BMC Neuroscience

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Article
Background Evidences indicate that inflammasome compounds participate in amyotrophic lateral sclerosis (ALS), a fatal progressive motoneuron degenerative disease. Researchers have observed the expressions of nucleotide oligomerization domain (NOD)-like receptor protein 3 (NLRP3) related inflammasome components in specific regions of the central nervous system in different ALS models, but the cellular spatiotemporal evolution of this canonical inflammasome pathway and pyroptosis during ALS progression are unclear. Methods The spinal cords of hSOD1 G93A mice (ALS mice) and age-matched littermates (CON mice) were dissected at pre-symptomatic stage (60 d), early- symptomatic stage (95 d), symptomatic stage (108 d) and late-symptomatic stage (122 d) of the disease. By using Nissl staining, double immunofluorescence labelling, qRT-PCR or western blot, we detected morphology change and the expression, cellular location of GSDMD, NLRP3, caspase-1 and IL-1β in the ventral horn of lumbar spinal cords over the course of disease. Results Neural morphology changes and GSDMD ⁺ /NeuN ⁺ double positive cells were observed in ventral horn from ALS mice even at 60 d of age, even though there were no changes of GSDMD mRNA and protein expressions at this stage compared with CON mice. With disease progression, compared with age-matched CON mice, increased expressions of GSDMD, NLRP3, activated caspase-1 and IL-1β were detected. Double immunofluorescence labeling revealed that NLRP3, caspase-1, IL-1β positive signals mainly localized in ventral horn neurons at pre- and early-symptomatic stages. From symptomatic stage to late-symptomatic stage, robust positive signals were co-expressed in reactive astrocytes and microglia. Conclusions Early activation of the canonical NLRP3 inflammasome induced pyroptosis in ventral horn neurons, which may participate in motor neuron degeneration and initiate neuroinflammatory processes during ALS progression.
 
Article
Background Microglia assume opposite phenotypes in response to ischemic brain injury, exerting neurotoxic and neuroprotective effects under different ischemic stages. Modulating M1/M2 polarization is a potential therapy for treating ischemic stroke. Repetitive transcranial magnetic stimulation (rTMS) held the capacity to regulate neuroinflammation and astrocytic polarization, but little is known about rTMS effects on microglia. Therefore, the present study aimed to examine the rTMS influence on microglia polarization and the underlying possible molecular mechanisms in ischemic stroke models. Methods Previously reported 10 Hz rTMS protocol that regulated astrocytic polarization was used to stimulate transient middle cerebral artery occlusion (MCAO) rats and oxygen and glucose deprivation/reoxygenation (OGD/R) injured BV2 cells. Specific expression levels of M1 marker iNOS and M2 marker CD206 were measured by western blotting and immunofluorescence. MicroRNA expression changes detected by high-throughput second-generation sequencing were validated by RT-PCR and fluorescence in situ hybridization (FISH) analysis. Dual-luciferase report assay and miRNA knock-down were applied to verify the possible mechanisms regulated by rTMS. Microglia culture medium (MCM) from different groups were collected to measure the TNF-α and IL-10 concentrations, and detect the influence on neuronal survival. Finally, TTC staining and modified Neurological Severity Score (mNSS) were used to determine the effects of MCM on ischemic stroke volume and neurological functions. Results The 10 Hz rTMS inhibited ischemia/reperfusion induced M1 microglia and significantly increased let-7b-5p level in microglia. HMGA2 was predicted and proved to be the target protein of let-7b-5p. HMGA2 and its downstream NF-κB signaling pathway were inhibited by rTMS. Microglia culture medium (MCM) collected from rTMS treated microglia contained lower TNF-α concentration but higher IL-10 concentration than no rTMS treated MCM, reducing ischemic volumes and neurological deficits of MCAO mice. However, knockdown of let-7b-5p by antagomir reversed rTMS effects on microglia phenotype and associated HMGA/NF-κB activation and neurological recovery. Conclusion High-frequency rTMS could alleviate ischemic stroke injury through inhibiting M1 microglia polarization via regulating let-7b-5p/HMGA2/NF-κB signaling pathway in MCAO models.
 
Article
During neurodegenerative diseases, the brain undergoes morphological and pathological changes; Iron deposits are one of the causes of pathological changes in the brain. The Quantitative susceptibility mapping (QSM) technique, a type of magnetic resonance (MR) image reconstruction, is one of the newest diagnostic methods for iron deposits to detect changes in magnetic susceptibility. Numerous research projects have been conducted in this field. The purpose of writing this review article is to identify the first deep brain nuclei that undergo magnetic susceptibility changes during neurodegenerative diseases such as Alzheimer's or Parkinson's disease. The purpose of this article is to identify the brain nuclei that are prone to iron deposition in any specific disorder. In addition to the mentioned purpose, this paper proposes the optimal scan parameters and appropriate algorithms of each QSM reconstruction step by reviewing the results of different articles. As a result, The QSM technique can identify nuclei exposed to iron deposition in various neurodegenerative diseases. Also, the selection of scan parameters is different based on the sequence and purpose; an example of the parameters is placed in the tables. The BET toolbox in FSL, Laplacian-based phase-unwrapping process, the V_SHARP algorithm, and morphology-enabled dipole inversion (MEDI) method are the most widely used algorithms in various stages of QSM reconstruction.
 
Article
Background Stem cell-based therapy has received considerable attention as a potential candidate in the treatment of ischemic stroke; however, employing an appropriate type of stem cells and an effective delivery route are still challenging. In the present study, we investigated the therapeutic effect of safe, noninvasive, and brain-targeted intranasal administration of hair follicle-derived stem cells (HFSCs) in a rat model of ischemic stroke. Methods Stem cells were obtained from the adult rat hair follicles. In experiment 1, stroke was induced by 30 min middle cerebral artery occlusion (MCAO) and stem cells were intranasally transplanted immediately after ischemia. In experiment 2, stroke was induced by 120 min MCAO and stem cells were administered 24 h after cerebral ischemia. In all experimental groups, neurological performance, short-term spatial working memory and infarct volume were assessed. Moreover, relative expression of major trophic factors in the striatum and cortex was evaluated by the quantitative PCR technique. The end point of experiment 1 was day 3 and the end point of experiment 2 was day 15. Results In both experiments, intranasal administration of HFSCs improved functional performance and decreased infarct volume compared to the MCAO rats. Furthermore, NeuN and VEGF expression were higher in the transplanted group and stem cell therapy partially prevented BDNF and neurotrophin-3 over-expression induced by cerebral ischemia. Conclusions These findings highlight the curative potential of HFSCs following intranasal transplantation in a rat model of ischemic stroke.
 
Silver staining in the corpus callosum and optic tracts. A Representative images of silver staining in mice receiving two sham operations, one sham operation and one mTBI, or two mTBIs in the corpus callosum (CC; top) and optic tract (OT; bottom). Scale bar represents 10 μm. B Quantified silver staining % area. * = p < 0.05. All data are presented as the mean ± the standard error of the mean. Sample sizes (1D = 1 day, 1X = 1 × mTBI, 2X = 2 × mTBI): CC-1D-1X: 12; CC-1D-2X: 16; CC-1 W-1X: 13; CC-1 W-2X: 16; CC-2 W-1X: 14; CC-2 W-2X: 14; OT-1D-1X: 13; OT-1D-2X: 15; OT-1 W-1X: 13; OT-1 W-2X: 15; OT-2 W-1X: 14; OT-2 W-2X: 15. Note that the extent of counter-staining (brownish-orange background) present on the tissue slices is an artifact of processing the tissue with the silver staining kit and is not related to the extent of silver deposited on the tissue itself (black specks and lines)
Silver staining outcomes stratified by sex in the corpus callosum or by impact depth in the optic tract. A–B Representative silver staining images in the corpus callosum A and optic tract B. C–D Quantified silver staining % area in the corpus callosum C and optic tract D. * = p < 0.05. Sample sizes (1D = 1 day, 1x = 1 × mTBI, 2x = 2 × mTBI, 1.2 = 1.2 mm depth, 1.6 = 1.6 mm depth): CC-1D-F: 15; CC-1D-M: 13; CC-1 W-F: 16; CC-1 W-M: 13; CC-2 W-F: 15; CC-2 W-M: 13; OT-1D-1.2: 13; OT-1D-1.6: 15; OT-1 W-1.2: 16; OT-1 W-1.6: 12; OT-2 W-1.2: 13; OT-2 W-1.6: 16. Note that the extent of counter-staining (brownish-orange background) present on the tissue slices is an artifact of processing the tissue with the silver staining kit and is not related to the extent of silver deposited on the tissue itself (black specks and lines)
Article
Background Following one mild traumatic brain injury (mTBI), there is a window of vulnerability during which subsequent mTBIs can cause substantially exacerbated impairments. Currently, there are no known methods to monitor, shorten or mitigate this window. Methods To characterize a preclinical model of this window of vulnerability, we first gave male and female mice one or two high-depth or low-depth mTBIs separated by 1, 7, or 14 days. We assessed brain white matter integrity using silver staining within the corpus callosum and optic tracts, as well as behavioural performance on the Y-maze test and visual cliff test. Results The injuries resulted in windows of white matter vulnerability longer than 2 weeks but produced no behavioural impairments. Notably, this window duration is substantially longer than those reported in any previous preclinical vulnerability study, despite our injury model likely being milder than the ones used in those studies. We also found that sex and impact depth differentially influenced white matter integrity in different white matter regions. Conclusions These results suggest that the experimental window of vulnerability following mTBI may be longer than previously reported. Additionally, this work highlights the value of including white matter damage, sex, and replicable injury models for the study of post-mTBI vulnerability and establishes important groundwork for the investigation of potential vulnerability mechanisms, biomarkers, and therapies.
 
Mean (± SEM) acceleration data (n = 28) for participants performing 30 min of ballistic motor training with concurrent active 0.75 Hz tACS (black bars) or sham tACS (grey bars). Acceleration measures were obtained prior to training (pre-training), 5 min after training and then 24 h after training. Training increased acceleration across both groups at the 5 min post-training time point. Acceleration 24 h after training was greater than baseline in both groups, however, acceleration in the active tACS was greater than sham stimulation at this time point (*)
Mean (± SEM) MEP data (n = 28) for participants performing 30 min of ballistic motor training with concurrent active 0.75 Hz tACS (black bars) or sham tACS (grey bars). MEP measures were obtained prior to training (pre-training), 5 min after training and then 24 h after training. There was no effect of training or tACS on MEP amplitude
Article
Physical exercise and neurorehabilitation involve repetitive training that can induce changes in motor performance arising from neuroplasticity. Retention of these motor changes occurs via an encoding process, during which rapid neuroplastic changes occur in response to training. Previous studies show that transcranial alternating current stimulation (tACS), a form of non-invasive brain stimulation, can enhance encoding of a cognitive learning task during wakefulness. However, the effect of tACS on motor processes in the awake brain is unknown. In this study, forty-two healthy 18–35 year old participants received either 0.75 Hz (active) tACS (or sham stimulation) for 30 min during a ballistic thumb abduction motor training task. Training-related behavioural effects were quantified by assessing changes in thumb abduction acceleration, and neuroplastic changes were quantified by measuring motor evoked potential (MEP) amplitude of the abductor pollicis brevis muscle. These measures were reassessed immediately after the motor training task to quantify short-term changes, and then 24 h later to assess longer-term changes. Thumb abduction acceleration in both active and sham stimulation conditions increased immediately after the motor learning, consistent with effective training. Critically, participants in the active group maintained significantly higher thumb acceleration 24 h later (t 40 = 2.810, P = 0.044). There were no significant changes or inter-group differences in MEPs for both conditions. The results suggest that 0.75 Hz tACS applied during motor training enhances the effectiveness of motor training, which manifests as enhancement in longer-term task benefits.
 
Article
Background Following Spinal Cord Injury (SCI), innumerable inflammatory and degenerative fluctuations appear in the injured site, and even remotely in manifold areas of the brain. Howbeit, inflammatory, degenerative, and oscillatory changes of motor cortices have been demonstrated to be due to SCI, according to recent studies confirming the involvement of cognitive areas of the brain, such as hippocampus and prefrontal cortex. Therefore, addressing SCI induced cognitive complications via different sights can be contributory in the treatment approaches. Results Herein, we used 16 male Wistar rats (Sham = 8, SCI = 8). Immunohistochemical results revealed that spinal cord contusion significantly increases the accumulation of alpha-synuclein and decreases the expression of Doublecortin (DCX) in the hippocampal regions like Cornu Ammonis1 (CA1) and Dentate Gyrus (DG). Theses degenerative manifestations were parallel with a low expression of Achaete-Scute Family BHLH Transcription Factor 1 (ASCL1), SRY (sex determining region Y)-box 2 (SOX2), and dopaminergic receptors (D1 and D5). Additionally, based on the TUNEL assay analysis, SCI significantly increased the number of apoptotic cells in the CA1 and DG regions. Cognitive function of the animals was assessed, using the O-X maze and Novel Object Recognition (NORT); the obtained findings indicted that after SCI, hippocampal neurodegeneration significantly coincides with the impairment of learning, memory and recognition capability of the injured animals. Conclusions Based on the obtained findings, herein SCI reduces neurogenesis, decreases the expression of D1 and D5, and increases apoptosis in the hippocampus, which are all associated with cognitive function deficits. Graphical Abstract
 
Article
Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is highly contagious, and the neurological symptoms of SARS-CoV-2 infection have already been reported. However, the mechanisms underlying the effect of SARS-CoV-2 infection on patients with central nervous system injuries remain unclear. Methods The high-throughput RNA sequencing was applied to analyze the transcriptomic changes in SK-N-SH cells after SARS-CoV-2 infection. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses were performed to identify the functions of differentially expressed genes and related pathways. Results A total of 820 mRNAs were significantly altered, including 671 upregulated and 149 downregulated mRNAs (showing an increase of ≥ 2-fold or decrease to ≤ 0.5-fold, respectively; p ≤ 0.05). Moreover, we verified the significant induction of cytokines, chemokines, and their receptors, as well as the activation of NF-κB, p38, and Akt signaling pathways, in SK-N-SH by SARS-CoV-2. Conclusions To our knowledge, this is the first time the transcriptional profiles of the host mRNAs involved in SARS-CoV-2 infection of SK-N-SH cells have been reported. These findings provide novel insight into the pathogenic mechanism of SARS-CoV-2 and might constitute a new approach for future prevention and treatment of SARS-CoV-2-induced central nervous system infection.
 
Article
Background Cervical spondylotic myelopathy (CSM) is a clinically symptomatic condition due to spinal cord compression, leading to spinal cord dysfunction. Surgical decompression is the main treatment of CSM, but the mechanisms of axonal regeneration after surgical decompression are still fragmentary. Methods In a rat model of CSM, the cacna2d2 (α2δ2) expression levels in anterior horn of spinal cord were observed following compression and decompression by western blot and immunofluorescence. The expression levels of 5 hydroxytryptamine (5HT) and GAP43 were also analyzed by immunofluorescence. Furthermore, gabapentin intervention was performed for 4 weeks after decompression to analyze the changes of behaviors and anterior horn of spinal cords. Results Following decompression, the expression levels of α2δ2 in the anterior horn of spinal cord were decreased, but the expression levels of 5HT andGAP43 were increased. Compared with the vehicle treated rats, gabapentin treatment for 4 weeks ameliorated the behaviors of rats and improved the damaged anterior horn of spinal cord. Besides, inhibition of α2δ2 through gabapentin intervention enhanced the axonal regeneration in the anterior horn of damaged spinal cord. Conclusions Inhibition of α2δ2 could enhance axonal recovery in anterior horn of damaged spinal cord induced by CSM after surgical decompression, providing a potential method for promoting axon regeneration following surgery.
 
Article
Exercise could prevent physical and psychological deteriorations, especially during pandemic times of lock-down scenarios and social isolation. But to meet both, the common exercise protocols require optimization based on holistic investigations and with respect to underlying processes. This study aimed to explore individual chronic and acute effects of continuous and interval running exercise on physical and cognitive performance, mood, and affect and underlying neurophysiological factors during a terrestrial simulated space mission. Six volunteers (three females) were isolated for 120 days. Accompanying exercise training consisted of a continuous and interval running protocol in a cross-over design. Incremental stage tests on a treadmill were done frequently to test physical performance. Actigraphy was used to monitor physical activity level. Cognitive performance, mood (MoodMeter®), affect (PANAS), brain-derived neurotrophic factor (BDNF), insulin-like growth factor 1 (IGF-1), vascular-endothelial growth factor (VEGF), and saliva cortisol were investigated prior to, four times during, and after isolation, pre- and post-exercise on two separate days, respectively. As a chronic effect, physical performance increased (and IGF-1 tended) in the course of isolation and training until the end of isolation. Subjective mood and affect state, as well as cognitive performance, basal BDNF and VEGF levels, were well-preserved across the intervention. No acute effects of exercise were detected, besides slower reaction time after exercise in two out of nine cognitive tests, testing sensorimotor speed and memory of complex figures. Consistently higher basal IGF-1 concentrations and faster reaction time in the psychomotor vigilance test were found for the continuous compared to the interval running protocol. The results suggest that 120 days of isolation and confinement can be undergone without cognitive and mental deteriorations. Regular, individual aerobic running training supporting physical fitness is hypothesized to play an important role in this regard. Continuous running exercise seems to trigger higher IGF-1 levels and vigilance compared to interval running. Systematic and prolonged investigations and larger sample size are required to follow up on exercise-protocol specific differences in order to optimize the exercise intervention for long-term psycho-physiological health and well-being.
 
Article
Background Previous research has shown that spatial performance (e.g. navigation, visuospatial memory, attention) can be influenced by acute stress; however, studies have produced mixed findings sometimes showing an improvement after stress, other times showing impairment or no overall effect. Some of these discrepancies may be related to: the type of stress system activated by the stressor (sympathetic adrenal medulla [SAM] or hypothalamic-pituitary-adrenocortical [HPA]); whether cortisol responders vs. nonresponders are analyzed subsequent to main effects; and sex differences in stress responses. In the present study, we examine the influence of HPA activation from an acute laboratory stressor (Socially Evaluated Cold Pressor test [SECPT]) on performance during two spatial tasks: Useful Field of View (UFOV; a measure of spatial attention) and virtual reality (VR) navigation. We assigned 31 males and 30 females to either the SECPT or a Non-Stress condition prior to the two spatial tasks. Cardiovascular measures including heart rate and blood pressure, and salivary cortisol biosamples were obtained at specific time points. Results Participants in the Stress condition showed increases in heart rate, systolic and diastolic blood pressure indicating sympathetic adrenal medulla (SAM) axis activation. Stress also led to increases in salivary cortisol, suggesting hypothalamic-pituitary-adrenocortical (HPA) activation. Stress did not influence overall performance in the spatial attention UFOV or the VR navigation task. However, a sex difference in spatial attention was detected when participants were divided into Stress-cortisol responders and non-responders in the UFOV task. Male Stress-cortisol responders (n = 9) showed better UFOV accuracy than female Stress-cortisol responders (n = 6); no sex differences were found among the Non-Stress control group. Furthermore, for females in the stress condition (n = 14), higher cortisol responses were associated with lower spatial attention performance. Conclusions Socially Evaluated Cold Pressor stress resulted in no change in speed or accuracy in a VR navigation task. For the spatial attention task, the SECPT led to a sex difference among Stress-cortisol responders with males showing improved accuracy over females. The relationship between HPA activation and prefrontal cortex activity may be necessary to understand sex differences in spatial attention performance.
 
Article
Background Corticotropin-releasing factor (CRF) is the major neuromodulator orchestrating the stress response, and is secreted by neurons in various regions of the brain. Cerebellar CRF is released by afferents from inferior olivary neurons and other brainstem nuclei in response to stressful challenges, and contributes to modulation of synaptic plasticity and motor learning behavior via its receptors. We recently found that CRF modulates facial stimulation-evoked molecular layer interneuron-Purkinje cell (MLI-PC) synaptic transmission via CRF type 1 receptor (CRF-R1) in vivo in mice, suggesting that CRF modulates sensory stimulation-evoked MLI-PC synaptic plasticity. However, the mechanism of how CRF modulates MLI-PC synaptic plasticity is unclear. We investigated the effect of CRF on facial stimulation-evoked MLI-PC long-term depression (LTD) in urethane-anesthetized mice by cell-attached recording technique and pharmacological methods. Results Facial stimulation at 1 Hz induced LTD of MLI-PC synaptic transmission under control conditions, but not in the presence of CRF (100 nM). The CRF-abolished MLI-PC LTD was restored by application of a selective CRF-R1 antagonist, BMS-763,534 (200 nM), but it was not restored by application of a selective CRF-R2 antagonist, antisauvagine-30 (200 nM). Blocking cannabinoid type 1 (CB1) receptor abolished the facial stimulation-induced MLI-PC LTD, and revealed a CRF-triggered MLI-PC long-term potentiation (LTP) via CRF-R1. Notably, either inhibition of protein kinase C (PKC) with chelerythrine (5 µM) or depletion of intracellular Ca²⁺ with cyclopiazonic acid (100 µM), completely prevented CRF-triggered MLI-PC LTP in mouse cerebellar cortex in vivo. Conclusions The present results indicated that CRF blocked sensory stimulation-induced opioid-dependent MLI-PC LTD by triggering MLI-PC LTP through CRF-R1/PKC and intracellular Ca²⁺ signaling pathway in mouse cerebellar cortex. These results suggest that activation of CRF-R1 opposes opioid-mediated cerebellar MLI-PC plasticity in vivo in mice.
 
Article
Critical development period of intestinal microbiota occurs concurrently with brain development, and their interaction is influenced by the microbiota–gut–brain axis. This study examined how antibiotics exposure affected gut microbiota and brain development and analyzed the possible benefits of heat-inactivated Lacticaseibacillus paracasei N1115 (N1115). Thirty neonatal male mice were randomly divided into three groups and treated with sterilized water (control), an antibiotic cocktail (Abx), or antibiotics plus heat-inactivated N1115 (Abx + N1115) for 84 days. We found that while the mRNA levels of GABA Aα1 , GABA b1 , and glucocorticoid receptor (GR) in the hippocampus and brain-derived neurotrophic factor (BDNF), GABA Aα1 , GABA b1 , and nerve growth factor (NGF) in the prefrontal cortex were higher, the mRNA levels of 5-HT 1A were lower in the Abx group. The Abx + N1115 group had lower mRNA levels of GABA Aα1 , GABA b1 , and GR in the hippocampus and BDNF, GABA b1 , and NGF in the prefrontal cortex than the Abx group. The latency period was longer in the Morris water maze test while longer rest time was seen in tail suspension test in the Abx group than the control and Abx + N1115 groups. In the open field test, the moving time and distance of the Abx group were reduced. Further, the alpha-diversity indexes of the Abx and Abx + N1115 groups were significantly lower than the control. Further, long-term exposure to antibiotics disrupted the intestinal microbiota as evidenced by decreased Bacteroides , Firmicutes , and Lactobacillus , and increased Proteobacteria and Citrobacter . However, N1115 significantly decreased the abundance of Citrobacter when compared with those in the Abx group. These results indicate that antibiotics can substantially damage the intestinal microbiota and cognitive function, causing anxiety and depression, which can be alleviated by heat-inactivated N1115 via modulation of the microbiota–gut–brain axis.
 
