Skip to main content

Fenamates Inhibit Human Sodium Channel Nav1.2 and Protect Glutamate-Induced Injury in SH-SY5Y Cells

Abstract

Voltage-gated sodium channels are crucial mediators of neuronal damage in ischemic and excitotoxicity disease models. Fenamates have been reported to have anti-inflammatory properties following a decrease in prostaglandin synthesis. Several researches showed that fenamates appear to be ion channel modulators and potential neuroprotectants. In this study, the neuroprotective effects of tolfenamic acid, flufenamic acid, and mefenamic acid were tested by glutamate-induced injury in SH-SY5Y cells. Following this, fenamates’ effects were examined on both the expression level and the function of hNav1.1 and hNav1.2, which were closely associated with neuroprotection, using Western blot and patch clamp. Finally, the effect of fenamates on the expression of apoptosis-related proteins in SH-SY5Y cells was examined. The results showed that both flufenamic acid and mefenamic acid exhibited neuroprotective effects against glutamate-induced injury in SH-SY5Y cells. They inhibited peak currents of both hNav1.1 and hNav1.2. However, fenamates exhibited decreased inhibitory effects on hNav1.1 when compared to hNav1.2. Correspondingly, the inhibitory effect of fenamates was found to be consistent with the level of neuroprotective effects in vitro. Fenamates inhibited glutamate-induced apoptosis through the modulation of the Bcl-2/Bax-dependent cell death pathways. Taken together, Nav1.2 might play a part in fenamates’ neuroprotection mechanism.

Graphic Abstract

Nav1.2 and NMDAR might take part in the neuroprotection mechanism of the fenamates. The fenamates inhibited glutamate-induced apoptosis through modulation of the Bcl-2/Bax-dependent cell death pathways.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Abbreviations

CNS:

Central nervous system

VGSC:

Voltage-gated sodium channel

NSAIDs:

Nonsteroidal anti-inflammatory drugs

DRG:

Dorsal root ganglion

ROS:

Reactive oxygen species

MCAO:

Middle cerebral artery occlusion

DMEM:

Dulbecco's modified Eagle medium

IMDM:

Iscove's modified Dulbecco's medium

CHO:

Chinese hamster ovary

FBS:

Fetal bovine serum

ATCC:

American Type Culture Collection

References

  1. Chen Q, Olney JW, Lukasiewicz PD, Almli T, Romano C (1998) Fenamates protect neurons against ischemic and excitotoxic injury in chick embryo retina. Neurosci Lett 242:163–166

    CAS  Article  Google Scholar 

  2. Daniels MJ, Rivers-Auty J, Schilling T, Spencer NG, Watremez W, Fasolino V, Booth SJ, White CS, Baldwin AG, Freeman S, Wong R, Latta C, Yu S, Jackson J, Fischer N, Koziel V, Pillot T, Bagnall J, Allan SM, Paszek P, Galea J, Harte MK, Eder C, Lawrence CB, Brough D (2016) Fenamate NSAIDs inhibit the NLRP3 inflammasome and protect against Alzheimer's disease in rodent models. Nat Commun 7:12504

    CAS  Article  Google Scholar 

  3. Florio SK, Loh C, Huang SM, Iwamaye AE, Kitto KF, Fowler KW, Treiberg JA, Hayflick JS, Walker JM, Fairbanks CA, Lai Y (2009) Disruption of nNOS-PSD95 protein-protein interaction inhibits acute thermal hyperalgesia and chronic mechanical allodynia in rodents. Br J Pharmacol 158:494–506

    CAS  Article  Google Scholar 

  4. Flower RJ (1974) Drugs which inhibit prostaglandin biosynthesis. Pharmacol Rev 26:33–67

    CAS  PubMed  Google Scholar 

  5. Flower R, Gryglewski R, Herbaczyn′ ska-Cedro K, Vane JR (1972) Effects of anti-inflammatory drugs on prostaglandin biosynthesis. Nat New Biol 238(104):106

    Google Scholar 

  6. Greenwood IA, Large WA (1995) Comparison of the effects of fenamates on Ca-activated chloride and potassium currents in rabbit portal vein smooth muscle cells. Br J Pharmacol 116:2939–2948

