Different Regions Of Mouse Brain Research Articles

Restraint stress-induced effects on learning, memory, cognition, and expression of transcripts in different brain regions of mice.

Stress is one of the prevalent factors influencing cognition. Several studies examined the effect of mild or chronic stress on cognition. However, most of these studies are limited to a few behavioral tests or the expression of selected RNA/proteins markers in a selected brain region. This study examined the effect of restraint stress on learning, memory, cognition, and expression of transcripts in key learning centers. Male mice were divided into three groups (n = 6/group)-control group, stress group (adult stressed group; S), and F1 group (parental stressed group). Stress group mice were subjected to physical restraint stress for 2h before light offset for 2weeks. The F1 group comprised adult male mice born of stressed parents. All animals were subjected to different tests and were sacrificed at the end. Transcription levels of Brain-Derived Neurotrophic Factor (Bdnf), Tyrosine kinase (TrkB), Growth Associated Protein 43 (Gap-43), Neurogranin (Ng), cAMP Response Element-Binding Protein (Creb), Glycogen synthase kinase-3β (Gsk3β), Interleukine-1 (IL-1) and Tumour necrosis factor-α (Tnf-α) were studied. Results show that both adult and parental stress negatively affect learning, memory and cognition, as reflected by taking longer time to achieve the task or showing reduced exploratory behavior. Expression of Bdnf, TrkB, Gsk3β and Ng was downregulated, while IL-1 and Tnf-α were upregulated in the brain's cortex, thalamus, and hippocampus region of stressed mice. These effects seem to be relatively less severe in the offspring of stressed parents. The findings suggest that physical restraint stress can alter learning, memory, cognition, and expression of transcripts in key learning centers of brain.

Early Effects of Alpha-Synuclein Depletion by Pan-Neuronal Inactivation of Encoding Gene on Electroencephalogram Coherence between Different Brain Regions in Mice.

We analyzed electroencephalogram (EEG) coherence ("functional connectivity") between the cortex (MC), putamen (Pt), and dopamine-producing brain regions (ventral tegmental area, VTA, and substantia nigra, SN) in two groups of two-month-old male mice. We compared EEG coherences in the conditional knockout Sncaflox/flox mice with those in their genetic background (C57Bl6J) one, two, and three months after chronic (for five days) intraperitoneal injections of TAM or the vehicle (corn oil). The EEG coherences in the TAM-treated group were compared with those in the alpha-synuclein knockout mice. A significant suppression of EEG coherence in the TAM-treated mice versus the vehicle group was observed in all inter-structural relations, with the exception of MC-VTA at one and three months and VTA-SN at two months after the injections. Suppressive changes in EEG coherence were observed in the alpha-synuclein knockout mice as well; the changes were similar to those in TAM-treated mice three months after treatment. our data demonstrate a combined time-dependent suppressive effect induced by TAM on intracerebral EEG coherence.

Datumetine Preferentially Upregulates N-methyl-D-aspartate Receptor Signalling Pathways in Different Brain Regions of Mice.

We previously reported that datumetine possesses binding affinity with N-methyl-D-aspartate receptor (NMDAR) and that 14-day exposure to datumetine altered NMDAR signaling by mimicking glutamate toxicity. Here, we investigated the potential neuroprotective effect of a single shot of a low dose of datumetine administration in BALB/c mice. 30 male adult BALB/c mice were used for the study. The mice were randomly divided into three groups of ten mice each with an intraperitoneal injection of 0.1 mL of 10% DMSO for the Vehicle group, Datumetine group were administered 0.1 mg/kg body weight (bw) of datumetine and MK-801+Datumetine group were administered 0.5 mg/kg bw of MK-801 (to block NMDAR) followed by 0.1 mg/kg bw of datumetine after 30 minutes. 24 hours after administration, mice were euthanized in an isoflurane chamber followed by perfusion with 1X PBS. Brains were excised and stored at -20°C till further processing. Mice designated for IHC were further perfused with 4% PFA and brain excised and stored in 4% PFA till further processing. NMDAR signalling molecules expression was evaluated in frozen brain samples and the fixed brain samples were stained for neuron, vGlut and NMDAR subtypes. Relative to vehicle (Veh), datumetine downregulate calcium calmodulin kinase II alpha (CamKIIα) expression in the hippocampus and prefrontal cortex (PFC) but not in the cerebellum, cyclic AMP response element binding protein (CREB) was also upregulated only in the PFC but phosphorylated CREB (pCREB) was also upregulated in three brain regions observed, while brain-derived neurotrophic factor (BDNF) was only upregulated in hippocampus and PFC of Datumetine relative to vehicle (Veh). On the other hand, dizocilpine (MK-801) reversed some of the effects of datumetine in the observed brain regions. No major histological alterations were observed in the different brain regions immunohistochemically. We conclude that a low dose of datumetine moderately enhances NMDAR activity. This showed the neuroprotective potentials of low datumetine exposure.