Withdrawal threshold in the CCI, vein wrapping (VW) and frozen vein wrapping (FVW) groups. Withdrawal threshold in the CCI, vein wrapping (VW) and frozen vein wrapping (FVW) groups. Data show mean ± standard error (n = 5, each time point). *p < 0.05
Effect of frozen vein wrapping on Hmox1 expression. Effect of frozen vein wrapping on Hmox1 messenger RNA (mRNA) levels in sciatic nerve after chronic constriction injury (CCI). Data show mean ± standard error (n = 5, each time point). *p < 0.05
bFGF concentration in vein and sciatic nerve. bFGF protein concentration (bFGF protein (ng)/total protein (mg)) in vein and sciatic nerve. Data show mean ± standard error (n = 5). *p < 0.05
Article
Background Autologous vein wrapping (VW) is used in the treatment of recurrent chronic constriction neuropathy and traumatic peripheral nerve injury. However, use of autologous veins is limited by the inability to obtain longer veins of sufficient length for larger sites. Frozen allograft tissue has several advantages, including its availability for large grafts, avoidance of donor-site morbidity, and shorter operation time. Here, we investigated the effect of frozen vein wrapping (FVW) in Wistar rats as a model of sciatic nerve injury. Results The rats were grouped by treatment as (i) untreated after chronic constriction injury surgery (CCI; control group), (ii) treated with vein wrapping using freshly isolated vein (VW), and (iii) treated with vein wrapping using frozen vein (FVW). Mechanical allodynia was assessed with von Frey filaments on postoperative days (PODs) 1, 3, 5, 7, and 14. Gene expression of HO-1 was evaluated by quantitative polymerase chain reaction (qPCR). The response of heme oxygenase-1 gene, Hmox-1, expression to VW and FVW was assessed by RT-PCR. Both VW and FVW significantly increased withdrawal threshold levels compared to the untreated control group on POD 1, 3, and 5. Both VW and FVW also showed increased HO-1 expression compared to the CCI group. Conclusions FVW increased the withdrawal threshold similar to VW in a rat CCI model for short periods. Frozen vein wrapping using vein allograft without donor site morbidity may be an alternative therapeutic option.
 
Examples of intracranial electrode localizations. Coronal MRI images (T1-weighted) of two patients. Red dots: Electrode contacts inside target structure. Orange dots: Electrode contacts outside target structure. Only data derived from electrode contacts inside target structure were analyzed. A Amygdala electrode contacts in patient 5, right hemisphere (Group M). B Hippocampus electrode contacts in patient 6, right hemisphere (Group O)
Event-related potentials. Grey areas mark the windows in which ERP amplitude was measured. Windows containing a significant categorical difference are marked with an asterisk (*). A ERPs for faces versus mosaics. In AM, N240 was higher for faces than mosaics in both groups, while N360 was higher only in Group O. Both groups showed higher N240 for faces in FU and higher N360 for faces in FU and PH. B ERPs for all face categories. In AM of Group O, N360 was higher for famous faces than wooden masks and veiled faces. In AM of Group M, N360 was higher for famous and dark-skinned faces than Caucasian faces, while N110 was lower for wooden masks than dark-skinned or veiled faces. In HI of Group M, N360 was lower for wooden masks than dark-skinned faces. In FU, wooden masks triggered lower amplitudes than all other face categories for N240 in both groups and N360 in Group O. In PH, N240 was higher for famous faces than dark-skinned faces and wooden masks in Group O. N360 was higher for famous faces than all other face categories in Group O and higher than wooden masks, Caucasian and veiled faces in Group M
Gamma frequency analysis. A1–D1, A2–D2 Power map in response to faces and mosaics in Group O and Group M. Mean gamma power was measured during 0–750 ms in AM, FU and PH and 250–1000 ms in HI after stimulus onset and applied to the linear mixed-effects model. Gamma power was higher for faces than mosaics in AM (A1) and FU (C1) of Group O. A3–D3, A4–D4 Power map showing the response to faces minus mosaics in Group O and Group M. A5–D5 Unthresholded t-statistics comparing the power map for faces minus mosaics in Group O and Group M. Positive t-statistics indicate higher power in Group O and negative t-statistics indicate higher power in Group M. A6–D6 Remaining clusters of t-statistics after voxel-based and cluster-based thresholding at p ≤ 0.05. Large clusters in the gamma frequency range remained in AM and FU, indicating higher gamma power in Group O
Phase-amplitude coupling and phase-slope index. A1–A4 PAC between AM and FU in patient 6, right hemisphere (Group O). Enhanced PAC (Z-score ≥ 2) was observed between FU theta/alpha phase and AM low gamma frequency amplitude when viewing faces. No enhanced PAC was observed when viewing mosaics. B Overlap between PSI and the area of enhanced PAC. Red area indicates modulation directionality from FU theta phase towards AM low gamma amplitude. Blue area indicates modulation directionality from AM low gamma amplitude towards FU alpha phase. C1–4 PAC between HI and PH in patient 9, right hemisphere (Group M). Enhanced PAC (Z-score ≥ 2) was calculated between PH alpha/beta phase and HI high gamma amplitude when viewing faces. No enhanced PAC was observed when viewing mosaics. D Overlap between PSI and the area of enhanced PAC. Red area indicates modulation directionality from PH alpha/beta phase towards HI high gamma amplitude. Blue areas indicate modulation directionality from HI high gamma amplitude towards PH alpha phase
Article
Background To examine the pathological effect of a mesial temporal seizure onset zone (SOZ) on local and inter-regional response to faces in the amygdala and other structures of the temporal lobe. Methods Intracranial EEG data was obtained from the amygdala, hippocampus, fusiform gyrus and parahippocampal gyrus of nine patients with drug-refractory epilepsy during visual stimulation with faces and mosaics. We analyzed event-related potentials (ERP), gamma frequency power, phase-amplitude coupling and phase-slope-index and compared the results between patients with versus without a mesial temporal SOZ. Results In the amygdala and fusiform gyrus, faces triggered higher ERP amplitudes compared to mosaics in both patient groups and higher gamma power in patients without a mesial temporal SOZ. In the hippocampus, famous faces triggered higher gamma power for both groups combined but did not affect ERPs in either group. The differentiated ERP response to famous faces in the parahippocampal gyrus was more pronounced in patients without a mesial temporal SOZ. Phase-amplitude coupling and phase-slope-index results yielded bidirectional modulation between amygdala and fusiform gyrus, and predominately unidirectional modulation between parahippocampal gyrus and hippocampus. Conclusions A mesial temporal SOZ was associated with an impaired response to faces in the amygdala, fusiform gyrus and parahippocampal gyrus in our patients. Compared to this, the response to faces in the hippocampus was impaired in patients with, as well as without, a mesial temporal SOZ. Our results support existing evidence for face processing deficits in patients with a mesial temporal SOZ and suggest the pathological effect of a mesial temporal SOZ on the amygdala to play a pivotal role in this matter in particular.
 
Article
Background Temporal envelope cues are conveyed by cochlear implants (CIs) to hearing loss patients to restore hearing. Although CIs could enable users to communicate in clear listening environments, noisy environments still pose a problem. To improve speech-processing strategies used in Chinese CIs, we explored the relative contributions made by the temporal envelope in various frequency regions, as relevant to Mandarin sentence recognition in noise. Methods Original speech material from the Mandarin version of the Hearing in Noise Test (MHINT) was mixed with speech-shaped noise (SSN), sinusoidally amplitude-modulated speech-shaped noise (SAM SSN), and sinusoidally amplitude-modulated (SAM) white noise (4 Hz) at a + 5 dB signal-to-noise ratio, respectively. Envelope information of the noise-corrupted speech material was extracted from 30 contiguous bands that were allocated to five frequency regions. The intelligibility of the noise-corrupted speech material (temporal cues from one or two regions were removed) was measured to estimate the relative weights of temporal envelope cues from the five frequency regions. Results In SSN, the mean weights of Regions 1–5 were 0.34, 0.19, 0.20, 0.16, and 0.11, respectively; in SAM SSN, the mean weights of Regions 1–5 were 0.34, 0.17, 0.24, 0.14, and 0.11, respectively; and in SAM white noise, the mean weights of Regions 1–5 were 0.46, 0.24, 0.22, 0.06, and 0.02, respectively. Conclusions The results suggest that the temporal envelope in the low-frequency region transmits the greatest amount of information in terms of Mandarin sentence recognition for three types of noise, which differed from the perception strategy employed in clear listening environments.
 
Article
Background Natural nutrition and physical training have been defined as non-pharmacochemical complementary and alternative medicines to prevent and treat various pathogenesis. Royal jelly possesses various pharmacological properties and is an effective therapeutic supplement for halting neurodegeneration. Multiple sclerosis is a prevalent neurodegenerative disorder that manifests as a progressive neurological condition. Inflammation, hypoxia, and oxidative stress have been identified as significant hallmarks of multiple sclerosis pathology. Results In the present study, based on artificial intelligence and bioinformatics algorithms, we marked hub genes, molecular signaling pathways, and molecular regulators such as non-coding RNAs involved in multiple sclerosis. Also, microRNAs as regulators can affect gene expression in many processes. Numerous pathomechanisms, including immunodeficiency, hypoxia, oxidative stress, neuroinflammation, and mitochondrial dysfunction, can play a significant role in the MSc pathogenesis that results in demyelination. Furthermore, we computed the binding affinity of bioactive compounds presented in Royal Jelly on macromolecules surfaces. Also, we predicted the alignment score of bioactive compounds over the pharmacophore model of candidate protein as a novel therapeutic approach. Based on the q-RT-PCR analysis, the expression of the Dnajb1/Dnajb1/Foxp1/Tnfsf14 and Hspa4 networks as well as miR-34a-5p and miR155-3p were regulated by the interaction of exercise training and 100 mg/kg Royal Jelly (ET-100RJ). Interestingly, characteristics, motor function, a proinflammatory cytokine, and demyelination were ameliorated by ET-100RJ. Discussion Here, we indicated that interaction between exercise training and 100 mg/kg Royal jelly had a more effect on regulating the microRNA profiles and hub genes in rats with Multiple sclerosis.
 
