Chelation of intracellular calcium reduces cell death after hyperglycemic in vitro ischemia in murine hippocampal slice cultures.
The aggravating effect of high glucose levels during cerebral ischemia has been extensively documented in clinical studies and in vivo models of global and focal ischemia. Detailed mechanistic studies of hyperglycemic ischemia have so far been hampered by the lack of in vitro models since glucose during anoxia in vitro is highly protective. We have previously reported glucose toxicity in murine hippocampal organotypic slice cultures exposed to anoxia in an acidotic medium containing high potassium and low calcium. In the present study, we compared the importance of calcium, nitric oxide and free radicals during in vitro ischemia (IVI) and hyperglycemic (40 mM) IVI. Extracellular calcium was a ubiquitous factor for cell death after IVI, but its removal from the medium had no effect on cell death after hyperglycemic IVI. When intracellular calcium was chelated by the 1,2-Bis(2-amino-5-fluorophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl) ester (BAPTA-AM) cell death appeared earlier but was mitigated in hyperglycemic IVI, while it was increased in glucose-free IVI. Addition of the nitric oxide synthase (NOS) inhibitor N(omega)-Nitro-L-arginine methyl ester hydrochloride (L-NAME) or the free radical scavengers N-tert-butyl-alpha-phenylnitrone (PBN), deferoxamine and N-acetyl-L-cysteine (NAC) did not affect cell damage in either paradigm. We conclude that the aggravating effect of hyperglycemia during in vitro ischemia is partially mediated by calcium ions released from intracellular stores.
Cronberg
Tobias
T
Laboratory for Experimental Brain Research, Wallenberg Neuroscience Center, Lund University, BMC A13, S-221 84 Lund, Sweden. Tobias.Cronberg@expbr.lu.se
Rytter
Anna
A
Wieloch
Tadeusz
T
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D016923:Cell Death / Q000187:drug effects; D002614:Chelating Agents / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D005609:Free Radicals / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism; D006624:Hippocampus / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D006943:Hyperglycemia / Q000150:complications / Q000378:metabolism* / Q000473:pathology; D066298:In Vitro Techniques; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism; D007511:Ischemia / Q000209:etiology / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D009569:Nitric Oxide / Q000378:metabolism*
D002614:Chelating Agents; D005609:Free Radicals; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D009569:Nitric Oxide; D004533:Egtazic Acid; D005947:Glucose; D002118:Calcium