    CAS  Article  Google Scholar 

  7. Grover GJ, D’Alonzo AJ, Sleph PG, Dzwonczyk S, Hess TA, Darbenzio RB (1994) The cardioprotective and electrophysiological effects of cromakalim are attenuated by meclofenamate through a cyclooxygenase-independent mechanism. J Pharmacol Exp Ther 269:536–540

    CAS  PubMed  Google Scholar 

  8. Han Z, Yang JL, Jiang SX, Hou ST, Zheng RY (2013) Fast, non-competitive and reversible inhibition of NMDA-activated currents by 2-BFI confers neuroprotection. PLoS ONE 8:e64894

    CAS  Article  Google Scholar 

  9. Jiang H, Zeng B, Chen GL, Bot D, Eastmond S, Elsenussi SE, Atkin SL, Boa AN, Xu SZ (2012) Effect of non-steroidal anti-inflammatory drugs and new fenamate analogues on TRPC4 and TRPC5 channels. Biochem Pharmacol 83:923–931

    CAS  Article  Google Scholar 

  10. Kahlig KM, Lepist I, Leung K, Rajamani S, George AL (2010) Ranolazine selectively blocks persistent current evoked by epilepsy-associated Nav1.1 mutations. Br J Pharmacol 161:1414–1426

    CAS  Article  Google Scholar 

  11. Khansari PS, Halliwell RF (2009) Evidence for neuroprotection by the fenamate NSAID, mefenamic acid. Neurochem Int 55:683–688

    CAS  Article  Google Scholar 

  12. Lee HM, Kim HI, Shin YK, Lee CS, Park M, Song JH (2003) Diclofenac inhibition of sodium currents in rat dorsal root ganglion neurons. Brain Res 992:120–127

    CAS  Article  Google Scholar 

  13. Lo EH, Dalkara T, Moskowitz MA (2003) Mechanisms, challenges and opportunities in stroke. Nat Rev Neurosci 4:399–415

    CAS  Article  Google Scholar 

  14. Lyden P, Wahlgren NG (2000) Mechanisms of action of neuroprotectants in stroke. J Stroke Cerebrovasc Dis 9:9–14

    CAS  Article  Google Scholar 

  15. Olney JW (1978) Neurotoxicity of excitatory amino acid. In: McGeer E, Olney JW, McGeer P (eds) Kainic acid as a tool in neurobiology. Raven, New York, pp 95–121

    Google Scholar 

  16. Rothman SM (1984) Synaptic release of excitatory amino acid neurotransmitter mediates anoxic neuronal death. J Neurosci 4:1884–1891

    CAS  Article  Google Scholar 

  17. Sandalon S, Könnecke B, Levkovitch-Verbin H, Simons M, Hein K, Sättler MB, Bähr M, Ofri R (2013) Functional and structural evaluation of lamotrigine treatment in rat models of acute and chronic ocular hypertension. Exp Eye Res 115:47–56

    CAS  Article  Google Scholar 

  18. Sanger GJ, Bennett A (1979) Fenamates may antagonize the actions of prostaglandin endoperoxides in human myometrium. Br J Clin Pharmacol 8:479–482

    CAS  Article  Google Scholar 

  19. Santos MS, Moreno AJ, Carvalho AP (1996) Relationships between ATP depletion, membrane potential, and the release of neurotransmitters in rat nerve terminals. An in vitro study under conditions that mimic anoxia, hypoglycemia, and ischemia. Stroke 27:941–950

    CAS  Article  Google Scholar 

  20. Shi HS, Lai K, Yin XL, Liang M, Ye HB, Shi HB, Wang LY, Yin SK (2019) Ca2+-dependent recruitment of voltage-gated sodium channels underlies bilirubin-induced overexcitation and neurotoxicity. Cell Death Dis 10:774

    Article  Google Scholar 

  21. Simon RP, Swan JH, Griffiths T, Meldrum BS (1984) Blockade of N-methyl-d-aspartate receptors may protect against ischemic damage in the brain. Science 226:850–852

