Reduced Connexin43 Expression Correlates with c-Src Activation, Proliferation, and Glucose Uptake in Reactive Astrocytes After an Excitotoxic Insult ESTER GANGOSO, 1 PASCAL EZAN, 2,3,4 JOS  E CARLOS VALLE-CASUSO, 1 SANDRA HERRERO-GONZ  ALEZ, 1 ANNETTE KOULAKOFF, 2,3,4 JOSE M. MEDINA, 1 CHRISTIAN GIAUME, 2,3,4 AND ARANTXA TABERNERO 1 * 1 Departamento de Bioqu  ımica y Biolog  ıa Molecular, Instituto de Neurociencias de Castilla y Le on (INCYL), Universidad de Salamanca, Spain 2 Colle `ge de France, Center for Interdisciplinary Research in Biology (CIRB)/Centre National de la Recherche Scientifique, Unit  e Mixte de Recherche 7241/ Institut National de la Sant  e et de la Recherche M edicale U1050, 75231 Paris Cedex 05, France 3 University Pierre et Marie Curie, ED, N°158, 75005 Paris, France 4 MEMOLIFE Laboratory of Excellence and Paris Science Lettre Research University, 75005 Paris, France KEY WORDS connexin; glia; brain; neuroprotection; injury ABSTRACT In diverse brain pathologies, astrocytes become reactive and undergo profound phenotypic changes. Connexin43 (Cx43), the main gap junction channel-forming protein in astrocytes, is one of the proteins modified in reactive astro- cytes. Downregulation of Cx43 in cultured astrocytes acti- vates c-Src, promotes proliferation, and increases the rate of glucose uptake; however, so far there have been no stud- ies examining whether this cascade of events takes place in reactive astrocytes. In this work, we analyzed this pathway after a cortical lesion induced by a kainic acid injection. As previously described, astrocytes reacted to the lesion with an increase in glial fibrillary acidic protein and a decrease in Cx43 expression. Some of these reactive astrocytes prolif- erated, as estimated by bromodeoxyuridine incorporation and cyclins D1 and D3 upregulation. In addition, the expression of the glucose transporter GLUT-3 and the enzyme responsible for glucose phosphorylation, Type II hexokinase (Hx-2), were induced in reactive astrocytes, sug- gesting an increased glucose uptake. Previous in vitro stud- ies reported that c-Src is the link between Cx43 and glucose uptake and proliferation in astrocytes. Here, we found that c-Src activity increased in the lesioned area. c-Src activa- tion and Cx43 downregulation preceded the peak of Hx-2 and cyclin D3 expression, suggesting that c-Src could medi- ate the effect of Cx43 on glucose uptake and proliferation in reactive astrocytes after an excitotoxic insult. Interest- ingly, we identify c-Src, GLUT-3, and Hx-2 in the signaling mechanisms involved in the reaction of astroglia to injury. Altogether these data contribute to identify new therapeutical targets to enhance astrocyte neuroprotective activities. V V C 2012 Wiley Periodicals, Inc. INTRODUCTION In diverse pathological situations, from acute injury to neurodegenerative diseases, astrocytes become reactive and undergo profound morphological and phenotypic changes (Giaume et al., 2010; Ridet et al., 1997). Modifi- cations in the expression of a variety of proteins have been described in reactive astrocytes, including connexin43 (Cx43), the main gap junction channel-form- ing protein in astrocytes (Koulakoff et al., 2008). Cx43 may be up- or downregulated depending on the type of brain pathology or the nature of injury, the time scale, and the distance from the lesioned area (Giaume et al., 2010; Rouach et al., 2002). Moreover, it has been shown that Cx43 expression is downregulated in cultured astrocytes submitted to inflammatory or injury media- tors, such as pro-inflammatory cytokines or peptides known to be produced during inflammation, such as endothelins (Hinkerohe et al., 2005; Meme et al., 2006; Rozyczka et al., 2005; Zhao et al., 2006), an effect that is potentiated in presence of Ab (Meme et al., 2006). In addition, excitotoxic treatments triggering neuronal death in vivo lead to a strong decrease of Cx43 immuno- reactivity in reactive astrocytes located within the area depleted of neurons (Hossain et al., 1994; Koulakoff et al., 2008; Ochalski et al., 1995; Sawchuk et al., 1995) and, at the ultrastructural level, to a virtual absence of gap junctions between astrocytes in the core of the lesion (Hossain et al., 1994; Ochalski et al., 1995). This pattern of labeling contrasts with the increased Cx43 immunoreactivity found in astrocytes located at the pe- riphery of the lesion (Hossain et al., 1994; Koulakoff et al., 2008; Ochalski et al., 1995). Changes in Cx43 expression affect gap junctional com- munication and astroglial networking but also have an impact on intracellular signaling (Giaume et al., 2010). In fact, the intracellular carboxy terminal tail of Cx43 interacts with a large number of signaling and scaffolding proteins (Giepmans, 2006; Herve et al., Additional Supporting Information may be found in the online version of this article. Grant sponsor: Ministerio de Econom  ıa y Competitividad; Grant number: FEDER BFU2011-27602 (A.T.); Grant sponsor: F ed eration de la Recherche sur le Cerveau (C.G.) Fellowships: E.G. (AP2008-03515) and S.H-G from the Ministerio de Econom ıa y Competividad. J.C.V-C. from the Junta de Castilla y Le on. *Correspondence to: Arantxa Tabernero, Departamento de Bioqu  ımica y Biolog  ıa Molecular, Instituto de Neurociencias de Castilla y Le on (INCYL), Universidad de Salamanca, C/ Pintor Fernando Gallego 1, 37007 Salamanca, Spain. E-mail: ataber@usal.es Received 14 May 2012; Accepted 23 August 2012 DOI 10.1002/glia.22418 Published online in Wiley Online Library (wileyonlinelibrary. com). GLIA 00:000–000 (2012) V V C 2012 Wiley Periodicals, Inc.