Metabolic alterations and cellular responses to ß-Hydroxybutyrate treatment in breast cancer cells.

BACKGROUND: The ketogenic diet (KD), based on high fat (over 70% of daily calories), low carbohydrate, and adequate protein intake, has become popular due to its potential therapeutic benefits for several diseases including cancer. Under KD and starvation conditions, the lack of carbohydrates promotes the production of ketone bodies (KB) from fats by the liver as an alternative source of metabolic energy. KD and starvation may affect the metabolism in cancer cells, as well as tumor characteristics. The aim of this study is to evaluate the effect of KD conditions on a wide variety of aspects of breast cancer cells in vitro. METHODS: Using two cancer and one non-cancer breast cell line, we evaluate the effect of ß-hydroxybutyrate (ßHb) treatment on cell growth, survival, proliferation, colony formation, and migration. We also assess the effect of KB on metabolic profile of the cells. Using RNAseq analysis, we elucidate the effect of ßHb on the gene expression profile. RESULTS: Significant effects were observed following treatment by ßHb which include effects on viability, proliferation, and colony formation of MCF7 cells, and different effects on colony formation of MDA-MB-231 cells, with no such effects on non-cancer HB2 cells. We found no changes in glucose intake or lactate output following ßHb treatment as measured by LC-MS, but an increase in reactive oxygen species (ROS) level was detected. RNAseq analysis demonstrated significant changes in genes involved in lipid metabolism, cancer, and oxidative phosphorylation. CONCLUSIONS: Based on our results, we conclude that differential response of cancer cell lines to ßHb treatment, as alternative energy source or signal to alter lipid metabolism and oncogenicity, supports the need for a personalized approach to breast cancer patient treatment.

Targeted Modulation of Interferon Response-Related Genes with IFN-Alpha/Lambda Inhibition.

Interferon (IFN) signaling resulting from external or internal inflammatory processes initiates the rapid release of cytokines and chemokines to target viral or bacterial invasion, as well as cancer and other diseases. Prolonged exposure to IFNs, or the overexpression of other cytokines, leads to immune exhaustion, enhancing inflammation and leading to the persistence of infection and promotion of disease. Hence, to control and stabilize an excessive immune response, approaches for the management of inflammation are required. The potential use of peptides as anti-inflammatory agents has been previously demonstrated. Our team discovered, and previously published, a 9-amino-acid cyclic peptide named ALOS4 which exhibits anti-cancer properties in vivo and in vitro. We suggested that the anti-cancer effect of ALOS4 arises from interaction with the immune system, possibly through the modulation of inflammatory processes. Here, we show that treatment with ALOS4 decreases basal cytokine levels in mice with chronic inflammation and prolongs the lifespan of mice with acute systemic inflammation induced by irradiation. We also show that pretreatment with ALOS4 reduces the expression of IFN alpha, IFN lambda, and selected interferon-response genes triggered by polyinosinic-polycytidylic acid (Poly I:C), a synthetic analog of viral double-stranded RNA, while upregulating the expression of other genes with antiviral activity. Hence, we conclude that ALOS4 does not prevent IFN signaling, but rather supports the antiviral response by upregulating the expression of interferon-response genes in an interferon-independent manner.

Anti-Cancer Effects of Cyclic Peptide ALOS4 in a Human Melanoma Mouse Model.

We examined the effects of ALOS4, a cyclic peptide discovered previously by phage library selection against integrin αvß3, on a human melanoma (A375) xenograft model to determine its abilities as a potential anti-cancer agent. We found that ALOS4 promoted healthy weight gain in A375-engrafted nude mice and reduced melanoma tumor mass and volume. Despite these positive changes, examination of the tumor tissue did not indicate any significant effects on proliferation, mitotic index, tissue vascularization, or reduction of αSMA or Ki-67 tumor markers. Modulation in overall expression of critical downstream αvß3 integrin factors, such as FAK and Src, as well as reductions in gene expression of c-Fos and c-Jun transcription factors, indirectly confirmed our suspicions that ALOS4 is likely acting through an integrin-mediated pathway. Further, we found no overt formulation issues with ALOS4 regarding interaction with standard inert laboratory materials (polypropylene, borosilicate glass) or with pH and temperature stability under prolonged storage. Collectively, ALOS4 appears to be safe, chemically stable, and produces anti-cancer effects in a human xenograft model of melanoma. We believe these results suggest a role for ALOS4 in an integrin-mediated pathway in exerting its anti-cancer effects possibly through immune response modulation.

