Astrocytic gap junction inhibition by carbenoxolone enhances the protective effects of ischemic preconditioning following cerebral ischemia

Di Ma,Xiang Yin,Yingying Cheng,Liangshu Feng,Li Cui,Jiachun Feng,Yulei Hao,Meiying Xin,Jiulin You

doi:10.1186/s12974-018-1230-5

Abstract

BackgroundStroke is the second leading cause of death worldwide and the most common cause of adult-acquired disability in many nations. Thus, attenuating the damage after ischemic injury and improving patient prognosis are of great importance. We have indicated that ischemic preconditioning (IP) can effectively reduce the damage of ischemia reperfusion and that inhibition of gap junctions may further reduce this damage. Although we confirmed that the function of gap junctions is closely associated with glutamate, we did not investigate the mechanism. In the present study, we aimed to clarify whether the blockade of cellular communication at gap junctions leads to significant reductions in the levels of glutamate released by astrocytes following cerebral ischemia.MethodsTo explore this hypothesis, we utilized the specific blocking agent carbenoxolone (CBX) to inhibit the opening and internalization of connexin 43 channels in an in vitro model of oxygen-glucose deprivation/re-oxygenation (OGD/R), following IP.ResultsOGD/R resulted in extensive astrocytic glutamate release following upregulation of hemichannel activity, thus increasing reactive oxygen species (ROS) generation and subsequent cell death. However, we observed significant increases in neuronal survival in neuron-astrocyte co-cultures that were subjected to IP prior to OGD/R. Moreover, the addition of CBX enhanced the protective effects of IP during the re-oxygenation period following OGD, by means of blocking the release of glutamate, increasing the level of the excitatory amino acid transporter 1, and downregulating glutamine expression.ConclusionsOur results suggest that combined use of IP and CBX represents a novel therapeutic strategy to attenuate damage from cerebral ischemia with minimal adverse side effects.

Highlights

Stroke is the second leading cause of death worldwide and the most common cause of adultacquired disability in many nations
CBX enhances the protective effects of ischemic preconditioning (IP) after 2 h of Oxygen-glucose deprivation (OGD) or 6 h of re-oxygenation We supposed that Gap junction (GJ) may exert a unique pathophysiological role during cerebral ischemia
As GJs are composed of connexin 43 (Cx43), we first investigated the expression of Cx43 in several astrocyte culture conditions [no treatment (NT), OGD/R, IP + OGD/R at different time points (1, 3, and 6 h)

Summary

Introduction

Stroke is the second leading cause of death worldwide and the most common cause of adultacquired disability in many nations. Recent studies have increasingly focused on the therapeutic potential of ischemic preconditioning (IP) in attenuating structural and functional deficits following ischemia reperfusion [1]. Several clinical trials have confirmed that such preconditioning strategies can attenuate the pathophysiological consequences of ischemia-reperfusion injury prior to cardiac bypass surgery [5, 6]. Gap junctions (GJs) are composed of hemichannels found at the corresponding position on the cell membrane where adjacent cells contact each other, forming channels which allow GJ-mediated intercellular communication (GJIC). This enables the synchronized information transfer between adjacent astrocytes, as well as metabolic substrate exchange and ion balance [8]. The regulation of the internal environment [9,10,11] is a function of astrocytes; Cx43, found both on the mitochondria and the cell membrane of astrocytes, as well as the hemichannels and gap junctions it forms, might play complex roles in ischemic injury [12, 13]

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