Balietti Marta, Fattorini Giorgia, Pugliese Arianna, Marcotulli Daniele, Bragina Luca, Conti Fiorenzo
Center for Neurobiology of Aging, INRCA, IRCCS, Ancona, Italy.
Department of Experimental and Clinical Medicine, Section of Neuroscience and Cell Biology, Università Politecnica delle Marche, Ancona, Italy.
Front Aging Neurosci. 2018 Apr 4;10:91. doi: 10.3389/fnagi.2018.00091. eCollection 2018.
The molecular substrate of age-associated cognitive decline (AACD) is still elusive. Evidence indicates that AACD is related to synaptic impairment in hippocampus, but different hippocampal regions play different roles, with the dorsal hippocampus (DH) associated to spatial learning, and the ventral hippocampus (VH) crucial for emotionality. If changes in hippocampal function contributes to AACD, this contribution may be reflected in alterations of synaptic protein levels. A commonly used approach to investigate this issue is western blotting. When this technique is applied to the entire hippocampus and the cognitive impairment is evaluated by a single task, changes in expression of a protein might undergo a "dilution effect", as they may occur only in a given hippocampal region. We show that two behavioral tests yield more accurate results than one test in evaluating the function of the whole rat hippocampus by studying the expression of synaptotagmin 1 (SYT1), a vesicular protein whose expression in aged hippocampus is reportedly inconsistent. Analysis of SYT1 levels in the whole hippocampus of rats selected by the Morris water maze (MWM) test only failed to highlight a difference, whereas analysis of SYT1 levels in the whole hippocampus of rats categorized by both the MWM and the step-through passive avoidance (STPA) tests demonstrated a significant increase of SYT1 level in impaired rats. These findings, besides showing that SYT1 increases in impaired aged rats, suggest that using the whole hippocampus in blotting studies may prevent false negative results only if animals are categorized with tests exploring both DH and VH.
衰老相关认知衰退（AACD）的分子底物仍然难以捉摸。有证据表明，AACD与海马体中的突触损伤有关，但不同的海马体区域发挥着不同的作用，背侧海马体（DH）与空间学习相关，腹侧海马体（VH）对情绪调节至关重要。如果海马体功能的变化导致了AACD，那么这种影响可能会反映在突触蛋白水平的改变上。研究这个问题常用的方法是蛋白质免疫印迹法。当将该技术应用于整个海马体，且通过单一任务评估认知障碍时，蛋白质表达的变化可能会受到“稀释效应”的影响，因为这些变化可能仅发生在特定的海马体区域。我们发现，通过研究突触结合蛋白1（SYT1）的表达，在评估整个大鼠海马体功能时，两项行为测试比一项测试能产生更准确的结果。据报道，SYT1是一种囊泡蛋白，其在衰老海马体中的表达并不一致。仅通过莫里斯水迷宫（MWM）测试选择的大鼠全海马体中SYT1水平的分析未能突出差异，而通过MWM和穿梭式被动回避（STPA）测试对大鼠进行分类后，对其全海马体中SYT1水平的分析表明，受损大鼠的SYT1水平显著升高。这些发现，除了表明受损衰老大鼠中SYT1增加外，还表明只有当动物通过探索DH和VH的测试进行分类时，在印迹研究中使用整个海马体才可能防止假阴性结果。
