Large-scale animal model study uncovers altered brain pH and lactate levels as a transdiagnostic endophenotype of neuropsychiatric disorders involving cognitive impairment.

Increased levels of lactate, an end-product of glycolysis, have been proposed as a potential surrogate marker for metabolic changes during neuronal excitation. These changes in lactate levels can result in decreased brain pH, which has been implicated in patients with various neuropsychiatric disorders. We previously demonstrated that such alterations are commonly observed in five mouse models of schizophrenia, bipolar disorder, and autism, suggesting a shared endophenotype among these disorders rather than mere artifacts due to medications or agonal state. However, there is still limited research on this phenomenon in animal models, leaving its generality across other disease animal models uncertain. Moreover, the association between changes in brain lactate levels and specific behavioral abnormalities remains unclear. To address these gaps, the International Brain pH Project Consortium investigated brain pH and lactate levels in 109 strains/conditions of 2294 animals with genetic and other experimental manipulations relevant to neuropsychiatric disorders. Systematic analysis revealed that decreased brain pH and increased lactate levels were common features observed in multiple models of depression, epilepsy, Alzheimer's disease, and some additional schizophrenia models. While certain autism models also exhibited decreased pH and increased lactate levels, others showed the opposite pattern, potentially reflecting subpopulations within the autism spectrum. Furthermore, utilizing large-scale behavioral test battery, a multivariate cross-validated prediction analysis demonstrated that poor working memory performance was predominantly associated with increased brain lactate levels. Importantly, this association was confirmed in an independent cohort of animal models. Collectively, these findings suggest that altered brain pH and lactate levels, which could be attributed to dysregulated excitation/inhibition balance, may serve as transdiagnostic endophenotypes of debilitating neuropsychiatric disorders characterized by cognitive impairment, irrespective of their beneficial or detrimental nature.

An Interaction between Brain-Derived Neurotrophic Factor and Stress-Related Glucocorticoids in the Pathophysiology of Alzheimer's Disease.

Both the brain-derived neurotrophic factor (BDNF) and glucocorticoids (GCs) play multiple roles in various aspects of neurons, including cell survival and synaptic function. BDNF and its receptor TrkB are extensively expressed in neurons of the central nervous system (CNS), and the contribution of the BDNF/TrkB system to neuronal function is evident; thus, its downregulation has been considered to be involved in the pathogenesis of Alzheimer's disease (AD). GCs, stress-related molecules, and glucocorticoid receptors (GRs) are also considered to be associated with AD in addition to mental disorders such as depression. Importantly, a growing body of evidence suggests a close relationship between BDNF/TrkB-mediated signaling and the GCs/GR system in the CNS. Here, we introduce the current studies on the interaction between the neurotrophic system and stress in CNS neurons and discuss their involvement in the pathophysiology of AD.

Involvement of brain-derived neurotrophic factor signaling in the pathogenesis of stress-related brain diseases.

Neurotrophins including brain-derived neurotrophic factor, BDNF, have critical roles in neuronal differentiation, cell survival, and synaptic function in the peripheral and central nervous system. It is well known that a variety of intracellular signaling stimulated by TrkB, a high-affinity receptor for BDNF, is involved in the physiological and pathological neuronal aspects via affecting cell viability, synaptic function, neurogenesis, and cognitive function. As expected, an alteration of the BDNF/TrkB system is suspected to be one of the molecular mechanisms underlying cognitive decline in cognitive diseases and mental disorders. Recent evidence has also highlighted a possible link between the alteration of TrkB signaling and chronic stress. Furthermore, it has been demonstrated that downregulation of the BDNF/TrkB system and chronic stress have a role in the pathogenesis of Alzheimer's disease (AD) and mental disorders. In this review, we introduce current evidence showing a close relationship between the BDNF/TrkB system and the development of cognition impairment in stress-related disorders, and the possible contribution of the upregulation of the BDNF/TrkB system in a therapeutic approach against these brain diseases.

