Chemogenetic Breakdown of the Dentate Gate Causes Seizures and Spatial Memory Deficits.

The dentate gyrus has often been posited to act as a gate that dampens highly active afferent input into the hippocampus. Effective gating is thought to prevent seizure initiation and propagation in the hippocampus and support learning and memory processes. Pathological changes to DG circuitry that occur in temporal lobe epilepsy (TLE) can increase DG excitability and impair its gating ability which can contribute to seizures and cognitive deficits.

Circuit-based interventions in the dentate gyrus rescue epilepsy-associated cognitive dysfunction.

Temporal lobe epilepsy is associated with significant structural pathology in the hippocampus. In the dentate gyrus, the summative effect of these pathologies is massive hyperexcitability in the granule cells, generating both increased seizure susceptibility and cognitive deficits. To date, therapeutic approaches have failed to improve the cognitive symptoms in fully developed, chronic epilepsy.

Altered NMDAR signaling underlies autistic-like features in mouse models of CDKL5 deficiency disorder.

CDKL5 deficiency disorder (CDD) is characterized by epilepsy, intellectual disability, and autistic features, and CDKL5-deficient mice exhibit a constellation of behavioral phenotypes reminiscent of the human disorder. We previously found that CDKL5 dysfunction in forebrain glutamatergic neurons results in deficits in learning and memory. However, the pathogenic origin of the autistic features of CDD remains unknown.

In Situ Hybridization for Detection of AAV-Mediated Gene Expression.

Techniques to localize vector transgenes in cells and tissues are essential in order to fully characterize gene therapy outcomes. In situ hybridization (ISH) uses synthesized complementary RNA or DNA nucleotide probes to localize and detect sequences of interest in fixed cells, tissue sections, or whole tissue mounts. Variations in techniques include adding labels to probes, such as fluorophores, which can allow for the simultaneous visualization of multiple targets.

Loss of CDKL5 in Glutamatergic Neurons Disrupts Hippocampal Microcircuitry and Leads to Memory Impairment in Mice.

Cyclin-dependent kinase-like 5 (CDKL5) deficiency is a neurodevelopmental disorder characterized by epileptic seizures, severe intellectual disability, and autistic features. Mice lacking CDKL5 display multiple behavioral abnormalities reminiscent of the disorder, but the cellular origins of these phenotypes remain unclear. Here, we find that ablating CDKL5 expression specifically from forebrain glutamatergic neurons impairs hippocampal-dependent memory in male conditional knock-out mice.

Massively augmented hippocampal dentate granule cell activation accompanies epilepsy development.

In a mouse model of temporal lobe epilepsy, multicellular calcium imaging revealed that disease emergence was accompanied by massive amplification in the normally sparse, afferent stimulation-induced activation of hippocampal dentate granule cells. Patch recordings demonstrated reductions in local inhibitory function within the dentate gyrus at time points where sparse activation was compromised. Mimicking changes in inhibitory synaptic function and transmembrane chloride regulation was sufficient to elicit the dentate gyrus circuit collapse evident during epilepsy development.

Normal and epilepsy-associated pathologic function of the dentate gyrus.

The dentate gyrus plays critical roles both in cognitive processing, and in regulation of the induction and propagation of pathological activity. The cellular and circuit mechanisms underlying these diverse functions overlap extensively. At the cellular level, the intrinsic properties of dentate granule cells combine to endow these neurons with a fundamental reluctance to activate, one of their hallmark traits.

Integrated analysis of proteome and transcriptome changes in the mucopolysaccharidosis type VII mouse hippocampus.

Mucopolysaccharidosis type VII (MPS VII) is a lysosomal storage disease caused by the deficiency of β-glucuronidase. In this study, we compared the changes relative to normal littermates in the proteome and transcriptome of the hippocampus in the C57Bl/6 mouse model of MPS VII, which has well-documented histopathological and neurodegenerative changes. A completely different set of significant changes between normal and MPS VII littermates were found in each assay.

Controlling AAV Tropism in the Nervous System with Natural and Engineered Capsids.

More than one hundred naturally occurring variants of adeno-associated virus (AAV) have been identified, and this library has been further expanded by an array of techniques for modification of the viral capsid. AAV capsid variants possess unique antigenic profiles and demonstrate distinct cellular tropisms driven by differences in receptor binding. AAV capsids can be chemically modified to alter tropism, can be produced as hybrid vectors that combine the properties of multiple serotypes, and can carry peptide insertions that introduce novel receptor-binding activity.

