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Neuronal protein synthesis is essential for long-term memory consolidation, and its dysregulation is implicated in various neurodegenerative disorders, including Alzheimer’s disease (AD). Cellular stress triggers the activation of protein kinases that converge on the phosphorylation of eukaryotic translation initiation factor 2α (eIF2α), which attenuates mRNA translation. This translational inhibition is one aspect of the integrated stress response (ISR). We found that postmortem brain tissue from AD patients showed increased phosphorylation of eIF2α and reduced abundance of eIF2B, another key component of the translation initiation complex. Systemic administration of the small-molecule compound ISRIB (which blocks the ISR downstream of phosphorylated eIF2α) rescued protein synthesis in the hippocampus, measures of synaptic plasticity, and performance on memory-associated behavior tests in wild-type mice cotreated with salubrinal (which inhibits translation by inducing eIF2α phosphorylation) and in both β-amyloid-treated and transgenic AD model mice. Thus, attenuating the ISR downstream of phosphorylated eIF2α may restore hippocampal protein synthesis and delay cognitive decline in AD patients.

Figure 1. Expression of AMPKα isoforms is dysregulated in AD hippocampus. (A) Hippocampus (Hip) lysate from sAD patients showed increased AMPKα1 and decreased AMPKα2 levels as compared with those of age-matched controls (CT). n = 10 with up to 4 technical replicates. *P = 0.0119; **P = 0.0014, unpaired t test. (B) Representative images of AMPKα isoform dysregulation in area CA1 of hippocampus in AD and age-matched control patients. Scale bar: 50 μm. Immunohistochemical experiments were replicated independently 3 times. (C) AMPKα isoform expression was unaffected in cerebellum (CER) samples from AD patients. n = 5 with up to 3 technical replicates. P = 0.8457 for AMPKα1; P = 0.9870 for AMPKα2, unpaired t test. (D) Hippocampal lysate from LBD patients had unaffected AMPKα isoform levels. n = 4 for control; n = 3 for LBD with 1 technical replicate. P = 0.9146 for AMPKα1; P = 0.5635 for AMPKα2, unpaired t test. (E) Levels of AMPK isoforms were unaltered in hippocampal tissue from FTD patients. n = 8 for control with 1 technical replicate; n = 5 for FTD with up to 3 technical replicates. P = 0.9283 for AMPKα1; P = 0.335 for AMPKα2, unpaired t test. (F) AMPKα1 levels were significantly increased in cortical lysates from FAD patients, while AMPKα2 levels were unaffected. n = 5 with 4 technical replicates. **P = 0.0060 for AMPKα1; P = 0.9412 for AMPKα2, unpaired t test. (G) AMPKα1 levels were significantly increased in hippocampal lysates from Tg19959 AD model mice compared with WT controls. AMPKα2 levels were unaffected. n = 7 with up to 2 technical replicates. **P = 0.0023 for AMPKα1; P = 0.9094 for AMPKα2, unpaired t test. Box-and-whisker plots represent the interquartile range, with the line across the box indicating the median. Whiskers show the highest and lowest values detected. (H) Immunofluorescent labeling of DAPI (blue) and AMPKα1 (red) distribution in area CA1 mouse hippocampal slices. n = 3. Scale bar: 200 μm.

Characterization of the molecular signaling pathways underlying protein synthesis-dependent forms of synaptic plasticity, such as late long-term potentiation (L-LTP), can provide insights not only into memory expression/maintenance under physiological conditions but also potential mechanisms associated with the pathogenesis of memory disorders. Here, we report in mice that L-LTP failure induced by the mammalian (mechanistic) target of rapamycin complex 1 (mTORC1) inhibitor rapamycin is reversed by brain-specific genetic deletion of PKR-like ER kinase, PERK (PERK KO), a kinase for eukaryotic initiation factor 2α (eIF2α). In contrast, genetic removal of general control non-derepressible-2, GCN2 (GCN2 KO), another eIF2α kinase, or treatment of hippocampal slices with the PERK inhibitor GSK2606414, does not rescue rapamycin-induced L-LTP failure, suggesting mechanisms independent of eIF2α phosphorylation. Moreover, we demonstrate that phosphorylation of eukaryotic elongation factor 2 (eEF2) is significantly decreased in PERK KO mice but unaltered in GCN2 KO mice or slices treated with the PERK inhibitor. Reduction in eEF2 phosphorylation results in increased general protein synthesis, and thus could contribute to the mTORC1-independent L-LTP in PERK KO mice. We further performed experiments on mutant mice with genetic removal of eEF2K (eEF2K KO), the only known kinase for eEF2, and found that L-LTP in eEF2K KO mice is insensitive to rapamycin. These data, for the first time, connect reduction in PERK activity with the regulation of translation elongation in enabling L-LTP independent of mTORC1. Thus, our findings indicate previously unrecognized levels of complexity in the regulation of protein synthesis-dependent synaptic plasticity. Read the Editorial Highlight for this article on page 119. Cover Image for this issue: doi: 10.1111/jnc.14185.