Neurotrophins and Other Growth Factors in the Pathogenesis of Alzheimer's Disease.

The involvement of the changed expression/function of neurotrophic factors in the pathogenesis of neurodegenerative diseases, including Alzheimer's disease (AD), has been suggested. AD is one of the age-related dementias, and is characterized by cognitive impairment with decreased memory function. Developing evidence demonstrates that decreased cell survival, synaptic dysfunction, and reduced neurogenesis are involved in the pathogenesis of AD. On the other hand, it is well known that neurotrophic factors, especially brain-derived neurotrophic factor (BDNF) and its high-affinity receptor TrkB, have multiple roles in the central nervous system (CNS), including neuronal maintenance, synaptic plasticity, and neurogenesis, which are closely linked to learning and memory function. Thus, many investigations regarding therapeutic approaches to AD, and/or the screening of novel drug candidates for its treatment, focus on upregulation of the BDNF/TrkB system. Furthermore, current studies also demonstrate that GDNF, IGF1, and bFGF, which play roles in neuroprotection, are associated with AD. In this review, we introduce data demonstrating close relationships between the pathogenesis of AD, neurotrophic factors, and drug candidates, including natural compounds that upregulate the BDNF-mediated neurotrophic system.

Enhanced social reward response and anxiety-like behavior with downregulation of nucleus accumbens glucocorticoid receptor in BALB/c mice.

Social anhedonia is a psychological state with difficulty in experiencing pleasure from social interactions and is observed in various diseases, such as depressive disorders. Although the relationships between social reward responses and anxiety- and depression-like behaviors have remained unclear, a social reward conditioned place preference (SCPP) test can be used to analyze the rewarding nature of social interactions. To elucidate these relationships, we used 5-week-old male mice of AKR, BALB/c, and C57BL/6J strains and conducted behavioral tests in the following order: elevated plus-maze test (EPM), open field test (OFT), SCPP, saccharin preference test (SPT), and passive avoidance test. The nucleus accumbens of these mice were collected 24 hr after these behavioral tests and were used for western blotting to determine the levels of receptors for brain-derived neurotrophic factors and glucocorticoids. BALB/c mice displayed the highest levels of anxiety-like behavior in EPM and OFT as well as physical anhedonia-like behaviors in SPT. They also showed increased responses to social rewards and huddling behaviors in SCPP, with downregulated glucocorticoid receptor (GR). Regression analysis results revealed positive influences of anxiety- and physical anhedonia-like behaviors and expressions of GR on social reward responses. Collectively, temperament associated with anxiety and physical anhedonia may affect social reward responses, which possibly is influenced by the expression of GR that can modify these psychological traits.

The Role of Neurotrophin Signaling in Age-Related Cognitive Decline and Cognitive Diseases.

Neurotrophins are a family of secreted proteins expressed in the peripheral nervous system and the central nervous system that support neuronal survival, synaptic plasticity, and neurogenesis. Brain-derived neurotrophic factor (BDNF) and its high affinity receptor TrkB are highly expressed in the cortical and hippocampal areas and play an essential role in learning and memory. The decline of cognitive function with aging is a major risk factor for cognitive diseases such as Alzheimer's disease. Therefore, an alteration of BDNF/TrkB signaling with aging and/or pathological conditions has been indicated as a potential mechanism of cognitive decline. In this review, we summarize the cellular function of neurotrophin signaling and review the current evidence indicating a pathological role of neurotrophin signaling, especially of BDNF/TrkB signaling, in the cognitive decline in aging and age-related cognitive diseases. We also review the therapeutic approach for cognitive decline by the upregulation of the endogenous BDNF/TrkB-system.

Brain-Derived Neurotrophic Factor Signaling in the Pathophysiology of Alzheimer's Disease: Beneficial Effects of Flavonoids for Neuroprotection.

