Alzheimer's disease (AD) is a progressive and incurable neurodegenerative disorder. Early in the pathophysiology of AD, synaptic function is disrupted by soluble Aβ oligomers, possibly through Aβ-mediated internalization of NMDA receptors. Striatal-enriched phosphatase (STEP) is a tyrosine phosphatase that regulates the internalization of NMDA receptors. Recent work shows that STEP is elevated in the prefrontal cortex of human AD patients and in animal models of AD. Here, we use genetic manipulations to reduce STEP activity in a triple transgenic AD mouse model and show that a decrease in STEP levels reverses cognitive and cellular deficits observed in these mice. Our results suggest that STEP inhibitors may prove therapeutic for this devastating disorder.β amyloid | glutamate receptor trafficking | protein tyrosine phosphatase | long-term potentiation A lzheimer's disease (AD) is the most common form of dementia. Considerable evidence implicates β amyloid (Aβ) peptides in the pathophysiology of AD (1). Aβ 1-42 is derived from sequential cleavage of amyloid procursor protein (APP) by β-and γ-secretases. A recent hypothesis suggests that soluble Aβ oligomers disrupt synaptic and cognitive function early in the disease process (2-4). This model is supported by the findings that synaptic function is disrupted and synapses are lost early in the disorder and in animal models, even before amyloid plaques are detected (5-7). Application of soluble Aβ preparations results in synapse loss, blocks long-term potentiation (LTP), and impairs cognitive function in animals (8-11). Furthermore, animals that express high levels of Aβ show impaired synaptic plasticity and learning (6,12,13).Striatal-enriched phosphatase 61 (STEP 61 ; protein tyrosine phosphatase non-receptor 5 [PTPN5]) is a brain-specific tyrosine phosphatase targeted to synaptic compartments in striatum, hippocampus, cortex, and related brain regions (14-16). STEP 61 associates with the NMDA subclass of glutamate receptors, decreases NMDA receptor (NMDAR) activity, and opposes the induction of LTP through dephosphorylation of Y 1472 on the NR2B receptor subunit, leading to endocytosis of NR1/NR2B receptors (17, 18). Acute reduction in STEP expression by RNAi leads to increased surface expression and function of NR1/NR2B receptors (19), and STEP knockout (STEP −/− ) mice have enhanced hippocampal LTP (SI Appendix, Fig. S1). In addition, STEP 61 dephosphorylates a regulatory tyrosine within the activation loop of two enzymes critical for the development of synaptic strengthening, ERK1/2 and Fyn, thereby inactivating them (20,21). Together, these findings support the current model that STEP activity opposes the development of synaptic strengthening (22).Elevated levels of STEP 61 are found in several transgenic AD mouse models as well as human AD prefrontal cortex (Tg-2576 and J20) (23, 24). The increase in STEP 61 levels and activity contributes to the removal of NR1/NR2B complexes from synapses (24). Given that STEP 61 regulates Aβ-induced internalization of NR1/...

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Glutamatergic signaling through N-methyl-D-aspartate receptors (NMDARs) is required for synaptic plasticity. Disruptions in glutamatergic signaling are proposed to contribute to the behavioral and cognitive deficits observed in schizophrenia (SZ). One possible source of compromised glutamatergic function in SZ is decreased surface expression of GluN2B-containing NMDARs. STEP61 is a brain-enriched protein tyrosine phosphatase that dephosphorylates a regulatory tyrosine on GluN2B, thereby promoting its internalization. Here, we report that STEP61 levels are significantly higher in the postmortem anterior cingulate cortex and dorsolateral prefrontal cortex of SZ patients, as well as in mice treated with the psychotomimetics MK-801 and phencyclidine (PCP). Accumulation of STEP61 after MK-801 treatment is due to a disruption in the ubiquitin proteasome system that normally degrades STEP61. STEP knockout mice are less sensitive to both the locomotor and cognitive effects of acute and chronic administration of PCP, supporting the functional relevance of increased STEP61 levels in SZ. In addition, chronic treatment of mice with both typical and atypical antipsychotic medications results in a protein kinase A-mediated phosphorylation and inactivation of STEP61 and, consequently, increased surface expression of GluN1/GluN2B receptors. Taken together, our findings suggest that STEP61 accumulation may contribute to the pathophysiology of SZ. Moreover, we show a mechanistic link between neuroleptic treatment, STEP61 inactivation and increased surface expression of NMDARs, consistent with the glutamate hypothesis of SZ.

A major question for gene therapy in brain concerns methods to administer therapeutic genes in a uniform manner over major portions of the brain. A second question in neuroimmunology concerns the extent to which monocytes migrate to the CNS in degenerative disorders. Here we show that CD11bϩ cells (largely monocytes) isolated from the bone marrow of GFP (green fluorescent protein)-expressing donors spontaneously home to compacted amyloid plaques in the brain. Injections of these cells as a single pulse show a rapid clearance from circulation (90 min half-life) and tissue residence half-lives of ϳ3 d. The uptake into brain was minimal in nontransgenic mice. In transgenic mice containing amyloid deposits, uptake was dramatically increased and associated with a corresponding decrease in monocyte uptake into peripheral organs comparedtonontransgeniclittermates.TwiceweeklyinfusionsoftheCD11bϩbonemarrowcellstransfectedwithageneticallyengineeredform of the protease neprilysin completely arrest amyloid deposition in an aggressively depositing transgenic model. Exploiting the natural homing properties of peripherally derived blood cells to deliver therapeutic genes has the advantages of access to the entire CNS, expression largely restricted to sites of injury, low risk of immune reactivity, and fading of expression if adverse reactions are encountered. These observations support the feasibility of testing autologous monocytes for application of therapeutic genes in human CNS disease. Moreover, these data support the results from bone marrow grafts that circulating CD11bϩ cells can enter the CNS without requiring the use of lethal irradiation.

