Article

Measurement of bioelectric current with a vibrating probe.

Dermatology, University of California, Davis, USA.
Journal of Visualized Experiments 01/2011; DOI: 10.3791/2358
Source: PubMed

ABSTRACT Electric fields, generated by active transport of ions, are present in many biological systems and often serve important functions in tissues and organs. For example, they play an important role in directing cell migration during wound healing. Here we describe the manufacture and use of ultrasensitive vibrating probes for measuring extracellular electric currents. The probe is an insulated, sharpened metal wire with a small platinum-black tip (30-35 μm), which can detect ionic currents in the μA/cm(2) range in physiological saline. The probe is vibrated at about 200 Hz by a piezoelectric bender. In the presence of an ionic current, the probe detects a voltage difference between the extremes of its movement. A lock-in amplifier filters out extraneous noise by locking on to the probe's frequency of vibration. Data are recorded onto computer. The probe is calibrated at the start and end of experiments in appropriate saline, using a chamber which applies a current of exactly 1.5 μA/cm(2). We describe how to make the probes, set up the system and calibrate. We also demonstrate the technique of cornea measurement, and show some representative results from different specimens (cornea, skin, brain).

1 Follower
 · 
104 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Cell polarity plays a key role in regulating cell-cell communication, tissue architecture, and development. Both internal and external cues participate in directing polarity and feedback onto each other for robust polarization. One poorly appreciated layer of polarity regulation comes from electrochemical signals spatially organized at the level of the cell or the tissue. These signals which include ion fluxes, membrane potential gradients, or even steady electric fields, emerge from the polarized activation of specific ion transporters, and may guide polarity in wound-healing, development or regeneration. How a given electrochemical cue may influence cytoskeletal elements and cell polarity remains unclear. Here, we review recent progress highlighting the role of electrochemical signals in cell and tissue spatial organization, and elucidating the mechanisms for how such signals may regulate cytoskeletal assembly for cell polarity. © 2012 Wiley Periodicals, Inc.
    Cytoskeleton 09/2012; 69(9):601-12. DOI:10.1002/cm.21047 · 3.01 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Endogenous electric fields and currents occur naturally at wounds and are a strong signal guiding cell migration into the wound to promote healing. Many cells involved in wound healing respond to small physiological electric fields in vitro. It has long been assumed that wound electric fields are produced by passive ion leakage from damaged tissue. Could these fields be actively maintained and regulated as an active wound response? What are the molecular, ionic and cellular mechanisms underlying the wound electric currents? Using rat cornea wounds as a model, we measured the dynamic timecourses of individual ion fluxes with ion-selective probes. We also examined chloride channel expression before and after wounding. After wounding, Ca(2+) efflux increased steadily whereas K(+) showed an initial large efflux which rapidly decreased. Surprisingly, Na(+) flux at wounds was inward. A most significant observation was a persistent large influx of Cl(-), which had a time course similar to the net wound electric currents we have measured previously. Fixation of the tissues abolished ion fluxes. Pharmacological agents which stimulate ion transport significantly increased flux of Cl(-), Na(+) and K(+). Injury to the cornea caused significant changes in distribution and expression of Cl(-) channel CLC2. These data suggest that the outward electric currents occurring naturally at corneal wounds are carried mainly by a large influx of chloride ions, and in part by effluxes of calcium and potassium ions. Ca(2+) and Cl(-) fluxes appear to be mainly actively regulated, while K(+) flux appears to be largely due to leakage. The dynamic changes of electric currents and specific ion fluxes after wounding suggest that electrical signaling is an active response to injury and offers potential novel approaches to modulate wound healing, for example eye-drops targeting ion transport to aid in the challenging management of non-healing corneal ulcers.
    PLoS ONE 02/2011; 6(2):e17411. DOI:10.1371/journal.pone.0017411 · 3.53 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The activity of ion channels and transporters generates ion-specific fluxes that encode electrical and/or chemical signals with biological significance. Even though it is long known that some of those signals are crucial for regeneration, only in recent years the corresponding molecular sources started to be identified using mainly invertebrate or larval vertebrate models. We used adult zebrafish caudal fin as a model to investigate which and how ion transporters affect regeneration in an adult vertebrate model. Through the combined use of biophysical and molecular approaches, we show that V-ATPase activity contributes to a regeneration-specific H+ ef`flux. The onset and intensity of both V-ATPase expression and H+ efflux correlate with the different regeneration rate along the proximal-distal axis. Moreover, we show that V-ATPase inhibition impairs regeneration in adult vertebrate. Notably, the activity of this H+ pump is necessary for aldh1a2 and mkp3 expression, blastema cell proliferation and fin innervation. To the best of our knowledge, this is the first report on the role of V-ATPase during adult vertebrate regeneration.
    PLoS ONE 03/2014; 9(3):e92594. DOI:10.1371/journal.pone.0092594 · 3.53 Impact Factor

Preview

Download
4 Downloads
Available from