Psychosocial stress and cannabinoid drugs affect acetylation of α-tubulin (K40) and gene expression in the prefrontal cortex of adult mice.

The dynamics of neuronal microtubules are essential for brain plasticity. Vesicular transport and synaptic transmission, additionally, requires acetylation of α-tubulin, and aberrant tubulin acetylation and neurobiological deficits are associated. Prolonged exposure to a stressor or consumption of drugs of abuse, like marihuana, lead to neurological changes and psychotic disorders. Here, we studied the effect of psychosocial stress and the administration of cannabinoid receptor type 1 drugs on α-tubulin acetylation in different brain regions of mice. We found significantly decreased tubulin acetylation in the prefrontal cortex in stressed mice. The impact of cannabinoid drugs on stress-induced microtubule disturbance was investigated by administration of the cannabinoid receptor agonist WIN55,212–2 and/or antagonist rimonabant. In both, control and stressed mice, the administration of WIN55,212–2 slightly increased the tubulin acetylation in the prefrontal cortex whereas administration of rimonabant acted antagonistically indicating a cannabinoid receptor type 1 mediated effect. The analysis of gene expression in the prefrontal cortex showed a consistent expression of ApoE attributable to either psychosocial stress or administration of the cannabinoid agonist. Additionally, ApoE expression inversely correlated with acetylated tubulin levels when comparing controls and stressed mice treated with WIN55,212–2 whereas rimonabant treatment showed the opposite.

Genistein suppresses microglial activation and inhibits apoptosis in different brain regions of hypoxia-exposed mice model of amnesia.

Genistein (GE) or 4',5,7-trihydroxyflavone, a plant derived isoflavone, is a biologically active compound having several beneficial properties. Studies showed that GE possesses anti-neoplastic, anti-tumor, anti-helminthic, anti-oxidant, and anti-inflammatory activities. Herein, we investigated the neuroprotective effects of GE in a mouse model of hypoxia-induced amnesia. Mice were exposed to hypoxic conditions (10% O2) in a designated hypoxia chamber and co-treated with GE (10, 20, or 30mg/kg) for 4weeks. Following this, behavioral tests were performed to evaluate memory performance. We assessed microglial activation in the hippocampus, amygdala, and pre-frontal cortex (PFC) regions by evaluating the Iba-1 and GFAP transcript levels, and MIP-1β, Cox-2, and IL6 protein levels. Apoptosis was assessed by evaluating Bax, BAD, and Bcl-2 mRNA levels, and caspase-3 activity. To uncover the underlying molecular mechanism, we evaluated the levels of Nrf2, HO-1, and NQO1 in different brain regions of mice from all groups. Results showed that hypoxia-exposed mice have reduced performance in the behavioral tests and GE treatment enhanced the memory performance in hypoxia-exposed mice. Moreover, hypoxia-exposed mice showed increased expression of microglial activation markers and enhanced apoptosis in the hippocampus, amygdala, and PFC. GE treatment suppressed microglial activation and prevented apoptosis in the brain of hypoxia-exposed mice. Furthermore, hypoxia-exposure reduced the expression of Nrf2, NQO1, and HO-1 while GE treatment ameliorated this decrease in different regions of hypoxia-exposed mice brain. In conclusion, GE prevents cognitive dysfunction by suppressing microglial activation and inhibiting apoptosis in the hypoxia-exposed mice brain.