Design of shRNA targeting Sirt1 for AAV packaging and hippocampal Sirt1 knockdown by stereotactic injection. A Validation of knockdown efficiency of Sirt1-shRNAs in GL261 cells by RT-qPCR. Abbreviations: GL261-WT (wide type GL261 cells), GL261-shSirt1A (GL261 cells infected by the first shRNA sequence targeting Sirt1), GL261-shSirt1B (GL261 cells infected by the second shRNA sequence targeting Sirt1). n = 3. ***p < 0.001 represents comparison between the GL261-WT and GL261-shSirt1A. #p < 0.05 represents comparison between the GL261-WT and GL261-shSirt1B. B Western blotting of Sirt1 levels after Sirt1 knockdown in G1261 cells. (C) Quantitative density values in B. D Illustration of plasmid construction for AAV packaging. Abbreviations: inverted terminal repeats (ITR), cytomegalovirus promoter (CMV), Sirt1 shRNA (shSirt1), and enhanced green fluorescent protein (EGFP) reporter gene. E Work flow of in vivo experiments performed on the control group, the sham group and the shSirt1 group. Abbreviations: Open Field test (OFT), Morris Water Maze (MWM), magnetic resonance imaging (MRI), and western blotting (WB). F Graphical illustration of stereotactic injection of AAV into bilateral CA1 regions of hippocampus. G Representative fluorescence image of frozen brain section
Volume analysis of hippocampus and its subregions based on 3T MR brain structural imaging. A Illustrative schemes for hippocampal segmentation based on Turone Mouse Brain Atlas and Template (TMBTA). B–G Statistical analysis of (B) total intracranial volume (TIV), (C) the volume of hippocampal formation region, (D) total hippocampal volume, and the volumes of hippocampal subregions including (E) dentate gyrus region, (F) CA1 region, (G) CA2 region, and (H) CA3 region. n = 9. *p < 0.05, and **p < 0.01 represent comparison between the control group and the shSirt1 group
Hippocampal Sirt1 knockdown caused cognitive impairment in middle-aged mice. A Percentage of time spent by mice in the center zone during the OFT. B Representative autonomous trajectory maps of mice in OFT. Green color indicates the defined center zone and the rest is defined peripheral zone. C The swimming speed of mice in MWM with visual platform before the learning phase. D Representative swimming paths of mice in the probe phase. E Escape latency of mice during the learning phase. F Time spent in target zone and (G) the number of platform crossings of mice in the probe phase. n = 12. *p < 0.05 and **p < 0.01 represent comparison between the control group and the shSirt1 group. #p < 0.05 represents comparison between the sham group and the shSirt1 group
Western blotting analysis of related protein levels after hippocampal Sirt1 knockdown. A Representative immunoblots. B–I Quantitative density values in A: (B)Sirt1, (C) T-tau, (D) P-tau at Ser396, (E) P-tau/T-tau, (F) PSD95, (G) Synaptophysin, (H) Synapsin1, (I) NMDAR2B. n = 3. *p < 0.05, and **p < 0.01 represent comparisons between the control group and the shSirt1 group. #p < 0.05, and ##p < 0.01 represent comparisons between the sham group and the shSirt1 group
Article
Background Sirtuin 1 ( Sirt1 ) is a recognized longevity gene and has been shown to be associated with aging and its related diseases. Hippocampal volume is considered to be the most sensitive brain imaging phenotype for cognition, but the effect of Sirt1 on hippocampal morphology during aging has not been reported. Results Herein, we investigated the effect of conditional Sirt1 knockdown on hippocampal volume in middle-aged mice, as well as its cognitive function and the underlying molecular mechanisms. Brain structural magnetic resonance imaging (MRI) showed that adeno-associated virus (AAV) mediated hippocampal Sirt1 knockdown caused hippocampal atrophy in 8-month-old mice. Open field test (OFT) and Morris Water Maze (MWM) test revealed that hippocampal Sirt1 knockdown significantly weakened spatial learning and memory of mice without effect on anxiety and exploratory behavior. Western blotting analysis showed that P-tau levels at serine 396 epitope were significantly increased with slightly decreased T-tau levels, while PSD95 and NMDAR2B levels were obviously reduced, indicating that hippocampal Sirt1 knockdown could activate tau hyperphosphorylation and synaptic damage. Conclusions This work revealed that Sirt1 is an important protective gene against hippocampal atrophy and its induced cognitive impairment during aging, providing potential therapeutic targets for the prevention and intervention of aging-related neuropsychic diseases.
 
Expression of Fmr1 exon 14 in the rat brain on E14, E19 or P2. A Full-length rat Fmr1 exon organization and correspondence to encoded FMRP domains: two N-terminal Agenet domains, three central hnRNP K-homology (KH) domains (KH-0, KH-1 and KH-2), and a C-terminal region RGG-box. NLS and NES correspond to the nuclear localization and export signal motifs, respectively. Arrows indicate RTqPCR primers location on exons, defining amplicons (black boxes) for exons 10 and 11, and messages specifically containing Fmr1 exon 14 or junctions between exons 13 and 15 by using the first (A), second (B), or third (C) splicing acceptor sites. B–E RTqPCR results of relative mRNA expression rates of the hippocampus (B), cerebral cortex (C), E14 telencephalic vesicle and E19 cerebral cortex (D), and cerebellum (E). Student´s t test: P < 0.05 (*), P < 0.01 (**), and P < 0.001(***)
BLAST RNA-Seq RPKM values quantifying Fmr1 sequences in the rat forebrain. Box plots of RPKM values for BLAST searches with (A) Fmr1 coding sequence from exon 1 through 11 [1–11]; or sequences spanning junctions between (B) exons 10 and 11, (C) 16 and 17B, (D) 13 and 14, (E) 14 and the summation of data for the three exon 15 sites. F RPKM ratios of 13–15A, 13–15B or 13–15C junction by exon 10 and 11 junction sequence reads. Significant Dunn´s post-hoc test results are indicated on graphs by lines with asterisks according to P values for comparisons to E14 RPKM values (*P < 0.05; **P < 0.01; ***P < 0.001). Other significant comparisons between embryonic ages or involving P3 are shown below each graph (A–E). N = 4 for each age group, except E14 and P7 (N = 3), and E17 (N = 6)
Knocking down UPF1 expression as a tool for assessing NMD-induced effects on FMR1 mRNA. A Schematic diagrams for FMR1 gene and transcripts. Full-length Fmr1 coding exons 1 to 17 (numbered boxes) and FMRP encoded domains, nuclear localization (NLS) and export (NES) signal motifs. FMR1 transcripts with or without exon 14 and selection of splice acceptor sites 15A, 15B or 15C are partially illustrated (exons 13 to 17). Location of translation termination codons is indicated by asterisks. B, C Western blot analysis of lysates of HEK293T transfected with UPF1 or control dsRNA, as indicated, treated with antibodies for UPF1 or α-tubulin, and plot of densitometry intensities. D RTqPCR data normalized by PPIA for HEK293T cells transfected with UPF1 or negative control dsRNAs, for SMG6 (positive control), FMR1 exons 10 and 11, exons 13 and 14, and amplicons 13–15A, 13–15B and 13–15C. Student´s t test: P < 0.05 (*), P < 0.01 (**), and P < 0.001(***). E Illustration of Fmr1 exon 15 sequence RNA (length legend in nucleotide, nt) and location of splice sites A, B and C, as well as the segment able to form a G-quartet structure stabilized by a counterion (K⁺). A suggested location of a UPF1 binding site is indicated between sites 15A and 15B
FMRP immunoblot of rat brain structures. A–C Panels of blots incubated with anti-vinculin or anti-FMRP, as indicated, corresponding to hippocampus (A), cerebral cortex (B) and cerebellum (C). Arrows indicate the blot areas that had the intensity estimated and labeled as bands 1 or 2. D–I Plots of band 1 (D–F) or band 2 (G–I) intensities normalized by the vinculin band intensity for hippocampus (D and G), cerebral cortex (E and H) and cerebellum (F and I)
Graphic scheme correlating the summary of the results to known neurodevelopmental aspects of FMRP. The column graphic style of the illustration presents the relative amount of total Fmr1 transcripts in the forebrain as columns, in a limited period of the development: the last two embryonic weeks (2 and 3) and the first three postnatal weeks (1–2 and 3). According to the data presented here, total Fmr1 mRNA reaches its highest values on the second embryonic and first postnatal weeks. Low levels of Fmr1 mRNA are observed in the third embryonic week as well as after the second postnatal week. In each pictured graph column of total Fmr1 mRNA, the most abundant exon 13 junction is represented on the top, while on the bottom part is the least frequently observed one in a fading color tone. A drawing of the predominant neuronal differentiation process that takes place in each of the illustrated developmental week highlights the first three graph columns. Each process is associated with a developmental period, representing neurogenesis that predominates in embryonic week 2, neuron migration in embryonic week 3; and synaptogenesis starting in postnatal weeks 1 and 2
Article
Background Fragile X syndrome, the major cause of inherited intellectual disability among men, is due to deficiency of the synaptic functional regulator FMR1 protein (FMRP), encoded by the FMRP translational regulator 1 ( FMR1 ) gene. FMR1 alternative splicing produces distinct transcripts that may consequently impact FMRP functional roles. In transcripts without exon 14 the translational reading frame is shifted. For deepening current knowledge of the differential expression of Fmr1 exon 14 along the rat nervous system development, we conducted a descriptive study employing quantitative RT-PCR and BLAST of RNA-Seq datasets. Results We observed in the rat forebrain progressive decline of total Fmr1 mRNA from E11 to P112 albeit an elevation on P3; and exon-14 skipping in E17–E20 with downregulation of the resulting mRNA. We tested if the reduced detection of messages without exon 14 could be explained by nonsense-mediated mRNA decay (NMD) vulnerability, but knocking down UPF1, a major component of this pathway, did not increase their quantities. Conversely, it significantly decreased FMR1 mRNA having exon 13 joined with either exon 14 or exon 15 site A. Conclusions The forebrain in the third embryonic week of the rat development is a period with significant skipping of Fmr1 exon 14. This alternative splicing event chronologically precedes a reduction of total Fmr1 mRNA, suggesting that it may be part of combinatorial mechanisms downregulating the gene’s expression in the late embryonic period. The decay of FMR1 mRNA without exon 14 should be mediated by a pathway different from NMD. Finally, we provide evidence of FMR1 mRNA stabilization by UPF1, likely depending on FMRP.
 
Relative expression of differentially expressed mRNA in rat whole blood in the microarray. a Hras were significantly down-regulated in the neuropathic pain group versus the control group after 2 weeks. b Jun was significantly down-regulated in the neuropathic pain group versus the control group after 2 weeks. c Pik3cb were significantly down-regulated in the neuropathic pain group versus the control group after 2 weeks. Data are presented as mean ± SE, *p < 0.05. NP group: neuropathic pain group. The KEGG pathway up regulation histogram. LGP (The P value is logarithmic and negative): the larger the (–LGP) value, the smaller the P value, indicating the higher the significance level of the pathway
The KEGG pathway down regulation histogram. LGP (The P value is logarithmic and negative): the larger the (–LGP) value, the smaller the P value, indicating the higher the significance level of the pathway
Relative expression of differentially expressed mRNA in rat whole blood in the microarray. a Hras were significantly down-regulated in the neuropathic pain group versus the control group after 2 weeks. b Jun was significantly down-regulated in the neuropathic pain group versus the control group after 2 weeks. c Pik3cb were significantly down-regulated in the neuropathic pain group versus the control group after 2 weeks. Data are presented as mean ± SE, *p < 0.05. NP group: neuropathic pain group
Article
The present study aimed to investigate the expression of mRNA in the brachial plexus avulsion neuropathic pain model and analyze biological functions. Microarray mRNA assay and reverse transcriptase quantitative polymerase chain reaction (RT-PCR) were conducted. The whole blood was collected from two groups for Microarray mRNA analysis. The predicted mRNA targets were studied by gene ontology analysis and pathway analysis. We identified 3 targeted mRNAs, including PIK3CB, HRAS, and JUN. The results showed that PIK3CB, HRAS, and JUN gene expression was increased in the control group but decreased in the neuropathic pain group. These findings indicate that certain genes may be important biomarkers for the potential targets for the prevention and treatment of brachial plexus avulsion caused neuropathic pain.
 