    CAS  Article  Google Scholar 

  22. Small DL, Morley P, Buchan AM (1999) Biology of ischemic cerebral cell death. Prog Cardiovasc Dis 42:185–207

    CAS  Article  Google Scholar 

  23. Stevens M, Timmermans S, Bottelbergs A, Hendriks JJ, Brône B, Baes M, Tytgat J (2013) Block of a subset of sodium channels exacerbates experimental autoimmune encephalomyelitis. J Neuroimmunol 261:21–28

    CAS  Article  Google Scholar 

  24. Sun JF, Xu YJ, Kong XH, Su Y, Wang ZY (2018) Fenamates inhibit human sodium channel Nav1.7 and Nav1.8. Neurosci Lett 696:67–73

    Article  Google Scholar 

  25. Waxman SG (2008) Mechanisms of disease: sodium channels and neuroprotection in multiple sclerosis-current status. Nat Clin Pract Neurol 4:159–169

    CAS  Article  Google Scholar 

  26. White MM, Aylwin M (1990) Niflumic and flufenamic acids are potent reversible blockers of Ca2+-activated Cl- channels in Xenopus oocytes. Mol Pharmacol 37:720–724

    CAS  PubMed  Google Scholar 

  27. Wu QJ, Tymianski M (2018) Targeting NMDA receptors in stroke: new hope in neuroprotection. Mol Brain 11:15

    Article  Google Scholar 

  28. Xu Y, Meng X, Hou X, Sun J, Kong X, Sun Y, Liu Z, Ma Y, Niu Y, Song Y, Cui Y, Zhao M, Zhang J (2017) A mutant of the Buthus martensii Karsch antitumor-analgesic peptide exhibits reduced inhibition to hNav1.4 and hNav1.5 channels while retaining analgesic activity. J Biol Chem 292:18270–18280

    CAS  Article  Google Scholar 

  29. Yau HJ, Baranauskas G, Martina M (2010) Flufenamic acid decreases neuronal excitability through modulation of voltage-gated sodium channel gating. J Physiol 588:3869–3882

    CAS  Article  Google Scholar 

  30. Zhang JM, Wang HK, Ye CQ, Ge W, Chen Y, Jiang ZL, Wu CP, Poo MM, Duan S (2003) ATP released by astrocytes mediates glutamatergic activity-dependent heterosynaptic suppression. Neuron 40:971–982

    CAS  Article  Google Scholar 

  31. Zhao ZW, Fan XX, Song JJ, Xu M, Chen MJ, Tu JF, Wu FZ, Zhang DK, Liu L, Chen L, Ying XH, Ji JS (2017) ShRNA knock-down of CXCR7 inhibits tumour invasion and metastasis in hepatocellular carcinoma after transcatheter arterial chemoembolization. J Cell Mol Med 21:1989–1999

    CAS  Article  Google Scholar 

Download references

Acknowledgements

We gratefully acknowledge the financial support from the Innovation Team Project of the Department of Education of Liaoning Province (LT2015010) and the PhD Start-up Fund of Natural Science Foundation of Liaoning Province (2019-BS-231).

Author information

Affiliations

Authors

Contributions

All authors contributed to the study conception and design. JS, YX, and XK carried out electrophysiology studies. JS carried out MTT assay. JS, YX, and YS carried out Western blot experiments and Annexin V-FITC/PI analysis. YX and ZW conceived and designed the experiments. JS, MZ, and YX wrote this manuscript, and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Yi-Jia Xu or Zhan-You Wang.

Ethics declarations

Ethical Approval

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.

Conflict of interest

The authors declare that they have no conflicts of interest.

Informed Consent

This article does not contain any studies with human participants performed by any of the authors.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Sun, JF., Zhao, MY., Xu, YJ. et al. Fenamates Inhibit Human Sodium Channel Nav1.2 and Protect Glutamate-Induced Injury in SH-SY5Y Cells. Cell Mol Neurobiol 40, 1405–1416 (2020). https://doi.org/10.1007/s10571-020-00826-1

Download citation

Keywords

  • Neuroprotection
  • Fenamates
  • Voltage-gated sodium channel
  • Apoptosis