Regional variability and genotypic and pharmacodynamic effects on PrP concentration in the CNS

Prion protein (PrP) concentration controls the kinetics of prion replication and is a genetically and pharmacologically validated therapeutic target for prion disease. In order to evaluate PrP concentration as a pharmacodynamic biomarker and assess its contribution to known prion disease risk factors, we developed and validated a plate-based immunoassay reactive for PrP across 6 species of interest and applicable to brain and cerebrospinal fluid (CSF). PrP concentration varied dramatically across different brain regions in mice, cynomolgus macaques, and humans. PrP expression did not appear to contribute to the known risk factors of age, sex, or common PRNP genetic variants. CSF PrP was lowered in the presence of rare pathogenic PRNP variants, with heterozygous carriers of P102L displaying 55%, and D178N just 31%, of the CSF PrP concentration of mutation-negative controls. In rodents, pharmacologic reduction of brain Prnp RNA was reflected in brain parenchyma PrP and, in turn in CSF PrP, validating CSF as a sampling compartment for the effect of PrP-lowering therapy. Our findings support the use of CSF PrP as a pharmacodynamic biomarker for PrP-lowering drugs and suggest that relative reduction from individual baseline CSF PrP concentration may be an appropriate marker for target engagement.

Asymmetric Opening of Mitochondrial Permeability Transition Pore in Mouse Brain Hemispheres: A Link to the Mitochondrial Calcium Uniporter Complex

The prolonged entry of large amounts of calcium into the mitochondria through the mitochondrial calcium uniporter complex (MCUC) may cause the permeability transition pore (mPTP) to open, which contributes to the pathogenesis of several diseases. Tissue-specific differences in mPTP opening due to variable expression of MCUC components may contribute to disease outcomes. We designed this study to determine differential mPTP opening in mitochondria isolated from different regions of mouse brain and kidney and to compare it with the expression of MCUC components. mPTP opening was measured using mitochondria isolated from the left/right brain hemispheres (LH/RH, respectively) and from kidney cortex/medulla, while the expression level of MCUC components was assessed from total cellular RNA. Interestingly, LH mitochondria showed less calcium-induced mPTP opening as compared to RH mitochondria at two different calcium concentrations. Conversely, mPTP opening was similar in the renal cortex and renal medulla mitochondria. However, the kidney mitochondria demonstrated bigger and faster mPTP opening as compared to the brain mitochondria. Furthermore, asymmetric mPTP opening in the LH and RH mitochondria was not associated with the expression of MCUC components. In brief, this study demonstrates thus far unreported asymmetric mPTP opening in mouse brain hemispheres that is not associated with the mRNA levels of MCUC components.

Antinociceptive effects of flower extracts and the active fraction from Styrax japonicus

Ethnopharmacological relevanceFlowers from Styrax japonicus sieb. et Zucc. have been used as a Chinese folk medicine to alleviate pain such as toothache and sore throat. Aim of the studyTo testify the analgesic effect of flowers from Styrax japonicus, analyze components of the active fraction, and investigate the mechanism of analgesia. Materials and methodsFlower extracts were obtained by ethanol, petroleum ether and hydrodistillation extraction. Different fractions of ethanol extracts (EE) were isolated by silica gel column chromatography and preparative liquid chromatography. Analgesic effects of EE, petroleum ether extracts (PEE), hydrodistillation extracts (HDE), and fractions of EE were evaluated using hot plate, acetic acid-induced writhing and formalin tests on mice. Components of the active fraction 1 (F1) were determined by the ultrahigh-performance liquid chromatography Q extractive mass spectrometry (UHPLC-QE-MS). Anti-inflammatory and sedative effects involving analgesic mechanisms were evaluated by carrageenan induced hind paw oedema and pentobarbital sodium sleep tests, respectively. In addition, antagonists including naloxone hydrochloride (NXH), flumazenil (FM), SCH23390 (SCH) and WAY100635 (WAY) were used to investigate the possible mechanism of analgesia. Contents of neurotransmitters and relevant metabolites in different brain regions of mice were also quantified by the ultraperformance liquid chromatography with a fluorescence detector (UPLC-FLD). ResultsEE rather than PEE and HDE at medium and high doses (150 mg/kg and 300 mg/kg) significantly prolonged the latency time of the response of mice to the thermal stimulation in the hot plate test. Moreover, EE significantly decreased number of writhes in the acetic acid-induced writhing test, and reduced licking time in both two phases of the formalin test in a dose-dependent manner. The F1 (50 mg/kg) showed effective antinociceptive responses in all mice models. However, fraction 2 (F2) and fraction 3 (F3) at 50 mg/kg performed no analgesic action. Kaempferol-3-O-rutinoside, isorhamnetin-3-O-rutinoside, pinoresinol-4-O-glucoside, forsythin and arctiin were identified from components of the F1. Furthermore, F1 (50 mg/kg) did not significantly affect hind paw oedema of mice induced by carrageenan but significantly shortened sleep latency and increased sleep duration in the pentobarbital sodium sleep test. In addition, the antinociceptive response of F1 was not affected by NXH in two mice models, but significantly blocked by FM and WAY in the hot plate test. In the formalin test, FM avoided the effect of F1 only in the first phase, while the analgesic activity of F1 was totally suppressed by WAY in both two phases. Otherwise, contents of 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) increased significantly in hippocampus and striatum of mice in the F1 group. ConclusionEE from flowers of Styrax japonicus, and F1, the active part isolated from EE, showed significant antinociceptive activities. The analgesic effect of F1 appeared to be related to the sedative effect, partially mediated by the GABAergic system, and highly involved in the serotonergic system. This was the first study confirming the analgesic effect of Styrax japonicus flower, which provided a candidate for the development of non-opioid analgesics.

Antibiotics-induced intestinal dysbacteriosis caused behavioral alternations and neuronal activation in different brain regions in mice

Antibiotics affect gut microbial composition, leading to Gut–Brain-Axis imbalance and neurobehavioral changes. However, the intestinal dysbacteriosis associated behavior changes are not consistently reported. It is not clear whether these changes are transient or permanent. The neuroprotective effect of probiotics against intestinal dysbacteriosis induced alternations needs to be determined either. In the present study, oral antibiotic mixture including Ampicillin, Streptomycin, and Clindamycin was utilized to induce intestinal dysbacteriosis in mice. Antibiotics application triggered mechanical allodynia in von frey test and spontaneous pain in open field test. It also resulted in increased anxiety and depressive-like behaviors and damaged spatial memory performance. After application of probiotics, the mechanical allodynia and spontaneous pain were alleviated significantly. The anxiety behaviors, depressive-like behaviors and recognitive performance were ameliorative as well. By using Fos protein as a marker, it is found that the sensory, emotion and memory related brain regions were activated in mice with intestinal dysbacteriosis. Our study is not only helpful for enriching our basic knowledge for understanding the changed pain responses and related brain disorders in antibiotics-induced dysbacteriosis mice, but also beneficial for providing a more comprehensive mechanistic explanation for the regulation of antibiotics and probiotics on gut microbiota and relevant alternations in animal neurological behaviors.

Chronic Neurobehavioral Impairments and Decreased Hippocampal Expression of Genes Important for Brain Glucose Utilization in a Mouse Model of Mild TBI.

