title: A model system using confocal fluorescence microscopy for examining real-time intracellular sodium ion regulation.
abstract: The gills of euryhaline fish are the ultimate ionoregulatory tissue, achieving ion homeostasis despite rapid and significant changes in external salinity. Cellular handling of sodium is not only critical for salt and water balance but is also directly linked to other essential functions such as acid-base homeostasis and nitrogen excretion. However, although measurement of intracellular sodium ([Na(+)]i) is important for an understanding of gill transport function, it is challenging and subject to methodological artifacts. Using gill filaments from a model euryhaline fish, inanga (Galaxias maculatus), the suitability of the fluorescent dye CoroNa Green as a probe for measuring [Na(+)]i in intact ionocytes was confirmed via confocal microscopy. Cell viability was verified, optimal dye loading parameters were determined, and the dye-ion dissociation constant was measured. Application of the technique to freshwater- and 100% seawater-acclimated inanga showed salinity-dependent changes in branchial [Na(+)]i, whereas no significant differences in branchial [Na(+)]i were determined in 50% seawater-acclimated fish. This technique facilitates the examination of real-time changes in gill [Na(+)]i in response to environmental factors and may offer significant insight into key homeostatic functions associated with the fish gill and the principles of sodium ion transport in other tissues and organisms.
authors.affiliation: School of Biological Sciences, University of Canterbury, Christchurch 8041, New Zealand.
authors.forename: Jacqueline A
authors.initials: JA
authors.lastname: Lee
authors.affiliation: School of Biological Sciences, University of Canterbury, Christchurch 8041, New Zealand; School of Environmental and Life Sciences, University of Newcastle, Callaghan, New South Wales 2308, Australia.
authors.forename: David A
authors.initials: DA
authors.lastname: Collings
authors.affiliation: School of Biological Sciences, University of Canterbury, Christchurch 8041, New Zealand; Faculty of Science and Technology and Athabasca River Basin Research Institute, Athabasca University, Athabasca, Alberta T9S 3A3, Canada; Department of Biological Sciences, University of Alberta, Edmonton, Alberta T6G 2R3, Canada. Electronic address: cglover@athabascau.ca.
authors.forename: Chris N
authors.initials: CN
authors.lastname: Glover
mesh_terms: D000818:Animals; D002470:Cell Survival; D005880:Gills / Q000166:cytology / Q000378:metabolism*; D017136:Ion Transport; D007477:Ions / Q000378:metabolism; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D036441:Osmeriformes / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*; D013997:Time Factors
chemical_list: D007477:Ions; D012964:Sodium
keywords: Amphidromous; Fish; Gill; Intracellular sodium concentration; Laser scanning confocal microscopy; Salinity acclimation