Brain Healthcare Quotient as a Tool for Standardized Approach in Brain Healthcare Interventions.

In addressing the challenge of assessing healthy brain aging across diverse interventions, this study introduces the use of MRI-derived Brain Healthcare Quotients (BHQ) for comprehensive evaluation. We analyzed BHQ changes in 319 participants aged 24-69, who were allocated into dietary (collagen peptide, euglena, matcha, isohumulone, xanthophyll) and physical activity (hand massage with lavender oil, handwriting, office stretching, pink lens, clinical art) groups, alongside a control group, over a month. These interventions were specifically chosen to test the efficacy of varying health strategies on brain health, measured through BHQ indices: GM-BHQ for gray matter volume, and FA-BHQ for white matter integrity. Notably, significant improvements in FA-BHQ were observed in the collagen peptide group, with marginal increases in the hand massage and office stretching groups. These findings highlight BHQ's potential as a sensitive tool for detecting brain health changes, offering evidence that low-intensity, easily implemented interventions can have beneficial effects on brain health. Moreover, BHQ allows for the systematic evaluation of such interventions using standard statistical approaches, suggesting its value in future brain healthcare research.

Disruption of SMC-related genes promotes recombinant cholesterol esterase production in Burkholderia stabilis.

Burkholderia stabilis strain FERMP-21014 secretes cholesterol esterase (BsChe), which is used in clinical settings to determine serum cholesterol levels. Previously, we constructed an expression plasmid with an endogenous constitutive promoter to enable the production of recombinant BsChe. In this study, we obtained one mutant strain with 13.1-fold higher BsChe activity than the wild type, using N-methyl-N'-nitro-N-nitrosoguanidine as a mutagen. DNA-sequencing analysis revealed that the strain had lost chromosome 3 (∆Chr3), suggesting that the genes hindering BsChe production may be encoded on Chr3. We also identified common mutations in the functionally unknown BSFP_068720/30 genes in the top 10 active strains generated during transposon mutagenesis. As BSFP_068720/30/40 comprised an operon on Chr3, we created the BSFP_068720/30/40 disruption mutant and confirmed that each disruption mutant containing the expression plasmid exhibited ~ 16.1-fold higher BsChe activity than the wild type. Quantitative PCR showed that each disruption mutant and ΔChr3 had a ~ 9.4-fold higher plasmid copy number than the wild type. Structural prediction models indicate that BSFP_068730/40 is structurally homologous to the structural maintenance of chromosomes (SMC) protein MukBE, which is responsible for chromosome segregation during cell division. Conversely, BSFP_068720/30/40 disruption did not lead to a Chr3 drop-out. These results imply that BSFP_068720/30/40 is not a SMC protein but is involved in destabilizing foreign plasmids to prevent the influx of genetic information from the environment. In conclusion, the disruption of BSFP_068720/30/40 improved plasmid stability and copy number, resulting in exceptionally high BsChe production. KEY POINTS: • Disruption of BSFP_068720/30/40 enabled mass production of Burkholderia Che/Lip. • BSFP_068730/40 is an SMC protein homolog not involved in chromosome retention. • BSFP_068720/30/40 is likely responsible for the exclusion of exogenous plasmids.

Oscillatory population-level activity of dorsal raphe serotonergic neurons is inscribed in sleep structure.

