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Evaluation of toxicity equivalent factors of paralytic shellfish poisoning toxins in seven human sodium channels types by an automated high throughput electrophysiology system

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Abstract

Although voltage-gated sodium channels (Na v ) are the cellular target of paralytic shellfish poisoning (PSP) toxins and that patch clamp electrophysiology is the most effective way of studying direct interaction of molecules with these channels, nowadays, this technique is still reduced to more specific analysis due to the difficulties of transforming it in a reliable throughput system. Actual functional methods for PSP detection are based in binding assays using receptors but not functional Na v channels. Currently, the availability of automated patch clamp platforms and also of stably transfected cell lines with human Na v channels allow us to introduce this specific and selective method for fast screenings in marine toxin detection. Taking advantage of the accessibility to pure PSP standards, we calculated the toxicity equivalent factors (TEFs) for nine PSP analogs obtaining reliable TEFs in human targets to fulfill the deficiencies of the official analytic methods and to verify automated patch clamp technology as a fast and reliable screening method for marine toxins that interact with the sodium channel. The main observation of this work was the large variation of TEFs depending on the channel subtype selected, being remarkable the variation of potency in the 1.7 channel subtype and the suitability of Na v 1.6 and 1.2 channels for PSP screening.

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References

  • AOAC (1990) Paralytic shellfish poison. Biological method. Final action. In: Hellrich EBK (ed) Official methods of analysis. Association of Official Methods of Analytical Chemists, Arlington, VA, pp 881–882

  • AOAC (2005) Method 2005.06: Paralytic shellfish poisoning toxins in shellfish. Prechromatographic oxidation and liquid chromatography with fluorescence detection. Official Methods of Analysis of the Association of Official Analytical Chemists., Method 2005.06, First Action.) (Commission, E. (2006). Commission Regulation (EC) No 1664/2006 of 6 November 2006 amending Regulation (EC) No 2074/2005 as regards implementing measures for certain products of animal origin intended for human consumption and repealing certain implementing measures. Off J Eur Union L 320:13–45

  • AOAC (2011a) AOAC Official method 2011.02 Determination of paralytic shellfish poisoning toxins in mussels, clams, oysters and scallops. Post-column oxidation method (PCOX). First action 2011. In MD Official Methods of Analysis of the AOAC (Association of Official Analytical Chemists) Gaithersburg, USA, Method 2011.02

  • AOAC (2011b) AOAC official method 2011.27. In Paralytic shellfish toxins (PSTs) in shellfish, receptor binding assay. Association of Official Methods of Analytical Chemists, Arlington, VA; AOAC International: AOAC International, Gaithersburg, MD), pp 881–882

  • Ben-Gigirey B et al (2012) A comparative study for PSP toxins quantification by using MBA and HPLC official methods in shellfish. Toxicon 60(5):864–873

    Article  CAS  PubMed  Google Scholar 

  • Botana LM (ed) (2014) Seafood and freshwater toxins. Pharmacology, physiology and detection, 3rd ed. CRC Press, Boca Raton

  • Botana LM, Vilariño N, Elliott CT, Campbell K, Alfonso A, Vale C, Louzao MC, Botana AM (2010) ‘The problem of toxicity equivalent factors in developping alternative methods to animal bioassays for marine toxin detection. Trends Anal Chem 29:1316–1325

    Article  CAS  Google Scholar 

  • Caldwell JH et al (2000) Sodium channel Na(v)1.6 is localized at nodes of ranvier, dendrites, and synapses. Proc Natl Acad Sci USA 97(10):5616–5620

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Castle N et al (2009) Sodium channel inhibitor drug discovery using automated high throughput electrophysiology platforms. Comb Chem High Throughput Screen 12(1):107–122

    Article  CAS  PubMed  Google Scholar 

  • Catterall WA, Goldin AL, Waxman SG (2005) International Union of Pharmacology. XLVII. Nomenclature and structure–function relationships of voltage-gated sodium channels. Pharmacol Rev 57(4):397–409

    Article  CAS  PubMed  Google Scholar 

  • Dunlop J et al (2008) High-throughput electrophysiology: an emerging paradigm for ion-channel screening and physiology. Nat Rev Drug Discov 7(4):358–368

    Article  CAS  PubMed  Google Scholar 

  • EFSA (2009a) Scientific opinion of the panel on contaminants in the food chainon: a request from the European Commission on marine biotoxins in shellfish: saxitoxin group (question no EFSA-Q-2006-065E). EFSA J 1019:1–76

    Google Scholar 

  • EFSA (2009b) Scientific opinion on marine biotoxins in shellfish–summary on regulated marine biotoxins. EFSA J 1306:1–23

    Google Scholar 

  • Fonfria ES et al (2007) Paralytic shellfish poisoning detection by surface plasmon resonance-based biosensors in shellfish matrixes. Anal Chem 79(16):6303–6311

    Article  CAS  PubMed  Google Scholar 

  • Fraga M et al (2012) Detection of paralytic shellfish toxins by a solid-phase inhibition immunoassay using a microsphere-flow cytometry system. Anal Chem 84(10):4350–4356

    Article  CAS  PubMed  Google Scholar 

  • Graef JD et al (2013) Validation of a high-throughput, automated electrophysiology platform for the screening of nicotinic agonists and antagonists. J Biomol Screen 18(1):116–127

    Article  CAS  PubMed  Google Scholar 

  • Hess P et al (2006) Three Rs Approaches in marine biotoxin testing. The Report and Recommendations of a joint ECVAM/DG SANCO workshop (ECVAM workshop 54). Altern Lab Anim 34(2):193–224