The function of the brain-derived neurotrophic factor (BDNF) via activation through its high-affinity receptor Tropomyosin receptor kinase B (TrkB) has a pivotal role in cell differentiation, cell survival, synaptic plasticity, and both embryonic and adult neurogenesis in central nervous system neurons. A number of studies have demonstrated the possible involvement of altered expression and action of the BDNF/TrkB signaling in the pathogenesis of neurodegenerative diseases, including Alzheimer's disease (AD). In this review, we introduce an essential role of the BDNF and its downstream signaling in neural function. We also review the current evidence on the deregulated the BDNF signaling in the pathophysiology of AD at gene, mRNA, and protein levels. Further, we discuss a potential usefulness of small compounds, including flavonoids, which can stimulate BDNF-related signaling as a BDNF-targeting therapy.

Rapid and Simplified Induction of Neural Stem/Progenitor Cells (NSCs/NPCs) and Neurons from Human Induced Pluripotent Stem Cells (hiPSCs).

Human induced pluripotent stem cells (iPSCs) and their progeny displaying tissue-specific characteristics have paved the way for regenerative medicine and research in various fields such as the elucidation of the pathological mechanism of diseases and the discovery of drug candidates. iPSC-derived neurons are particularly valuable as it is difficult to analyze neural cells obtained from the central nervous system in humans. For neuronal induction with iPSCs, one of the commonly used approaches is the isolation and expansion of neural rosettes, following the formation of embryonic bodies (EBs). However, this process is laborious, inefficient, and requires further purification of the cells. To overcome these limitations, we have developed an efficient neural induction method that allows for the generation of neural stem/progenitor cells (NSCs/NPCs) from iPSCs within 7 days and of functional mature neurons. Our method yields a PAX6-positive homogeneous cell population, a cortical NSCs/NPCs, and the resultant NSCs/NPCs can be cryopreserved, expanded, and differentiated into functional mature neurons. Moreover, our protocol will be less expensive than other methods since the protocol requires fewer neural supplements during neural induction. This article also presents the FM1-43 imaging assay, which is useful for the presynaptic assessment of the iPSCs-derived human neurons. This protocol provides a quick and simplified way to generate NSCs/NPCs and neurons, enabling researchers to establish in vitro cellular models to study brain disease pathology.

Small noncoding vault RNA modulates synapse formation by amplifying MAPK signaling.

The small noncoding vault RNA (vtRNA) is a component of the vault complex, a ribonucleoprotein complex found in most eukaryotes. Emerging evidence suggests that vtRNAs may be involved in the regulation of a variety of cellular functions when unassociated with the vault complex. Here, we demonstrate a novel role for vtRNA in synaptogenesis. Using an in vitro synapse formation model, we show that murine vtRNA (mvtRNA) promotes synapse formation by modulating the MAPK signaling pathway. mvtRNA is transported to the distal region of neurites as part of the vault complex. Interestingly, mvtRNA is released from the vault complex in the neurite by a mitotic kinase Aurora-A-dependent phosphorylation of MVP, a major protein component of the vault complex. mvtRNA binds to and activates MEK1 and thereby enhances MEK1-mediated ERK activation in neurites. These results suggest the existence of a regulatory mechanism of the MAPK signaling pathway by vtRNAs as a new molecular basis for synapse formation.

An iPSC-based neural model of sialidosis uncovers glycolytic impairment-causing presynaptic dysfunction and deregulation of Ca2+ dynamics.

