Self-referencing, non-invasive, ion selective electrode for single cell detection of trans-plasma membrane calcium flux

Smith, Peter J. S.; Hammar, Katherine; Porterfield, D. Marshall; Sanger, Richard H.; Trimarchi, James R.

doi:10.1002/(sici)1097-0029(19990915)46:6<398::aid-jemt8>3.0.co;2-h

Cited by 155 publications

(143 citation statements)

References 72 publications

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“…For self referencing ion selective probe (SERIS) measurements of proton flux (Smith et al, 1999), healthy Xenopus embryos were cultured in 0.1ϫ MMR at pH 7.0. H + flux measurements were made to the left and right, equidistant from the ventral midline, near the AV midline, slightly on the animal side, and approximately in the middle of the left-ventral quadrant and the right-ventral quadrant of the embryo.…”

Section: Self Referencing Ion Selective Probe Measurementsmentioning

confidence: 99%

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Early, H+-V-ATPase-dependent proton flux is necessary for consistent left-right patterning of non-mammalian vertebrates

Adams

Robinson

Fukumoto³

et al. 2006

Development

251

306

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Biased left-right asymmetry is a fascinating and medically important phenomenon. We provide molecular genetic and physiological characterization of a novel, conserved, early, biophysical event that is crucial for correct asymmetry: H + flux. A pharmacological screen implicated the H + -pump H + -V-ATPase in Xenopus asymmetry, where it acts upstream of early asymmetric markers. Immunohistochemistry revealed an actin-dependent asymmetry of H + -V-ATPase subunits during the first three cleavages. H + -flux across plasma membranes is also asymmetric at the four-and eight-cell stages, and this asymmetry requires H + -V-ATPase activity. Abolishing the asymmetry in H + flux, using a dominant-negative subunit of the H + -V-ATPase or an ectopic H + pump, randomized embryonic situs without causing any other defects. To understand the mechanism of action of H + -V-ATPase, we isolated its two physiological functions, cytoplasmic pH and membrane voltage (V mem ) regulation. Varying either pH or V mem , independently of direct manipulation of H + -V-ATPase, caused disruptions of normal asymmetry, suggesting roles for both functions. V-ATPase inhibition also abolished the normal early localization of serotonin, functionally linking these two early asymmetry pathways. The involvement of H + -V-ATPase in asymmetry is conserved to chick and zebrafish. Inhibition of the H + -V-ATPase induces heterotaxia in both species; in chick, H + -V-ATPase activity is upstream of Shh; in fish, it is upstream of Kupffer's vesicle and Spaw expression. Our data implicate H + -V-ATPase activity in patterning the LR axis of vertebrates and reveal mechanisms upstream and downstream of its activity. We propose a pH-and V mem -dependent model of the early physiology of LR patterning. Development 133, 1657Development 133, -1671Development 133, (2006 DEVELOPMENT 1658 necessary to characterize the endogenous behavior of the relevant pumps in embryos and to place their function in the context of known LR patterning mechanisms. Here, we explore the properties of H + -V-ATPase function in several vertebrate embryos. Through endogenous localization of the H + -V-ATPase and gain-and loss-offunction experiments in chick, frog and zebrafish, we identify the H + -V-ATPase as a novel, conserved, obligate component of LR patterning upstream of asymmetric gene expression. KEY WORDS: Left-right asymmetry, H + -V-ATPase, V-ATPase, Xenopus, Chick, Zebrafish, Axial patterning, Cytoplasmic pH, Membrane voltage MATERIALS AND METHODS Animal husbandryXenopus embryos were collected according to standard protocols (Sive et al., 2000) in 0.1ϫ Modified Marc's Ringers (MMR) pH 7.8 + 0.1% Gentamicin. Xenopus embryos were staged according to Nieuwkoop and Faber (Nieuwkoop and Faber, 1967). Chick embryos from Charles River Laboratories, maintained at 38°C, were staged according to Hamburger and Hamilton (Hamburger and Hamilton, 1992). Zebrafish embryos (Westerfield, 1995) were maintained at 28.5°C in water containing 1 drop per gallon Methyl Blue. Assaying organ situsXenopus e...

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Section: Self Referencing Ion Selective Probe Measurementsmentioning

confidence: 99%

“…Using a SERIS probe (Hotary et al, 1992;Smith et al, 1999;Smith and Trimarchi, 2001), we detected a large net efflux of protons from the cleavage furrow of the two-cell stage. This efflux averaged 12.7±22 pmole cm -2 s -1 (n=5) at about the midpoint of cleavage.…”

Section: Early Xenopus Embryos Drive H + -V-atpasedependent Asymmetrmentioning

confidence: 99%

Early, H+-V-ATPase-dependent proton flux is necessary for consistent left-right patterning of non-mammalian vertebrates

