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Use of ion-channel modulating agents to study cyanobacterial Na+-K+ fluxes

Abstract

Here we describe an experimental design aimed to investigate changes in total cellular levels of Na+ and K+ ions in cultures of freshwater filamentous cyanobacteria. Ion concentrations were measured in whole cells by flame photometry. Cellular Na+ levels increased exponentially with rising alkalinity, with K+ levels being maximal for optimal growth pH (8). At standardized pH conditions, the increase in cellular Na+, as induced by NaCl at 10 mM, was coupled by the two sodium channel-modulating agents lidocaine hydrochloride at 1 µM and veratridine at 100 µM. Both the channel-blockers amiloride (1 mM) and saxitoxin (1 µM), decreased cell-bound Na+ and K+ levels. Results presented demonstrate the robustness of well-defined channel blockers and channel-activators in the study of cyanobacterial Na+- K+ fluxes.

Abbreviations

STX:

saxitoxin

VTD:

veratridine

References

  1. Horikoshi K. Microorganisms in alkaline environments. New York: VCH;1991. p. 110.

    Google Scholar 

  2. Krulwich TA, Ito M, Guffanti AA. The Na+-dependence of alkaliphily in Bacillus. Biochim Biophys Acta 2001; 1505:158–168.

    Article  PubMed  CAS  Google Scholar 

  3. Lengeler JW, Drews G, Schlegel HG. Biology of the Prokaryotes. Oxford: Blackwell Science; 1999. p. 269.

    Google Scholar 

  4. Maestri O, Joset F. Regulation by external pH and stationary growth phase of the acetolactate synthase from Synechocystis PCC6803. Mol Microbiol 2000; 37:828–838.

    Article  PubMed  CAS  Google Scholar 

  5. Waditee R, Hibino T, Tanaka Y, Nakamura T, Incharoensakdi A, Takabe T. Halotolerant cyanobacterium Aphanothece halophytica contains an Na+/H+ antiporter, homologous to eukaryotic ones, with novel ion specificity affected by C-terminal tail. J Biol Chem 2001; 276:36931–36938.

    Article  PubMed  CAS  Google Scholar 

  6. Apte SK, Thomas J. Membrane electrogenesis and sodium transport in filamentous nitrogen-fixing cyanobacteria. Eur J Biochem 1986; 154:395–401.

    Article  PubMed  CAS  Google Scholar 

  7. Sonoda M, Katoh H, Vermaas W, Schmetterer G, Ogawa T. Photosynthetic electron transport involved in PxcA-dependent proton extrusion in Synechocystis sp. strain PCC6803: effect of pxcA inactivation on CO2, HCO3−, and NO3− uptake. J Bacteriol 1998; 180:3799–3803.

    PubMed  CAS  Google Scholar 

  8. Miller AG, Turpin DH, Canvin DT. Na+ requirement for growth, photosynthesis, and pH regulation in the alkalotolerant cyanobacterium Synechococcus leopoliensis. J Bacteriol 1984; 159:100–106.

    PubMed  CAS  Google Scholar 

  9. Ren D, Navarro B, Xu H, Yue L, Shi Q, Clapham DE. A prokaryotic voltage-gated sodium channel. Science 2001; 294:2306–2308.

    Article  Google Scholar 

  10. Koishi R, Xu H, Ren D, Navarro B, Spiller BW, Shi Q, Clapham DE. A superfamily of voltage-gated sodium channels in bacteria. J Biol Chem 2004; 279:9532–9538.

    Article  PubMed  CAS  Google Scholar 

  11. Humpage AR, Rositano J, Bretag A, Brown R, Baker P, Nicholson BC, Steffensen DA. Paralytic shellfish poisons from Australian cyanobacterial blooms. Aust J Mar Freshwater Res 1994; 45:761–771.