Dorsal raphe (DR) 5-HT neurons regulate sleep-wake transitions. Previous studies demonstrated that single-unit activity of DR 5-HT neurons is high during wakefulness, decreases during non-rapid eye movement (NREM) sleep, and ceases during rapid eye movement (REM) sleep. However, characteristics of the population-level activity of DR 5-HT neurons, which influence the entire brain, are largely unknown. Here, we measured population activities of 5-HT neurons in the male and female mouse DR across the sleep-wake cycle by ratiometric fiber photometry. We found a slow oscillatory activity of compound intracellular Ca2+ signals during NREM sleep. The trough of the concave 5-HT activity increased across sleep progression, but 5-HT activity always returned to that seen during the wake period. When the trough reached a minimum and remained there, REM sleep was initiated. We also found a unique coupling of the oscillatory 5-HT activity and wide-band EEG power fluctuation. Furthermore, optogenetic activation of 5-HT neurons during NREM sleep triggered a high EMG power and induced wakefulness, demonstrating a causal role of 5-HT neuron activation. Optogenetic inhibition induced REM sleep or sustained NREM, with an EEG power increase and EEG fluctuation, and pharmacological silencing of 5-HT activity using a selective serotonin reuptake inhibitor led to sustained NREM, with an EEG power decrease and EEG fluctuation. These inhibitory manipulations supported the association between oscillatory 5-HT activity and EEG fluctuation. We propose that NREM sleep is not a monotonous state, but rather it contains dynamic changes that coincide with the oscillatory population-level activity of DR 5-HT neurons.SIGNIFICANT STATEMENTPrevious studies have demonstrated single-cell 5-HT neuronal activity across sleep-wake conditions. However, population-level activities of these neurons are not well understood. We monitored dorsal raphe (DR) 5-HT population activity using a fiber photometry system in mice and found that activity was highest during wakefulness and lowest during rapid eye movement (REM) sleep. Surprisingly, during non-REM sleep, the 5-HT population activity decreased with an oscillatory pattern, coinciding with EEG fluctuations. EEG fluctuations persisted when DR 5-HT neuron activity was silenced by either optogenetic or pharmacological interventions during non-REM sleep, suggesting an association between the two. Although oscillatory DR 5-HT neuron activity did not generate EEG fluctuations, it provides evidence that non-REM sleep exhibits at least binary states.

Rhythmic activation of excitatory neurons in the mouse frontal cortex improves the prefrontal cortex-mediated cognitive function.

The prefrontal cortex (PFC) plays essential roles in cognitive processes. Previous studies have suggested the layer and the cell type-specific activation for cognitive enhancement. However, the mechanism by which a temporal pattern of activation affects cognitive function remains to be elucidated. Here, we investigated whether the specific activation of excitatory neurons in the superficial layers mainly in the PFC according to a rhythmic or nonrhythmic pattern could modulate the cognitive functions of normal mice. We used a C128S mutant of channelrhodopsin 2, a step function opsin, and administered two light illumination patterns: (i) alternating pulses of blue and yellow light for rhythmic activation or (ii) pulsed blue light only for nonrhythmic activation. Behavioral analyses were performed to compare the behavioral consequences of these two neural activation patterns. The alternating blue and yellow light pulses, but not the pulsed blue light only, significantly improved spatial working memory and social recognition without affecting motor activity or the anxiety level. These results suggest that the rhythmic, but not the nonrhythmic, activation could enhance cognitive functions. This study indicates that not only the population of neurons that are activated but also the pattern of activation plays a crucial role in the cognitive enhancement.

Chronic social defeat stress impairs goal-directed behavior through dysregulation of ventral hippocampal activity in male mice.

Chronic stress is a risk factor for a variety of psychiatric disorders, including depression. Although impairments to motivated behavior are a major symptom of clinical depression, little is known about the circuit mechanisms through which stress impairs motivation. Furthermore, research in animal models for depression has focused on impairments to hedonic aspects of motivation, whereas patient studies suggest that impairments to appetitive, goal-directed motivation contribute significantly to motivational impairments in depression. Here, we characterized goal-directed motivation in repeated social defeat stress (R-SDS), a well-established mouse model for depression in male mice. R-SDS impaired the ability to sustain and complete goal-directed behavior in a food-seeking operant lever-press task. Furthermore, stress-exposed mice segregated into susceptible and resilient subpopulations. Interestingly, susceptibility to stress-induced motivational impairments was unrelated to stress-induced social withdrawal, another prominent effect of R-SDS in mouse models. Based on evidence that ventral hippocampus (vHP) modulates sustainment of goal-directed behavior, we monitored vHP activity during the task using fiber photometry. Successful task completion was associated with suppression of ventral hippocampal neural activity. This suppression was diminished after R-SDS in stress-susceptible but not stress-resilient mice. The serotonin selective reuptake inhibitor (SSRI) escitalopram and ketamine both normalized vHP activity during the task and restored motivated behavior. Furthermore, optogenetic vHP inhibition was sufficient to restore motivated behavior after stress. These results identify vHP hyperactivity as a circuit mechanism of stress-induced impairments to goal-directed behavior and a putative biomarker that is sensitive to antidepressant treatments and that differentiates susceptible and resilient individuals.