Huntington’s disease (HD) is an autosomal dominant neurodegenerative disorder caused by a CAG trinucleotide repeat expansion in the huntingtin gene. Major pathological hallmarks of HD include inclusions of mutant huntingtin (mHTT) protein, loss of neurons predominantly in the caudate nucleus, and atrophy of multiple brain regions. However, the early sequence of histological events that manifest in region- and cell-specific manner has not been well characterized. Here we use a high-content histological approach to precisely monitor changes in HTT expression and characterize deposition dynamics of mHTT protein inclusion bodies in the recently characterized zQ175 knock-in mouse line. We carried out an automated multi-parameter quantitative analysis of individual cortical and striatal cells in tissue slices from mice aged 2–12 months and confirmed biochemical reports of an age-associated increase in mHTT inclusions in this model. We also found distinct regional and subregional dynamics for inclusion number, size and distribution with subcellular resolution. We used viral-mediated suppression of total HTT in the striatum of zQ175 mice as an example of a therapeutically-relevant but heterogeneously transducing strategy to demonstrate successful application of this platform to quantitatively assess target engagement and outcome on a cellular basis.

The use of recombinant adeno-associated viral (rAAV) vectors as a means of gene delivery to the central nervous system has emerged as a potentially viable method for the treatment of several types of degenerative brain diseases. However, a limitation of typical intracranial injections into the adult brain parenchyma is the relatively restricted distribution of the delivered gene to large brain regions such as the cortex, presumably due to confined dispersion of the injected particles. Optimizing the administration techniques to maximize gene distribution and gene expression is an important step in developing gene therapy studies. Here, we have found additive increases in distribution when 3 methods to increase brain distribution of rAAV were combined. The convection enhanced delivery (CED) method with the step-design cannula was used to deliver rAAV vector serotypes 5, 8 and 9 encoding GFP into the hippocampus of the mouse brain. While the CED method improved distribution of all 3 serotypes, the combination of rAAV9 and CED was particularly effective. Systemic mannitol administration, which reduces intracranial pressure, also further expanded distribution of GFP expression, in particular, increased expression on the contralateral hippocampi. These data suggest that combining advanced injection techniques with newer rAAV serotypes greatly improves viral vector distribution, which could have significant benefits for implementation of gene therapy strategies.

The brain-specific tyrosine phosphatase, STEP (STriatal-Enriched protein tyrosine Phosphatase) is an important regulator of synaptic function. STEP normally opposes synaptic strengthening by increasing N-methyl D-aspartate glutamate receptors (NMDARs) internalization through dephosphorylation of GluN2B and inactivation of the kinases ERK1/2 and Fyn. Here we show that STEP61 is elevated in the cortex in the Nrg1+/− knockout mouse model of SZ. Genetic reduction or pharmacological inhibition of STEP prevents the loss of NMDA receptors from synaptic membranes and reverses behavioral deficits in Nrg1+/− mice. STEP61 protein is also increased in cortical lysates from the CNS-specific ErbB2/4 mouse model of SZ, as well as in human induced pluripotent stem cell (hiPSC)-derived forebrain neurons and Ngn2-induced excitatory neurons from two independent SZ patient cohorts. In these selected SZ models, increased STEP61 protein levels likely reflect reduced ubiquitination and degradation. These convergent findings from mouse and hiPSC SZ models provide evidence for STEP61 dysfunction in SZ.

Reduction of Aβ deposition is a major therapeutic strategy in Alzheimer's disease (AD). The concentration of Aβ in the brain is modulated, not only by Aβ production, but also by its degradation. One protease involved in the degradation of Aβ peptides is endothelin converting enzyme (ECE). In the current study, we investigated the effects of an intracranial administration of a recombinant adeno-associated viral vector (rAAV) containing the ECE-1 gene on amyloid deposition in amyloid precursor protein (APP) plus presenilin-1 (PS1) transgenic mice. The recombinant AAV vector was injected unilaterally into the right anterior cortex and hippocampus of six-month-old mice while control mice received an AAV vector expressing GFP. Immunohistochemistry for the haemagglutinin tag appended to ECE revealed strong expression in areas surrounding the injection sites but minimal expression in the contralateralregions. Immunohistochemistry for Aß decreased in the anterior cortex and hippocampus of mice receiving ECE cDNA. Further, decreases in Congo red positive deposits were also observed in both regions. These results indicate that increasing the expression of β-amyloid degrading enzymes through gene therapy is a promising therapeutic avenue through which to treat AD.