine in rat brain slices. Eur J Pharmacol 172(4–5):413–416

    Article  CAS  PubMed  Google Scholar 

  21. Prast H, Philippu A (2001) Nitric oxide as modulator of neuronal function. Prog Neurobiol 64(1):51–68

    Article  CAS  PubMed  Google Scholar 

  22. Hopper RA, Garthwaite J (2006) Tonic and phasic nitric oxide signals in hippocampal long-term potentiation. J Neurosci 26(45):11513–11521. https://doi.org/10.1523/JNEUROSCI.2259-06.2006

    Article  PubMed  CAS  Google Scholar 

  23. Laranjinha J, Ledo A (2007) Coordination of physiologic and toxic pathways in hippocampus by nitric oxide and mitochondria. Front Biosci 12:1094–1106

    Article  CAS  PubMed  Google Scholar 

  24. Zhuo M, Hawkins RD (1995) Long-term depression: a learning-related type of synaptic plasticity in the mammalian central nervous system. Rev Neurosci 6:259–277

    Article  CAS  PubMed  Google Scholar 

  25. Zorumski CF, Izumi Y (1998) Modulation of LTP induction by NMDA receptor activation and nitric oxide release. Prog Brain Res 118:173–182

    Article  CAS  PubMed  Google Scholar 

  26. Amaral DG, Witter MP (1989) The three-dimensional organization of the hippocampal formation: a review of anatomical data. Neuroscience 31(3):571–591

    Article  CAS  PubMed  Google Scholar 

  27. Anderson P, Bliss TV, Skrede KK (1971) Lamellar organization of hippocampal pathways. Exp Brain Res 13(2):222–238

    Google Scholar 

  28. Bliss TV, Collingridge GL (1993) A synaptic model of memory: long-term potentiation in the hippocampus. Nature 361(6407):31–39. https://doi.org/10.1038/361031a0

    Article  PubMed  CAS  Google Scholar 

  29. Gerhardt GA, Oke AF, Nagy G, Moghaddam B, Adams RN (1984) Nafion-coated electrodes with high selectivity for CNS electrochemistry. Brain Res 290(2):390–395

    Article  CAS  PubMed  Google Scholar 

  30. Santos RM, Lourenco CF, Piedade AP, Andrews R, Pomerleau F, Huettl P, Gerhardt GA, Laranjinha J, Barbosa RM (2008) A comparative study of carbon fiber-based microelectrodes for the measurement of nitric oxide in brain tissue. Biosens Bioelectron 24(4):704–709. https://doi.org/10.1016/j.bios.2008.06.034

    Article  PubMed  CAS  Google Scholar 

  31. Ferreira NR, Ledo A, Frade JG, Gerhardt GA, Laranjinha J, Barbosa RM (2005) Electrochemical measurement of endogenously produced nitric oxide in brain slices using Nafion/o-phenylenediamine modified carbon fiber microelectrodes. Anal Chim Acta 535(1–2):1–7

    Article  CAS  Google Scholar 

  32. Ledo A, Barbosa RM, Gerhardt GA, Cadenas E, Laranjinha J (2005) Concentration dynamics of nitric oxide in rat hippocampal subregions evoked by stimulation of the NMDA glutamate receptor. Proc Natl Acad Sci U S A 102(48):17483–17488. https://doi.org/10.1073/pnas.0503624102

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  33. Dias C, Lourenço CF, Ferreiro E, Barbosa RM, Laranjinha J, Ledo A (2016) Age-dependent changes in the glutamate-nitric oxide pathway in the hippocampus of the triple transgenic model of Alzheimer's disease: implications for neurometabolic regulation. Neurobiol Aging 46:84–95. https://doi.org/10.1016/j.neurobiolaging.2016.06.012

    Article  PubMed  CAS  Google Scholar 

  34. Ledo A, Lourenco CF, Caetano M, Barbosa RM, Laranjinha J (2015) Age-associated changes of nitric oxide concentration dynamics in the central nervous system of fisher 344 rats. Cell Mol Neurobiol 35(1):33–44. https://doi.org/10.1007/s10571-014-0115-0

    Article  PubMed  CAS  Google Scholar 

  35. Lourenço CF, Santos R, Barbosa RM, Gerhardt G, Cadenas E, Laranjinha J (2011) In vivo modulation of nitric oxide concentration dynamics upon glutamatergic neuronal activation in the hippocampus. Hippocampus 21(6):622–630

    Article  CAS  PubMed  Google Scholar 

  36. Ledo A, Lourenco CF, Laranjinha J, Brett CM, Gerhardt GA, Barbosa RM (2017) Ceramic-based multisite platinum microelectrode arrays: morphological characteristics and electrochemical performance for extracellular oxygen measurements in brain tissue. Anal Chem 89(3):1674–1683. https://doi.org/10.1021/acs.analchem.6b03772

    Article  PubMed  CAS  Google Scholar 

  37. Jiang C, Agulian S, Haddad GG (1991) O2 tension in adult and neonatal brain slices under several experimental conditions. Brain Res 568(1–2):159–164

    Article  CAS  PubMed  Google Scholar 

  38. Mulkey DK, Henderson RA 3rd, Olson JE, Putnam RW, Dean JB (2001) Oxygen measurements in brain stem slices exposed to normobaric hyperoxia and hyperbaric oxygen. J Appl Physiol 90(5):1887–1899

    Article  CAS  PubMed  Google Scholar 

  39. Barbosa RM, Lopes Jesus AJ, Santos RM, Pereira CL, Marques CF, Rocha BS, Ferreira NR, Ledo A, Laranjinha J (2011) Preparation, standardization and measurement of nitric oxide solutions. Global J Anal Chem 2(6):272–284

    CAS  Google Scholar 

  40. Friedemann MN, Robinson SW, Gerhardt GA (1996) o-Phenylenediamine-modified carbon fiber electrodes for the detection of nitric oxide. Anal Chem 68(15):2621–2628

    Article  CAS  PubMed  Google Scholar 

  41. Sander R (2015) Compilation of Henry's law constants (version 4.0) for water as solvent. Atmos Chem Phys 15(8):4399–4981. https://doi.org/10.5194/acp-15-4399-2015

    Article  CAS  Google Scholar 

  42. Gerhardt GA, Hoffman AF (2001) Effects of recording media composition on the responses of Nafion-coated carbon fiber microelectrodes measured using high-speed chronoamperometry. J Neurosci Methods 109(1):13–21

    Article  CAS  PubMed  Google Scholar 

  43. Rice ME (2000) Ascorbate regulation and its neuroprotective role in the brain. Trends Neurosci 23(5):209–216. https://doi.org/10.1016/S0166-2236(99)01543-X

    Article  PubMed  CAS  Google Scholar 

  44. Ferreira NR, Santos RM, Laranjinha J, Barbosa RM (2013) Real time in vivo measurement of ascorbate in the brain using carbon nanotube-modified microelectrodes. Electroanalysis 25(7):1757–1763. https://doi.org/10.1002/elan.201300053

    Article  CAS  Google Scholar 

  45. Ferreira NR, Lourenço CF, Barbosa RM, Laranjinha J (2015) Coupling of ascorbate and nitric oxide dynamics in vivo in the rat hippocampus upon glutamatergic neuronal stimulation: a novel functional interplay. Brain Res Bull 114:13–19. https://doi.org/10.1016/j.brainresbull.2015.03.002

    Article  PubMed  CAS  Google Scholar 

  46. Erecińska M, Silver IA (2001) Tissue oxygen tension and brain sensitivity to hypoxia. Respir Physiol 128(3):263–276

    Article  PubMed  Google Scholar 

Download references

Acknowledgments

This work was funded by FEDER via the COMPETE 2020 Program and National Funds via FCT (Fundação para a Ciência e Tecnologia, Portugal) through the strategical project POCI-01-0145-FEDER-007440.

Author information

Authors and Affiliations

  1. Center for Neurosciences and Cell Biology, University of Coimbra, Rua Larga, Coimbra, 3004-504, Portugal

    Ana Ledo, Rui M. Barbosa & João Laranjinha

  2. Faculty of Pharmacy, University of Coimbra, Health Science Campus, Azinhaga de Santa Comba, Coimbra, 3000-548, Portugal

    Rui M. Barbosa & João Laranjinha

Authors
  1. Ana Ledo
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Rui M. Barbosa
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. João Laranjinha
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ana Ledo .

Editor information

Editors and Affiliations

  1. Department of Life Sciences, University of Coimbra, Coimbra, Portugal

    Carlos M. Palmeira

  2. Department of Life Sciences, University of Coimbra, Coimbra, Portugal

    António J. Moreno

Rights and permissions

Reprints and permissions

Copyright information

© 2018 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Ledo, A., Barbosa, R.M., Laranjinha, J. (2018). Modulation of Cellular Respiration by Endogenously Produced Nitric Oxide in Rat Hippocampal Slices. In: Palmeira, C., Moreno, A. (eds) Mitochondrial Bioenergetics. Methods in Molecular Biology, vol 1782. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7831-1_5

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-7831-1_5

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7830-4

  • Online ISBN: 978-1-4939-7831-1

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation