Glutamate levels control HT22 murine hippocampal cell death by regulating biphasic patterns of Erk1/2 activation: role of metabolic glutamate receptor 5

Extracellular glutamate concentration is a critical determinant of neuronal cell fate. We recently demonstrated that HT22 murine hippocampal cell viability was reduced by exposure to high concentrations of glutamate, whereas low concentrations promoted cell survival. Extracellular signal-regulated kinase (Erk)1/2 activation by glutamate is important for both glutamate-induced cell death and survival. In this study, we investigated the role of glutamate-induced or hydrogen peroxide (H₂O₂)-induced Erk1/2 activation in HT22 cell fate determination. Glutamate and H₂O₂ treatment similarly induced early (<1 h) Erk1/2 phosphorylation regardless of concentration. On the other hand, persistent Erk1/2 phosphorylation (16–24 h) was observed only in the presence of excess glutamate. Only the latter contributed to glutamate-induced cell death, which involved metabolic glutamate receptor 5. Our findings suggest that glutamate concentration modulates two distinct phases of Erk1/2 activation, which can explain the glutamate concentration-dependent determination of HT22 cell fate.     

Growth and nodulation in <Emphasis Type="Italic">Alnus sieboldiana</Emphasis> in response to <Emphasis Type="Italic">Frankia</Emphasis> inoculation and nitrogen treatments

Key message

We investigated a Frankia – Alnus sieboldiana symbiosis, including the minimum inoculum dose for constant nodulation, the period of time to nodulation after inoculation, and the effects of N on nodulation.

Abstract

Frankia is a nitrogen-fixing actinomycete that forms root nodules in some dicotyledonous plants, which are called actinorhizal. We studied nodule formation in Alnus sieboldiana, an actinorhizal plant, after inoculation with a Frankia isolate to establish techniques for Frankia inoculation and the cultivation of inoculated plants. Root nodules formed on seedlings of A. sieboldiana by 2 weeks after inoculation, and N₂ fixation measured by acetylene reduction activity started 3 weeks after inoculation. Nodulation was observed after inoculation with a Frankia isolate at 0.001 μL packed cell volume (pcv). The number of nodules formed on the seedlings inoculated with Frankia at more than 0.05 μL pcv was not significantly different. Nodule development and N₂ fixation were reduced when inoculated seedlings were treated weekly with 15 mM NH₄NO₃-N. In contrast, treatment with 3.75 or 0.9375 mM NH₄NO₃-N did not inhibit nodule development or N₂ fixation of inoculated seedlings by 15 weeks of N treatment.

     

Specific role of the truncated betaIV-spectrin Sigma6 in sodium channel clustering at axon initial segments and nodes of ranvier

At axon initial segments and nodes of Ranvier in neurons, the spectrin membrane skeleton plays roles in physically stabilizing the plasma membrane integrity and in clustering voltage-gated sodium channels for proper conduction of the action potential. betaIV-Spectrin, an essential component of the membrane skeleton at these sites, has an N-terminal-truncated isoform, Sigma6, which is expressed at much higher levels than the full-length isoform Sigma1. To investigate the role of betaIV-spectrin Sigma6, we generated Sigma1-deficient mice with a normal level of Sigma6 expression (Sigma1(-/-) mice), and compared their phenotypes with those of previously generated mice lacking both Sigma1 and Sigma6(Sigma1Sigma6(-/-) mice). The gross neurological defects observed in Sigma1Sigma6(-/-) mice, such as hindleg contraction, were apparently ameliorated in Sigma1(-/-) mice. At cellular levels, Sigma1Sigma6(-/-) and Sigma1(-/-) neurons similarly exhibited waving and swelling of the plasma membrane at axon initial segments and nodes of Ranvier. By contrast, the levels of ankyrin G and voltage-gated sodium channels at these sites, which are significantly reduced in Sigma1Sigma6(-/-) mice, were substantially recovered in Sigma1(-/-) mice. We conclude that the truncated betaIV-spectrin isoform Sigma6 plays a specific role in clustering voltage-gated sodium channels, whereas it is dispensable for membrane stabilization at axon initial segments and nodes of Ranvier.     

Quantitative proteomic analysis of rat liver for carcinogenicity prediction in a 28-day repeated dose study

The potential of quantitative proteomic analysis to predict carcinogenicity of chemical compounds was investigated. Using 2D-DIGE, we analyzed the effects of 63 chemical compounds on protein expression in the rat liver after 28 daily doses. Types of carcinogens were categorized depending on the species and organ specificity. The carcinogen characteristic proteins for each classification were identified by Welch's t value. For evaluation of the predictive concordance we used support vector machines. The rat hepatic carcinogen-specific classification gave higher concordance than the other classification. The generalization performance was measured by leave-one-out cross-validation. For genotoxic and non-genotoxic compounds, a concordance of 79.3 and 76.5%, respectively, was obtained by the top 30 ranked proteins with Welch's t value. Furthermore, we found that the increase of the expression level of the stress response proteins as the common feature of poorly predicted chemical compounds in the leave-20%-out cross-validation. Quantitative proteomics could be promising technique for developing biomarker panels that can be used for carcinogenicity prediction. The list of proteins identified in this study and the zoomed gel images of the top ranked proteins in statistic analysis are provided in Supplementary Data.