Biology subjects

Kubota, K.

Publications and source records attributed to Kubota, K..

8 recordsLinked to original sources

bioRxiv2024

Mitigating nitrous oxide emission by an ultra-fast bioprocess enabling the removal of high concentration N2O

Nitrous oxide (N2O) is known as a greenhouse gas as well as an ozone-depleting substance. Wastewater treatment process is one of the sources of N2O emission, and the high concentrations of N2O in off-gas were reported from an anaerobic ammonium oxidation process. This study developed a novel N2O removal process using a down-flow hanging sponge reactor to remove high concentrations of N2O. More than 96% removal efficiencies were achieved for up to 300 ppm N2O with 3 min gas retention time (GRT), and more than 99% removal efficiency was obtained for 2,000 ppm N2O with 18 min GRT. A maximum removal rate of 161 {+/-} 26 mg-N/L-reactor/day was achieved, that was over 10 times faster than the pioneering process. Kinetic analysis indicated that the N2O dissolution rate is a crucial factor in determining the N2O removal rate in the reactor. Various N2O reducers belonging to both clade I and II were detected in reactors, and Azonexus was thought to play a key role.

microbiology↗

bioRxiv2024

5-Deazaflavin (TND1128) and its hybrid analogs are cytoprotective against hydrogen peroxide (H2O2)-induced oxidative stress.

Increased production of reactive oxygen species (ROS) and oxidative stress are implicated in mitochondrial dysfunction, contributing to the pathogenesis of many neurodegenerative diseases. Research is ongoing into a new treatment approach for neurodegeneration, focusing on reactivating dysfunctional mitochondria. Some 5-deazaflavins, such as 10-ethyl-3-methylpyrimido[4,5-b]quinoline-2,4(3H,10H)-dione (TND1128), and four analogs of 5-deazaflavin, including {beta}-nicotinamide mononucleotide ({beta}-NMN), demonstrate efficient self-redox abilities similar to {beta}-NMN, making them potential activators of mitochondrial energy synthesis. This study examines whether TND1128 and its analogs have protective effects against cellular impairment induced by oxidative stress. These compounds exhibit proliferative potential against normal cells. Moreover, TND1128 and its analogs significantly improved cell viability against hydrogen peroxide (H2O2)-induced oxidative stress injury. Our study confirms the cytoprotective effect of these 5-deazaflavins through mitochondrial activation. We anticipate TND1128 and its analogs will serve as mitochondria-stimulating drugs capable of rescuing deteriorating neurons in aging or diseases.

pharmacology and toxicology↗

bioRxiv2023

Location analysis of presynaptically active and silent synapses in single-cultured hippocampal neurons

A morphologically present but non-functioning synapse is termed a silent synapse. Silent synapses are categorized into "postsynaptically silent synapses," where AMPA receptors are either absent or non-functional, and "presynaptically silent synapses," where neurotransmitters cannot be released from nerve terminals. The presence of presynaptically silent synapses remains enigmatic, and their physiological significance is highly intriguing. In this study, we examined the distribution and developmental changes of presynaptically active and silent synapses in individual neurons. Our findings show a gradual increase in the number of excitatory synapses, along with a corresponding decrease in the percentage of presynaptically silent synapses during neuronal development. To pinpoint the distribution of presynaptically active and silent synapses, i.e., their positional information, we enhanced the traditional Sholl analysis and introduced a novel method termed "donut analysis." Our results indicate that the distribution of presynaptically silent synapses within a single neuron does not exhibit a distinct pattern during synapse development in different areas. However, irrespective of neuronal development, the proportion of presynaptically silent synapses tends to rise as the projection site moves farther from the cell body, suggesting that synapses near the cell body may exhibit higher synaptic transmission efficiency. This study represents the first observation of changes in the distribution of presynaptically active and silent synapses within a single neuron. Additionally, we propose that donut analysis can serve as a valuable analytical tool for evaluating synaptic positional information. Scope statementA morphologically present but non-functioning synapse is termed a silent synapse. The presence of presynaptically silent synapses remains enigmatic, and their physiological significance is highly intriguing. This study focused on the distribution and developmental changes of presynaptically active and silent synapses in individual neurons. To pinpoint the distribution of presynaptically active and silent synapses, we enhanced the traditional Sholl analysis and introduced a novel method termed "donut analysis." We found that the distribution of presynaptically silent synapses within a single neuron does not exhibit a distinct pattern during synapse development in different areas. However, irrespective of neuronal development, the proportion of presynaptically silent synapses tends to rise as the projection site moves farther from the cell body. This is a new paper that applies "Sholl analysis," a method invented 70 years ago that is now the gold standard of morphological analysis of the single neuron. This study represents the first observation of changes in the distribution of presynaptically active and silent synapses within a single neuron. Additionally, we propose that donut analysis can serve as a valuable analytical tool for evaluating synaptic positional information for the design of "synaptic maps" in neural circuits.