    Article  CAS  Google Scholar 

  12. Hawkins PR, Chandrasena NR, Jones GJ, Humpage AR, Falconer IR. Isolation and toxicity of Cylindrospermopsis raciborskii from an ornamental lake. Toxicon 1997; 35:341–346.

    Article  PubMed  CAS  Google Scholar 

  13. Lagos N, Onodera H, Zagatto PA, Andrinolo D, Azevedo SMFQ, Oshima Y. The first evidence of paralytic shellfish toxins in the freshwater cyanobacterium Cylindrospermopsis raciborskii, isolated from Brazil. Toxicon 1999; 37:1359–1373.

    Article  PubMed  CAS  Google Scholar 

  14. Bowling LC, Baker PD. Major cyanobacterial bloom in the Barwon-Darling River, Australia, in 1991, and underlying limnological conditions. Mar Freshwater Res 1996; 47:643–657.

    Article  CAS  Google Scholar 

  15. Gorham PR, McLachlan J, Hammer UT, Kim WK. Isolation and culture of toxic strains of Anabaena flos-aquae (Lyngb.) de Breb. Verh. Int Verein Theor Angew Limnol 1964; 15:796–804.

    Google Scholar 

  16. Hille B. Ionic Channels of Excitable Membranes. 2nd Edition. Sunderland: Sinauer Associates; 1997. p. 224.

    Google Scholar 

  17. Quintero MJ, Muro-Pastor AM, Herrero A, Flores E. Arginine catabolism in the cyanobacterium Synechocystis sp. strain PCC6803 involves the urea cycle and arginase pathway. J Bacteriol 2000; 182:1008–1015.

    Article  PubMed  CAS  Google Scholar 

  18. Kanesaki Y, Suzuki I, Allakhverdiev SI, Mikami K, Murata N. Salt stress and hyperosmotic stress regulate the expression of different sets of genes in Synechocystis sp. PCC 6803. Biochem Biophys Res Commun 2002; 290:339–348.

    Article  PubMed  CAS  Google Scholar 

  19. Pomati F, Neilan BA, Manarolla G, Suzuki T, Rossetti C. Enhancement of intracellular saxitoxin accumulation by lidocaine hydrochloride in the cyanobacterium Cylindrospermopsis raciborskii T3 (Nostocales). J Phycol 2003; 39:535–542.

    Article  CAS  Google Scholar 

  20. Catteral WA, Nirenberg M. Sodium uptake associated with activation of action potential ionophores of cultured neuroblastoma and muscle cells. Proc Natl Acad Sci USA 1973; 70:3759–3763.

    Article  Google Scholar 

  21. Catterall WA. Neurotoxins that act on voltage-sensitive sodium channels in excitable membranes. Annu Rev Pharmacol Toxicol 1980; 20:15–43.

    Article  PubMed  CAS  Google Scholar 

  22. Kim D, Smith TW. Effects of amiloride and ouabain on contractile state, Ca and Na fluxes, and Na content in cultured chick heart cells. Mol Pharmacol 1986; 29:363–371.

    PubMed  CAS  Google Scholar 

  23. Shibamoto S, Hori T, Hayakawa M, Nango M, Cragoe EJ Jr, Oku N, Ito F. Inhibitory effect of local anesthetics on Na+/H+ antiporter in brush border membrane-reconstituted vesicles. Life Sci 1990; 47:1129–1133.

    Article  PubMed  CAS  Google Scholar 

  24. Schaefer L, Sakai H, Mattei MG, Lazdunski M, Lingueglia E. Molecular cloning, functional expression and chromosomal localization of an amiloride-sensitive Na+ channel from human small intestine. FEBS Letters 2001; 471:205–210.

    Article  Google Scholar 

  25. Rowbury RJ, Goodson M, Humphrey TJ. Sodium chloride induces an NhaA/NhaR-independent acid sensitivity at neutral external pH in Escherichia coli. Appl Environ Microbiol 1994; 60:1630–1634.

