Antitumor Effects of an Anthocyanin-Rich Grain Diet in a Mouse Model of Lewis Lung Carcinoma.

Functional foods enriched with plant polyphenol anthocyanins attract particular attention due to their health-promoting properties, including antitumor activity. We evaluated the effects of a grain diet rich in anthocyanins in a mouse model of Lewis lung carcinoma. Mice of the C57BL/6 strain were fed with wheat of near-isogenic lines differing in the anthocyanin content for four months prior to tumor transplantation. Although a significant decrease in the size of the tumor and the number of metastases in the lungs was revealed in the groups with both types of grain diet, the highest percentage of animals without metastases and with attenuated cell proliferation in the primary tumor were observed in the mice with the anthocyanin-rich diet. Both grain diets reduced the body weight gain and spleen weight index. The antitumor effects of the grain diets were associated with the activation of different mechanisms: immune response of the allergic type with augmented interleukin(IL)-9 and eotaxin serum levels in mice fed with control grain vs. inhibition of the IL-6/LIF system accompanied by a decrease in the tumor-associated M2 macrophage marker arginase 1 gene mRNA levels and enhanced autophagy in the tumor evaluated by the mRNA levels of Beclin 1 gene. Thus, anthocyanin-rich wheat is suggested as a promising source of functional nutrition with confirmed in vivo antitumor activity.

Zebrafish models of impulsivity and impulse control disorders.

Impulse control disorders (ICDs) are characterized by generalized difficulty controlling emotions and behaviors. ICDs are a broad group of the central nervous system (CNS) disorders including conduct disorder, intermittent explosive, oppositional-defiant disorder, antisocial personality disorder, kleptomania, pyromania and other illnesses. Although they all share a common feature (aberrant impulsivity), their pathobiology is complex and poorly understood. There are also currently no ICD-specific therapies to treat these illnesses. Animal models are a valuable tool for studying ICD pathobiology and potential therapies. The zebrafish (Danio rerio) has become a useful model organism to study CNS disorders due to high genetic and physiological homology to mammals, and sensitivity to various pharmacological and genetic manipulations. Here, we summarize experimental models of impulsivity and ICD in zebrafish and highlight their growing translational significance. We also emphasize the need for further development of zebrafish ICD models to improve our understanding of their pathogenesis and to search for novel therapeutic treatments.

Understanding neurobehavioral genetics of zebrafish.

Due to its fully sequenced genome, high genetic homology to humans, external fertilization, fast development, transparency of embryos, low cost and active reproduction, the zebrafish (Danio rerio) has become a novel promising model organism in biomedicine. Zebrafish are a useful tool in genetic and neuroscience research, including linking various genetic mutations to brain mechanisms using forward and reverse genetics. These approaches have produced novel models of rare genetic CNS disorders and common brain illnesses, such as addiction, aggression, anxiety and depression. Genetically modified zebrafish also foster neuroanatomical studies, manipulating neural circuits and linking them to different behaviors. Here, we discuss recent advances in neurogenetics of zebrafish, and evaluate their unique strengths, inherent limitations and the rapidly growing potential for elucidating the conserved roles of genes in neuropsychiatric disorders.

Effects of Grape Polyphenols on the Life Span and Neuroinflammatory Alterations Related to Neurodegenerative Parkinson Disease-Like Disturbances in Mice.

Functional nutrition is a valuable supplementation to dietary therapy. Functional foods are enriched with biologically active substances. Plant polyphenols attract particular attention due to multiple beneficial properties attributed to their high antioxidant and other biological activities. We assessed the effect of grape polyphenols on the life span of C57BL/6 mice and on behavioral and neuroinflammatory alterations in a transgenic mouse model of Parkinson disease (PD) with overexpression of the A53T-mutant human α-synuclein. C57BL/6 mice were given a dietary supplement containing grape polyphenol concentrate (GPC-1.5 mL/kg/day) with drinking water from the age of 6-8 weeks for life. Transgenic PD mice received GPC beginning at the age of 10 weeks for four months. GPC significantly influenced the cumulative proportion of surviving and substantially augmented the average life span in mice. In the transgenic PD model, the grape polyphenol (GP) diet enhanced memory reconsolidation and diminished memory extinction in a passive avoidance test. Behavioral effects of GP treatment were accompanied by a decrease in α-synuclein accumulation in the frontal cortex and a reduction in the expression of neuroinflammatory markers (IBA1 and CD54) in the frontal cortex and hippocampus. Thus, a GP-rich diet is recommended as promising functional nutrition for aging people and patients with neurodegenerative disorders.