Sialidosis is a neuropathic lysosomal storage disease caused by a deficiency in the NEU1 gene-encoding lysosomal neuraminidase and characterized by abnormal accumulation of undigested sialyl-oligoconjugates in systemic organs including brain. Although patients exhibit neurological symptoms, the underlying neuropathological mechanism remains unclear. Here, we generated induced pluripotent stem cells (iPSCs) from skin fibroblasts with sialidosis and induced the differentiation into neural progenitor cells (NPCs) and neurons. Sialidosis NPCs and neurons mimicked the disease-like phenotypes including reduced neuraminidase activity, accumulation of sialyl-oligoconjugates and lysosomal expansions. Functional analysis also revealed that sialidosis neurons displayed two distinct abnormalities, defective exocytotic glutamate release and augmented α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptor (AMPAR)-mediated Ca2+ influx. These abnormalities were restored by overexpression of the wild-type NEU1 gene, demonstrating causative role of neuraminidase deficiency in functional impairments of disease neurons. Comprehensive proteomics analysis revealed the significant reduction of SNARE proteins and glycolytic enzymes in synaptosomal fraction, with downregulation of ATP production. Bypassing the glycolysis by treatment of pyruvate, which is final metabolite of glycolysis pathway, improved both the synaptsomal ATP production and the exocytotic function. We also found that upregulation of AMPAR and L-type voltage dependent Ca2+ channel (VDCC) subunits in disease neurons, with the restoration of AMPAR-mediated Ca2+ over-load by treatment of antagonists for the AMPAR and L-type VDCC. Our present study provides new insights into both the neuronal pathophysiology and potential therapeutic strategy for sialidosis.

Novel Drug Candidates Improve Ganglioside Accumulation and Neural Dysfunction in GM1 Gangliosidosis Models with Autophagy Activation.

GM1 gangliosidosis is a lysosomal storage disease caused by loss of lysosomal ß-galactosidase activity and characterized by progressive neurodegeneration due to massive accumulation of GM1 ganglioside in the brain. Here, we generated induced pluripotent stem cells (iPSCs) derived from patients with GM1 gangliosidosis, and the resultant neurons showed impaired neurotransmitter release as a presynaptic function and accumulation of GM1 ganglioside. Treatment of normal neurons with GM1 ganglioside also disturbed presynaptic function. A high-content drug-screening system was then established and identified two compounds as drug candidates for GM1 gangliosidosis. Treatment of the patient-derived neurons with the candidate agents activated autophagy pathways, reducing GM1 ganglioside accumulation in vitro and in vivo, and restoring the presynaptic dysfunction. Our findings thus demonstrated the potential value of patient-derived iPSC lines as cellular models of GM1 gangliosidosis and revealed two potential therapeutic agents for future clinical application.

Presynaptic Dysfunction in Neurons Derived from Tay-Sachs iPSCs.

Tay-Sachs disease (TSD) is a GM2 gangliosidosis lysosomal storage disease caused by a loss of lysosomal hexosaminidase-A (HEXA) activity and characterized by progressive neurodegeneration due to the massive accumulation of GM2 ganglioside in the brain. Here, we generated iPSCs derived from patients with TSD, and found similar potential for neural differentiation between TSD-iPSCs and normal iPSCs, although neural progenitor cells (NPCs) derived from the TSD-iPSCs exhibited enlarged lysosomes and upregulation of the lysosomal marker, LAMP1, caused by the accumulation of GM2 ganglioside. The NPCs derived from TSD-iPSCs also had an increased incidence of oxidative stress-induced cell death. TSD-iPSC-derived neurons showed a decrease in exocytotic activity with the accumulation of GM2 ganglioside, suggesting deficient neurotransmission in TSD. Our findings demonstrated that NPCs and mature neurons derived from TSD-iPSCs are potentially useful cellular models of TSD and are useful for investigating the efficacy of drug candidates in the future.

Actions of Brain-Derived Neurotrophin Factor in the Neurogenesis and Neuronal Function, and Its Involvement in the Pathophysiology of Brain Diseases.

It is well known that brain-derived neurotrophic factor, BDNF, has an important role in a variety of neuronal aspects, such as differentiation, maturation, and synaptic function in the central nervous system (CNS). BDNF stimulates mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK), phosphoinositide-3kinase (PI3K), and phospholipase C (PLC)-gamma pathways via activation of tropomyosin receptor kinase B (TrkB), a high affinity receptor for BDNF. Evidence has shown significant contributions of these signaling pathways in neurogenesis and synaptic plasticity in in vivo and in vitro experiments. Importantly, it has been demonstrated that dysfunction of the BDNF/TrkB system is involved in the onset of brain diseases, including neurodegenerative and psychiatric disorders. In this review, we discuss actions of BDNF and related signaling molecules on CNS neurons, and their contributions to the pathophysiology of brain diseases.