    PubMed  CAS  Google Scholar 

  26. Nosaka S, Ohkawa TA, Okihara K, Yoshikawa K. Effects of local anesthetics on the Chara plasmalemma. Biochim Biophys Acta 1992; 1106:325–334.

    Article  PubMed  CAS  Google Scholar 

  27. Bidani A, Heming TA. Effects of lidocaine on cytosolic pH regulation and stimulus-induced effector functions in alveolar macrophages. Lung 1997; 175:349–361.

    Article  PubMed  CAS  Google Scholar 

  28. Strichartz G. Relative potencies of several derivates of saxitoxin: electrophysiological and toxin binding studies. Biophys J 1981; 33:209–216.

    Google Scholar 

  29. Kaneko T, Sato S, Kotani H, Tanaka A, Asamizu E, Nakamura Y, Miyajima N, Hirosawa M, Sugiura M, Sasamoto S, Kimura T, Hosouchi T, Matsuno A, Musaraki A, Nakazaki N, Nauro K, Okumura S, Shimpo S, Takeuchi C, Wada T, Watanabe A, Yamada M, Yasuda M, Tabata S. Sequence analysis of the genome of the unicellular cyanobacterium Synechocystis sp. PCC6803: II. Sequence determination of the entire genome and assignment of potential protein coding regions. DNA Res Suppl 1996; 3:741–747.

    Google Scholar 

  30. Murata T, Takase K, Yamato I, Igarashi K, Kakiuma Y. The ntpJ gene in the Enterococcus hirae ntp operon encodes a component of KtrII potassium transport system functionally independent of vacuolar Na+-ATPase. J Biol Chem 1996; 271:10042–10047.

    Article  PubMed  CAS  Google Scholar 

  31. Nakamura T, Yuda R, Unemoto T, Bakker EP. KtrAB, a new type of bacterial K+-uptake system from Vibrio alginolyticus. J Bacteriol 1998; 180:3491–3494.

    PubMed  CAS  Google Scholar 

  32. Neilan BA, Saker ML, Fastner J, Torokne A, Burns BP. Phylogeography of the invasive cyanobacterium Cylindrospermopsis raciborskii. Mol Ecol 2003; 12:133–140.

    Article  PubMed  CAS  Google Scholar 

  33. Martins CA, Kulis D, Franca S, Anderson DM. The loss of PSP toxin production in a formerly toxic Alexandrium lusitanicum clone. Toxicon 2004; 43:195–205.

    Article  PubMed  CAS  Google Scholar 

  34. Shibata M, Katoh H, Sonoda M, Ohkawa H, Shimoyama M, Fukuzawa H, Kaplan A, Ogawa T. Genes Essential to Sodium-dependent Bicarbonate Transport in Cyanobacteria. Function and phylogenetic analysis. J Biol Chem 2002; 277:18658–18664.

    Article  PubMed  CAS  Google Scholar 

  35. Kaneko T, Nakamura Y, Wolk CP, Kuritz T, Sasamoto S, Watanabe A, Iriguchi M, Ishikawa A, Kawashima K, Kimura T, Kishida Y, Kohara M, Matsumoto M, Matsuno A, Muraki A, Nakazaki N, Shimpo S, Sugimoto M, Takazawa M, Yamada M, Yasuda M, Tabata S. Complete genomic sequence of the filamentous nitrogen-fixing cyanobacterium Anabaena sp. strain PCC 7120. DNA Res 2001; 8:205–213.

    Article  PubMed  CAS  Google Scholar 

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Correspondence to Brett A. Neilan.

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Published: June 29, 2004.

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Pomati, F., Burns, B.P. & Neilan, B.A. Use of ion-channel modulating agents to study cyanobacterial Na+-K+ fluxes. Biol. Proced. Online 6, 137–143 (2004). https://doi.org/10.1251/bpo82

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  • DOI: https://doi.org/10.1251/bpo82

Indexing terms

  • Cyanobacteria
  • Sodium
  • Potassium