Clinical Improvement of Alpha-mannosidosis Cat Following a Single Cisterna Magna Infusion of AAV1.

Lysosomal storage diseases (LSDs) are debilitating neurometabolic disorders for most of which long-term effective therapies have not been developed. Gene therapy is a potential treatment but a critical barrier to treating the brain is the need for global correction. We tested the efficacy of cisterna magna infusion of adeno-associated virus type 1 (AAV1) expressing feline alpha-mannosidase gene in the postsymptomatic alpha-mannosidosis (AMD) cat, a homologue of the human disease.

Ex vivo gene therapy using patient iPSC-derived NSCs reverses pathology in the brain of a homologous mouse model.

Neural stem cell (NSC) transplantation is a promising strategy for delivering therapeutic proteins in the brain. We evaluated a complete process of ex vivo gene therapy using human induced pluripotent stem cell (iPSC)-derived NSC transplants in a well-characterized mouse model of a human lysosomal storage disease, Sly disease. Human Sly disease fibroblasts were reprogrammed into iPSCs, differentiated into a stable and expandable population of NSCs, genetically corrected with a transposon vector, and assessed for engraftment in NOD/SCID mice.

Adeno-associated virus serotypes 1, 8, and 9 share conserved mechanisms for anterograde and retrograde axonal transport.

Adeno-associated virus (AAV) vectors often undergo long-distance axonal transport after brain injection. This leads to transduction of brain regions distal to the injection site, although the extent of axonal transport and distal transduction varies widely among AAV serotypes. The mechanisms driving this variability are poorly understood.

High-resolution magnetic resonance microscopy and diffusion tensor imaging to assess brain structural abnormalities in the murine mucopolysaccharidosis VII model.

High-resolution microscopic magnetic resonance imaging (μMRI) and diffusion tensor imaging (DTI) were performed to characterize brain structural abnormalities in a mouse model of mucopolysaccharidosis type VII (MPS VII). Microscopic magnetic resonance imaging demonstrated a decrease in the volume of anterior commissure and corpus callosum and a slight increase in the volume of the hippocampus in MPS VII versus wild-type mice. Diffusion tensor imaging indices were analyzed in gray and white matter.

Long-distance axonal transport of AAV9 is driven by dynein and kinesin-2 and is trafficked in a highly motile Rab7-positive compartment.

Adeno-associated virus (AAV) vectors can move along axonal pathways after brain injection, resulting in transduction of distal brain regions. This can enhance the spread of therapeutic gene transfer and improve treatment of neurogenetic disorders that require global correction. To better understand the underlying cellular mechanisms that drive AAV trafficking in neurons, we investigated the axonal transport of dye-conjugated AAV9, utilizing microfluidic primary neuron cultures that isolate cell bodies from axon termini and permit independent analysis of retrograde and anterograde axonal transport.

Engraftment of nonintegrating neural stem cells differentially perturbs cortical activity in a dose-dependent manner.

Neural stem cell (NSC) therapy represents a potentially powerful approach for gene transfer in the diseased central nervous system. However, transplanted primary, embryonic stem cell- and induced pluripotent stem cell-derived NSCs generate largely undifferentiated progeny. Understanding how physiologically immature cells influence host activity is critical to evaluating the therapeutic utility of NSCs.

Transplantation of CD15-enriched murine neural stem cells increases total engraftment and shifts differentiation toward the oligodendrocyte lineage.

Neural stem cell (NSC) transplantation is a promising therapeutic approach for neurological diseases. However, only a limited number of cells can be transplanted into the brain, resulting in relatively low levels of engraftment. This study investigated the potential of using a cell surface marker to enrich a primary NSC population to increase stable engraftment in the recipient brain.

Dysregulation of gene expression in a lysosomal storage disease varies between brain regions implicating unexpected mechanisms of neuropathology.

The characteristic neurological feature of many neurogenetic diseases is intellectual disability. Although specific neuropathological features have been described, the mechanisms by which specific gene defects lead to cognitive impairment remain obscure. To gain insight into abnormal functions occurring secondary to a single gene defect, whole transcriptome analysis was used to identify molecular and cellular pathways that are dysregulated in the brain in a mouse model of a lysosomal storage disorder (LSD) (mucopolysaccharidosis [MPS] VII).

Formation and function of the lytic NK-cell immunological synapse.