Zebrafish models for personalized psychiatry: Insights from individual, strain and sex differences, and modeling gene x environment interactions.

Currently becoming widely recognized, personalized psychiatry focuses on unique physiological and genetic profiles of patients to best tailor their therapy. However, the role of individual differences, as well as genetic and environmental factors, in human psychiatric disorders remains poorly understood. Animal experimental models are a valuable tool to improve our understanding of disease pathophysiology and its molecular mechanisms. Due to high reproduction capability, fully sequenced genome, easy gene editing, and high genetic and physiological homology with humans, zebrafish (Danio rerio) are emerging as a novel powerful model in biomedicine. Mounting evidence supports zebrafish as a useful model organism in CNS research. Robustly expressed in these fish, individual, strain, and sex differences shape their CNS responses to genetic, environmental, and pharmacological manipulations. Here, we discuss zebrafish as a promising complementary translational tool to further advance patient-centered personalized psychiatry.

Modulation of the expression of genes related to the system of amyloid-beta metabolism in the brain as a novel mechanism of ceftriaxone neuroprotective properties.

BACKGROUND: The dominant hypothesis about the pathogenesis of Alzheimer's disease (AD) is the "amyloid cascade" concept and modulating the expression of proteins involved in the metabolism of amyloid-beta (Aß) is proposed as an effective strategy for the prevention and therapy of AD. Recently, we found that an antibiotic ceftriaxone (CEF), which possesses neuroprotective activity, reduced cognitive deficits and neurodegenerative changes in OXYS rats, a model of sporadic AD. The molecular mechanisms of this effect are not completely clear, we suggested that the drug might serve as the regulator of the expression of the genes involved in the metabolism of Aß and the pathogenesis of AD. The study was aimed to determine the effects of CEF on mRNA levels of Bace1 (encoding ß-secretase BACE1 involved in Aß production), Mme, Ide, Ece1, Ace2 (encoding enzymes involved in Aß degradation), Epo (encoding erythropoietin related to endothelial function and clearance of Aß across the blood brain barrier) in the frontal cortex, hippocampus, striatum, hypothalamus, and amygdala of OXYS and Wistar (control strain) male rats. Starting from the age of 14 weeks, animals received CEF (100 mg/kg/day, i.p., 36 days) or saline. mRNA levels were evaluated with RT-qPCR method. Biochemical parameters of plasma were measured for control of system effects of the treatment. RESULTS: To better understand strain variations studied here, we compared the gene expression between untreated OXYS and Wistar rats. This comparison showed a significant decrease in mRNA levels of Ace2 in the frontal cortex and hypothalamus, and of Actb in the amygdala of untreated OXYS rats. Analysis of potential effects of CEF revealed its novel targets. In the compound-treated OXYS cohort, CEF diminished mRNA levels of Bace1 and Ace2 in the hypothalamus, and Aktb in the frontal cortex. Furthermore, CEF augmented Mme, Ide, and Epo mRNA levels in the amygdala as well as the levels of Ece1 and Aktb in the striatum. Finally, CEF also attenuated the activity of ALT and AST in plasma of OXYS rats. CONCLUSION: Those findings disclosed novel targets for CEF action that might be involved into neuroprotective mechanisms at early, pre-plaque stages of AD-like pathology development.

NKCC1 mediates traumatic brain injury-induced hippocampal neurogenesis through CREB phosphorylation and HIF-1α expression.