Aragonite-Polylysine: Neuro-Regenerative Scaffolds with Diverse Effects on Astrogliosis.

Biomaterials, especially when coated with adhesive polymers, are a key tool for restorative medicine, being biocompatible and supportive for cell adherence, growth, and function. Aragonite skeletons of corals are biomaterials that support survival and growth of a range of cell types, including neurons and glia. However, it is not known if this scaffold affects neural cell migration or elongation of neuronal and astrocytic processes, prerequisites for initiating repair of damage in the nervous system. To address this, hippocampal cells were aggregated into neurospheres and cultivated on aragonite skeleton of the coral Trachyphyllia geoffroyi (Coral Skeleton (CS)), on naturally occurring aragonite (Geological Aragonite (GA)), and on glass, all pre-coated with the oligomer poly-D-lysine (PDL). The two aragonite matrices promoted equally strong cell migration (4.8 and 4.3-fold above glass-PDL, respectively) and axonal sprouting (1.96 and 1.95-fold above glass-PDL, respectively). However, CS-PDL had a stronger effect than GA-PDL on the promotion of astrocytic processes elongation (1.7 vs. 1.2-fold above glass-PDL, respectively) and expression of the glial fibrillary acidic protein (3.8 vs. and 1.8-fold above glass-PDL, respectively). These differences are likely to emerge from a reaction of astrocytes to the degree of roughness of the surface of the scaffold, which is higher on CS than on GA. Hence, CS-PDL and GA-PDL are scaffolds of strong capacity to derive neural cell movements and growth required for regeneration, while controlling the extent of astrocytic involvement. As such, implants of PDL-aragonites have significant potential as tools for damage repair and the reduction of scar formation in the brain following trauma or disease.

TRAIN (Transcription of Repeats Activates INterferon) in response to chromatin destabilization induced by small molecules in mammalian cells.

Cellular responses to the loss of genomic stability are well-established, while how mammalian cells respond to chromatin destabilization is largely unknown. We previously found that DNA demethylation on p53-deficient background leads to transcription of repetitive heterochromatin elements, followed by an interferon response, a phenomenon we named TRAIN (Transcription of Repeats Activates INterferon). Here, we report that curaxin, an anticancer small molecule, destabilizing nucleosomes via disruption of histone/DNA interactions, also induces TRAIN. Furthermore, curaxin inhibits oncogene-induced transformation and tumor growth in mice in an interferon-dependent manner, suggesting that anticancer activity of curaxin, previously attributed to p53-activation and NF-kappaB-inhibition, may also involve induction of interferon response to epigenetic derepression of the cellular 'repeatome'. Moreover, we observed that another type of drugs decondensing chromatin, HDAC inhibitor, also induces TRAIN. Thus, we proposed that TRAIN may be one of the mechanisms ensuring epigenetic integrity of mammalian cells via elimination of cells with desilenced chromatin.

Role of Chromatin Damage and Chromatin Trapping of FACT in Mediating the Anticancer Cytotoxicity of DNA-Binding Small-Molecule Drugs.

Precisely how DNA-targeting chemotherapeutic drugs trigger cancer cell death remains unclear, as it is difficult to separate direct DNA damage from other effects in cells. Recent work on curaxins, a class of small-molecule drugs with broad anticancer activity, shows that they interfere with histone-DNA interactions and destabilize nucleosomes without causing detectable DNA damage. Chromatin damage caused by curaxins is sensed by the histone chaperone FACT, which binds unfolded nucleosomes becoming trapped in chromatin. In this study, we investigated whether classical DNA-targeting chemotherapeutic drugs also similarly disturbed chromatin to cause chromatin trapping of FACT (c-trapping). Drugs that directly bound DNA induced both chromatin damage and c-trapping. However, chromatin damage occurred irrespective of direct DNA damage and was dependent on how a drug bound DNA, specifically, in the way it bound chromatinized DNA in cells. FACT was sensitive to a plethora of nucleosome perturbations induced by DNA-binding small molecules, including displacement of the linker histone, eviction of core histones, and accumulation of negative supercoiling. Strikingly, we found that the cytotoxicity of DNA-binding small molecules correlated with their ability to cause chromatin damage, not DNA damage. Our results suggest implications for the development of chromatin-damaging agents as selective anticancer drugs.Significance: These provocative results suggest that the anticancer efficacy of traditional DNA-targeting chemotherapeutic drugs may be based in large part on chromatin damage rather than direct DNA damage. Cancer Res; 78(6); 1431-43. ©2018 AACR.

Effect of social interactions on hippocampal protein expression in animal dominant and submissive model of behavioral disorders.

PURPOSE: Psychiatric conditions, in many cases, arise from social interactions necessary for optimal mental functioning. Dominance and submissiveness are two opposite poles of behavior, stemming from processes of social interactions between members inside one group or species. Extreme dominance and submissiveness expressions in humans is accompanied by mental impairments, including mania and depression. Here, taking advantage of animals bred selectively for traits of dominance and submissiveness, we assess protein expression profiles in dominant and submissive mice in the context of social interaction. EXPERIMENTAL DESIGN: Proteins extracted from hippocampi of naïve and social interaction subjected dominant, submissive and wild type mice (15 mice per each group) are quantified using label-free quantitative LC/MS/MS analysis. Complexity of social interaction-related protein expression is resolved by factor analysis and enriched with GO and protein-protein interaction functional network analyses. RESULTS: In total, 1146 proteins exhibiting expression changes in the wild type mice, as well as dominant and submissive mice are enriched in protein datasets responsible for: 1) socially triggered dominance (90 proteins), 2) inherent submissiveness (75 proteins), 3) socially triggered submissiveness (117 proteins), and 4) social interaction triggered protein expression changes, related to resilience/adaptation to stress (69 proteins). Among the most enriched categories, extensive changes are found in proteins related to presynaptic release, ion channel regulation, circadian rhythm, MAPK, ErbB and NF-kB pathways. CONCLUSION: Data extracted from this first extensive proteomic study of a social interaction paradigm may facilitate decoding of molecular mechanisms responsible for pathogenesis of psychiatric disorders.

FACT is a sensor of DNA torsional stress in eukaryotic cells.

Transitions of B-DNA to alternative DNA structures (ADS) can be triggered by negative torsional strain, which occurs during replication and transcription, and may lead to genomic instability. However, how ADS are recognized in cells is unclear. We found that the binding of candidate anticancer drug, curaxin, to cellular DNA results in uncoiling of nucleosomal DNA, accumulation of negative supercoiling and conversion of multiple regions of genomic DNA into left-handed Z-form. Histone chaperone FACT binds rapidly to the same regions via the SSRP1 subunit in curaxin-treated cells. In vitro binding of purified SSRP1 or its isolated CID domain to a methylated DNA fragment containing alternating purine/pyrimidines, which is prone to Z-DNA transition, is much stronger than to other types of DNA. We propose that FACT can recognize and bind Z-DNA or DNA in transition from a B to Z form. Binding of FACT to these genomic regions triggers a p53 response. Furthermore, FACT has been shown to bind to other types of ADS through a different structural domain, which also leads to p53 activation. Thus, we propose that FACT acts as a sensor of ADS formation in cells. Recognition of ADS by FACT followed by a p53 response may explain the role of FACT in DNA damage prevention.

Fast detection of deletion breakpoints using quantitative PCR.