Basic fibroblast growth factor increased glucocorticoid receptors in cortical neurons through MAP kinase pathway.

Prolonged and intense stress chronically increases blood concentration of glucocorticoids, which in turn causes downregulation of glucocorticoid receptor (GR) in the central nervous system (CNS). This process has been suggested to be involved in the pathogenesis of major depressive disorder (MDD). Here, we found that basic fibroblast growth factor (bFGF) increased the expression of GR in the rat cerebral cortex and cultured cortical neurons and restored the reduced GR expression caused by glucocorticoid exposure. Among intracellular signaling pathways stimulated by bFGF, extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) pathway was responsible for the upregulation of GR. The bFGF-induced GR was functional as a transcription factor to enhance transcription of a target gene. Because high stress augments bFGF levels in the brain, it is likely that bFGF plays a compensating role for reduced GR expression after stress and thus should be studied as a therapeutic target for the treatment of MDD.

The Sox2 promoter-driven CD63-GFP transgenic rat model allows tracking of neural stem cell-derived extracellular vesicles.

Extracellular vesicles (EVs) can modulate microenvironments by transferring biomolecules, including RNAs and proteins derived from releasing cells, to target cells. To understand the molecular mechanisms maintaining the neural stem cell (NSC) niche through EVs, a new transgenic (Tg) rat strain that can release human CD63-GFP-expressing EVs from the NSCs was established. Human CD63-GFP expression was controlled under the rat Sox2 promoter (Sox2/human CD63-GFP), and it was expressed in undifferentiated fetal brains. GFP signals were specifically observed in in vitro cultured NSCs obtained from embryonic brains of the Tg rats. We also demonstrated that embryonic NSC (eNSC)-derived EVs were labelled by human CD63-GFP. Furthermore, when we examined the transfer of EVs, eNSC-derived EVs were found to be incorporated into astrocytes and eNSCs, thus implying an EV-mediated communication between different cell types around NSCs. This new Sox2/human CD63-GFP Tg rat strain should provide resources to analyse the cell-to-cell communication via EVs in NSC microenvironments.

Actions of Brain-Derived Neurotrophic Factor and Glucocorticoid Stress in Neurogenesis.

Altered neurogenesis is suggested to be involved in the onset of brain diseases, including mental disorders and neurodegenerative diseases. Neurotrophic factors are well known for their positive effects on the proliferation/differentiation of both embryonic and adult neural stem/progenitor cells (NSCs/NPCs). Especially, brain-derived neurotrophic factor (BDNF) has been extensively investigated because of its roles in the differentiation/maturation of NSCs/NPCs. On the other hand, recent evidence indicates a negative impact of the stress hormone glucocorticoids (GCs) on the cell fate of NSCs/NPCs, which is also related to the pathophysiology of brain diseases, such as depression and autism spectrum disorder. Furthermore, studies including ours have demonstrated functional interactions between neurotrophic factors and GCs in neural events, including neurogenesis. In this review, we show and discuss relationships among the behaviors of NSCs/NPCs, BDNF, and GCs.

Impact of glucocorticoid on neurogenesis.

Neurogenesis is currently an area of great interest in neuroscience. It is closely linked to brain diseases, including mental disorders and neurodevelopmental disease. Both embryonic and adult neurogeneses are influenced by glucocorticoids secreted from the adrenal glands in response to a variety of stressors. Moreover, proliferation/differentiation of the neural stem/progenitor cells (NSPCs) is affected by glucocorticoids through intracellular signaling pathways such as phosphoinositide 3-kinase (PI3K)/Akt, hedgehog, and Wnt. Our review presents recent evidence of the impact of glucocorticoids on NSPC behaviors and the underlying molecular mechanisms; this provides important information for understanding the pathological role of glucocorticoids on neurogenesis-associated brain diseases.