Traumatic brain injury (TBI) is one of the most prevalent causes of worldwide mortality and morbidity. We previously had evidenced that TBI induced Na-K-2Cl co-transporter (NKCC1) upregulation in hippocampus. Here, we aim to investigate the role of NKCC1 in TBI-induced neurogenesis and the detailed mechanisms. The TBI-associated alternations in the expression of NKCC1, HIF-1α, VEGF, MAPK cascade, and CREB phosphorylation were analyzed by Western blot. TBI-induced neurogenesis was determined by immuno-fluorescence labeling. Chromatin immunoprecipitation was used to elucidate whether HIF-1α would activate VEGF gene after TBI. We found that the level of hippocampal NKCC1 and VEGF began to rise 8 h after TBI, and both of them reached maxima at day 7. Along with the upregulation of NKCC1 and VEGF, MAPK cascade was activated and hippocampal neurogenesis was promoted. Administration of CREB antisense oligonucleotide significantly attenuated the expression of HIF-1α, while HIF-1α antisense oligonucleotide exhibited little effect on the expression of CREB. However, HIF-1α antisense oligonucleotide administration did effectively suppress the expression of VEGF. Our results of the chromosome immunoprecipitation also indicated that HIF-1α could directly act on the VEGF promoter and presumably would elevate the VEGF expression after TBI. All these results have illustrated the correlation between NKCC1 upregulation and TBI-associated neurogenesis. The pathway involves the activation of Raf/MEK/ERK cascade, CREB phosphorylation, and HIF-1α upregulation, and finally leads to the stimulation of VEGF expression and the induction of neurogenesis.

Improving Bone Microarchitecture in Aging with Diosgenin Treatment: A Study in Senescence-Accelerated OXYS Rats.

Osteoporosis is a major disease associated with aging. We have previously demonstrated that diosgenin prevents osteoporosis in both menopause and D-galactose-induced aging rats. OXYS rats reveal an accelerated senescence and are used as a suitable model of osteoporosis. The aim of the present study was to analyze microarchitecture and morphological changes in femur of OXYS rats using morphological tests and microcomputed tomography scanning, and to evaluate the effects of oral administration of diosgenin at 10 and 50 mg/kg/day on femur in OXYS rats. The result showed that, compared with age-matched Wistar rats, the femur of OXYS rats revealed lower bone length, bone weight, bone volume, frame volume, frame density, void volume, porosity, external and internal diameters, cortical bone area, BV/TV, Tb.N, and Tb.Th, but higher Tb.Sp. Eight weeks of diosgenin treatment decreased porosity and Tb.Sp, but increased BV/TV, cortical bone area, Tb.N and bone mineral density, compared with OXYS rats treated with vehicle. These data reveal that microarchitecture and morphological changes in femur of OXYS rats showed osteoporotic aging features and suggest that diosgenin may have beneficial effects on aging-induced osteoporosis.

Effects of chronic treatment with diosgenin on bone loss in a D-galactose-induced aging rat model.

D-galactose is known to cause oxidative stress and induce aging-related diseases. Our previous study demonstrated that diosgenin can prevent osteoporosis in menopausal rats. The aim of the present study was to determine the effects of oral administration of diosgenin on bone loss in a D-galactose-induced aging rat model. Three groups of twelve-week-old male Wistar rats received a daily injection of D-galactose (150 mg/kg/day, i.p.) and orally administered diosgenin (0, 10, or 50 mg/kg/day) for eight weeks, while a control group received saline injection (1 ml/kg/day, i.p.), then the femurs were taken to measure mechanical and morphological properties. The results showed that frame volume and femur volume decreased and porosity and frame density increased in the D-galactose-induced aging rats compared to controls and that these effects were prevented by co-administration of diosgenin. This suggests that diosgenin might prevent bone loss during aging and provide beneficial effects in osteoporosis in the elderly.

Social isolation during peri-adolescence or adulthood: effects on sexual motivation, testosterone and corticosterone response under conditions of sexual arousal in male rats.

Reproductive functions in adult organism are known to be affected by different factors. Effects of social environment at the postnatal ontogenesis attract particular attention since it has deep impact on the development of physiological and emotional state of an individual. Effects of chronic social isolation at different ages on male sexual motivation, testosterone and corticosterone response under conditions of sexual arousal were studied in Wistar rats. After weaning at the 21st [corrected] day of age, rats of one group were isolated for six weeks and after that they were housed in groups of five per cage for ten weeks (Iso3-9). Rats of the second group were housed in groups of five animals per cage till 13 weeks of age, and then they were isolated for six weeks (Iso13-19). Rats of the control group were housed in groups during the experiment. Adult 19 week- old male rats were tested under conditions of sexual arousal. The expression of sexual motivation was estimated as the behavioral activity of a male at the transparent perforated partition separating a receptive female. Isolation of adult male rats reduced the number of approaches to the partition, while the period of time a male spent at the partition was not changed and testosterone response was enhanced as compared to control rats. Chronic social isolation during peri-adolescence reduced sexual motivation and prevented arousal-induced elevation of testosterone. Plasma corticosterone increases at sexual arousal in the two groups of isolated rats did not differ from that in controls. Our results are evidence that social isolation during the post-maturity stage (Iso13-19) did not diminish the manifestation of sexual motivation and hormonal response to a receptive female, while isolation during peri-adolescence attenuated behavioral and hormonal expression of sexual arousal in adult males.

Understanding CNS Effects of Antimicrobial Drugs Using Zebrafish Models.

Antimicrobial drugs represent a diverse group of widely utilized antibiotic, antifungal, antiparasitic and antiviral agents. Their growing use and clinical importance necessitate our improved understanding of physiological effects of antimicrobial drugs, including their potential effects on the central nervous system (CNS), at molecular, cellular, and behavioral levels. In addition, antimicrobial drugs can alter the composition of gut microbiota, and hence affect the gut-microbiota-brain axis, further modulating brain and behavioral processes. Complementing rodent studies, the zebrafish (Danio rerio) emerges as a powerful model system for screening various antimicrobial drugs, including probing their putative CNS effects. Here, we critically discuss recent evidence on the effects of antimicrobial drugs on brain and behavior in zebrafish, and outline future related lines of research using this aquatic model organism.

Comparative analysis of early neurodegeneration signs in a mouse model of Alzheimer's disease-like pathology induced by two types of the central (Intracerebroventricular vs. Intrahippocampal) administration of Aß25-35 oligomers.

Animal models of Alzheimer's disease (AD) induced by intracerebroventricular (ICV) or intrahippocampal (IH) administration of amyloid-beta (Aß) are widely used in current research. It remains unclear whether these models provide similar outcomes or mimic pathological mechanisms of AD equally. The aim of the work was to compare two models induced by ICV or IH administration of Aß25-35 oligomers to C57BL/6 mice. Parameters characterizing cognitive function (passive avoidance test), protein expression (IBA1, Aß, LC3-II) and expression of genes for neuroinflammation (Aif1, Lcn2, Nrf2), autophagy (Atg8, Becn1, Park2), or markers of neurodegeneration (Cst3, Insr, Vegfa) were analyzed. Сognitive deficits, amyloid accumulation, and neuroinflammatory response in the brain evaluated by the microglial activation were similar in both models. Thus, both ways of Aß administration appear to be equally suitable for modelling AD-like pathology in mice. Our findings strongly support the key role of Aß load and neuroinflammatory response in the hippocampus and frontal cortex for the progression of AD-like pathology and development of cognitive deficits. There were certain minor differences between the models in the mRNA level of genes involved in the processes of neuroinflammation, neurodegeneration, and autophagy. Modulating effects of the central administration of Aß25-35 on the mRNA expression of Aif1, Lcn2, Park2, and Vegfa genes in different brain structures were revealed. The effects occurred to be more pronounced with the ICV method compared with the IH method. These findings give insight into the processes at initial stages of Aß-induced pathology depending on a primary location of Aß oligomers in the brain.

Single-cell electric lysis on an electroosmotic-driven microfluidic chip with arrays of microwells.

Accurate analysis at the single-cell level has become a highly attractive tool for investigating cellular content. An electroosmotic-driven microfluidic chip with arrays of 30-µm-diameter microwells was developed for single-cell electric lysis in the present study. The cellular occupancy in the microwells when the applied voltage was 5 V (82.4%) was slightly higher than that at an applied voltage of 10 V (81.8%). When the applied voltage was increased to 15 V, the cellular occupancy in the microwells dropped to 64.3%. More than 50% of the occupied microwells contain individual cells. The results of electric lysis experiments at the single-cell level indicate that the cells were gradually lysed as the DC voltage of 30 V was applied; the cell was fully lysed after 25 s. Single-cell electric lysis was demonstrated in the proposed microfluidic chip, which is suitable for high-throughput cell lysis.

