Intracellular magnesium-dependent modulation of gap junction channels formed by neuronal connexin36

J Neurosci. 2013 Mar 13;33(11):4741-53. doi: 10.1523/JNEUROSCI.2825-12.2013.

Abstract

Gap junction (GJ) channels composed of Connexin36 (Cx36) are widely expressed in the mammalian CNS and form electrical synapses between neurons. Here we describe a novel modulatory mechanism of Cx36 GJ channels dependent on intracellular free magnesium ([Mg(2+)]i). We examined junctional conductance (gj) and its dependence on transjunctional voltage (Vj) at different [Mg(2+)]i in cultures of HeLa or N2A cells expressing Cx36. We found that Cx36 GJs are partially inhibited at resting [Mg(2+)]i. Thus, gj can be augmented or reduced by lowering or increasing [Mg(2+)]i, respectively. Similar changes in gj and Vj-gating were observed using MgATP or K2ATP in pipette solutions, which increases or decreases [Mg(2+)]i, respectively. Changes in phosphorylation of Cx36 or in intracellular free calcium concentration were not involved in the observed Mg(2+)-dependent modulation of gj. Magnesium ions permeate the channel and transjunctional asymmetry in [Mg(2+)]i resulted in asymmetric Vj-gating. The gj of GJs formed of Cx26, Cx32, Cx43, Cx45, and Cx47 was also reduced by increasing [Mg(2+)]i, but was not increased by lowering [Mg(2+)]i; single-channel conductance did not change. We showed that [Mg(2+)]i affects both open probability and the number of functional channels, likely through binding in the channel lumen. Finally, we showed that Cx36-containing electrical synapses between neurons of the trigeminal mesencephalic nucleus in rat brain slices are similarly affected by changes in [Mg(2+)]i. Thus, this novel modulatory mechanism could underlie changes in neuronal synchronization under conditions in which ATP levels, and consequently [Mg(2+)]i, are modified.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Animals
  • Animals, Newborn
  • Biophysical Phenomena / drug effects
  • Biophysical Phenomena / physiology
  • Cations, Divalent / metabolism
  • Cell Line, Tumor
  • Chelating Agents / pharmacology
  • Connexins / genetics
  • Connexins / physiology*
  • Dose-Response Relationship, Drug
  • Egtazic Acid / analogs & derivatives
  • Egtazic Acid / pharmacology
  • Female
  • Gap Junctions / drug effects
  • Gap Junctions / physiology*
  • Green Fluorescent Proteins / genetics
  • Humans
  • In Vitro Techniques
  • Intracellular Fluid / metabolism*
  • Ion Channel Gating / drug effects
  • Ion Channel Gating / physiology*
  • Magnesium / metabolism*
  • Magnesium / pharmacology
  • Male
  • Membrane Potentials / drug effects
  • Membrane Potentials / physiology
  • Mice
  • Neurons / cytology
  • Neurons / metabolism*
  • Patch-Clamp Techniques
  • Phosphorylation
  • Rats
  • Rats, Sprague-Dawley
  • Tegmentum Mesencephali / cytology
  • Transfection

Substances

  • Cations, Divalent
  • Chelating Agents
  • Connexins
  • GJB2 protein, human
  • connexin 36
  • Green Fluorescent Proteins
  • Egtazic Acid
  • Adenosine Triphosphate
  • Magnesium
  • 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid