Local Ca2+ rise near store operated Ca2+ channels inhibits cell coupling during capacitative Ca2+ influx

Kenneth Dakin, Wen Hong Li

Research output: Contribution to journalArticle

9 Scopus citations

Abstract

Using a new fluorescence imaging technique, LAMP, we recently reported that Ca2+ influx through store operated Ca2+ channels (SOCs) strongly inhibits cell coupling in primary human fibroblasts (HF) expressing Cx43. To understand the mechanism of inhibition, we studied the involvement of cytosolic pH (pHi) and Ca2+ ([Ca2+]i) in the process by using fluorescence imaging and ion clamping techniques. During the capacitative Ca2+ influx, there was a modest decline of pHi measured by BCECF. Decreasing pHi below neutral using thioacetate had little effect by itself on cell coupling, and concomitant pHi drop with thioacetate and bulk [Ca2+ rise with ionomycin was much less effective in inhibiting cell coupling than Ca2+ influx. Moreover, clamping pHi with a weak acid and a weak base during Ca2+ influx largely suppressed bulk pHi drop, yet the inhibition of cell coupling was not affected. In contrast, buffering [Ca2+]i with BAPTA, but not EGTA, efficiently prevented cell uncoupling by Ca2+ influx. We concluded that local Ca2+ elevation subjacent to the plasma membrane is the primary cause for closing Cx43 channels during capacitative Ca2+ influx. To assess how Ca2+ influx affects junctional coupling mediated by other types of connexins, we applied the LAMP assay to Hela cells expressing Cx26. Capacitative Ca2+ influx also caused a strong reduction of cell coupling, suggesting that the inhibitory effect by Ca2+ influx may be a more general phenomenon.

Original languageEnglish (US)
Pages (from-to)29-39
Number of pages11
JournalCell Communication and Adhesion
Volume13
Issue number1-2
DOIs
StatePublished - Jan 1 2006

Keywords

  • Capacitative Ca influx
  • Connexin 43
  • Fluorescence imaging
  • Gap junction coupling
  • LAMP assay
  • Photoactivatable fluorophores

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Cell Biology

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