Structural and functional characteristics of the hippocampus in depressive-like recipients after transplantation of in vitro caffeine-modulated immune cells.

The hippocampus is a key anatomical brain region associated with depression. On the other hand, immune cells and their releasing cytokines play an essential role in stress and depression. Noteworthy that the most of psychoactive drugs produce unidirectional effects on the cells of both nervous and immune systems. This suggests the immunotherapy for behavioral disorders based on the treatment with autologous immune cells in which functional activity was modulated ex vivo by a psychoactive drug. Here, we treated the immune cells of depressive-like mice in vitro with caffeine (100 µg per 15 × 106 cells). The effects of caffeine-treated immune cells transplantation on neuronal density, production of brain-derived neurotrophic factor (BDNF) and a number of cytokines in the hippocampus of depressive-like syngeneic animals were studied. In depressive-like recipients, an increase in the density of pyramidal neurons in CA1 and CA3 hippocampal regions, accompanied with augmented level of BDNF, decreased levels of pro-inflammatory (IL-1ß, IL-6, INF-γ, and TNF-α) and increased levels of anti-inflammatory (IL-10 and IL-4) cytokines was found. The mechanisms of the revealed structural and functional alterations in the hippocampus of depressive-like recipients after transplantation of caffeine-treated immune cells are discussed.

Combined induction of mTOR-dependent and mTOR-independent pathways of autophagy activation as an experimental therapy for Alzheimer's disease-like pathology in a mouse model.

Alzheimer's disease (AD) is associated with amyloid-ß (Aß) accumulation that might be hindered by autophagy. There are two ways to induce autophagy: through mTOR-dependent and mTOR-independent pathways (here, by means of rapamycin and trehalose, respectively). The aim of this study was to evaluate the contribution of these pathways and their combination to the treatment of experimental AD. Mice were injected bilaterally intracerebroventricularly with an Aß fragment (25-35) to set up an AD model. Treatment with rapamycin (10 mg/kg, every other day), trehalose consumption with drinking water (2 mg/mL, ad libitum), or their combination started 2 days after the surgery and lasted for 2 weeks. Open-field, plus-maze, and passive avoidance tests were used for behavioral phenotyping. Neuronal density, Aß accumulation, and the expression of autophagy marker LC3-II and neuroinflammatory marker IBA1 were measured in the frontal cortex and hippocampus. mRNA levels of autophagy genes (Atg8, Becn1, and Park2) were assessed in the hippocampus. Trehalose but not rapamycin caused pronounced prolonged autophagy induction and transcriptional activation of autophagy genes. Both drugs effectively prevented Aß deposition and microglia activation. Autophagy inhibitor 3-methyladenine significantly attenuated autophagy activation and disturbed the effect of the inducers on Aß load. The inducers substantially reversed behavioral and neuronal deficits in Aß-injected mice. In many cases, the best outcomes were achieved with the combined treatment. Thus, trehalose alone or combined autophagy activation by the two inducers may be a promising treatment approach to AD-like neurodegeneration. Some aspects of interaction between mTOR-dependent and mTOR-independent pathways of autophagy are discussed.

Trehalose Activates Hepatic and Myocardial Autophagy and Has Anti-Inflammatory Effects in db/db Diabetic Mice.

Db/db mice (carrying a mutation in the gene encoding leptin receptor) show autophagy suppression. Our aim was to evaluate the effect of autophagy inducer trehalose on liver and heart autophagy in db/db mice and to study inflammation dysregulation and the suitability of chitinases' expression levels as diabetes markers. Thirty-eight male db/db mice and C57/BL mice (control) were used. The db/db model manifested inflammation symptoms: overexpression of TNF-α in the spleen and underexpression of IL-10 in the liver and spleen (cytokine imbalance). Simultaneously, we revealed decreased expression of chitotriosidase (CHIT1) and acid mammalian chitinase (CHIA) in the liver of db/db mice. CHIA expression in db/db mice is significantly lower only in the spleen. Trehalose treatment significantly reduced blood glucose concentration and glycated hemoglobin. Treatment of db/db mice by trehalose was followed by increased autophagy induction in the heart and liver (increased autolysosomes volume density studied by morphometric electron-microscopic method). Trehalose exerted beneficial cardiac effects possibly via increased lipophagy (uptake of lipid droplets). The autophagy activation by trehalose had several positive effects on the heart and liver of db/db mice; therefore, lipophagy activation seems to be a promising therapy for diabetes.

