Characterization of Regulatory Volume Behavior by Fluorescence Quenching in Human Corneal Epithelial Cells

Characterization of Regulatory Volume Behavior by Fluorescence Quenching in Human Corneal... An in-depth understanding of the mechanisms underlying regulatory volume behavior in corneal epithelial cells has been in part hampered by the lack of adequate methodology for characterizing this phenomenon. Accordingly, we developed a novel approach to characterize time-dependent changes in relative cell volume induced by anisosmotic challenges in calcein-loaded SV40-immortalized human corneal epithelial (HCE) cells with a fluorescence microplate analyzer. During a hypertonic challenge, cells shrank rapidly, followed by a temperature-dependent regulatory volume increase (RVI), τc = 19 min. In contrast, a hypotonic challenge induced a rapid (τc = 2.5 min) regulatory volume decrease (RVD). Temperature decline from 37 to 24°C reduced RVI by 59%, but did not affect RVD. Bumetanide (50 μM), ouabain (1 mM), DIDS (1 mM), EIPA (100 μM), or Na+-free solution reduced the RVI by 60, 61, 39, 32, and 69%, respectively. K+, Cl− channel and K+-Cl− cotransporter (KCC) inhibition obtained with either 4-AP (1 mM), DIDS (1 mM), DIOA (100 μM), high K+ (20 mM) or Cl−-free solution, suppressed RVD by 42, 47, 34, 52 and 58%, respectively. KCC activity also affects steady-state cell volume, since its inhibition or stimulation induced relative volume alterations under isotonic conditions. Taken together, K+ and Cl− channels in parallel with KCC activity are important mediators of RVD, whereas RVI is temperature-dependent and is essentially mediated by the Na+-K+-2Cl− cotransporter (Na+-K+-2Cl−) and the Na+-K+ pump. Inhibition of K+ and Cl− channels and KCC but not Na+-K+-2Cl− affect steady-state cell volume under isotonic conditions. This is the first report that KCC activity is required for HCE cell volume regulation and maintenance of steady-state cell volume. The Journal of Membrane Biology Springer Journals

Characterization of Regulatory Volume Behavior by Fluorescence Quenching in Human Corneal Epithelial Cells

Loading next page...
Copyright © 2005 by Springer Science+Business Media, Inc.
Life Sciences; Human Physiology; Biochemistry, general
Publisher site
See Article on Publisher Site


You’re reading a free preview. Subscribe to read the entire article.

DeepDyve is your
personal research library

It’s your single place to instantly
discover and read the research
that matters to you.

Enjoy affordable access to
over 12 million articles from more than
10,000 peer-reviewed journals.

All for just $49/month

Explore the DeepDyve Library

Unlimited reading

Read as many articles as you need. Full articles with original layout, charts and figures. Read online, from anywhere.

Stay up to date

Keep up with your field with Personalized Recommendations and Follow Journals to get automatic updates.

Organize your research

It’s easy to organize your research with our built-in tools.

Your journals are on DeepDyve

Read from thousands of the leading scholarly journals from SpringerNature, Elsevier, Wiley-Blackwell, Oxford University Press and more.

All the latest content is available, no embargo periods.

See the journals in your area

Monthly Plan

  • Read unlimited articles
  • Personalized recommendations
  • No expiration
  • Print 20 pages per month
  • 20% off on PDF purchases
  • Organize your research
  • Get updates on your journals and topic searches


Start Free Trial

14-day Free Trial

Best Deal — 39% off

Annual Plan

  • All the features of the Professional Plan, but for 39% off!
  • Billed annually
  • No expiration
  • For the normal price of 10 articles elsewhere, you get one full year of unlimited access to articles.



billed annually
Start Free Trial

14-day Free Trial