The routine detection of large and medium copy number variants (CNVs) is well established. Hemizygotic deletions or duplications in the large Duchenne muscular dystrophy DMD gene responsible for Duchenne and Becker muscular dystrophies are routinely identified using multiple ligation probe amplification and array-based comparative genomic hybridization. These methods only map deleted or duplicated exons, without providing the exact location of breakpoints. Commonly used methods for the detection of CNV breakpoints include long-range PCR and primer walking, their success being limited by the deletion size, GC content and presence of DNA repeats. Here, we present a strategy for detecting the breakpoints of medium and large CNVs regardless of their size. The hemizygous deletion of exons 45-50 in the DMD gene and the large autosomal heterozygous PARK2 deletion were used to demonstrate the workflow that relies on real-time quantitative PCR to narrow down the deletion region and Sanger sequencing for breakpoint confirmation. The strategy is fast, reliable and cost-efficient, making it amenable to widespread use in genetic laboratories.

Novel synthetic cyclic integrin αvß3 binding peptide ALOS4: Antitumor activity in mouse melanoma models.

ALOS4, a unique synthetic cyclic peptide without resemblance to known integrin ligand sequences, was discovered through repeated biopanning with pIII phage expressing a disulfide-constrained nonapeptide library. Binding assays using a FITC-labeled analogue demonstrated selective binding to immobilized αvß3 and a lack of significant binding to other common proteins, such as bovine serum albumin and collagen. In B16F10 cell cultures, ALOS4 treatment at 72 h inhibited cell migration (30%) and adhesion (up to 67%). Immunofluorescent imaging an ALOS4-FITC analogue with B16F10 cells demonstrated rapid cell surface binding, and uptake and localization in the cytoplasm. Daily injections of ALOS4 (0.1, 0.3 or 0.5 mg/kg i.p.) to mice inoculated with B16F10 mouse melanoma cells in two different cancer models, metastatic and subcutaneous tumor, resulted in reduction of lung tumor count (metastatic) and tumor mass (subcutaneous) and increased survival of animals monitored to 45 and 60 days, respectively. Examination of cellular activity indicated that ALOS4 produces inhibition of cell migration and adhesion in a concentration-dependent manner. Collectively, these results suggest that ALOS4 is a structurally-unique selective αvß3 integrin ligand with potential anti-metastatic activity.

Dynamics of Hippocampal Protein Expression During Long-term Spatial Memory Formation.

Spatial memory depends on the hippocampus, which is particularly vulnerable to aging. This vulnerability has implications for the impairment of navigation capacities in older people, who may show a marked drop in performance of spatial tasks with advancing age. Contemporary understanding of long-term memory formation relies on molecular mechanisms underlying long-term synaptic plasticity. With memory acquisition, activity-dependent changes occurring in synapses initiate multiple signal transduction pathways enhancing protein turnover. This enhancement facilitates de novo synthesis of plasticity related proteins, crucial factors for establishing persistent long-term synaptic plasticity and forming memory engrams. Extensive studies have been performed to elucidate molecular mechanisms of memory traces formation; however, the identity of plasticity related proteins is still evasive. In this study, we investigated protein turnover in mouse hippocampus during long-term spatial memory formation using the reference memory version of radial arm maze (RAM) paradigm. We identified 1592 proteins, which exhibited a complex picture of expression changes during spatial memory formation. Variable linear decomposition reduced significantly data dimensionality and enriched three principal factors responsible for variance of memory-related protein levels at (1) the initial phase of memory acquisition (165 proteins), (2) during the steep learning improvement (148 proteins), and (3) the final phase of the learning curve (123 proteins). Gene ontology and signaling pathways analysis revealed a clear correlation between memory improvement and learning phase-curbed expression profiles of proteins belonging to specific functional categories. We found differential enrichment of (1) neurotrophic factors signaling pathways, proteins regulating synaptic transmission, and actin microfilament during the first day of the learning curve; (2) transcription and translation machinery, protein trafficking, enhancement of metabolic activity, and Wnt signaling pathway during the steep phase of memory formation; and (3) cytoskeleton organization proteins. Taken together, this study clearly demonstrates dynamic assembly and disassembly of protein-protein interaction networks depending on the stage of memory formation engrams.

Blood BDNF level is gender specific in severe depression.

Though the role of brain derived neurotrophic factor (BDNF) as a marker for major depressive disorder (MDD) and antidepressant efficacy has been widely studied, the role of BDNF in distinct groups of patients remains unclear. We evaluated the diagnostic value of BDNF as a marker of disease severity measured by HAM-D scores and antidepressants efficacy among MDD patients. Fifty-one patients who met DSM-IV criteria for MDD and were prescribed antidepressants and 38 controls participated in this study. BDNF in serum was measured at baseline, 1st, 2nd and 8th treatment weeks. Depression severity was evaluated using the Hamilton Rating Scale for Depression (HAM-D). BDNF polymorphism rs6265 (val66met) was genotyped. We found a positive correlation between blood BDNF levels and severity of depression only among untreated women with severe MDD (HAM-D>24). Serum BDNF levels were lower in untreated MDD patients compared to control group. Antidepressants increased serum BDNF levels and reduced between-group differences after two weeks of treatment. No correlations were observed between BDNF polymorphism, depression severity, duration of illness, age and BDNF serum levels. Further supporting the role of BDNF in the pathology and treatment of MDD, we suggest that it should not be used as a universal biomarker for diagnosis of MDD in the general population. However, it has diagnostic value for the assessment of disease progression and treatment efficacy in individual patients.

Synapsin IIb as a functional marker of submissive behavior.

Dominance and submissiveness are important functional elements of the social hierarchy. By employing selective breeding based on a social interaction test, we developed mice with strong and stable, inheritable features of dominance and submissiveness. In order to identify candidate genes responsible for dominant and submissive behavior, we applied transcriptomic and proteomic studies supported by molecular, behavioral and pharmacological approaches. We clearly show here that the expression of Synapsin II isoform b (Syn IIb) is constitutively upregulated in the hippocampus and striatum of submissive mice in comparison to their dominant and wild type counterparts. Moreover, the reduction of submissive behavior achieved after mating and delivery was accompanied by a marked reduction of Syn IIb expression. Since submissiveness has been shown to be associated with depressive-like behavior, we applied acute SSRI (Paroxetine) treatment to reduce submissiveness in studied mice. We found that reduction of submissive behavior evoked by Paroxetine was paired with significantly decreased Syn IIb expression. In conclusion, our findings indicate that submissiveness, known to be an important element of depressive-like behavioral abnormalities, is strongly linked with changes in Syn IIb expression.

Early onset of cognitive impairment is associated with altered synaptic plasticity and enhanced hippocampal GluA1 expression in a mouse model of depression.

Memory deficit is a common manifestation of age-related cognitive impairment, of which depression is a frequently occurring comorbidity. Previously, we developed a submissive (Sub) mouse line, validated as a model of depressive-like behavior. Using learning paradigms testing hippocampus-dependent spatial and nonspatial memory, we demonstrate here that Sub mice developed cognitive impairments at earlier age (3 months), compared with wild-type mice. Furthermore, acute hippocampal slices from Sub animals failed to display paired-pulse facilitation, whereas primed burst stimulation elicited significantly enhanced long-term potentiation in region CA1, relative to control mice. Changes in synaptic plasticity were accompanied by markedly reduced hippocampal messenger RNA expression of insulin-like growth factor and brain-derived neurotrophic factor. Finally, we identified markedly elevated protein levels of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunit GluA1 in the hippocampi of Sub mice, which was exacerbated with age. Taken together, the results point to a linkage between depressive-like behavior and the susceptibility to develop age-related cognitive impairment, potentially by hippocampal α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor-mediated glutamatergic signaling.

Exploring diazepam's effect on hemodynamic responses of mouse brain tissue by optical spectroscopic imaging.