Activation of ventral CA1 hippocampal neurons projecting to the lateral septum during feeding.

A number of studies have reported the involvement of the ventral hippocampus (vHip) and the lateral septum (LS) in negative emotional responses. Besides these well-documented functions, they are also thought to control feeding behavior. In particular, optogenetic and pharmacogenetic interventions to LS-projecting vHip neurons have demonstrated that the vHip→LS neural circuit exerts an inhibition on feeding behavior. However, there have been no reports of vHip neuronal activity during feeding. Here, we focused on LS-projecting vCA1 neurons (vCA1→LS ) and monitored their activity during feeding behaviors in mice. vCA1→LS neurons were retrogradely labeled with adeno-associated virus carrying a ratiometric Ca2+ indicator and measured compound Ca2+ dynamics by fiber photometry. We first examined vCA1→LS activity in random food-exploring behavior and found that vCA1→LS activation seemed to coincide with food intake; however, our ability to visually confirm this during freely moving behaviors was not sufficiently reliable. We next examined vCA1→LS activity in a goal-directed, food-seeking lever-press task which temporally divided the mouse state into preparatory, effort, and consummatory phases. We observed vCA1→LS activation in the postprandial period during the consummatory phase. Such timing- and pathway-specific activation was not observed from pan-vCA1 neurons. In contrast, reward omission eliminated this activity, indicating that vCA1→LS activation is contingent on the food reward. Sated mice pressed the lever significantly fewer times but still ate food; however, vCA1→LS neurons were not activated, suggesting that vCA1→LS neurons did not respond to habitual behavior. Combined, these results suggest that gastrointestinal interoception rather than food-intake motions or external sensations are likely to coincide with vCA1→LS activity. Accordingly, we propose that vCA1→LS neurons discriminate between matched or unmatched predictive bodily states in which incoming food will satisfy an appetite. We also demonstrate that vCA1→LS neurons are activated in aversive/anxious situations in an elevated plus maze and tail suspension test. Future behavioral tests utilizing anxious conflict and food intake may reconcile the multiple functions of vCA1→LS neurons.

Bacterial triacylglycerol lipase is a potential cholesterol esterase: Identification of a key determinant for sterol-binding specificity.

Cholesterol esterase (Che) from Burkholderia stabilis (BsChe) is a homolog of well-characterized and industrially relevant bacterial triacylglycerol lipases (Lips). BsChe is a rare bacterial Lip enzyme that exhibits practical Che activity and is currently used in clinical applications to determine total serum cholesterol levels. To investigate the sterol specificity of BsChe, we determined the X-ray structure of BsChe. We discovered a local structural change in the active-site cleft, which might be related to substrate binding and product release. We also performed molecular docking studies by using the X-ray models of BsChe and cholesterol linoleate (CLL), the most favorable substrate for BsChe. The results showed that the sterol moieties of reasonable CLL docking poses localized to a specific active-site cleft surface formed by Leu266 and Ile287, which are unconserved among Burkholderia Lip homologs. Site-directed mutagenesis identified these residues as essential for the Che activity of BsChe, and Leu or Ile substitution conferred marked Che activity to Burkholderia Lips. In particular, Burkholderia cepacia and Burkholderia ubonensis Lips with the V266L/L287I double mutation exhibited ~50-fold and 500-fold higher Che activities than those of the wild-type enzymes, respectively. These results provide new insights into the substrate-binding mechanisms and selectivities of bacterial Lips.

Opposing Ventral Striatal Medium Spiny Neuron Activities Shaped by Striatal Parvalbumin-Expressing Interneurons during Goal-Directed Behaviors.