Glucose is an essential cellular fuel for maintaining normal brain functions. Traumatic brain injury (TBI) decreases brain glucose utilization in both human and experimental animals during the acute or subacute phase of TBI. It remains unclear as to how the damages affect brain glucose utilization and its association with persistent neurobehavioral impairments in the chronic phase of mild TBI (mTBI). Accordingly, we compared expression of selected genes important to brain glucose utilization in different brain regions of mice during the chronic phase in mTBI vs. sham operated mice. These genes included hexokinase-1 (HK1), phosphofructokinase (PFK), pyruvate kinase (PK), pyruvate dehydrogenase (PDH), capillary glucose transporter (Glut-1), neuron glucose transporter (Glut-3), astrocyte lactate transpor1 (MCT-1), neuron lactate transporter (MCT-2), lactate receptor (GPR81), and Hexokinase isoform-2 (HK2). Young adult male C57BL/6J mice were brain injured with repetitive closed-head concussions. Morris water maze (MWM), elevated plus maze (EPM), and neurological severity score test (NSS) were performed for evaluation of mice neurobehavioral impairments at 2, 4, and 6 months post mTBI. Two days after completion of the last behavioral test, the frontal cortex, hippocampus, brainstem, hypothalamus, and cerebellum were collected for gene expression measurements. The expression of the mRNAs encoding PK, and PDH, two critical enzymes in glucose metabolism, was decreased at all-time points only in the hippocampus, but was unchanged in the brainstem, hypothalamus, and cortex in mTBI mice. mTBI mice also exhibited the following behavioral alterations: (1) decreased spatial learning and memory 2, 4, and 6 months after the injury, (2) increased proportion of time spent on open vs. closed arms determined by EPM, and (3) accelerated reduction in motor activity observed at 4 months, two months earlier than observed in the sham group, during the EPM testing. There were no significant differences in NSS between injury and sham groups at any of the three time points. Thus, mTBI in male mice led to persistent decreased hippocampal expression of mRNAs that encode critical glucose utilization related enzymes in association with long-term impairments in selected neurobehavioral outcomes.

Chronic Psychosocial Stress Causes Increased Anxiety-Like Behavior and Alters Endocannabinoid Levels in the Brain of C57Bl/6J Mice

Introduction: Chronic stress causes a variety of physiological and behavioral alterations, including social impairments, altered endocrine function, and an increased risk for psychiatric disorders. Thereby, social stress is one of the most effective stressful stimuli among mammals and considered to be one of the major risk factors for the onset and progression of neuropsychiatric diseases. For analyzing the effects of social stress in mice, the resident/intruder paradigm of social defeat is a widely used model. Although the chronic social defeat stress model has been extensively studied, little is known about the effects of repeated or chronic social defeat stress on the endocannabinoid system (ECS). The present study aimed to understand the effects of chronic social stress on anxiety behavior and the levels of endocannabinoids (ECs) and two N-acylethanolamines (NAEs) in different brain regions of mice. Materials and Methods: Two-month-old, male C57Bl/6J mice were exposed to chronic psychosocial stress for 3 weeks. The effects of stress on anxiety behavior were measured using the light-dark box and hole board test. The EC levels of 2-arachidonoyl glycerol (2-AG) and anandamide (N-arachidonoylethanolamine [AEA]), as well as the levels of two NAEs (oleoylethanolamide [OEA] and palmitoylethanolamide), were analyzed by liquid chromatography-tandem mass spectrometry in the hippocampus, cerebellum, and cortex. Results: In comparison with control mice (n=12), mice exposed to social defeat stress (n=11) showed increased anxiety behaviors in the light-dark box and hole board test and gained significantly more weight during the experimental period. Additionally, chronic social stress induced differential alterations in the brain levels of 2-AG and AEA. More precisely, 2-AG levels were higher in the cortex and cerebellum, whereas reduced AEA levels were found in the hippocampus. Furthermore, we observed lower OEA levels in the hippocampus. Conclusion: The current study confirms that the ECS plays an essential role in stress responses, whereby its modulation seems to be brain region dependent.

Tissue- and Region-Specific Accumulation of Arsenic Species, Especially in the Brain of Mice, After Long-term Arsenite Exposure in Drinking Water.