Study design. A total of 68 rat pups were divided into 18 experimental groups of 5–7 pups/group, which received from one to nine daily injections of thyroxine (T4) or vehicle, starting from postnatal day 2 (PND2). Groups 1 and 2 received T4 or vehicle, respectively, on PND2 and were sacrificed on PND3, groups 3 and 4 received T4 or vehicle, respectively, on PND2 and PND3 and were sacrificed on PND4, etc. Animals were sampled daily from PND3 to PND11, and brain hemispheres were collected for IHC and RR-ELISA experiments. Diffusion tensor MRI (DTI) was performed on PND10 for vehicle and T4 rat pups (group 17 and group 18)
Diffusion tensor MRI on major white matter structures. a Fractional anisotropy (FA) map with highlighted 10 major white matter brain regions obtained by diffusion tensor imaging in a representative thyroxine (T4)-treated rat at PND10. Scaling of the FA map is between 0 and 0.7. b FA values in 10 different areas of the white matter in rat pups at PND10 following nine daily administrations of T4 (0.1 mg/kg, n = 6) or vehicle (water, n = 6). Data are presented as the mean ± SEM. Statistical significance is indicated as follows: **P < 0.01 (passed correction for the false discovery rate at q set at 0.1). fmi forceps minor of corpus callosum, gcc genu of corpus callosum, bcc body of corpus callosum, scc splenium of corpus callosum, fmj forceps major of corpus callosum, ec external capsule, aca anterior part of anterior commissure, ic internal capsule, opt optic tract, cp. cerebellar peduncle
Global distribution of fractional anisotropy (FA) values in the brains of rat pups at PND10. a Group averages of normalized FA histograms for thyroxine (T4) and vehicle rat pups at PND10; distribution of T4 group FA values shows an apparent shift to higher values. b Analysis of the FA shift by the comparison of Burr distribution fit medians for individual subjects. Data are presented as the mean (thick solid line) ± SEM. *P < 0.05 (Welch’s t-test). The T4 group (n = 6) received T4 at 0.1 mg/kg and the vehicle group (n = 6) received water
T4 facilitates myelination in the developing rat brain. The developmental increase in brain MBP levels in thyroxine (T4)- and vehicle-treated rats aged 3–11 days old was determined by Resonance Raman Spectroscopy ELISA (RRS-ELISA) in homogenized brain lysates prepared from brain hemispheres. a The relationships between brain lysate MBP level and age in the two treatment groups are illustrated. T4 exhibited age-dependent positive effect on MBP levels. Data are presented as the mean ± SEM (n = 5–7). Statistical significance of differences between MBP levels in the two treatment groups at the analyzed age points is indicated as follows: **P < 0.01; ***P < 0.001 (Tukey’s multiple comparisons test following two-way ANOVA). b A significant positive correlation between brain lysate MBP levels and median FA distribution value in rat pups that underwent DTI. Red empty circles and black filled squares denote T4-treated and vehicle-treated pups, respectively
Analysis of immunohistochemical staining for MBP in coronal brain slices of vehicle- and T4-treated rat pups. a Representative examples of MBP protein staining after treatment with thyroxine (T4) or vehicle at PND5–11. b Quantification of the MBP signal optical density at PND5 and PND7–PND11. Six sagittal adjacent brain sections containing corpus callosum rostral to the hippocampus from at least two animals per group were used for this analysis. T4 treatment significantly increased MBP expression levels at PND 9 and 11. Data are presented as the mean ± SEM (n = 6). For the data quantification analysis, the regions of T4 vs. vehicle were compared to using the Student’s t-test and the correction for multiple comparisons was done by controlling for the false discovery rate (FDR; set at q = 0.1) by using the Benjamini–Krieger–Yekutieli procedure implemented in Prism 9. Data are presented as the mean ± SEM. Differences were considered statistically significant if adjusted P < 0.05. Asterisk (*) denotes P values of T4 compared to vehicle for each postnatal day,*P < 0.05 (Student’s t-test). Black and white bars denote T4-treated and vehicle-treated pups, respectively
Article
Background Therapeutic agents stimulating the process of myelination could be beneficial for the treatment of demyelinating diseases, such as multiple sclerosis. The efficient translation of compounds promoting myelination in vitro to efficacy in vivo is inherently time-consuming and expensive. Thyroid hormones accelerate the differentiation and maturation of oligodendrocytes, thereby promoting myelination. Systemic administration of the thyroid hormone thyroxine (T4) accelerates brain maturation, including myelination, during early postnatal development. The objective of this study was to validate an animal model for rapid testing of promyelinating therapeutic candidates for their effects on early postnatal development by using T4 as a reference compound. Methods Daily subcutaneous injections of T4 were given to Sprague Dawley rat pups from postnatal day (PND) 2 to PND10. Changes in white matter were determined at PND10 using diffusion tensor magnetic resonance imaging (DTI). Temporal changes in myelination from PND3 to PND11 were also assessed by quantifying myelin basic protein (MBP) expression levels in the brain using the resonance Raman spectroscopy/enzyme-linked immunosorbent assay (RRS-ELISA) and quantitative immunohistochemistry. Results DTI of white matter tracts showed significantly higher fractional anisotropy in the internal capsule of T4-treated rat pups. The distribution of total FA values in the forebrain was significantly shifted towards higher values in the T4-treated group, suggesting increased myelination. In vivo imaging data were supported by in vitro observations, as T4 administration significantly potentiated the developmental increase in MBP levels in brain lysates starting from PND8. MBP levels in the brain of animals that received treatment for 9 days correlated with the FA metric determined in the same pups in vivo a day earlier. Furthermore, accelerated developmental myelination following T4 administration was confirmed by immunohistochemical staining for MBP in coronal brain sections of treated rat pups. Conclusions T4-treated rat pups had increased MBP expression levels and higher MRI fractional anisotropy values, both indications of accelerated myelination. This simple developmental myelination model affords a rapid test of promyelinating activity in vivo within several days, which could facilitate in vivo prescreening of candidate therapeutic compounds for developmental hypomyelinating diseases. Further research will be necessary to assess the utility of this platform for screening promyelination compounds in more complex demyelination disease models, such us multiple sclerosis.
 
Acute epidural intracranial pressure (ICP) elevation in obstructive hydrocephalic (OHC) animals compared to control animals. A Mean curves showing the postoperative mean continuous epidural ICP (dots representing ± SEM). B Raw ICP trace from the overnight continuous recording in an OHC animal (X-axis in hours and Y-axis in mmHg). C Epidural ICP values from each animal in both groups are plotted at day 0, 1 and 7 (mean is showed with SEM error bars, **** = p < 0.0001). D Epidural ICP correlated with the ventricular ICP values at day 7 (Pearson’s r = 0.89, p < 0.0001)
Optic disc photos showing peripapillary changes in an obstructive hydrocephalic (OHC) animal. A + B ODP from a left eye displaying a normal optic disc at baseline (A) and after 1 week with OHC (B). White arrow is pointing out paleness in the upper temporal peripapillary area. C + D ODP from a left eye displaying a normal optic disc at baseline and after 1 week after sham injection. No considerable changes were observed. A–D Left eyes are illustrated in this figure, but both eyes were evaluated in all animals. E Bar diagram presenting distribution of normal and deformed optic discs in context of hydrocephalus
Ventricular dilation after kaolin insertion in the obstructive hydrocephalic (OHC) animal. A Midbrain slice showing ventricular dilation in an OHC animal 1 week after kaolin insertion. Note the deposition of kaolin at the base of the brain. B Midbrain slice of a control animal 1 week after sham insertion. C Bar diagram presenting distribution of normal and enlarged lateral ventricles in sham (control group) and kaolin (hydrocephalus group) injected animals
Article
Background: The kaolin induced obstructive hydrocephalus (OHC) model is well known for its ability to increase intracranial pressure (ICP) in experimental animals. Papilledema (PE) which is a predominant hallmark of elevated ICP in the clinic has not yet been studied in this model using high-resolution digital fundus microscopy. Further, the long-term effect on ICP and optic nerve head changes have not been fully demonstrated. In this study we aimed to monitor epidural ICP after induction of OHC and to examine changes in the optic disc. In addition, we validated epidural ICP to intraventricular ICP in this disease model. Method: Thirteen male Sprague-Dawley rats received an injection into the cisterna magna containing either kaolin-Ringer's lactate suspension (n = 8) or an equal amount of Ringer's lactate solution (n = 5). Epidural ICP was recorded post-operatively, and then continuously overnight and followed up after 1 week. The final epidural ICP value after 1 week was confirmed with simultaneous ventricular ICP measurement. Optic disc photos (ODP) were obtained preoperatively at baseline and after one week and were assessed for papilledema. Results: All animals injected with kaolin developed OHC and had significant higher epidural ICP (15.49 ± 2.47 mmHg) compared to control animals (5.81 ± 1.33 mmHg) on day 1 (p < 0.0001). After 1 week, the epidural ICP values were subsided to normal range in hydrocephalus animals and there was no significant difference in epidural ICP between the groups. Epidural ICP after 1 week correlated with the ventricular ICP with a Pearson's r = 0.89 (p < 0.0001). ODPs from both groups showed no signs of acute papilledema, but 5 out of 8 (62.5%) of the hydrocephalus animals were identified with peripapillary changes. Conclusions: We demonstrated that the raised ICP at day 1 in the hydrocephalus animals was completely normalized within 1 week and that epidural ICP measurements are valid method in this model. No acute papilledema was identified in the hydrocephalus animals, but the peripapillary changes indicate a potential gliosis formation or an early state of a growing papilledema in the context of lateral ventricle dilation and increased ICP.
 
Article
Background Psychological disorders including depression, anxiety, and stress comprise a huge public health problem. The aim of this cross-sectional study is to assess the relationship between dietary glycemic index (DGI) and glycemic load (DGL) and mental disorders. Method Participants (n = 10,000) aged 20–69 were randomly selected from 200 clusters in Yazd from the recruitment phase of Yazd Health Study. The dietary intake of study participants was collected by a reliable and validated food frequency questionnaire consisting of 178 food items. DGI and DGL were calculated from the FFQ data using previously published reference values. To assess psychological disorders an Iranian validated short version of a self-reported questionnaire Depression Anxiety Stress Scales 21 was used. Results There were no significant associations between DGI and DGL with odds of depression or anxiety in crude and adjusted models. However, individuals in the highest quartiles of DGL had the lowest odds of stress (OR: 0.69; 95% CI 0.47–1, P-trend = 0.023). This association remained significant after adjustment for potential confounding variables in model I (OR: 0.45; 95% CI 0.22–0.9, P-trend = 0.023), model II (OR: 0.46; 95% CI 0.22–0.96, P-trend = 0.039) and model III (OR: 0.46; 95% CI 0.22–0.96, P-trend = 0.042). Conclusion In conclusion, consumption of foods with higher GL was associated with lower odds of stress; however, no significant association was found between DGI or DGL and risk of depression and anxiety. Performing further studies with longitudinal design is suggested to confirm these results.
 
Article
Background Auditory temporal processing plays an important role in speech comprehension. Usually, behavioral tests that require subjects to detect silent gaps embedded within a continuous sound are used to assess the ability of auditory temporal processing in humans. To evaluate auditory temporal processing objectively, the present study aimed to measure the auditory steady state responses (ASSRs) elicited by silent gaps of different lengths embedded within a broadband noise. We presented a broadband noise with 40-Hz silent gaps of 3.125, 6.25, and 12.5 ms. Results The 40-Hz silent gaps of 3.125, 6.25, and 12.5 ms elicited clear ASSRs. Longer silent gaps elicited larger ASSR amplitudes and ASSR phases significantly differed between conditions. Conclusion The 40 Hz gap-evoked ASSR contributes to our understanding of the neural mechanisms underlying auditory temporal processing and may lead to the development of objective measures of auditory temporal acuity in humans.
 
Article
Background Transcranial magneto-acoustical stimulation (TMAS) is a noninvasive technique that has advantages in spatial resolution and penetration depth. It changes the firing properties of neurons through the current generated by focused ultrasound and a static magnetic field. Spike-frequency adaptation is an important dynamic characteristic of neural information processing. Methods To address the effects of TMAS on neural spike-frequency adaptation, this study employs some ultrasound and magnetic field parameters, such as magnetic flux density, ultrasonic intensity, fundamental ultrasonic frequency, modulation frequency, and duty cycle. Using these different ultrasound and magnetic field parameters, membrane potential curves, spike-frequency curves, and adapted onset spike-frequency curves are exhibited and analyzed. Results The results show that spike-frequency adaptation is strongly dependent on ultrasonic intensity and magnetic flux density and is rarely affected by other parameters. However, modulation frequency and duty cycle influence membrane potentials and spike frequencies to some degree. Conclusions This study reveals the mechanism of the effects of TMAS on neural spike-frequency adaptation and serves as theoretical guidance for TMAS experiments.
 
