Huntington's disease (HD) is an inherited neurodegenerative disorder caused by an expanded stretch of CAG trinucleotide repeats that results in neuronal dysfunction and death. Here, the HD consortium reports the generation and characterization of 14 induced pluripotent stem cell (iPSC) lines from HD patients and controls. Microarray profiling revealed CAG expansion-associated gene expression patterns that distinguish patient lines from controls, and early onset versus late onset HD. Differentiated HD neural cells showed disease associated changes in electrophysiology, metabolism, cell adhesion, and ultimately cell death for lines with both medium and longer CAG repeat expansions. The longer repeat lines were however the most vulnerable to cellular stressors and BDNF withdrawal using a range of assays across consortium laboratories. The HD iPSC collection represents a unique and well-characterized resource to elucidate disease mechanisms in HD and provides a novel human stem cell platform for screening new candidate therapeutics.
Mammalian target of rapamycin (mTOR) is a key regulator of translational capacity. The mTOR inhibitor rapamycin can prevent forms of protein synthesis-dependent synaptic plasticity such as long-term facilitation in Aplysia and late-phase long-term potentiation (L-LTP) in the hippocampal CA1 region of rodents. In the latter model, two issues remain to be addressed: defining the L-LTP phase sensitive to rapamycin and identifying the site of rapamycinsensitive protein synthesis. Here, we show that L-LTP is sensitive to application of rapamycin only during the induction paradigm, whereas rapamycin application after the establishment of L-LTP was ineffective. Second, we observed that Thr-389-phosphorylated p70 S6 kinase (p70 S6K ), the main active phosphoform of the mTOR effector p70 S6K , was induced in an N-methyl-D-aspartateand phosphatidylinositol 3-kinase-dependent manner throughout the dendrites but not in the cell bodies of CA1 neurons in hippocampal slices after L-LTP induction. A similar dendrite-wide activation of p70 S6K was induced in primary hippocampal neurons by depolarization with KCL or glutamate. In primary hippocampal neurons, the sites of dendritic activation of p70 S6K appeared as discrete compartments along dendritic shafts like the hotspots for fast dendritic translation. Conversely, only a subset of dendritic spines also displayed activated p70 S6K . Taken together, the present data suggest that the N-methyl-D-aspartate-, phosphatidylinositol 3-kinase-dependent dendritic activation of the mTOR-p70 S6K pathway is necessary for the induction phase of protein synthesisdependent synaptic plasticity. Newly synthesized proteins in dendritic shafts could be targeted selectively to activity-tagged synapses. Thus, coordinated activation of dendrite-wide translation and synaptic-specific activation is likely to be necessary for long-term synaptic plasticity. F orms of long-term synaptic plasticity that require protein synthesis are believed to be cellular counterparts of longterm memory storage, whereas forms of synaptic plasticity that do not require protein synthesis are believed to be counterparts of short-term memory (1). In particular, dendritic protein synthesis is believed to play a crucial role in long-term synaptic plasticity and memory (1-4). Mammalian target of rapamycin (mTOR) regulates the translation initiation complex in a rapamycin-sensitive manner. It does so primarily through its downstream targets, the kinase p70 S6 kinase (p70 S6K ) and the elongation factor binding protein 4E-BP1. p70 S6K is a major regulator of translation under the control of multiple signal transduction pathways including phosphatidylinositol 3-kinase (PI3K) (5). It increases translational capacity by promoting the expression of several members of the translational machinery whose mRNAs display oligopyrimidine tracts at their 5Ј ends (6). 4E-BP1 is an inhibitor of the cap binding protein eukaryotic initiation factor 4E. 4E-BP1 phosphorylation by mTOR leads to increased translation of capped mRNAs (7). The major d...
Neural cultures derived from Huntington’s disease (HD) patient-derived induced pluripotent stem cells were used for ‘omics’ analyses to identify mechanisms underlying neurodegeneration. RNA-seq analysis identified genes in glutamate and GABA signaling, axonal guidance and calcium influx whose expression was decreased in HD cultures. One-third of gene changes were in pathways regulating neuronal development and maturation. When mapped to stages of mouse striatal development, the profiles aligned with earlier embryonic stages of neuronal differentiation. We observed a strong correlation between HD-related histone marks, gene expression and unique peak profiles associated with dysregulated genes, suggesting a coordinated epigenetic program. Treatment with isoxazole-9, which targets key dysregulated pathways, led to amelioration of expanded polyglutamine repeat-associated phenotypes in neural cells and of cognitive impairment and synaptic pathology in HD model R6/2 mice. These data suggest that mutant huntingtin impairs neurodevelopmental pathways that could disrupt synaptic homeostasis and increase vulnerability to the pathologic consequence of expanded polyglutamine repeats over time.
Background-CB 1 cannabinoid receptors in the brain are known to participate in the regulation of reward-based behaviors, however, the contribution of each of the endocannabinoid transmitters, anandamide and 2-arachidonoylglycerol (2-AG), to these behaviors remains undefined. To address this question, we assessed the effects of URB597, a selective anandamide deactivation inhibitor, as a reinforcer of drug-seeking and drug-taking behavior in squirrel monkeys.
Summary Monoacylglycerol lipase (MGL) is a serine hydrolase involved in the biological deactivation of the endocannabinoid 2-arachidonoyl-sn-glycerol (2-AG). Previous efforts to design MGL inhibitors have focused on chemical scaffolds that irreversibly block the activity of this enzyme. Here, we describe two naturally occurring terpenoids, pristimerin and euphol, which inhibit MGL activity with high potency (median effective concentration, IC50 = 93 nM and 315 nM, respectively) through a reversible mechanism. Mutational and modeling studies suggest that the two agents occupy a common hydrophobic pocket located within the putative lid domain of MGL, and each reversibly interact with one of two adjacent cysteine residues (Cys201 and Cys208) flanking such pocket. This previously unrecognized regulatory region may offer a novel molecular target for potent and reversible inhibitors of MGL.
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