The natural killer (NK)-cell immunological synapse is the dynamic interface formed between an NK cell and its target cell. Formation of the NK-cell immunological synapse involves several distinct stages, from the initiation of contact with a target cell to the directed delivery of lytic-granule contents for target-cell lysis. Progression through the individual stages is regulated, and this tight regulation underlies the precision with which NK cells select and kill susceptible target cells (including virally infected cells and cancerous cells) that they encounter during their routine surveillance of the body.

Selective serotonin reuptake inhibitor and substance P antagonist enhancement of natural killer cell innate immunity in human immunodeficiency virus/acquired immunodeficiency syndrome.

Background: Natural killer (NK) cells play an important role in innate immunity and are involved in the host defense against human immunodeficiency virus (HIV) infection. This study examines the potential role of three underlying regulatory systems that have been under investigation in central nervous system research as well as immune and viral research: serotonin, neurokinin, and glucocorticoid systems.

Methods: Fifty-one HIV-seropositive subjects were recruited to achieve a representative sample of depressed and nondepressed women.

Selective roles for alpha-PKC in positive signaling for O-(2) generation and calcium mobilization but not elastase release in differentiated HL60 cells.

Protein kinase C (PKC) isotypes and Ca2+ mobilization have been implicated in phagocytic cell functions such as O(-)(2) generation. Ca/DG-dependent alpha-PKC and beta-PKC have similar substrate specificities and cofactor requirements in vitro. However it is not known if these isotypes play redundant or unique roles in the intact cell.

Natural killer cell inhibits human immunodeficiency virus replication in chronically infected immune cells.

Natural killer (NK) cells are a crucial component of the host innate immune system. We investigated the noncytolytic anti-human immunodeficiency virus (HIV) activity of NK cells in chronically HIV-infected immune cells. Supernatants collected from NK cell cultures (both primary NK cells and NK cell lines, YTS and NK 92) inhibited HIV activation in peripheral blood mononuclear cells (PBMCs) from HIV-infected subjects.

A non-peptide substance P antagonist (CP-96,345) inhibits morphine-induced NF-kappa B promoter activation in human NT2-N neurons.

Opioids and the neuropeptide substance P (SP) modulate the expression of inflammatory cytokines and chemokines, which are under the control of nuclear factor kappaB (NF-kappaB). We investigated whether the neurokinin-1 receptor (SP receptor) pathway is biologically involved in morphine-mediated modulation of NF-kappaB promoter activation in a human neuronal cell line (NT2-N) that expresses both the mu-opioid receptor (MOR) and the SP receptor. Morphine significantly enhanced NF-kappaB promoter-directed luciferase activity in NT2-N neurons.

Interleukin-1beta stimulates macrophage inflammatory protein-1alpha and -1beta expression in human neuronal cells (NT2-N).

Chemokines are important mediators in immune responses and inflammatory processes of neuroimmunologic and infectious diseases. Although chemokines are expressed predominantly by cells of the immune system, neurons also express chemokines and chemokine receptors. We report herein that human neuronal cells (NT2-N) produce macrophage inflammatory protein-1alpha and -1beta (MIP-1alpha and MIP-1beta), which could be enhanced by interleukin (IL)-1beta at both mRNA and protein levels.

Progesterone withdrawal reduces paired-pulse inhibition in rat hippocampus: dependence on GABA(A) receptor alpha4 subunit upregulation.

Withdrawal from the endogenous steroid progesterone (P) after chronic administration increases anxiety and seizure susceptibility via declining levels of its potent GABA-modulatory metabolite 3alpha-OH-5alpha-pregnan-20-one (3alpha,5alphaTHP). This 3alpha,5alpha-THP withdrawal also results in a decreased decay time constant for GABA-gated current assessed using whole cell patch-clamp techniques on pyramidal cells acutely dissociated from CA1 hippocampus. The purpose of this study was to test the hypothesis that the decreases in total integrated GABA-gated current observed at the level of the isolated pyramidal cell would be manifested as a reduced GABA inhibition at the circuit level following hormone withdrawal.

Substance P up-regulates macrophage inflammatory protein-1beta expression in human T lymphocytes.

Substance P (SP) is an important modulator of neuroimmunoregulation. We have demonstrated that human T lymphocytes express SP and neurokinin-1 receptor (NK-1R), a primary SP receptor. In the present study, we investigated whether SP stimulates synthesis of macrophage inflammatory protein-1beta (MIP-1beta) in human T lymphocytes.