Arsenic is identified as a known carcinogen and ubiquitously exists in nature. It appears that accumulation of inorganic arsenic (iAs) and its methylated metabolites in various tissues is closely correlated with the long-term toxicity and carcinogenicity of this metalloid. In this study, various arsenic species in murine tissues, especially in the cerebral cortex, cerebellum, and hippocampus, were determined after long-term exposure to 25, 50, 100, and 200mg/L sodium arsenite in drinking water for 1 and 12months. Our data showed that the amount of total arsenic (TAs) increased in an obvious dose-dependent manner in various tissues, and TAs levels were in the order of urinary bladder > brain > lung > liver > kidney > spleen. Furthermore, iAsIII and DMA could be observed in all tissues and brain regions with DMA being the predominant metabolite. The bladder, brain, and lung orderly contained the higher levels of DMA, while the liver, kidney, and spleen accumulated the higher proportion of iAsIII. MMA was preferentially accumulated in the lung and bladder of mice regardless of arsenic exposure doses or duration. What's more, amazingly higher levels of MMA were observed in the hippocampus, which was distinguished from the cerebral cortex and cerebellum. Together with these results, our study clearly demonstrates that the accumulation of iAs and its methylated metabolites is tissue-specific and even not homogeneous among different brain regions in mice by long-term exposure to arsenite. Our study thus provides crucial information for recognizing arsenical neurotoxicity, and reducing the uncertainty in the risk assessment for this toxic metalloid.

Neural control of fasting-induced torpor in mice

Torpor is a peculiar mammalian behaviour, characterized by the active reduction of metabolic rate, followed by a drop in body temperature. To enter torpor, the activation of all thermogenic organs that could potentially defend body temperature must be prevented. Most of these organs, such as the brown adipose tissue, are controlled by the key thermoregulatory region of the Raphe Pallidus (RPa). Currently, it is not known which brain areas mediate the entrance into torpor. To identify these areas, the expression of the early gene c-Fos at torpor onset was assessed in different brain regions in mice injected with a retrograde tracer (Cholera Toxin subunit b, CTb) into the RPa region. The results show a network of hypothalamic neurons that are specifically activated at torpor onset and a direct torpor-specific projection from the Dorsomedial Hypothalamus to the RPa that could putatively mediate the suppression of thermogenesis during torpor.

Aberrant Expression of Collagen Gene Family in the Brain Regions of Male Mice with Behavioral Psychopathologies Induced by Chronic Agonistic Interactions.

Chronic agonistic interactions promote the development of experimental psychopathologies in animals: a depression-like state in chronically defeated mice and the pathology of aggressive behavior in the mice with repeated wins. The abundant research data indicate that such psychopathological states are associated with significant molecular and cellular changes in the brain. This paper aims to study the influence of a 20-day period of agonistic interactions on the expression patterns of collagen family genes encoding the proteins which are basic components of extracellular matrix (ECM) in different brain regions of mice using the RNA-Seq database. Most of differentially expressed collagen genes were shown to be upregulated in the hypothalamus and striatum of chronically aggressive and defeated mice and in the hippocampus of defeated mice, whereas downregulation of collagen genes was demonstrated in the ventral tegmental areas in both experimental groups. Aberrant expression of collagen genes induced by chronic agonistic interactions may be indicative of specific ECM defects in the brain regions of mice with alternative social experience. This is the first study demonstrating remodeling of ECM under the development of experimental disorders.

Low-Fat Diet With Caloric Restriction Reduces White Matter Microglia Activation During Aging.

Rodent models of both aging and obesity are characterized by inflammation in specific brain regions, notably the corpus callosum, fornix, and hypothalamus. Microglia, the resident macrophages of the central nervous system, are important for brain development, neural support, and homeostasis. However, the effects of diet and lifestyle on microglia during aging are only partly understood. Here, we report alterations in microglia phenotype and functions in different brain regions of mice on a high-fat diet (HFD) or low-fat diet (LFD) during aging and in response to voluntary running wheel exercise. We compared the expression levels of genes involved in immune response, phagocytosis, and metabolism in the hypothalamus of 6-month-old HFD and LFD mice. We also compared the immune response of microglia from HFD or LFD mice to peripheral inflammation induced by intraperitoneal injection of lipopolysaccharide (LPS). Finally, we investigated the effect of diet, physical exercise, and caloric restriction (40% reduction compared to ad libitum intake) on microglia in 24-month-old HFD and LFD mice. Changes in diet caused morphological changes in microglia, but did not change the microglia response to LPS-induced systemic inflammation. Expression of phagocytic markers (i.e., Mac-2/Lgals3, Dectin-1/Clec7a, and CD16/CD32) in the white matter microglia of 24-month-old brain was markedly decreased in calorically restricted LFD mice. In conclusion, LFD resulted in reduced activation of microglia, which might be an underlying mechanism for the protective role of caloric restriction during aging-associated decline.