Adams

Robinson

Fukumoto³

et al. 2006

Development

251

306

View full text Add to dashboard Cite

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“…Under such controversy, we raised the question of whether there is any in vivo evidence to demonstrate the acid-secretion function in cells expressing V-type H + -ATPase. An in vivo approach, the scanning ion-selective electrode technique (SIET), was developed to enable non-invasive measurement of ion fluxes from single cells and epithelial structures (Smith et al, 1999;Smith and Trimarchi, 2001), and it was initially used to detect Cl -excretion from MR cells in isolated opercular epithelium of seawater (SW) fish in a landmark study on fish MR cells (Foskett and Scheffey, 1982). After SIET measurements, the intact fish can be directly subjected to subsequent analysis to identify cells measured by SIET.…”

Section: H + -Atpase Rich (Hr) Cellsmentioning

confidence: 99%

Ion uptake and acid secretion in zebrafish (Danio rerio)

Hwang

2009

Journal of Experimental Biology

169

165

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SummaryTransepithelial transport is one of the major processes involved in the mechanism of homeostasis of body fluids in vertebrates including fish. The current models of ion regulation in fish gill ionocytes have been proposed mainly based on studies in traditional model species like salmon, trout, tilapia, eel and killifish, but the mechanisms are still being debated due to the lack of convincing molecular physiological evidence. Taking advantage of plentiful genetic databases for zebrafish, we studied the molecular/cellular mechanisms of ion regulation in fish skin/gills. In our recently proposed model, there are at least three subtypes of ionocytes in zebrafish skin/gills: Na + -K + -ATPase-rich (NaR), Na + -Cl -cotransporter (NCC) and H + -ATPase-rich (HR) cells. Specific isoforms of transporters and enzymes have been identified as being expressed by these ionocytes: zECaC, zPMCA2 and zNCX1b by NaR cells; zNCC gill form by NCC cells; and zH + -ATPase, zNHE3b, zCA2-like a and zCA15a by HR cells. Serial molecular physiological experiments demonstrated the distinct roles of these ionocytes in the transport of various ions: HR, NaR and NCC cells are respectively responsible for acid secretion/Na + uptake, Ca 2+ uptake and Cl -uptake. The expression, regulation and function of transporters in HR and NaR cells are much better understood than those in NCC cells. The basolateral transport pathways in HR and NCC cells are still unclear, and the driving forces for the operations of apical NHE and NCC are another unresolved issue. Studies on zebrafish skin/gill ionocytes are providing new insights into fish ion-regulatory mechanisms, but the zebrafish model cannot simply be applied to other species because of species differences and a lack of sufficient molecular physiological evidence in other species.

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“…The construction of liquid-membrane ISMEs has been previously described in detail (Smith et al 1994(Smith et al , 1999Kühtreibe and Jaffe 1990). Various ionophore cocktails were prepared based on a Pb 2+ ionophore and a Cu 2+ ionophore previously described (Kamata and Onoyama 1991;Ceresa and Pretsch 1999).…”

Section: Construction and Characterization Of Pb 2+ And Cu 2+ Ismesmentioning

confidence: 99%

Determining the fluxes of ions (Pb2+, Cu2+ and Cd2+) at the root surface of wetland plants using the scanning ion-selective electrode technique

Peijnenburg

et al. 2016

Plant Soil

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Background and aims Measuring specific ion fluxes from different regions of the root under practical physiological conditions is crucial for understanding metal uptake mechanisms by plants. Methods We developed and tested a neutral carrierbased liquid-membrane Pb 2+ and Cu 2+ ion selective microelectrode (ISME) to investigate ion-transport processes along the roots of three common wetland plant species. Results The Pb 2+ and Cu 2+ ISME exhibited a Nernstian response with Pb 2+ and Cu 2+ activities as low as 1.0 nM and 1.0 μM in deionized water and simulated soil solution, respectively. Phragmites australis had a region of Cu 2+ release for approximately the first 200 μm, while it exhibited Pb 2+ and Cd 2+ outward net flux up to the first 500 μm. Although in older sections of the root of Phragmites australis there were areas of influx of Cu 2+ , Pb 2+ and Cd 2+ , the overall influx was much smaller than that of Typha latifolia or Canna indica. Such a reduced uptake and/or an increased efflux of metal ions across the root-cell plasma-membrane might explain the higher resistance of Phragmites australis to metals, at least in part. Conclusions The Pb 2+ and Cu 2+ ISMEs are shown to permit detailed investigation of heavy-metal ion transport in plant roots, especially for plants used for phytoremediation.

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Self-referencing, non-invasive, ion selective electrode for single cell detection of trans-plasma membrane calcium flux

Cited by 155 publications

References 72 publications

Early, H+-V-ATPase-dependent proton flux is necessary for consistent left-right patterning of non-mammalian vertebrates

Early, H+-V-ATPase-dependent proton flux is necessary for consistent left-right patterning of non-mammalian vertebrates

Ion uptake and acid secretion in zebrafish (Danio rerio)

Determining the fluxes of ions (Pb2+, Cu2+ and Cd2+) at the root surface of wetland plants using the scanning ion-selective electrode technique

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