关键词: aging, memory, synaptic plasticity, LTP, protein synthesis, eEF2, mTOR, Alzheimer&rsquo, s disease

Abstract: Objective: Understanding of the molecular mechanisms underlying age-associated cognitive impairments will not only contribute to our general knowledge about “aging” biology, but also provide insights for more effective strategies to prevent and improve the quality of life for both normal aging and pathological aging such as Alzheimer’s disease (AD). multiple lines of evidence suggest that subtle morphological and/or biochemical neuronal changes, instead of profound loss of neurons, is responsible for aging-related impairments of cognition and synaptic plasticity, which is often measured in vertebrates as long-term potentiation (LTP), a synaptic model for memory. A substantial body of evidence demonstrates that de novo protein synthesis (mRNA translation) is indispensable to maintain long-lasting forms of memory and synaptic plasticity. Of interest, activities of translational factors involved into various stages of protein synthesis and synthesis of translational machinery per se (i.e. translational capacity) are known to be regulated in synaptic plasticity and memory formation by various signaling pathways.
Methods: Breeders for C57BL/6J mice were purchased from the Jackson Laboratory (Bar Harbor, ME USA). All mice were housed in the barrier Mouse Facility at Wake Forest School of Medicine Animal Facility. Mice were kept in compliance with the National Institute of Health (NIH) guide for Care and Use of Laboratory Animals. The facility kept a 12 hour light/dark cycle with regular feeding, cage cleaning, and 24 hour access to water. Male and female mice, aged 3-5 or 19-21 months, were used for these experiments. Mice were subjected to the following behavioral tasks: hidden platform Morris water maze, which was consisted of 4 trials each day for 5 consecutive days, with probe trial being carried out 2 hours after training day 5; visible platform task, which was consisted of 4 trials each day for 2 consecutive days with the escape platform marked by a visible cue and moves randomly between four locations; novel object recognition, in which the amount of time spent exploring the novel object was normalized by the total time spent exploring both objects to yield a preference index to calculate percent object preference; reversal Y water maze: mice were trained to pick up one side of the maze, where a platform was hidden. The memory test phase began after a delay of 24 hours, which included 5 trials.  For mice chose the right arm, the escape platform was switched to the opposite arm, and the mice were trained to learn the new location of the platform. For electrophysiology experiments, acute 400 µm transverse hippocampal slices were prepared using a Leica VT1200S vibratome. Late long-term potentiation (LTP) was induced using high-frequency stimulation (HFS) consisting of two 1-sec 100 Hz trains separated by 60 sec, each delivered at 70-80% of the intensity that evoked spiked fEPSPs. Early LTP was induced using one-train HFS (100 Hz) delivered at 25-30% of the intensity that evoked spiked fEPSPs. Mouse brain tissue was harvested for biochemical experiments as described before (Ma et al., Nature Neuroscience, 2013).
Results: Here we first assessed and compared the performance of cognition and synaptic plasticity in young (3-5 month old) and aged c57BL/6J mice (19-21 months old). Findings from behavioral tests demonstrated that old mice, compared to young mice, displayed impairments in spatial learning/memory, working memory, and behavioral flexibility. Further, synaptic electrophysiology experiments on hippocampal slices revealed that early form LTP (a synaptic model for memory formation) was inhibited in old mice.   At the molecular level, biochemical assays on brain tissue showed dysregulation of signaling pathways controlling protein synthesis capacity including: up-regulation of AKT-mTORC1-p70S6K signaling, which is associated with translation of terminal oligopyrimidine (TOP) class of mRNAs that encode translational machinery; hyper-phosphorylation of mRNA translational elongation factor 2 (eEF2) and its upstream regulator AMP-activated protein kinase (AMPK), indicating repression of general protein synthesis. Moreover, young and old mice exhibited similar brain levels of translational initiation factor 2α (eIF2α) phosphorylation, which is known to be increased in AD and linked to the disease pathophysiology.
Conclusion: Our findings provide evidence at the molecular level to highlight the similarity and difference between normal and pathological aging, which may contribute to future studies on diagnostic/prognostic biomarkers for aging-related dementia syndromes.

Key words: aging, memory, synaptic plasticity, LTP, protein synthesis, eEF2, mTOR, Alzheimer&rsquo, s disease