Three-dimensional analysis revealed that microglia have different structural features in different brain regions of mice

Microglia are immune cells of the central nervous system, and can constantly monitor the changes in surrounding environment by extending or retracting their ramified processes. To investigate the differences in morphological features of microglia in different brain regions, brain slices of transgenic mice expressing green fluorescent protein (GFP) were prepared and the structure of microglia in the cortex, striatum and polymorph layer of the dentate gyrus (PoDG) were imaged and quantified by combining techniques of tissue clearing, laser scanning confocal microscopy and three-dimensional reconstruction. The individual microglia with complex ramified processes was segmented based on the fact that microglial processes are extended from the cell body and have three-dimensional continuity. The results revealed that the occupied volume of a microglia in the striatum was significantly higher than that in the cortex or PoDG, but there was no significant difference in the occupied volume between the cortex and PoDG. The results of three-dimensional analysis indicated that the total length, the total number, the number of branching points, the total number of endpoints, and the maximum branching level of the microglial processes were significantly different between the cortex, striatum and PoDG. The tissue clearing technique combined with three-dimensional reconstruction can be used to analyze the features of microglial structures from multiple perspectives, and provide a novel approach for further analysis of the relationship between microglial structures and functions and their changes under different pathological conditions.

Expression of HDAC9 in different brain regions in mice with cerebral ischemic stroke

To investigate the expression and the subcellular localization of HDAC9 in different brain regions of mice after cerebral ischemic injury and explore the association between HDAC9 and ischemic stroke. Twenty-one male C57BL/6 mice were randomly divided into sham-operated group (n=9) and operated group (n=12). In the latter group, the mice with Zea-Longa neurological deficit scores of 2 or 3 following middle cerebral artery occlusion (MCAO) were assigned into MCAO group (n=9). Immunofluorescence was performed to investigate the subcellular localization of HDAC9 in the brain tissues on day 3 after MCAO. Western blotting and qRT-PCR were used to analyze the expression of HDAC9 in different regions of the brain. Results Immunofluorescence showed more intense HDAC9 expressions in the brain tissues around the infarct focus, and in the cells surrounding the infarct, HDAC9 expression was obviously increased in the cytoplasm and reduced in the cell nuclei. Compared with the other brain regions, the ipsilesional cortex with MCAO showed more abundant HDAC9 expressions at both the mRNA and protein levels (P<0.05). HDAC9 may be closely related to cerebral ischemic injury and participate in the pathophysiological process of ischemic stroke.

Establish and optimize the model of experimental allergic encephalomyelitis in mice

Objective To explore the appropriate dosage of drugs inducing experimental allergic encephalomyelitis (EAE) in mice, and evaluate the modified model mice. Methods Different doses of myelin oligodendrocyte glycoprotein (MOG35-55: 200 μg, 100 μg, 50 μg, 25 μg), together with different doses of inactivation of mycobacterium tuberculosis (H37RA: 800 μg, 250 μg, 100 μg) and pertussis toxin(500 ng, 200 ng), were used to induce the EAE model. After immunized, the clinical disease severity of EAE mice was measured by the standard EAE grading clinical score daily. The open field test was used to detect the locomotion of mice. The Western blot and immunofluorescence were used to detect the level of myelin basic proteins (MBP) in different brain regions of mice. Results Compared with the EAE mice induced with high-dose drugs, the mice with low-dose drugs (25 μg MOG35-55, 100 μg H37RA, 200 ng pertussis toxin) had low neurological scores. And they displayed normal locomotion compared with the control mice (day 16: group EAE(8.885±0.772)cm/s vs control group (8.933±0.567)cm/s, P>0.05; day 31: group EAE (11.130±0.630)cm/s vs control group (10.670±0.959)cm/s, P>0.05; day 55: group EAE (7.686±0.428)cm/s vs control group (8.313±0.918)cm/s, P>0.05). Moreover, there was a significant decrease of MBP in the parahippocampal cortex (PHC) and fimbria-fornix of EAE mice induced with low-dose of drugs (PHC: group EAE (0.369±0.096) vs control group (1.000±0.163), P<0.05; fimbria-fornix: group EAE (0.494±0.071) vs control group (1.000±0.143), P<0.05). Conclusion The EAE mice induced with low-dose drugs(25 μg MOG35-55, 100 μg H37RA, 200 ng pertussis toxin) have low neurological scores, normal locomotion, and myelin impairment in the central neuronal system. And it can be used in the cognitive behavioral research of demyelination disease, such as multiple sclerosis. Key words: Experimental allergic encephalomyelitis (EAE); Neurological score; Locomotion; Myelin impairment

