Gating and regulation of connexin 43 (Cx43) hemichannels

Jorge E. Contreras, Juan C. Sáez, Feliksas F. Bukauskas, Michael V. L. Bennett

Research output: Contribution to journalArticle

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Abstract

Connexin 43 (Cx43) nonjunctional or "unapposed" hemichannels can open under physiological or pathological conditions. We characterize hemichannels comprised of Cx43 or Cx43-EGFP (Cx43 with enhanced GFP fused to the C terminus) expressed in HeLa cells. Channel opening was induced at potentials greater than +60 mV. Open probability appeared to be very low. No comparable opening was detected in the parental, nontransfected HeLa cells. Conductance of fully open single hemichannels was ≈220 pS, which is approximately double that of Cx43 cell-cell channels. Cx43 hemichannels exhibited two types of gating: fast transitions (<1 ms) between the fully open state and a substate of ≈75 pS and slow transitions (>5 ms) between either open state and the fully closed state. Cx43-EGFP hemichannels exhibited only slow transitions (>5 ms) between closed and fully open states. These properties resemble those of the corresponding Cx43 and Cx43-EGFP cell-cell channels. Cx43 with EGFP on the N terminus (EGFP-Cx43) inserted into the surface and formed plaques but did not form hemichannels or cell-cell channels. Hemichannel blockers, 18β-glycyrrhetinic acid of La3+, blocked depolarization-induced currents. Uptake of ethidium bromide (i) was faster in Cx43 and Cx43-EGFP than parental and EGFP-Cx43 cells, (ii) was directly correlated with Cx43-EGFP expression, (iii) was reduced by hemichannel blockers, and (iv) occurred at the same low rate in EGFP-Cx43 and parental cells. Although hemichannel opening was not detected electrophysiologically at the resting potential, infrequent or brief opening could account for ethidium bromide uptake. Opening of Cx43 hemichannels may mediate normal signaling or be deleterious.

Original languageEnglish (US)
Pages (from-to)11388-11393
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume100
Issue number20
DOIs
StatePublished - Sep 30 2003

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Connexin 43
Ethidium
HeLa Cells
Glycyrrhetinic Acid

Keywords

  • Connexon
  • Dye uptake
  • Gap junction
  • Permeation

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

Gating and regulation of connexin 43 (Cx43) hemichannels. / Contreras, Jorge E.; Sáez, Juan C.; Bukauskas, Feliksas F.; Bennett, Michael V. L.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 100, No. 20, 30.09.2003, p. 11388-11393.

Research output: Contribution to journalArticle

Contreras, Jorge E. ; Sáez, Juan C. ; Bukauskas, Feliksas F. ; Bennett, Michael V. L. / Gating and regulation of connexin 43 (Cx43) hemichannels. In: Proceedings of the National Academy of Sciences of the United States of America. 2003 ; Vol. 100, No. 20. pp. 11388-11393.
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AB - Connexin 43 (Cx43) nonjunctional or "unapposed" hemichannels can open under physiological or pathological conditions. We characterize hemichannels comprised of Cx43 or Cx43-EGFP (Cx43 with enhanced GFP fused to the C terminus) expressed in HeLa cells. Channel opening was induced at potentials greater than +60 mV. Open probability appeared to be very low. No comparable opening was detected in the parental, nontransfected HeLa cells. Conductance of fully open single hemichannels was ≈220 pS, which is approximately double that of Cx43 cell-cell channels. Cx43 hemichannels exhibited two types of gating: fast transitions (<1 ms) between the fully open state and a substate of ≈75 pS and slow transitions (>5 ms) between either open state and the fully closed state. Cx43-EGFP hemichannels exhibited only slow transitions (>5 ms) between closed and fully open states. These properties resemble those of the corresponding Cx43 and Cx43-EGFP cell-cell channels. Cx43 with EGFP on the N terminus (EGFP-Cx43) inserted into the surface and formed plaques but did not form hemichannels or cell-cell channels. Hemichannel blockers, 18β-glycyrrhetinic acid of La3+, blocked depolarization-induced currents. Uptake of ethidium bromide (i) was faster in Cx43 and Cx43-EGFP than parental and EGFP-Cx43 cells, (ii) was directly correlated with Cx43-EGFP expression, (iii) was reduced by hemichannel blockers, and (iv) occurred at the same low rate in EGFP-Cx43 and parental cells. Although hemichannel opening was not detected electrophysiologically at the resting potential, infrequent or brief opening could account for ethidium bromide uptake. Opening of Cx43 hemichannels may mediate normal signaling or be deleterious.

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