It is imperative to develop novel therapeutic strategies for Alzheimer's disease (AD) and related dementia syndromes based on solid mechanistic studies. Maintenance of memory and synaptic plasticity relies on de novo protein synthesis, which is partially regulated by phosphorylation of eukaryotic elongation factor 2 (eEF2) via its kinase eEF2K. Abnormally increased eEF2 phosphorylation and impaired mRNA translation have been linked to AD. We recently reported that prenatal genetic suppression of eEF2K is able to prevent aging-related cognitive deficits in AD model mice, suggesting the therapeutic potential of targeting eEF2K/eEF2 signaling in AD. Here, we tested two structurally distinct small-molecule eEF2K inhibitors in two different lines of AD model mice after the onset of cognitive impairments. Our data revealed that treatment with eEF2K inhibitors improved AD-associated synaptic plasticity impairments and cognitive dysfunction, without altering brain amyloid β (Aβ) and tau pathology. Furthermore, eEF2K inhibition alleviated AD-associated defects in dendritic spine morphology, post-synaptic density formation, protein synthesis, and dendritic polyribosome assembly. Our results may offer critical therapeutic implications for AD, and the proof-of-principle study indicates translational implication of inhibiting eEF2K for AD and related dementia syndromes.

2-Hydroxypropyl-β-cyclodextrin (HP-β-CD) is an experimental therapy for Niemann-Pick disease type C (NPC) that reduced neuronal cholesterol and ganglioside storage, reduced Purkinje cell death, and increased lifespan in npc1−/− mice and NPC1 cats. In this study, tissue distribution was investigated in normal cats that received a single 120-mg dose of [ 14 C]-HP-β-CD (approximately 200 μCi/cat) via the cerebellomedullary cistern (CBMC) and lumbar cistern. One cat was euthanized at each of various time points up to 24 hours postdose for subsequent processing and quantitative whole-body autoradiographic analysis. HP-β-CD-derived radioactivity absorbed from the CBMC was widely distributed to cat tissues; most tissues were observed to have reached their highest concentration at 1 hour postdose. HP-β-CD-derived radioactivity penetrated into the deeper parts of the central nervous system with the highest concentration at 4 hours (403 μg Eq/g or 0.28 mM) and remained high (49.7 μg Eq/g or 0.03 mM) at 24 hours. The relatively long halflife (11-30 hours) in cerebral ventricles and the subarachnoid space surrounding the brain and spinal cord might contribute to the efficacy of HP-β-CD in NPC1 cats. Other tissues with high concentrations of radioactivity were nasal turbinates, pituitary gland, and urinary bladder, while relatively low concentrations were observed in blood and bile. K E Y W O R D S animal model, brain, cholesterol, drug therapy, inborn errors of metabolism, Niemann Pick type C, storage diseases Abbreviations: AUC, area under the tissue concentration-time curve; AUC inf , AUC from time 0 to infinity with extrapolation of the terminal phase; AUC last , AUC from time 0 to the time correspondent to the last quantifiable

Neuronal protein synthesis is essential for long-term memory consolidation. Conversely, dysregulation of protein synthesis has been implicated in a number o neurodegenerative disorders, including Alzheimer's disease (AD). Several types of cellular stress trigger the activation of protein kinases that converge on the phosphorylation of eukaryotic translation initiation facor 2 α (eIF2α-P). This leads to attenuation of cap-dependent mRNA translation, a component of the integrated stress response (ISR). We show that AD brains exhibit increased eIF2α-P and reduced eIF2B, key components of the eIF2 translation initiation complex. We further demonstrate that attenuating the ISR wit the small molecule compound ISRIB (ISR inhibitor) rescues hippocampal protein synthesis and corrects impaired synaptic plasticity and memory in mouse models of AD. Our findings suggest that attenuating eIF2α-P-mediated translational inhibition may comprise an effective approach to alleviate cognitive decline in AD.

It is imperative to develop novel therapeutic strategies for Alzheimer's disease (AD) and related dementia syndromes based on solid mechanistic studies. Maintenance of memory and synaptic plasticity relies on de novo protein synthesis, which is partially regulated by phosphorylation of eukaryotic elongation factor 2 (eEF2) via its kinase eEF2K. Abnormally increased eEF2 phosphorylation and impaired mRNA translation have been linked to AD. We recently reported that prenatal genetic suppression of eEF2K is able to prevent aging-related cognitive deficits in AD model mice, suggesting the therapeutic potential of targeting eEF2K/eEF2 signaling in AD. Here, we tested two structurally-distinct small-molecule eEF2K inhibitors in two different lines of AD model mice after onset of cognitive impairments. Our data revealed that treatment with eEF2K inhibitors improved AD-associated synaptic plasticity impairments and cognitive dysfunction, without altering brain amyloid (Aβ) and tau pathology. Furthermore, eEF2K inhibition alleviated AD-associated defects in dendritic spine morphology, postsynaptic density formation, protein synthesis, and dendritic polyribosome assembly. Our results may offer critical therapeutic implications for AD, and the proof-of-principle study indicates translational implication of inhibiting eEF2K for AD and related dementia syndromes.