neuroscience↗

bioRxiv2023

Microscopic and metatranscriptomic analyses revealed unique cross-domain symbiosis between Candidatus Patescibacteria/candidate phyla radiation (CPR) and methanogenic archaea in anaerobic ecosystems

To verify the parasitic lifestyle of Candidatus Patescibacteria in the enrichment cultures derived from a methanogenic bioreactor, we applied multifaceted approaches combining cultivation, microscopy, metatranscriptomic, and protein structure prediction analyses. Cultivation experiments with the addition of exogenous methanogenic archaea with acetate, amino acids, and nucleoside monophosphates and 16S rRNA gene sequencing confirmed the increase in the relative abundance of Ca. Patescibacteria and methanogens. The predominant Ca. Patescibacteria were Ca. Yanofskybacteria and 32-520 lineages (to which belongs to class Ca. Paceibacteria) and positive linear relationships (r2 [≥] 0.70) between the relative abundance of Ca. Yanofskybacteria and Methanothrix, suggesting that the tendency of the growth rate is similar to that of the host. By fluorescence in situ hybridization (FISH) observations, the FISH signals of Methanothrix and Methanospirillum cells with Ca. Yanofskybacteria and with 32-520 lineages, respectively, were significantly lower than those of the methanogens without Ca. Patescibacteria, suggesting their parasitic interaction. The TEM and SEM observations also support parasitism in that the cell walls and plugs of these methanogens associated with submicron cells were often deformed. In particular, some Methanothrix-like filamentous cells were dented where the submicron cells were attached. Metatranscriptomic and protein structure prediction analyses identified highly expressed secreted genes from the genomes of Ca. Yanofskybacteria and 32-520, and these genes contain adhesion-related domains to the host cells. Considering the results through the combination of microscopic observations, gene expression, and computational protein modeling, we propose that the interactions between Ca. Yanofskybacteria and 32-520 belonging to class Ca. Paceibacteria and methanogenic archaea are parasitism.

microbiology↗

bioRxiv2022

Symbiosis between Patescibacteria and Archaea discovered in wastewater-treating bioreactors

Each prokaryotic domain, Bacteria and Archaea, contains a large and diverse group of organisms characterized with ultrasmall cell size and symbiotic lifestyles - Patescibacteria (also known as Candidate Phyla Radiation/CPR) and DPANN archaea. Cultivation-based approaches have revealed that Patesibacteria and DPANN symbiotically interact with bacterial and archaeal partners/hosts respectively, but cross-domain symbiosis/parasitism has never been observed. Here, we discovered physical interaction between ultramicrobacterial Patescibacteria and methanogenic archaea using cultures from anaerobic wastewater treatment sludge. In the cultures, we observed physical attachment of ultramicrobial cells to cells resembling Methanothrix and Methanospirillum using transmission electron microscopy and successfully detected physical association of Ca. Yanofskybacteria and Methanothrix using fluorescence in situ hybridization (FISH) (other ultramicrosized bacterial cells, presumably Patescibacteria, were also observed to attach on Methanospirillum). This was further confirmed to be a symbiosis rather than simple aggregation based on the observation that most ultramicrobacterial cells attached to Methanothrix were Ca. Yanofskybacteria and positive correlation (p < 0.05) between the relative abundance of Patescibacteria lineages and methanogenic archaea (e.g., Ca. Yanofskybacteria-Methanothrix and uncultured clade 32-520-Methanospirillum). The results shed light on a novel cross-domain symbiosis and inspire potential strategies for culturing CPR/DPANN.