Understanding early-life pain and its effects on adult human and animal emotionality: Translational lessons from rodent and zebrafish models.

Critical for organismal survival, pain evokes strong physiological and behavioral responses in various sentient species. Clinical and preclinical (animal) studies markedly increase our understanding of biological consequences of developmental (early-life) adversity, as well as acute and chronic pain. However, the long-term effects of early-life pain exposure on human and animal emotional responses remain poorly understood. Here, we discuss experimental models of nociception in rodents and zebrafish, and summarize mounting evidence of the role of early-life pain in shaping emotional traits later in life. We also call for further development of animal models to probe the impact of early-life pain exposure on behavioral traits, brain disorders and novel therapeutic treatments.

Using zebrafish (Danio rerio) models to understand the critical role of social interactions in mental health and wellbeing.

Social behavior represents a beneficial interaction between conspecifics that is critical for maintaining health and wellbeing. Dysfunctional or poor social interaction are associated with increased risk of physical (e.g., vascular) and psychiatric disorders (e.g., anxiety, depression, and substance abuse). Although the impact of negative and positive social interactions is well-studied, their underlying mechanisms remain poorly understood. Zebrafish have well-characterized social behavior phenotypes, high genetic homology with humans, relative experimental simplicity and the potential for high-throughput screens. Here, we discuss the use of zebrafish as a candidate model organism for studying the fundamental mechanisms underlying social interactions, as well as potential impacts of social isolation on human health and wellbeing. Overall, the growing utility of zebrafish models may improve our understanding of how the presence and absence of social interactions can differentially modulate various molecular and physiological biomarkers, as well as a wide range of other behaviors.

Modeling neurodegenerative disorders in zebrafish.

Neurodegeneration is a major cause of Alzheimer's, Parkinson's, Huntington's, multiple and amyotrophic lateral sclerosis, pontocerebellar hypoplasia, dementia and other related brain disorders. Their complex pathogenesis commonly includes genetic and neurochemical deficits, misfolded protein toxicity, demyelination, apoptosis and mitochondrial dysfunctions. Albeit differing in specific underlying mechanisms, neurodegenerative disorders typically display evolutionarily conserved mechanisms across taxa. Here, we review the role of zebrafish models in recapitulating major human and rodent neurodegenerative conditions, demonstrating this species as a highly relevant experimental model for research on neurodegenerative diseases, and discussing how these fish models can further clarify the underlying genetic, neurochemical, neuroanatomical and behavioral pathogenic mechanisms.

A Novel Laser-Based Zebrafish Model for Studying Traumatic Brain Injury and Its Molecular Targets.

Traumatic brain injury (TBI) is a major public health problem. Here, we developed a novel model of non-invasive TBI induced by laser irradiation in the telencephalon of adult zebrafish (Danio rerio) and assessed their behavior and neuromorphology to validate the model and evaluate potential targets for neuroreparative treatment. Overall, TBI induced hypolocomotion and anxiety-like behavior in the novel tank test, strikingly recapitulating responses in mammalian TBI models, hence supporting the face validity of our model. NeuN-positive cell staining was markedly reduced one day, but not seven days, after TBI, suggesting increased neuronal damage immediately after the injury, and its fast recovery. The brain-derived neurotrophic factor (Bdnf) level in the brain dropped immediately after the trauma, but fully recovered seven days later. A marker of microglial activation, Iba1, was elevated in the TBI brain, albeit decreasing from Day 3. The levels of hypoxia-inducible factor 1-alpha (Hif1a) increased 30 min after the injury, and recovered by Day 7, further supporting the construct validity of the model. Collectively, these findings suggest that our model of laser-induced brain injury in zebrafish reproduces mild TBI and can be a useful tool for TBI research and preclinical neuroprotective drug screening.