Cronberg Tobias, Rytter Anna, Wieloch Tadeusz
Laboratory for Experimental Brain Research, Wallenberg Neuroscience Center, Lund University, BMC A13, S-221 84 Lund, Sweden.
Brain Res. 2005 Jul 5;1049(1):120-7. doi: 10.1016/j.brainres.2005.05.003.
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.
高血糖水平在脑缺血期间的加重作用已在临床研究以及全脑缺血和局灶性缺血的体内模型中得到广泛记载。由于体外缺氧时葡萄糖具有高度保护作用，缺乏体外模型阻碍了对高血糖缺血的详细机制研究。我们之前报道过，在含有高钾和低钙的酸性培养基中暴露于缺氧状态的小鼠海马器官型脑片培养物中存在葡萄糖毒性。在本研究中，我们比较了体外缺血（IVI）和高血糖（40 mM）IVI期间钙、一氧化氮和自由基的重要性。细胞外钙是IVI后细胞死亡的普遍因素，但从培养基中去除它对高血糖IVI后的细胞死亡没有影响。当用1,2-双（2-氨基-5-氟苯氧基）乙烷-N,N,N',N'-四乙酸四（乙酰氧基甲基）酯（BAPTA-AM）螯合细胞内钙时，细胞死亡出现得更早，但在高血糖IVI中有所减轻，而在无糖IVI中则增加。添加一氧化氮合酶（NOS）抑制剂盐酸N（ω）-硝基-L-精氨酸甲酯（L-NAME）或自由基清除剂N-叔丁基-α-苯基硝酮（PBN）、去铁胺和N-乙酰-L-半胱氨酸（NAC）在两种模式下均不影响细胞损伤。我们得出结论，体外缺血期间高血糖的加重作用部分由细胞内储存释放的钙离子介导。