In this study, a simple duel-optical spectroscopic imaging apparatus capable of simultaneously determining relative changes in brain oxy-and deoxy-hemoglobin concentrations was used following administration of the anxiolytic compound diazepam in mice with strong dominant (Dom) and submissive (Sub) behavioral traits. Three month old mice (n = 30) were anesthetized and after 10 min of baseline imaging, diazepam (1.5 mg/kg) was administered and measurements were taken for 80 min. The mouse head was illuminated by white light based LED's and diffused reflected light passing through different channels, consisting of a bandpass filter and a CCD camera, respectively, was collected and analyzed to measure the hemodynamic response. This work's major findings are threefold: first, Dom and Sub animals showed statistically significant differences in hemodynamic response to diazepam administration. Secondly, diazepam was found to more strongly affect the Sub group. Thirdly, different time-series profiles were observed post-injection, which can serve as a possible marker for the groups' differentiation. To the best of our knowledge, this is the first report on the effects of an anxiolytic drug on brain hemodynamic responses in mice using diffused light optical imaging.

Chronic food administration of Salvia sclarea oil reduces animals' anxious and dominant behavior.

Recent studies indicate that an oil extract from Salvia sclarea may provide clinical benefits in various pathological conditions. In comparison to extracts from other Salvia species, S. sclarea oil contains twice as much omega-3 fatty acids, which are involved in eicosanoid synthesis pathways, and has been found to contain significant levels of the psychoactive monoterpane linalool. In the present study, we examined the mood stabilizing and anxiolytic-like effects of chronic food administration of S. sclarea oil extract on behavioral and physiological parameters of mice with prominent dominant and submissive features in behavioral assays used to test mood stabilizing and antidepressant drugs. Experimental animals received oil supplemented food from the age of 4 weeks or from conception via their pregnant dams. Each age group received either S. sclarea oil- or sunflower oil-enriched feed. Dominant animals, whose pregnant mothers received S. sclarea oil-enriched feed from the date of conception, showed a significant reduction of dominant and anxiety-like behavior, in comparison to their sunflower oil-treated counterparts. S. sclarea oil-treated submissive animals exhibited a similar tendency, and showed a significant reduction in blood corticosterone levels. These findings enforce the hypothesis that S. sclarea oil possesses anxiolytic properties.

Differential responses to distinct psychotropic agents of selectively bred dominant and submissive animals.

Dominance and submissiveness are two opposite poles of behavior representing important functional elements in the development of social interactions. We previously demonstrated the inheritability of these traits by selective breeding based upon the dominant-submissive relationships (DSR) food competition paradigm. Continued multigenerational behavioral selection of Sabra mice yielded animal populations with strong and stable features of dominance and submissiveness. We found that these animals react differentially to stressogenic triggers, antidepressants and mood stabilizing agents. The anxiolytic compound diazepam (1.5mg/kg, i.p.) reduced anxiety-like behavior of submissive animals, but showed anxiogenic effects among dominant animals. In the Forced Swim test, the antidepressant paroxetine (1, 3 and 10mg/kg, i.p.) markedly reduced immobility of submissive animals, demonstrating antidepressant-like effect. In contrast, when administered to dominant animals, paroxetine caused extreme (frenetic) activity. The mood stabilizer lithium (0.4%, p.o.) selectively influenced dominant mice, without affecting the behavior of submissive animals. In summary, we describe here two distinct animal populations possessing strong dominant and submissive phenotypes. We suggest that these populations hold potential as tools for studying the molecular basis and pharmacogenetics of dominant and submissive behavior.

Diffuse near-infrared reflectance spectroscopy during heatstroke in a mouse model: pilot study.

