Brain Tools (United States)

Previously we showed that the vitamin A metabolite all-trans retinoic acid (atRA) induces synaptic plasticity in acute brain slices prepared from the mouse and human neocortex (Lenz et al., 2021). Depending on the brain region studied, distinct effects of atRA on excitatory and inhibitory neurotransmission have been reported. Here, we used intraperitoneal injections of atRA (10 mg/kg) in adult C57BL/6J mice to study the effects of atRA on excitatory and inhibitory neurotransmission in the mouse fascia dentata-a brain region implicated in memory acquisition. No major changes in synaptic transmission were observed in the ventral hippocampus while a significant increase in both spontaneous excitatory postsynaptic current frequencies and synapse numbers were evident in the dorsal hippocampus 6 hr after atRA administration. The intrinsic properties of hippocampal dentate granule cells were not significantly different and hippocampal transcriptome analysis revealed no essential neuronal changes upon atRA treatment. In light of these findings, we tested for the metaplastic effects of atRA, that is, for its ability to modulate synaptic plasticity expression in the absence of major changes in baseline synaptic strength. Indeed, in vivo long-term potentiation (LTP) experiments demonstrated that systemic atRA treatment improves the ability of dentate granule cells to express LTP. The plasticity-promoting effects of atRA were not observed in synaptopodin-deficient mice, therefore, extending our previous results regarding the relevance of synaptopodin in atRA-mediated synaptic strengthening in the mouse prefrontal cortex. Taken together, our data show that atRA mediates synaptopodin-dependent metaplasticity in mouse dentate granule cells.

Therapeutic modalities designed specifically to inhibit COVID-19 infection and replication would limit progressive COVID-19-associated pulmonary disease in infected patients and prevent or limit systemic disease. If effective, antivirals could reduce viral transmission rates by reducing viral burden and allow time for immune clearance. For individuals infected with acute-stage disease, antivirals in support of the existing vaccines could reduce COVID-19 hospitalizations and deaths. Here, we evaluate MRCV-19, a phosphorodiamidate morpholino oligo with delivery dendrimer (Vivo-Morpholino), to prevent coronavirus infection in a cell culture model. This is a novel antiviral that effectively inhibits SARS-CoV-2 replication in vitro. By design, MRCV-19 targets the SARS-CoV-2 5’UTR and overlaps the pp1a start site of translation in order to block access of the translation initiation complex to the start. MRCV-19 testing is conducted in a high-throughput, 384-well plate format with a 10-point dose-response curve (common ratio of 2) assayed in duplicate with parallel cytotoxicity evaluations. MRCV-19 was shown to be more effective than hydroxychloroquine and remdesivir in our CPE reduction assay with low toxicity. The clinical translational impact of this study is providing the basis for evaluating MRCV-19 on a large scale in an appropriate infection model for toxicity and systemic high-level inhibition of SARS-CoV-2, which could lead in time to phase I testing in humans.

Toxoplasma gondii, an Apicomplexan parasite, causes significant morbidity and mortality, including severe disease in immunocompromised hosts and devastating congenital disease, with no effective treatment for the bradyzoite stage. To address this, we used the Tropical Disease Research database, crystallography, molecular modeling, and antisense to identify and characterize a range of potential therapeutic targets for toxoplasmosis. Phosphoglycerate mutase II (PGMII), nucleoside diphosphate kinase (NDK), ribulose phosphate 3-epimerase (RPE), ribose-5-phosphate isomerase (RPI), and ornithine aminotransferase (OAT) were structurally characterized. Crystallography revealed insights into the overall structure, protein oligomeric states and molecular details of active sites important for ligand recognition. Literature and molecular modeling suggested potential inhibitors and druggability. The targets were further studied with vivoPMO to interrupt enzyme synthesis, identifying the targets as potentially important to parasitic replication and, therefore, of therapeutic interest. Targeted vivoPMO resulted in statistically significant perturbation of parasite replication without concomitant host cell toxicity, consistent with a previous CRISPR/Cas9 screen showing PGM, RPE, and RPI contribute to parasite fitness. PGM, RPE, and RPI have the greatest promise for affecting replication in tachyzoites. These targets are shared between other medically important parasites and may have wider therapeutic potential.

Although the molecular basis of many inherited metabolic diseases has been defined, the availability of effective therapies in such disorders remains problematic. Menkes disease is a fatal neurodegenerative disorder due to loss-of-function mutations in the ATP7A gene encoding a copper-transporting P-type Atpase. To develop therapeutic approaches in affected patients, we have identified a zebrafish model of Menkes disease termed calamity that results from splicing defects in the zebrafish orthologue of the ATP7A gene. Embryonic-recessive lethal mutants have impaired copper homeostasis that results in absent melanin pigmentation, impaired notochord formation, and hindbrain neurodegeneration. In this current study, we have attempted to rescue these striking phenotypic alterations by using a series of antisense morpholino oligonucleotides directed against the splice-site junctions of two mutant calamity alleles. Our findings reveal a robust and complete correction of the copper-deficient defects of calamity in association with the generation of the WT Menkes protein in all rescued mutants. Interestingly, a quantitative analysis of atp7a-specific transcripts suggests that competitive translational regulation may account for the synthesis of WT protein in these embryos. This in vivo correction of Menkes disease through the rescue of aberrant splicing may provide therapeutic options in this fatal disease and illustrates the potential for zebrafish models of human genetic disease in the development of treatments based on the principles of interactions of synthetic oligonucleotide analogues with mRNA.

Efforts to understand and model the dynamics of the upper ocean would be significantly advanced given the ability to rapidly determine mixed layer depths (MLDs) over large regions. Remote sensing technologies are an ideal choice for achieving this goal. This study addresses the feasibility of estimating MLDs from optical properties. These properties are strongly influenced by suspended particle concentrations, which generally reach a maximum at pycnoclines. The premise therefore is to use a gradient in beam attenuation at 660 nm (c660) as a proxy for the depth of a particle‐scattering layer. Using a global data set collected during World Ocean Circulation Experiment cruises from 1988–1997, six algorithms were employed to compute MLDs from either density or temperature profiles. Given the absence of published optically based MLD algorithms, two new methods were developed that use c660 profiles to estimate the MLD. Intercomparison of the six hydrographically based algorithms revealed some significant disparities among the resulting MLD values. Comparisons between the hydrographical and optical approaches indicated a first‐order agreement between the MLDs based on the depths of gradient maxima for density and c660. When comparing various hydrographically based algorithms, other investigators reported that inherent fluctuations of the mixed layer depth limit the accuracy of its determination to 20 m. Using this benchmark, we found a ∼70% agreement between the best hydrographical‐optical algorithm pairings.

This article asks and addresses thequestion ``what is `optimal' maintenance inirrigation.'' It examines alternativedefinitions of irrigation maintenance andexplores pathways of system deteriorationunder alternative input levels. It thendefines maintenance as a service, where thefunctions of arranging, providing, payingfor, and using the service are considered,and discusses the arrangements andrelationships necessary to provide thisservice. Key to this approach is theconcept of maintaining a specified level ofservice – a concept drawn from the assetmanagement approach to systems maintenance. A contingency matrix is developed whichdefines basic maintenance strategies underdifferent levels of cost and risk. Thepaper concludes that optimality ofirrigation system maintenance is somethingwhich cannot be defined on technicalgrounds, or economic ones, or even oninstitutional grounds alone. Rather it mustbe defined as a process operating within aset of framework conditions and technical,economic and institutional constraints.