Article
Background Hashimoto’s thyroiditis (HT) is an autoimmune illness that renders individuals vulnerable to neuropsychopathology even in the euthyroid state, the mechanisms involved remain unclear. We hypothesized that activated microglia might disrupt synapses, resulting in cognitive disturbance in the context of euthyroid HT, and designed the present study to test this hypothesis. Methods Experimental HT model was induced by immunizing NOD mice with thyroglobulin and adjuvant twice. Morris Water Maze was measured to determine mice spatial learning and memory. The synaptic parameters such as the synaptic density, synaptic ultrastructure and synaptic-markers (SYN and PSD95) as well as the interactions of microglia with synapses were also determined. Results HT mice had poorer performance in Morris Water Maze than controls. Concurrently, HT resulted in a significant reduction in synapse density and ultrastructure damage, along with decreased synaptic puncta visualized by immunostaining with synaptophysin and PSD-95. In parallel, frontal activated microglia in euthyroid HT mice showed increased engulfment of PSD95 and EM revealed that the synaptic structures were visible within the microglia. These functional alterations in microglia corresponded to structural increases in their attachment to neuronal perikarya and a reduction in presynaptic terminals covering the neurons. Conclusion Our results provide initial evidence that HT can induce synaptic loss in the euthyroid state with deficits might be attributable to activated microglia, which may underlie the deleterious effects of HT on spatial learning and memory.
 
Results of behavior measurement for both groups. A The level of perceptual expertise in the domain of radiological images as assessed by the Radiological Expertise Task. The radiology interns group had a significantly larger AUC than the normal control group (p<0.001, Mann-Whitney test), indicating better visual recognition ability in radiological images; B Response time of both groups in the Radiological Expertise Task; C The level of perceptual expertise in the domain of faces as measured by the Cambridge Face Memory Test. RET Radiological Expertise Task, RIG radiology interns group, NECG normal control group, CMFT Cambridge Face Memory Test. *Indicates significant group differences (p < 0.05)
Samples of behavioral tests. A User interface of in-house software for the Radiological Expertise Task; B Stimulus used in Cambridge Face Memory Test
The ALFF differences between the radiology interns group (n = 22) and the normal control group (n = 22) (p < 0.05, alphasim corrected, RIG v.s. NECG) and voxel-wise correlation map between ALFF and the level of perceptual expertise in the domain of radiological images as assessed by the AUC of Radiological Expertise Task for the inter RIG group. A The intern radiologists group showed higher ALFF in the left OFC (displayed in sagittal view); B The intern radiologists group showed higher ALFF in the right fusiform gyrus (displayed in axial view); C Significant correlation between ALFF and visual recognition expertise was found the in the right fusiform gyrus (p < 0.05, multiple comparison corrected); D The scatter plot map computed as ALFF of the peak voxel (40, 56, − 16) in the correlation analysis and RET scores. Please note that this map is only for illustration purpose, otherwise there would be the risk of a circular analysis. OFC the orbitofrontal cortex, FG the fusiform gyrus
Article
Background visual expertise and experience modulate evoked brain activity in response to training-related stimuli. However, few studies have considered how the visual experience is represented in the resting state brain activity. This study tried to investigate the way visual experience, i.e., visual recognition expertise, modulates baseline brain neuronal activity in the resting state using the model of radiologists. Methods The amplitude of low-frequency (< 0.08 Hz) fluctuation (ALFF) was used as the metric of baseline brain activity and a visual expertise model of radiologists to investigated this question. The visual recognition skill enables them to accurately identify pathological information in medical images. After the behavior measurement, a cohort group of radiology interns (n = 22) and a group of matched layperson (n = 22) were selected for inclusion in the study. The resting state functional magnetic resonance imaging (fMRI) scans were performed for all of the subjects. Results Higher ALFF in the right fusiform gyrus and the left orbitofrontal cortex were observed, and the ALFF in the fusiform gyrus was correlated with the intern radiologists’ behavioral expertise(all results corrected for multiple comparisons). Conclusions Visual experience modulates the baseline brain activity in both high-level visual cortex and high-order cognitive cortex, indicating the engagement of both top-down and bottom-up facilitation. We provide a novel perspective to how visual experience modulated cortical brain activity by introducing the resting state changes. Also, we propose that our current study may provide novel ideas for the development of new training protocols in medical school.
 
Experimental design. Transient middle cerebral artery occlusion (tMCAO) was induced in female Wistar rats at day 0 and the in vivo part of the study was designed for a period of two weeks. tMCAO was induced in animals that were under low influence of 17β-estradiol during their estrous cycle. Animals were acclimatized with animal facility conditions for 5 days before any in vivo experiments. At day 14 after reperfusion, the animals were euthanized and decapitated. CCA common carotid artery; ECA external carotid artery; ICA internal carotid artery; MCA middle cerebral artery
Spontaneous functional recovery after ischemic stroke. A Spontaneous recovery from day 2 to 14 after tMCAO was observed by 28-point test B neurological score of animals from 0 for healthy function to 5 for death overnight is shown. Neurological function of the animals was significantly affected by tMCAO, however, the animals showed significant functional improvement at day 14 after tMCAO. *Significance level at p < 0.05 for comparison between day -1 (pre-stroke) and day 14. #Significance level at p < 0.05 for comparison between day 1 (for 6-point test) and 2 (for 28-point test) vs day 14. Number of rats in both tests; n = 8. C Infarct distribution in representative silver infarct staining of brain sections. Lesioned area is marked in red on right hemisphere
ERK1/2 and Akt pathways are affected at day 14 after tMCAO. A Representative western blots of p-ERK1/2 and t-ERK1/2 for peri-infarct and ischemic core vs. contralateral samples. B Quantification of p-ERK1/2 (normalized to t-ERK1/2 and β-actin—left graph) and t-ERK1/2 (normalized to β-actin—right graph) in peri-infarct and ischemic core compared to contralateral. C Representative western blots of p-Akt and Akt for peri-infarct vs contralateral (left panel) and ischemic core vs contralateral (right panel). D Quantification of p-Akt (normalized to Akt and β-actin) in peri-infarct and ischemic core showed significant increase compared to their related contralateral samples (left graph). Akt expression was significantly lower in peri-infarct and ischemic core compared to contralateral (right graph). Number of animals for ischemic core; n = 7, number of animals for peri-infarct; n = 5. AU arbitrary units, *Significant differences (p < 0.05). The presented western blot images are cropped and the full-length blots/gels are presented in Additional file 2: Figure S1
Expression of Nestin, NeuN, Tie-2, IL-10 and TGF-β in brain at day 14 after tMCAO. A, B Representative western blots and quantification of NeuN, nestin and TGF-β in peri-infarct and ischemic core vs. contralateral side. C, D Representative western blots and quantification of Tie-2 and IL-10 in peri-infarct and ischemic core vs contralateral side. All markers were normalized to β-actin. Ischemic core; n = 7, peri-infarct; n = 5. AU arbitrary units, *Significant differences between peri-infarct or ischemic core and contralateral (p < 0.05). The presented western blot images are cropped and the full-length blots/gels are presented in Additional file 3: Figure S2 (Nestin and NeuN) and Additional file 4: Figure S3 (Tie-2, IL-10 and TGF-β)
Ischemia significantly increased expression of TGF-β in the smooth muscle cell layer of occluded MCAs. A Representative image of TGF-β expression at day 14 after tMCAO in the occluded and non-occluded MCAs. B Quantification of TGF-β mean fluorescence intensity in occluded vs non-occluded MCAs at day 14 after tMCAO; n = 5, *p < 0.05
Article
Background Some degree of spontaneous recovery is usually observed after stroke. Experimental studies have provided information about molecular mechanisms underlying this recovery. However, the majority of pre-clinical stroke studies are performed in male rodents, and females are not well studied. This is a clear discrepancy when considering the clinical situation. Thus, it is important to include females in the evaluation of recovery mechanisms for future therapeutic strategies. This study aimed to evaluate spontaneous recovery and molecular mechanisms involved in the recovery phase two weeks after stroke in female rats. Methods Transient middle cerebral artery occlusion was induced in female Wistar rats using a filament model. Neurological functions were assessed up to day 14 after stroke. Protein expression of interleukin 10 (IL-10), transforming growth factor (TGF)-β, neuronal specific nuclei protein (NeuN), nestin, tyrosine-protein kinase receptor Tie-2, extracellular signal-regulated kinase (ERK) 1/2, and Akt were evaluated in the peri-infarct and ischemic core compared to contralateral side of the brain at day 14 by western blot. Expression of TGF-β in middle cerebral arteries was evaluated by immunohistochemistry. Results Spontaneous recovery after stroke was observed from day 2 to day 14 and was accompanied by a significantly higher expression of nestin, p-Akt, p-ERK1/2 and TGF-β in ischemic regions compared to contralateral side at day 14. In addition, a significantly higher expression of TGF-β was observed in occluded versus non-occluded middle cerebral arteries. The expression of Tie-2 and IL-10 did not differ between the ischemic and contralateral sides. Conclusion Spontaneous recovery after ischemic stroke in female rats was coincided by a difference observed in the expression of molecular markers. The alteration of these markers might be of importance to address future therapeutic strategies.
 
Article
Background The preventive role of muscular strength on diminishing neuroinflammation is yet unknown. In this study, the role of the prophylactic muscular strength exercise was investigated in order to verify whether it would diminish cognitive alterations and modify the antioxidant intracellular scenery in an animal neuroinflammatory model in of the CA1 region of the hippocampus. Methods The animals received muscular strength training (SE) three times a week for eight weeks. Subsequently, the stereotaxic surgery was performed with an intra-hippocampal infusion of either saline solution (SAL) or lipopolysaccharide (LPS). Next, we performed the behavioral tests: object recognition and social recognition. Then, the animals were euthanized, and their hippocampus and prefrontal cortex were collected. In another moment, we performed the dosage of the antioxidant activity and histological analysis. Results The results showed that the muscular strength exercises could show a beneficial prophylactic effect in the cognitive deficiencies caused by acute neuroinflammation. Regarding oxidative stress, there was an increase in catalase enzyme activity (CAT) in the group (SE + LPS) compared to the control groups (p < 0.05). As for the cognitive alterations, there were found in the (SE + LPS) group, diminishing the mnemonic hazard of the discriminative and social memories compared to the control groups (p < 0.05). Conclusion We concluded, therefore, that the exercise performed prophylactically presents a protective effect capable of minimizing such mnemonic deficits and increasing catalase enzyme activity in rats that suffered a local neuroinflammatory process in the hippocampus.
 
