Oxygen-Glucose Deprivation in Mouse Astrocytes is Associated with Ultrastructural Changes in Connexin 43 Gap Junctions.

Affiliation

Department of Anatomy and Cell Biology, Saarland University, 66421 Homburg, Saar, Germany. Electronic address: [Email]

Abstract

In the intact brain, astrocytes play an important role in a number of physiological functions like spatial buffering of potassium, maintenance of calcium homeostasis, neurotransmitter release, regulation of the cerebral blood flow, and many more. As pathophysiological events upon hypoxic-ischemic brain injury include excitotoxicity by glutamate release as well as oxidative stress, astrocytes and their gap junction-based syncytium are of major relevance for regulating the extent of resulting brain damage. The gap junction protein Connexin (Cx) 43 contributes mainly to the astrocytic intercellular communication. As little is known about the ultrastructural assemblage of Cx43 and its changes in response to hypoxic events, we chose temporary oxygen and glucose deprivation with subsequent reoxygenation (OGD-R) as a metabolic inhibition model of hypoxia in primary murine astrocytes. Gap junction morphology and assembly/disintegration were analyzed at the ultrastructural level using freeze-fracture replica immunolabeling. The exposure of cultured astrocytes to short-term OGD-R resulted in the activation of ERK1/2 (p44/p42), downregulation of Cx43 protein expression, and the rearrangement of Cx43 particles within the cell membrane and within gap junctions. These changes in gap junction morphology were associated with phosphorylation of Cx43 at Serine 368. Analysis of the nearest-neighbor distance within gap junction plaques revealed the loosening of Cx43 particle clusters. Together with the observation of additional connexons being present in the vicinity of gap junction plaques after OGD-R treatment, our study indicates that changes in gap junction assembly are associated with the early phase of hypoxic cell damage.

Keywords

Cx43,FRIL,OGD-R,astrocytes,freeze fracture,ultrastructure,