Heatstroke, a form of hyperthermia, is a life-threatening condition characterized by an elevated core body temperature that rises above 40°C (104°F) and central nervous system dysfunction that results in delirium, convulsions, or coma. Without emergency treatment, the victim lapses into a coma and death soon follows. The study presented was conducted with a diffuse reflectance spectroscopy (DRS) setup to assess the effects of brain dysfunction that occurred during heatstroke in mice model (n=6). It was hypothesized that DRS can be utilized in small animal studies to monitor change in internal brain tissue temperature during heatstroke injury since it induces a sequence of pathologic changes that change the tissue composition and structure. Heatstroke was induced by exposure of the mice body under general anesthesia, to a high ambient temperature. A type of DRS in which the brain tissue was illuminated through the intact scalp with a broadband light source and diffuse reflected spectra was employed, taking in the spectral region between 650 and 1000 nm and acquired at an angle of 90 deg at a position on the scalp ∼12 mm from the illumination site. The temperature at the onset of the experiment was ∼34°C (rectal temperature) with increasing intervals of 1°C until mouse death. The increase in temperature caused optical scattering signal changes consistent with a structural alteration of brain tissue, ultimately resulting in death. We have found that the peak absorbance intensity and its second derivative at specific wavelengths correlate well with temperature with an exponential dependence. Based on these findings, in order to estimate the influence of temperature on the internal brain tissue a reflectance-temperature index was established and was seen to correlate as well with measured temperature. Overall, results indicate variations in neural tissue properties during heatstroke and the feasibility to monitor and assess internal temperature variations using DRS. Although several approaches have described the rise in temperature and its impact on tissue, to the best of our knowledge no information is available describing the ability to monitor temperature during heatstroke with DRS. The motivation of this study was to successfully describe this ability.

Incensole acetate reduces depressive-like behavior and modulates hippocampal BDNF and CRF expression of submissive animals.

Incensole acetate (IA), a constituent of Boswellia resin ('frankincense'), was previously demonstrated to exhibit an antidepressive-like effect in the Forced Swim Test (FST) in mice following single dose administration (50 mg/kg). Here, we show that acute administration of considerably lower dose (10 mg/kg) IA to selectively bred mice, showing prominent submissive behavior, exerted significant antidepressant-like effects in the FST. Furthermore, chronic administration of 1 or 5 mg/kg per day of IA for three consecutive weeks dose- and time-dependently reduced the submissiveness of the mice in the Dominant-Submissive Relationship test, developed to screen the chronic effect of antidepressants. This behavioral effect was concomitant to reduced serum corticosterone levels, dose-dependent down-regulation of corticotropin releasing factor and up-regulation of brain derived neurotrophic factor transcripts IV and VI expression in the hippocampus. These data suggest that IA modulates the hypothalamic-pituitary-adrenal (HPA) axis and influences hippocampal gene expression, leading to beneficial behavioral effects supporting its potential as a novel treatment of depressive-like disorders.

Comparative analysis of the behavioral and biomolecular parameters of four mouse strains.

The use of mice as experimental models in pharmacological and biochemical research began over 100 years ago, during which time different mice strains with specific features were developed. Numerous studies demonstrate that the pharmacological efficacy of various compounds significantly varies among different animal strains, a factor which must be considered when analyzing experimental data. The Sabra strain, developed more than 35 years ago, is widely used for research in Israel but has an unclear origin and is not characterized as well as other strains. Comparative analyses of the molecular characteristics of Sabra and other strains should help to understand their characteristics and to enhance the validity of their experimental use. Thus, four mouse strains-outbred ICR and Sabra as well as inbred C57Bl/6J and Balb/c were compared. Animals' weight, blood corticosterone and hippocampal BDNF mRNA levels were measured, and animals' behavior was compared using the EPM, open field, FST, and hot plate tests. We found that although Sabra mice are bigger and heavier than other tested lines, this is not reflected in behavior or in biomolecular features, wherein Sabra mice lay within the diapason of other tested animals. Thus, behavioral tests of anxiety-like behavior and locomotor activity revealed that Sabra mice scored close to the mean of all tested lines. Analysis of blood corticosterone levels did not show significant differences among tested strains. We also found a correlation between general and locomotor activity of the tested strains and their hippocampal BDNF mRNA expression. In summary, we may conclude that Sabra mice have traits similar to the better known lines, and therefore they are good subjects for neuroscience research.