Medium spiny neurons (MSNs) of mice show opposing activities upon the initiation of a food-seeking lever press task. Ventromedial striatal (VMS)-MSNs are inhibited but ventrolateral striatal (VLS)-MSNs are activated; these activities mediate action selection and action initiation, respectively. To understand what input shapes the opposing MSN activities, here, we monitor cortical input activities at the cell population level and artificially reverse them. We demonstrate that the ventral hippocampus (vHP) and the insular cortex (IC) are major inputs to the VMS and VLS, both projections show silencing at the trial start time, and the vHP-VMS and IC-VLS pathways form functionally coupled input-output units during the task. Of note, the upstream IC silencing is converted to the downstream VLS-MSN activation. We find biased localization of striatal parvalbumin-expressing interneurons (PV INs) and verify PV IN-dependent feedforward architecture in the VLS. Our results reveal a distinct mode of cortico-striatal signal conveyance via feedforward disinhibition in behaving animals.

[Optical-Observation and -Manipulation Reveals the Neural Mechanisms of Motivated Behavior].

Motivated behavior consists of a directional component that enables subjects to select a behavior, which leads to an outcome, and an activational component that initiates and sustains actions directed toward goals. How does the brain regulate different components underlying motivated behaviors? This review describes the investigation of specific components of motivated behaviors using optical-observation and -manipulation, which allows for neurocircuit identification.

Behavioral and electrophysiological evidence for a neuroprotective role of aquaporin-4 in the 5xFAD transgenic mice model.

Aquaporin-4 (AQP4) has been suggested to be involved in the pathogenesis of neurodegenerative diseases including Alzheimer's disease (AD), which may be due to the modulation of neuroinflammation or the impairment of interstitial fluid bulk flow system in the central nervous system. Here, we show an age-dependent impairment of several behavioral outcomes in 5xFAD AQP4 null mice. Twenty-four-hour video recordings and computational analyses of their movement revealed that the nighttime motion of AQP4-deficient 5xFAD mice was progressively reduced between 20 and 36 weeks of age, with a sharp deterioration occurring between 30 and 32 weeks. This reduction in nighttime motion was accompanied by motor dysfunction and epileptiform neuronal activities, demonstrated by increased abnormal spikes by electroencephalography. In addition, all AQP4-deficient 5xFAD mice exhibited convulsions at least once during the period of the analysis. Interestingly, despite such obvious phenotypes, parenchymal amyloid ß (Aß) deposition, reactive astrocytosis, and activated microgliosis surrounding amyloid plaques were unchanged in the AQP4-deficient 5xFAD mice relative to 5xFAD mice. Taken together, our data indicate that AQP4 deficiency greatly accelerates an age-dependent deterioration of neuronal function in 5xFAD mice associated with epileptiform neuronal activity without significantly altering Aß deposition or neuroinflammation in this mouse model. We therefore propose that there exists another pathophysiological phase in AD which follows amyloid plaque deposition and neuroinflammation and is sensitive to AQP4 deficiency.

Peculiarities of biofilm formation by Paracoccus denitrificans.

Most bacteria form biofilms, which are thick multicellular communities covered in extracellular matrix. Biofilms can become thick enough to be even observed by the naked eye, and biofilm formation is a tightly regulated process. Paracoccus denitrificans is a non-motile, Gram-negative bacterium that forms a very thin, unique biofilm. A key factor in the biofilm formed by this bacterium is a large surface protein named biofilm-associated protein A (BapA), which was recently reported to be regulated by cyclic diguanosine monophosphate (cyclic-di-GMP or c-di-GMP). Cyclic-di-GMP is a major second messenger involved in biofilm formation in many bacteria. Though cyclic-di-GMP is generally reported as a positive regulatory factor in biofilm formation, it represses biofilm formation in P. denitrificans. Furthermore, quorum sensing (QS) represses biofilm formation in this bacterium, which is also reported as a positive regulator of biofilm formation in most bacteria. The QS signal used in P. denitrificans is hydrophobic and is delivered through membrane vesicles. Studies on QS show that P. denitrificans can potentially form a thick biofilm but maintains a thin biofilm under normal growth conditions. In this review, we discuss the peculiarities of biofilm formation by P. denitrificans with the aim of deepening the overall understanding of bacterial biofilm formation and functions.