    CAS  PubMed  Google Scholar 

  • Kohrman DC et al (1996) A missense mutation in the sodium channel Scn8a is responsible for cerebellar ataxia in the mouse mutant jolting. J Neurosci 16(19):5993–5999

    CAS  PubMed  Google Scholar 

  • Leffler A et al (2005) Pharmacological properties of neuronal TTX-resistant sodium channels and the role of a critical serine pore residue. Pflugers Arch 451(3):454–463

    Article  CAS  PubMed  Google Scholar 

  • Lehane L (2001) Paralytic shellfish poisoning: a potential public health problem. Med J Aust 175(1):29–31

    CAS  PubMed  Google Scholar 

  • Munday R et al (2013) Acute toxicities of saxitoxin, neosaxitoxin, decarbamoyl saxitoxin and gonyautoxins 1&4 and 2&3 to mice by various routes of administration. Toxicon 76:77–83

    Article  CAS  PubMed  Google Scholar 

  • Oshima Y (1995) Postcolumnderivatization liquid chromatographic method for paralytic shellfish toxins. J AOAC Int 78:528–532

    CAS  Google Scholar 

  • Perez S et al (2011) Determination of toxicity equivalent factors for paralytic shellfish toxins by electrophysiological measurements in cultured neurons. Chem Res Toxicol 24(7):1153–1157

    Article  CAS  PubMed  Google Scholar 

  • Savio-Galimberti E, Gollob MH, Darbar D (2012) Voltage-gated sodium channels: biophysics, pharmacology, and related channelopathies. Front Pharmacol 3:124

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Schaller KL, Caldwell JH (2003) Expression and distribution of voltage-gated sodium channels in the cerebellum. Cerebellum 2(1):2–9

    Article  CAS  PubMed  Google Scholar 

  • Sivilotti L et al (1997) A single serine residue confers tetrodotoxin insensitivity on the rat sensory-neuron-specific sodium channel SNS. FEBS Lett 409(1):49–52

    Article  CAS  PubMed  Google Scholar 

  • Spencer CI et al (2012) Ion channel pharmacology under flow: automation via well-plate microfluidics. Assay Drug Dev Technol 10(4):313–324

    Article  CAS  PubMed  Google Scholar 

  • Turner AD, Hatfield RG (2012) Refinement of AOAC official method 2005.06 liquid chromatography-fluorescence detection method to improve performance characteristics for the determination of paralytic shellfish toxins in king and queen scallops. J AOAC Int 95:129–142

    Article  CAS  PubMed  Google Scholar 

  • Vale P (2014) Saxitoxin and analogs: ecobiology, origin, chemistry and detection. In: Botana LM (ed) Seafood and freshwater toxins. Pharmacology, physiology and detection, 3rd edn. CRC Press, Boca Raton

    Google Scholar 

  • Vale C et al (2008) In vitro and in vivo evaluation of paralytic shellfish poisoning toxin potency and the influence of the pH of extraction. Anal Chem 80(5):1770–1776

    Article  CAS  PubMed  Google Scholar 

  • van den Top HJ et al (2011) Surface plasmon resonance biosensor screening method for paralytic shellfish poisoning toxins: a pilot interlaboratory study. Anal Chem 83(11):4206–4213

    Article  PubMed  Google Scholar 

  • Velez P et al (2001) A functional assay for paralytic shellfish toxins that uses recombinant sodium channels. Toxicon 39(7):929–935

    Article  CAS  PubMed  Google Scholar 

  • Walker JR et al (2012) Marked difference in saxitoxin and tetrodotoxin affinity for the human nociceptive voltage-gated sodium channel (Na v 1.7) [corrected]. Proc Natl Acad Sci U S A 109(44):18102–18107

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Wiese M et al (2010) Neurotoxic alkaloids: saxitoxin and its analogs. Mar Drugs 8(7):2185–2211

    Article  PubMed Central  CAS  PubMed  Google Scholar 

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Acknowledgments

The research leading to these results has received funding from the following FEDER cofounded-grants. From CDTI and Technological Funds, supported by Ministerio de Economía y Competitividad, AGL2012-40185-CO2-01 and Consellería de Cultura, Educación e OrdenaciónUniversitaria, GRC2013-016, and through AxenciaGalega de Innovación, Spain, ITC-20133020 SINTOX, IN852A 2013/16-3 MYTIGAL. From CDTI under ISIP Programme, Spain, IDI-20130304 APTAFOOD. From the European Union’s Seventh Framework Programme managed by REA—Research Executive Agency (FP7/2007–2013) under Grant Agreement Nos. 265409 µAQUA, 315285 CIGUATOOLS, and 312184 PHARMASEA.

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The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Departamento de Farmacología, Facultad de Veterinaria, Universidad de Santiago de Compostela, 27002, Lugo, Spain

    Eva Alonso, Amparo Alfonso & Luis M. Botana

  2. Departamento de Fisiología, Facultad de Veterinaria, Universidad de Santiago de Compostela, 27003, Lugo, Spain

    Mercedes R. Vieytes

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  1. Eva Alonso
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  2. Amparo Alfonso
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  3. Mercedes R. Vieytes
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  4. Luis M. Botana
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Correspondence to Luis M. Botana.

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Alonso, E., Alfonso, A., Vieytes, M.R. et al. Evaluation of toxicity equivalent factors of paralytic shellfish poisoning toxins in seven human sodium channels types by an automated high throughput electrophysiology system. Arch Toxicol 90, 479–488 (2016). https://doi.org/10.1007/s00204-014-1444-y

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