Gating strategies and detection of LAG-3 expression by FITC conjugated LAG-3 antibody. A Gating strategy of isolating single CD11⁺/CD45low cells from the brains of PBS injected mice. Single-cell suspension from the brains of naïve mice involved pre-gating by forward scatter area (FSC-A) vs. side scatter area (SSC-A) plot, and forward scatter height (FSC-H) vs. FSC-A density plot to select singlets. Microglia were then identified by the high expression of CD11b and the low expression of CD45. B Distribution of fluorescence intensities from FITC conjugated LAG-3 antibody and isotype control antibody treated microglia. C Gating strategy of isolating single CD11⁺/CD45low microglia cells and CD11⁻/CD45high lymphocytes from the brains of LPS injected mice. D, E Distribution of fluorescence intensities from FITC conjugated LAG-3 and isotype control antibody-treated microglia (D) and lymphocytes (E) from LPS injected mice. All data were replicated in four independent experiments
PE-conjugated antibody outperforms FITC and APC conjugated antibodies in detecting microglia LAG3. A Distribution of fluorescence intensities from FITC conjugated isotype control antibody labeled microglia with the previous blocking by mouse IgG, CD16/CD32/CD16.2 antibodies, or the combination of mouse IgG and CD16/CD32/CD16.2 antibodies. B Distribution of fluorescence intensities from nontreated microglia and isotype control antibody treated microglia. C Distribution of fluorescence intensities from nontreated microglia and FITC, PE, and APC conjugated LAG-3 antibodies and their respective isotype control antibodies treated microglia. D Median fluorescence intensity (MFI) of microglia as treated in (C). E Comparison of MFI LAG-3/MFI IgG of FITC, PE, and APC conjugated LAG-3 antibodies (n = 4 mice). Bars represent the means ± standard deviation (SD). Comparisons were made by one-way ANOVA test with Tukey’s post hoc multiple comparisons test. *P < 0.05; **P < 0.01; NS, not significantly different
PE-conjugated antibody outperforms FITC and APC conjugated antibodies in detecting microglia TIM-3. A Distribution of fluorescence intensities from nontreated microglia, FITC, PE, and APC conjugated TIM-3 antibodies treated, and their respective isotype control antibody-treated microglia. B The MFI of microglia treated as in (A). C Comparison of MFI TIM-3/MFI IgG of FITC, PE, and APC conjugated TIM-3 antibodies (n = 5 mice). Bars represent means ± SD. Comparisons were made by one-way ANOVA test with Tukey’s post hoc multiple comparisons test. *P < 0.05; **P < 0.01; NS, not significantly different
Imaging of autofluorescent granules in microglia on fixed brain slices. A Diagram illustrates the imaging of hippocampus CA1 microglia on fixed brain slices. B Representative 3D reconstructed images of a microglia from 2 months old Cx3cr1GFP mice with autofluorescence (AF) imaged in TRITC channel (561 nm laser, 570–620 nm band pass filter). C Representative 3D reconstructed images of a microglia stained with CD68 from Cx3cr1GFP mice. The arrow points to a CD68⁺ lysosome containing an autofluorescent granule in soma (AF +); while the arrowhead points to CD68⁺ lysosomes located in process without autofluorescent granules (AF-). D Volume of AF⁺ and AF⁻ CD68 + lysosomes as in panel (C) (n = 15 cells from 3 mice, Mann–Whitney test). E Images of an autofluorescent granule from a GFP expressing microglia through continuous emission filter excited by 566 nm laser and its corresponding emission spectrum (n = 8 cells from 3 mice). F Images of an autofluorescent granule from a microglia in wild type mouse labeled by Iba-1 antibody through continuous emission filter excited by 488 nm laser and its corresponding emission spectrum. The emission spectrum of FITC, PE, and APC were shown for comparison (n = 8 cells from 3 mice). G Z projected stack images of microglia from Cx3cr1GFP mouse with autofluorescence imaged as in panel (B). H Z projected stack images of microglia on slices with the treatment of TrueBlack® lipofuscin autofluorescence quencher before imaging. I Values of autofluorescence in microglial somata as in (G) and (H) (n = 17 cells from 3 mice, 2 tailed t-test). Bars represent means ± SD. ***P < 0.001
Imaging of autofluorescent granules in microglia on live slices and in vivo. A Diagram illustrates the strategy to image calcium signals in hippocampus CA1 microglia cells on live brain slices; B A representative Z-projected image of 60 GCaMP6s frames during 15 min imaging of a microglia cell on Cx3cr1CreER: GCaMP6s mouse brain slice. The arrow points to the soma of a microglia cell, which contains an autofluorescent granule. ROI1 circles the soma of microglia, excluding the autofluorescent granule, while ROI2 including it. Right top: the GCaMP6s image in basal condition. Right bottom: GCaMP6s image during application of 100 µM UDP. C F/F0 in ROI1 and ROI2 during 15 min imaging, at 5 min of which 100 µM UDP was applied. D The peak of F/F0 of GCaMP6s signals during UDP application in ROIs either excluding (ROI1) or including (ROI2) the autofluorescent granules (n = 5 cells from 3 mice, paired t-test). E Emission spectrum of the autofluorescent puncta as in panel (B) by spectral imaging (n = 5 cells from 3 mice). F Representative images of a microglia from Cx3cr1GFP mice with GFP and autofluorescence simultaneously imaged by 2-photon microscopy through cranial window. The white arrows point to the autofluorescent granule in different time points. Bars represent means ± SD
Article
Background Microglia, the resident immune cells in the central nervous system, accrue autofluorescent granules inside their cytoplasm throughout their lifespan. In this report, we studied the impacts of autofluorescence on widely used fluorescence-based techniques to study microglia, including flow cytometry, immunofluorescence staining, and live imaging. Results The failed attempt of using fluorescein isothiocyanate (FITC) conjugated antibody to detect lymphocyte-activation gene 3 protein in microglia prompted us to compare the sensitivity of FITC, phycoerythrin (PE) and allophycocyanin (APC) conjugated antibodies to detect surface protein expression in microglia. We found that PE outperformed FITC and APC as the fluorophore conjugated to antibody for flow cytometry by overcoming the interference from microglia autofluorescence. To identify the location and source of microglia autofluorescence, we did confocal imaging and spectral analysis of microglia autofluorescence on fixed brain tissues, revealing that microglia autofluorescence emitted from cytoplasmic granules and displayed a multi-peak emission spectrum. We recommended removing autofluorescence by lipofuscin removing agents when staining intracellular proteins in microglia with the immunofluorescence techniques. On live brain slices, autofluorescent granules reduced the amplitudes of calcium signals in microglial somata derived from GCaMP6s fluorescence and thus needed to be excluded when selecting regions of interest (ROI). Conclusions In conclusion, autofluorescence is a critical factor to consider when designing experiments and interpreting results based on fluorescence-based techniques to study microglia.
 
Article
Background C-C motif chemokine ligand 2 (CCL2) is reported to be involved in the pathogenesis of various neurological and/or psychiatric diseases. Tissue or cellular expression of CCL2, in normal or pathological condition, may play an essential role in recruiting monocytes or macrophages into targeted organs, and be involved in a certain pathogenic mechanism. However, few studies focused on tissue and cellular distribution of the CCL2 peptide in brain grey and white matters (GM, WM), and the changes of the GM and WM cellular CCL2 level in septic or endotoxic encephalopathy was not explored. Hence, the CCL2 cellular distribution in the front brain cortex and the corpus callosum (CC) was investigated in the present work by using immunofluorescent staining. Results (1) CCL2 like immunoreactivity (CCL2-ir) in the CC is evidently higher than the cortex. When the measurement includes ependymal layer attached to the CC, CCL2-ir intensity is significantly higher than cortex. (2) Structures in perivascular areas, most of them are GFAP positive, contribute major CCL2-ir positive profiles in both GM and WM, but apparently more in the CC, where they are bilaterally distributed in the lateral CC between the cingulate cortex and ventricles. (3) The neuron-like CCL2-ir positive cells in cortex are significantly more than in the CC, and that number is significantly increased in the cortex following systemic lipopolysaccharide (LPS), but not in the CC. (4) In addition to CCL2-ir positive perivascular rings, more CCL2-ir filled cashew shape elements are observed, probably inside of microvasculature, especially in the CC following systemic LPS. (5) Few macrophage/microglia marker-Iba-1 and CCL2-ir co-labeled structures especially the soma is found in normal cortex and CC; the co-localizations are significantly augmented following systemic LPS, and co-labeled amoeba like somata are presented. (6) CCL2-ir and astrocyte marker GFAP or Iba-1 double labeled structures are also observed within the ependymal layer. No accumulation of neutrophils was detected. Conclusion There exist differences in the cellular distribution of the CCL2 peptide in frontal cortex GM and subcortical WM–CC, in both the physiological condition and experimental endotoxemia. Which might cause different pathological change in the GM and WM.
 
Article
Background Deoxythymidine triphosphate (dTTP) is an essential building block of DNA, and defects in enzymes involved in dTTP synthesis cause neurodegenerative disorders. For instance, mutations in DTYMK, the gene coding for thymidylate kinase (TMPK), cause severe microcephaly in human. However, the mechanism behind this is not well-understood. Here we used the zebrafish model and studied (i) TMPK, an enzyme required for both the de novo and the salvage pathways of dTTP synthesis, and (ii) thymidine kinases (TK) of the salvage pathway in order to understand their role in neuropathology. Results Our findings reveal that maternal-stored dNTPs are only sufficient for 6 cell division cycles, and the levels of dNTPs are inversely correlated to cell cycle length during early embryogenesis. TMPK and TK activities are prominent in the cytosol of embryos, larvae and adult fish and brain contains the highest TMPK activity. During early development, TMPK activity increased gradually from 6 hpf and a profound increase was observed at 72 hpf, and TMPK activity reached its maximal level at 96 hpf, and remained at high level until 144 hpf. The expression of dtymk encoded Dtymk protein correlated to its mRNA expression and neuronal development but not to the TMPK activity detected. However, despite the high TMPK activity detected at later stages of development, the Dtymk protein was undetectable. Furthermore, the TMPK enzyme detected at later stages showed similar biochemical properties as the Dtymk enzyme but was not recognized by the Dtymk specific antibody. Conclusions Our results suggest that active dNTP synthesis in early embryogenesis is vital and that Dtymk is essential for neurodevelopment, which is supported by a recent study of dtymk knockout zebrafish with neurological disorder and lethal outcomes. Furthermore, there is a novel TMPK-like enzyme expressed at later stages of development.
 
Article
Background Hematoma expansion can be related to increased mortality and poor clinical outcomes in patients with intracerebral hemorrhage (ICH). So, early identification and prevention of hematoma expansion can be considered as an important therapeutic aim. This study aimed to evaluate the hypothesis that the neutrophil to lymphocyte ratio (NLR) is associated with hematoma expansion in ICH patients. Methods We retrospectively evaluated the clinical data of a total of 221 patients with ICH who were treated in our department between April 2018 and April 2021. The demographic, clinical, radiological, and laboratory test data including the NLR upon admission were investigated. A binary logistic regression analysis was used to assess the independent associations between different variables and hematoma expansion. Results A total of 221 patients with ICH were included. There were 122 (55.2%) males and 99 (44.8%) females. The mean age (years) at admission was 66.43 ± 8.28. The hematoma expansion occurred in 57 (25.8%) cases. The results of the multivariate analysis showed that hematoma volume at baseline (OR, 3.12; 95% CI 1.78–5.02; P < 0.001), admission systolic blood pressure (OR, 2.87; 95% CI 1.79–4.34; P = 0.013), Glasgow Coma Scale (GCS) (OR, 1.94; 95% CI 1.45–2.93; P = 0.020), and NLR (OR, 1.74; 95% CI 1.16–2.60; P = 0.032) were correlated with hematoma expansion in these patients. Conclusions Our findings suggest that NLR can be a predictor of hematoma expansion in patients with ICH. This cost-effective and easily available biomarker could be used to early prediction of hematoma expansion in these patients.
 
Results of neural connectivity between the VN of pons and vestibular-related areas (parietal lobe, PIVC, thalamus, and cerebellum) in patients with cerebellar injury, at thresholds of 10 streamlines as determined by DTI. The control showed a subject out of six
Results of neural connectivity between the VN of medulla oblongata and vestibular-related areas (parietal lobe, PIVC, thalamus, and cerebellum) in patients with cerebellar injury, at thresholds of 10 streamlines as determined by DTI. The control showed a subject out of six
Article
Objective Cerebellar injury can not only cause gait and postural instability, nystagmus, and vertigo but also affect the vestibular system. However, changes in connectivity regarding the vestibular projection pathway after cerebellar injury have not yet been reported. Therefore, in the current study, we investigated differences in the connectivity of the vestibular projection pathway after cerebellar injury using diffusion tensor imaging (DTI) tractography. Methods We recruited four stroke patients with cerebellar injury. Neural connectivity in the vestibular nucleus (VN) of the pons and medulla oblongata in patients with cerebellar injury was measured using DTI. Connectivity was defined as the incidence of connection between the VN on the pons and medulla oblongata and target brain regions such as the cerebellum, thalamus, parieto-insular vestibular cortex (PIVC), and parietal lobe. Results At thresholds of 10 and 30, there was lower connectivity in the ipsilateral hemisphere between the VN at the medullar level and thalamus in the patients than in healthy adults. At a threshold of 1 and 10, the patient group showed lower VN connectivity with the PIVC than healthy adults. At a threshold of 1, VN connectivity with the parietal lobe in the contralateral hemisphere was lower in the patients than in healthy adults. Additionally, at a threshold of 30, VN connectivity at the pons level with the cerebellum was lower in healthy adults than in the patients. Conclusion Cerebellar injury seems to be associated with decreased vestibular projection pathway connectivity, especially in the ipsilateral thalamus, PIVC, and contralateral parietal lobe.
 