Cerebral White Matter Hyperintensity as a Healthcare Quotient.

To better understand the risk factors and optimal therapeutic strategies of cerebral white matter hyperintensity (WMH), we examined a large population of adults with and without various vascular risk factors (VRFs) or vascular risk conditions (VRCs), such as hypertension (HT), diabetes mellitus (DM), and dyslipidemia (DLP), including the comorbidities. We assessed two participant groups having no medical history of stroke or dementia that underwent brain checkup using magnetic resonance imaging (MRI): 5541 participants (2760 men, 2781 women) without VRCs and 1969 participants (1169 men, 800 women) who had received drug treatments for VRCs and the combination of comorbidities. For data analysis, we constructed WMH-brain healthcare quotient (WMH-BHQ) based on the percentile rank of WMH volume. This metric has an inverse relation to WMH. Multiple linear regression analysis of 5541 participants without VRCs revealed that age, systolic blood pressure (SBP), Brinkman index (BI), and female sex were significant factors lowering WMH-BHQ, whereas body mass index (BMI), male sex, fasting blood sugar, and triglyceride levels were increasing factors. The Kruskal-Wallis test and Dunn tests showed that WMH-BHQs significantly increased or decreased with BMI or SBP and with BI classification, respectively. Regarding the impact of impaired fasting glucose and abnormal lipid metabolism, there were almost no significant relationships. For 1969 participants who had HT, DM, and DLP, as well as their comorbidities, we found that DLP played a substantial role in increasing WMH-BHQ for some comorbidities, whereas the presence of HT and DM alone tended to decrease it. Cerebral WMH can be used as a healthcare quotient for quantitatively evaluating VRFs and VRCs and their comorbidities.

Production of recombinant extracellular cholesterol esterase using consistently active promoters in Burkholderia stabilis.

Burkholderia stabilis FERMP-21014 produces highly active cholesterol esterase in the presence of fatty acids. To develop an overexpression system for cholesterol esterase production, we carried out RNA sequencing analyses to screen strongly active promoters in FERMP-21014. Based on gene expression consistency analysis, we selected nine genes that were consistently expressed at high levels, following which we constructed expression vectors using their promoter sequences and achieved overproduction of extracellular cholesterol esterase under fatty acid-free conditions. Of the tested promoters, the promoter of BSFP_0720, which encodes the alkyl hydroperoxide reductase subunit AhpC, resulted in the highest cholesterol esterase activity (24.3 U mL-1). This activity level was 243-fold higher than that of the wild-type strain under fatty acid-free conditions. We confirmed that cholesterol esterase was secreted without excessive accumulation within the cells. The gene expression consistency analysis will be useful to screen promoters applicable to the overexpression of other industrially important enzymes.

Genome Sequence of Rhodococcus erythropolis Type Strain JCM 3201.

Rhodococcus erythropolis JCM 3201 can express several recombinant proteins that are difficult to express in Escherichia coli It is used as one of the hosts for protein expression and bioconversion. Here, we report the draft genome sequence of R. erythropolis JCM 3201.

Serotonin-mediated inhibition of ventral hippocampus is required for sustained goal-directed behavior.

The ability to sustain goal-directed action is essential for success in many domains, but little is known about the corresponding neural substrates. Using fiber photometry to monitor population neural activity, we demonstrate that engagement in sustained food- or punishment-motivated behavior is associated with suppression of ventral but not dorsal hippocampal activity. Using optogenetic stimulation, we demonstrate that this suppression is required for goal-directed behavior, whereas optogenetic suppression of the ventral hippocampus (vHP) enhances the ability to sustain goal-directed behavior. Suppression of vHP during sustained goal-directed behavior was accompanied by increased activity in median but not dorsal raphe, implicating serotonergic signaling through Htr3a as a mechanism of vHP suppression during successful goal-directed behavior. Sustainment of goal-directed action may require suppression of vHP because of the structure's well-documented role in behavioral inhibition.