Generation of a novel transgenic rat model for tracing extracellular vesicles in body fluids.

Extracellular vesicles (EVs) play an important role in the transfer of biomolecules between cells. To elucidate the intercellular transfer fate of EVs in vivo, we generated a new transgenic (Tg) rat model using green fluorescent protein (GFP)-tagged human CD63. CD63 protein is highly enriched on EV membranes via trafficking into late endosomes and is often used as an EV marker. The new Tg rat line in which human CD63-GFP is under control of the CAG promoter exhibited high expression of GFP in various body tissues. Exogenous human CD63-GFP was detected on EVs isolated from three body fluids of the Tg rats: blood serum, breast milk and amniotic fluid. In vitro culture allowed transfer of serum-derived CD63-GFP EVs into recipient rat embryonic fibroblasts, where the EVs localized in endocytic organelles. These results suggested that this Tg rat model should provide significant information for understanding the intercellular transfer and/or mother-child transfer of EVs in vivo.

Chronic glucocorticoid exposure suppressed the differentiation and survival of embryonic neural stem/progenitor cells: Possible involvement of ERK and PI3K/Akt signaling in the neuronal differentiation.

Growing evidence suggests that excess glucocorticoids (GCs) exposure during the pregnancy results in behavioral abnormality in offspring. Although research using animal models has demonstrated that systemic GCs treatment impairs development of fetal brain, direct impact of GCs on the phenotype of embryonic neural stem/progenitor cells (eNSPCs) and its mechanism has not been fully understood. Here, we investigated the effect of chronic GCs exposure on cell proliferation, differentiation, and survival of eNSPCs in vitro. Corticosterone (CORT, a murine GC) treatment did not affect the proliferation of eNSPCs. On the other hand, decreased expression of neuronal, synaptic, and astroglial marker proteins were observed when the differentiation of eNSPCs was induced in the presence of CORT. CORT also reduced the survival rate of eNSPCs after the differentiation. Moreover, CORT inhibited extracellular signal-regulated kinase (ERK) and phosphatidylinositol 3-kinase/Akt (PI3K/Akt) signaling pathways, which were activated during cell differentiation of eNSPCs. Inhibiting these signaling pathways reduced neural differentiation and eNSPCs viability, indicating their essential roles in the eNSPCs differentiation. Furthermore, IGF-I, a potent PI3K/Akt and ERK signaling stimulator, partially restored the adverse effect of CORT on eNSPCs, suggesting a possible involvement of the repression of these intracellular signaling in the GCs-caused eNSPCs impairment.

Negative regulation of microRNA-132 in expression of synaptic proteins in neuronal differentiation of embryonic neural stem cells.

MicroRNAs (miRs) play important roles in neuronal differentiation, maturation, and synaptic function in the central nervous system. They have also been suggested to be implicated in the pathogenesis of neurodegenerative and psychiatric diseases. Although miR-132 is one of the well-studied brain-specific miRs, which regulates synaptic structure and function in the postnatal brain, its function in the prenatal brain is still unclear. Here, we investigated miR-132 function during differentiation of rat embryonic neural stem cells (eNSCs). We found that miR-132 expression significantly increased during the fetal rat brain development and neural differentiation of eNSCs in vitro. Furthermore, miR-132 expression was increased during differentiation through MAPK/ERK1/2 pathway. Inhibition of ERK1/2 activation resulted in increased levels of synaptic proteins including PSD-95, GluR1 and synapsin I. Silencing of miR-132 also increased PSD-95 and GluR1. Considering that miR-132 increases synaptic proteins in differentiated cortical neurons, our result shows a novel function of miR-132 as a negative regulator for synaptic maturation in the neuronal differentiation of eNSCs.