A viscoelastic analysis of the P56 mouse brain under large-deformation dynamic indentation.

We present the first mechanical characterization of the viscoelastic response for different regions of mouse brain. Force-relaxation tests are performed under large strain dynamic micro-indentation, and viscoelastic models are used subsequently, providing time-dependent mechanical properties of brain tissue under loading conditions comparable to what is experienced in TBI. The unique mechanical properties of different brain regions are highlighted, with substantial variations in the viscoelastic properties and damping ratio of each region. Cortex and pons were the stiffest regions, while the thalamus and medulla were most compliant. The cerebellum and thalamus had highest damping ratio values and those of the medulla were lowest. The reported material parameters can be implemented into finite element computer models of the mouse to investigate the effects of trauma on individual brain regions.

Heat Shock Protein 90 Regulates μ‐Opioid Receptor Signaling In Vitro and in Different Regions of Mouse Brain

Heat shock protein 90 (Hsp90) is a ubiquitous and highly expressed regulator for various signaling pathways and proteins. Only 2 previous studies suggested that Hsp90 might be involved in the regulation of opioid receptor signaling, which is one of the primary targets for drug discovery for chronic pain. This study was designed to determine the regulatory activities of Hsp90 on μ‐opioid receptor (MOR) signaling in various types of MOR expressing cells and in mouse brain. MOR expressing CHO, HEK, U2OS, and SH‐SY5Y neuroblastoma cells were treated with an Hsp90 specific inhibitor (17‐AAG) for 24 hrs followed by treatment with the MOR full agonist DAMGO. The expression and activation of signaling proteins was investigated by Western blot. We found that the expression of MOR, βarrestin2, and Akt was reduced in some cell types by inhibiting Hsp90. In contrast, the expression of Hsp70 was increased by the treatment of 17‐AAG in all cell types, as expected. The activation of ERK MAPK was also altered by Hsp90 inhibition in a cell context dependent manner – whether increasing DAMGO stimulated activation, increasing the baseline, or blunting activation. The strongly context dependent nature of Hsp90 regulation indicated that we would need to determine the role of Hsp90 in regulating the MOR in vivo. We thus injected CD‐1 mice intracerebroventricularly with 17‐AAG for 24 hrs, followed by injection of vehicle or DAMGO into the brain. Interestingly, we found an increased ERK signaling baseline and abolished DAMGO induction in the periaqueductal grey, along with increased Hsp70 expression. Preliminary results in the brain stem suggest that ERK MAPK activation in response to DAMGO is abolished by 17‐AAG, without increasing the baseline. Furthermore, 17‐AAG treatment showed a modest decrease on the acute anti‐nociceptive tail‐flick response to DAMGO, and increased urine and feces output after naloxone precipitated withdrawal in models of acute and chronic dependence. These findings demonstrate that Hsp90 has a strong regulatory role in MOR signaling both in vitro and in vivo, and suggest that Hsp90 activators or other modulators could be potentially used as a co‐therapy for improving the therapeutic index of opioid drugs.Support or Funding InformationWork supported by institutional startup funds from the University of Arizona, along with an internal UNE Mini‐Grant and a Pilot Grant from a UNE Centers for Biomedical Research and Excellent (COBRE) grant (Ian Meng PI, #P20GM103643).