Complete Genome Sequence of an Efficient Vitamin D3-Hydroxylating Bacterium, Pseudonocardia autotrophica NBRC 12743.

Pseudonocardia autotrophica NBRC 12743 contains a cytochrome P450 vitamin D3 hydroxylase, and it is used as a biocatalyst for the commercial production of hydroxyvitamin D3, a valuable compound for medication. Here, we report the complete genome sequence of P. autotrophica NBRC 12743, which could be useful for improving the productivity of hydroxyvitamin D3.

Nitric Oxide Production from Nitrite Reduction and Hydroxylamine Oxidation by Copper-containing Dissimilatory Nitrite Reductase (NirK) from the Aerobic Ammonia-oxidizing Archaeon, Nitrososphaera viennensis.

Aerobic ammonia-oxidizing archaea (AOA) play a crucial role in the global nitrogen cycle by oxidizing ammonia to nitrite, and nitric oxide (NO) is a key intermediate in AOA for sustaining aerobic ammonia oxidation activity. We herein heterologously expressed the NO-forming, copper-containing, dissimilatory nitrite reductase (NirK) from Nitrososphaera viennensis and investigated its enzymatic properties. The recombinant protein catalyzed the reduction of 15NO2- to 15NO, the oxidation of hydroxylamine (15NH2OH) to 15NO, and the production of 14-15N2O from 15NH2OH and 14NO2-. To the best of our knowledge, the present study is the first to document the enzymatic properties of AOA NirK.

Heparan Sulfate Organizes Neuronal Synapses through Neurexin Partnerships.

Synapses are fundamental units of communication in the brain. The prototypical synapse-organizing complex neurexin-neuroligin mediates synapse development and function and is central to a shared genetic risk pathway in autism and schizophrenia. Neurexin's role in synapse development is thought to be mediated purely by its protein domains, but we reveal a requirement for a rare glycan modification. Mice lacking heparan sulfate (HS) on neurexin-1 show reduced survival, as well as structural and functional deficits at central synapses. HS directly binds postsynaptic partners neuroligins and LRRTMs, revealing a dual binding mode involving intrinsic glycan and protein domains for canonical synapse-organizing complexes. Neurexin HS chains also bind novel ligands, potentially expanding the neurexin interactome to hundreds of HS-binding proteins. Because HS structure is heterogeneous, our findings indicate an additional dimension to neurexin diversity, provide a molecular basis for fine-tuning synaptic function, and open therapeutic directions targeting glycan-binding motifs critical for brain development.

Optogenetic astrocyte activation evokes BOLD fMRI response with oxygen consumption without neuronal activity modulation.

Functional magnetic resonance imaging (fMRI) based on the blood oxygenation level-dependent (BOLD) signal has been used to infer sites of neuronal activation in the brain. A recent study demonstrated, however, unexpected BOLD signal generation without neuronal excitation, which led us to hypothesize the presence of another cellular source for BOLD signal generation. Collective assessment of optogenetic activation of astrocytes or neurons, fMRI in awake mice, electrophysiological measurements, and histochemical detection of neuronal activation, coherently suggested astrocytes as another cellular source. Unexpectedly, astrocyte-evoked BOLD signal accompanied oxygen consumption without modulation of neuronal activity. Imaging mass spectrometry of brain sections identified synthesis of acetyl-carnitine via oxidative glucose metabolism at the site of astrocyte-, but not neuron-evoked BOLD signal. Our data provide causal evidence that astrocytic activation alone is able to evoke BOLD signal response, which may lead to reconsideration of current interpretation of BOLD signal as a marker of neuronal activation.

Striatonigral direct pathway activation is sufficient to induce repetitive behaviors.

Pharmacological intervention in the substantia nigra is known to induce repetitive behaviors in rodents, but a direct causal relationship between a specific neural circuit and repetitive behavior has not yet been established. Here we demonstrate that optogenetic activation of dopamine D1 receptor-expressing MSNs terminals in the substantia nigra pars reticulata resulted in sustained and chronic repetitive behaviors. These data show for the first time that activation of the striatonigral direct pathway is sufficient to generate motor stereotypies.