microbiology↗

bioRxiv2021

Astrocyte Ca2+ Signaling is Facilitated in an Scn1a+/- Mouse Model of Dravet Syndrome

Dravet syndrome (DS) is an infantile-onset epileptic encephalopathy. More than 80% of DS patients have a heterozygous mutation in SCN1A, which encodes a subunit of the voltage-gated sodium channel, Nav1.1, in neurons. The roles played by astrocytes, the most abundant glial cell type in the brain, have been investigated in the pathogenesis of epilepsy; however, the specific involvement of astrocytes in DS has not been clarified. In this study, we evaluated Ca2+ signaling in astrocytes using genetically modified mice that have a loss-of-function mutation in Scn1a. We found that the slope of spontaneous Ca2+ spiking was increased without a change in amplitude in Scn1a+/- astrocytes. In addition, ATP-induced transient Ca2+ influx and the slope of Ca2+ spiking were also increased in Scn1a+/- astrocytes. These data indicate that perturbed Ca2+ dynamics in astrocytes may be involved in the pathogenesis of DS.

neuroscience↗

bioRxiv2021

The excitatory/inhibitory balance of synaptic transmission is impaired at higher extracellular Ca2+ concentrations in Scn1a+/- mouse model of Dravet syndrome

Dravet syndrome (DS) is an intractable form of childhood epilepsy that occurs in infancy. More than 80% of all patients have a heterozygous abnormality in the SCN1A gene, which encodes a subunit of Na+ channels in the brain. However, the detailed pathogenesis of DS remains unclear. This study investigated the synaptic pathogenesis of this disease in terms of excitatory/inhibitory balance using a mouse model of DS. We show that excitatory postsynaptic currents were similar between Scn1a knock-in neurons (Scn1a+/- neurons) and wild-type neurons, but inhibitory postsynaptic currents were significantly lower in Scn1a+/- neurons. Moreover, both the vesicular release probability and the number of inhibitory synapses were significantly lower in Scn1a+/- neurons compared with wild-type neurons. There was no proportional increase in inhibitory postsynaptic current amplitude in response to increased extracellular Ca2+ concentrations. Our study revealed that the number of inhibitory synapses is significantly reduced in Scn1a+/- neurons, while the sensitivity of inhibitory synapses to extracellular Ca2+ concentrations is markedly increased. These data suggest that Ca2+ tethering in inhibitory nerve terminals may be disturbed following the synaptic burst, likely leading to epileptic symptoms.

neuroscience↗

bioRxiv2020

Presynaptically silent synapses are modulated by the density of surrounding astrocytes

The astrocyte, a major glial cell type, is involved in formation and maturation of synapses, and thus contributes to sustainable synaptic transmission between neurons. Given that the animals in the higher phylogenetic tree have brains with higher density of glial cells with respect to neurons, there is a possibility that the relative astrocytic density directly influences synaptic transmission. However, the notion has not been tested thoroughly. Here we addressed it, by using a primary culture preparation where single hippocampal neurons are surrounded by a variable but countable number of cortical astrocytes in dot-patterned microislands, and recording synaptic transmission by patch-clamp electrophysiology. Neurons with a higher astrocytic density showed a higher amplitude of evoked excitatory postsynaptic current (EPSC) than that of neurons with a lower astrocytic density. The size of readily releasable pool of synaptic vesicles per neuron was significantly higher. The frequency of spontaneous synaptic transmission (miniature EPSC) was higher, but the amplitude was unchanged. The number of morphologically identified glutamatergic synapses was unchanged, but the number of functional ones was increased, indicating a lower ratio of presynaptically silent synapses. Taken together, the higher astrocytic density enhanced excitatory synaptic transmission by increasing the number of functional synapses through presynaptic un-silencing.

neuroscience↗