Abstract
Astrocyte death may occur in neurodegenerative disorders and complicates the outcome of brain ischemia, a condition associated with high extracellular levels of adenosine and glutamate. We show that pharmacological activation of A1 adenosine and mGlu3 metabotropic glutamate receptors with N6-chlorocyclopentyladenosine (CCPA) and (–)2-oxa-4-aminocyclo-[3.1.0]hexane-4,6-dicarboxylic acid (LY379268), respectively, protects cultured astrocytes against apoptosis induced by a 3-h exposure to oxygen/glucose deprivation (OGD). Protection by CCPA and LY379268 was less than additive and was abrogated by receptor blockade with selective competitive antagonists or pertussis toxin. Both in control astrocytes and in astrocytes exposed to OGD, CCPA and LY379268 induced a rapid activation of the phosphatidylinositol-3-kinase (PI3K) and extracellular signal-regulated kinases 1 and 2 (ERK1/2)/mitogen-activated protein kinase (MAPK) pathways, which are known to support cell survival. In cultures exposed to OGD, CCPA and LY379268 reduced the activation of c-Jun N-terminal kinase and p38/MAPK, reduced the levels of the proapoptotic protein Bad, increased the levels of the antiapoptotic protein Bcl-XL, and were highly protective against apoptotic death, as shown by nuclear 4′-6-diamidino-2-phenylindole staining and measurements of caspase-3 activity. All of these effects were attenuated by treatment with 1,4-diamino-2,3-dicyano-1,4-bis(methylthio)butadiene (U0126) and 2-(4-morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride (LY294002), which inhibit the MAPK and the PI3K pathways, respectively. These data suggest that pharmacological activation of A1 and mGlu3 receptors protects astrocytes against hypoxic/ischemic damage by stimulating the PI3K and ERK1/2 MAPK pathways.
Footnotes
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This research was supported by funds (to R.C. and F.C.) from the Centre of Excellence on Aging of the University of Chieti and the Italian Ministry of Education, University and Research.
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R.C. and I.D.A. contributed equally to this article.
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Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
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doi:10.1124/mol.106.031617.
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ABBREVIATIONS: CCPA, N6-chlorocyclopentyladenosine; ASK1, apoptosis-signal-regulating kinase 1; DAPI, 4′-6-diamidino-2-phenylindole; DMEM, Dulbecco's modified Eagle's medium; DPCPX, 1,3-dipropyl-8-cyclopentyl xanthine; ERK1/2, extracellular signal-regulated kinases 1 and 2; HRP, horseradish peroxidase; JNK, c-Jun N-terminal kinase; LY294002, 2-(4-morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride; LY341495, (2S, 1′S, 2′S)-2(9-xanthylmethyl)-2-(2′-carboxycyclo-propyl)glycine; LY379268, (–)2-oxa-4-aminocyclo-[3.1.0] hexane-4,6-dicarboxylic acid; MAPK, mitogen-activated protein kinase; mGluR, metabotropic glutamate receptor; MTS, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium; PBS, phosphate-buffered saline; PI3K, phosphatidylinositol-3 kinase; OGD, oxygen glucose deprivation; PTX, pertussis toxin; PKB, protein kinase B; U0126, 1,4-diamino-2,3-dicyano-1,4-bis(2-aminophenylthio)-butadiene; ANOVA, analysis of variance; RFLU, relative fluorescence units; MEK, mitogen-activated protein kinase kinase; solvent A, KH2PO4 and tetrabutylammonium phosphate; solvent B, methanol plus KH2PO4 and tetrabutylammonium phosphate.
- Received October 11, 2006.
- Accepted February 9, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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