Article
Background Spinal cord injury (SCI) is a life-threatening traumatic disorder. Paeonol has been confirmed to be involved in a variety of diseases. The purpose of this study is to investigate the role of paeonol on SCI progression. Methods Sprague Dawley (SD) rat was used for the establishment of SCI model to explore the anti-inflammation, anti-oxidation, and neuroprotective effects of paeonol (60 mg/kg) on SCI in vivo. For in vitro study, mouse primary microglial cells (BV-2) were induced by lipopolysaccharide (LPS)/adenosine triphosphate (ATP) treatment. The effect of paeonol on the polarization of LPS/ATP-induced BV-2 cells was determined by detection the expression inducible nitric oxide synthase (iNOS), tumour necrosis factor alpha (TNF-α), arginase-1 (Arg-1), and interleukin (IL)-10 using qRT-PCR. ELISA was used to assess the levels of IL-1β, IL-18, TNF-α, malondialdehyde (MDA), and glutathione (GSH). Western blotting was conducted to determine the levels of NLRP3 inflammasomes and TLR4/MyD88/NF-κB (p65) pathway proteins. Results Paeonol promoted the recovery of locomotion function and spinal cord structure, and decreased spinal cord water content in rats following SCI. Meanwhile, paeonol reduced the levels of apoptosis-associated speck-like protein (ASC), NLRP3, active caspase 1 and N-gasdermin D (N-GSDMD), repressed the contents of IL-1β, IL-18, TNF-α and MDA, and elevated GSH level. In vitro, paeonol exerted similarly inhibiting effects on pyroptosis and inflammation. Meanwhile, paeonol promoted BV-2 cells M2 polarization. In addition, paeonol also inactivated the expression of TLR4/MyD88/NF-κB (p65) pathway. Conclusion Paeonol may regulate NLRP3 inflammasomes and pyroptosis to alleviate SCI, pointing out the potential for treating SCI in clinic.
 
Brain areas exhibited significantly different FCs with the right A1 in PAs (postoperative vs preoperative)
Brain areas exhibited significantly different FCs with the broca in PAs (postoperative vs preoperative)
Brain areas exhibited significantly different FCs with the DMN in PAs (postoperative vs preoperative)
Brain areas exhibited significantly different FCs with the ECN in PAs (postoperative vs preoperative)
Brain areas exhibited significantly different FCs with the SN in PAs (postoperative vs preoperative)
Article
Background Pituitary adenoma (PA) may compress the optic apparatus, resulting in impaired vision. Some patients can experience improved vision rapidly after surgery. During the early period after surgery, however, the change in neurofunction in the extravisual cortex and higher cognitive cortex has yet to be explored. Objective Our study focused on the changes in the extravisual resting-state networks in patients with PA after vision restoration. Methods We recruited 14 patients with PA who experienced visual improvement after surgery. The functional connectivity (FC) of 6 seeds [auditory cortex (A1), Broca’s area, posterior cingulate cortex (PCC) for the default mode network (DMN), right caudal anterior cingulate cortex for the salience network (SN) and left dorsolateral prefrontal cortex for the executive control network (ECN)] were evaluated. A paired t test was conducted to identify the differences between two groups of patients. Results Compared with their preoperative counterparts, patients with PA with improved vision exhibited decreased FC with the right A1 in the left insula lobule, right middle temporal gyrus and left postcentral gyrus and increased FC in the right paracentral lobule; decreased FC with the Broca in the left middle temporal gyrus and increased FC in the left insula lobule and right thalamus; decreased FC with the DMN in the right declive and right precuneus; increased FC in right Brodmann area 17, the left cuneus and the right posterior cingulate; decreased FC with the ECN in the right posterior cingulate, right angular and right precuneus; decreased FC with the SN in the right middle temporal gyrus, right hippocampus, and right precuneus; and increased FC in the right fusiform gyrus, the left lingual gyrus and right Brodmann area 19. Conclusions Vision restoration may cause a response of cross-modal plasticity and multisensory systems related to A1 and the Broca. The DMN and SN may be involved in top-down control of the subareas within the visual cortex. The precuneus may be involved in the DMN, ECN and SN simultaneously.
 
The mRNA levels of MYD88±SEM in the blood of AT (N = 28) and WT (N = 22) mice (FC = 2.04, p < 0.001). Comparison between mice groups was performed with the Mann Whitney test, and differences were considered significant for p < 0.05
MYD88 mRNA levels ±SEM in the blood of WT (N = 22) and AT (N = 28) mice in different age categories. Comparison between groups was performed using the Mann Whitney test, and differences were considered significant for p < 0.05
MYD88 mRNA blood levels at different time points before and during Rivastigmine treatment: T0-before treatment, T1-at 20 days, T2-at 50 days after treatment initiation. The mean±SD of mice age at the investigated time points during therapy is mentioned. Bars represent the mean of MYD88 levels±SEM. Comparison among T0, T1 and T2 was performed using Repeated Measures ANOVA with the Bonferroni Corrected t-tests, and differences were considered significant for p < 0.05. RIV Rivastigmine
Article
Background Neuroinflammation plays a prominent role in Alzheimer’s disease (AD), both in pathogenesis and disease progression. It has been shown that TLR/MYD88 signaling is involved in the chronic low-grade sterile inflammation associated with AD. Several studies have evidenced high levels of MYD88 in the brain of patients and animal models of AD, but no study has assessed so far its levels in blood. Methods In this study we evaluated the blood mRNA levels of MYD88 in a mouse model of AD, and also the putative effect of Rivastigmine treatment on MYD88 expression. Twenty-eight transgenic APP/TAU mice (AT) and twenty-two control C57/BL6j mice (WT) were included in this study, out of which five transgenic AT and five WT mice were treated with Rivastigmine. Results Increased MYD88 transcript in the whole blood from AT mice as compared to WT controls was found, which seems to increase in time due to disease progression and not to aging. This finding suggests that blood leukocytes are primed to develop TLR/MYD-mediated inflammatory processes. Moreover, results indicate that MYD88 blood levels were not modulated by the diseases-specific treatment with Rivastigmine. Conclusions Our results suggest that MYD88 might be a promising blood biomarker to monitor AD progression.
 
Self-grooming times for WT, cHET and cKO mice in a home cage and novel environment. Self-grooming times are different in novel versus home environments overall (p = 0.0002, two-way ANOVA). WT and cHET mice exhibit differences in grooming behavior in a novel versus home environment (** p = 0.0049, * p = 0.0143, Šidák's multiple comparisons) while cKO mice do not (p = 0.6896, Šidák's multiple comparisons). Insets: Female (top) and male (bottom) grooming times in WT, cHET and cKO groups. See Table 1 and Additional file 2: Table S1 for means ± SEM
Elevated plus maze (EPM), open field test (OFT) and marble burying test (MBT) results among WT, cHET and cKO mice. All p > 0.13 (two-way ANOVA); see Table 2 for means ± SEM
Sociability test: time spent within 1 cm of a novel object (Obj) or a stranger mouse (S1). Times spent near Obj vs S1 are different overall (p = 0.0002, two-way ANOVA). WT and cHET mice exhibit differences in time spent with Obj vs S1 (i.e., “are sociable”; **p = 0.0026, *p = 0.0132, Šidák's multiple comparisons) while cKO mice do not (p = 0.7917, Šidák's multiple comparisons). See Table 3 for means ± SEM. Inset: Schematic (not to scale) of the experimental setup for sociability experiments. An object (Obj) was placed in one of the circles marked “α” or “β”, and a stranger mouse (S1) was kept inside the other circle. Experimental mice had the freedom to move around all three chambers
Article
Background Autism spectrum disorder (ASD) is characterized by repetitive behaviors, deficits in communication, and overall impaired social interaction. Of all the integrin subunit mutations, mutations in integrin β3 ( Itgb3 ) may be the most closely associated with ASD. Integrin β3 is required for normal structural plasticity of dendrites and synapses specifically in excitatory cortical and hippocampal circuitry. However, the behavioral consequences of Itgb3 function in the forebrain have not been assessed. We tested the hypothesis that behaviors that are typically abnormal in ASD—such as self-grooming and sociability behaviors—are disrupted with conditional Itgb3 loss of function in forebrain circuitry in male and female mice. Methods We generated male and female conditional knockouts (cKO) and conditional heterozygotes (cHET) of Itgb3 in excitatory neurons and glia that were derived from Emx1 -expressing forebrain cells during development. We used several different assays to determine whether male and female cKO and cHET mice have repetitive self-grooming behaviors, anxiety-like behaviors, abnormal locomotion, compulsive-like behaviors, or abnormal social behaviors, when compared to male and female wildtype (WT) mice. Results Our findings indicate that only self-grooming and sociability are altered in cKO, but not cHET or WT mice, suggesting that Itgb3 is specifically required in forebrain Emx1 -expressing cells for normal repetitive self-grooming and social behaviors. Furthermore, in cKO (but not cHET or WT), we observed an interaction effect for sex and self-grooming environment and an interaction effect for sex and sociability test chamber. Limitations While this study demonstrated a role for forebrain Itgb3 in specific repetitive and social behaviors, it was unable to determine whether forebrain Itgb3 is required for a preference for social novelty, whether cHET are haploinsufficient with respect to repetitive self-grooming and social behaviors, or the nature of the interaction effect for sex and environment/chamber in affected behaviors of cKO. Conclusions Together, these findings strengthen the idea that Itgb3 has a specific role in shaping forebrain circuitry that is relevant to endophenotypes of autism spectrum disorder.
 
Article
Background Mitoxantrone has proved efficacy in treatment of multiple sclerosis (MS). The fact that physical exercise could slow down the progression of disease and improve performance is still a debatable issue, hence; we aimed at studying whether combining mitoxantrone with exercise is of value in the management of MS. Methods Thirty-six male rats were divided into sedentary and exercised groups. During a 14-day habituation period rats were subjected to exercise training on a rotarod (30 min/day) before Experimental Autoimmune Encephalomyelitis (EAE) induction and thereafter for 17 consecutive days. On day 13 after induction, EAE groups (exercised &sedentary) were divided into untreated and mitoxantrone treated ones. Disease development was evaluated by motor performance and EAE score. Cerebrospinal fluid (CSF) was used for biochemical analysis. Brain stem and cerebellum were examined histopathological and immunohistochemically. Results Exercise training alone did not add a significant value to the studied parameters, except for reducing Foxp3 immunoreactivity in EAE group and caspase-3 in the mitoxantrone treated group. Unexpectedly, exercise worsened the mitoxantrone effect on EAE score, Bcl2 and Bax. Mitoxantrone alone decreased EAE/demyelination/inflammation scores, Foxp3 immunoreactivity, and interleukin-6, while increased the re-myelination marker BDNF without any change in tumor necrosis factor-α. It clearly interrupted the apoptotic pathway in brain stem, but worsened EAE mediated changes of the anti-apoptotic Bcl2 and pro-apoptotic marker Bax in the CSF. Conclusions The neuroprotective effect of mitoxantrone was related with remyelination, immunosuppressive and anti-inflammatory potentials. Exercise training did not show added value to mitoxantrone, in contrast, it disrupts the apoptotic pathway.
 
Sequence of steps involved in evaluating calmodulin binding domains in suspect proteins. Putative domains and binding motifs in suspect proteins were determined by performing a Calmodulin Target Database scan followed by a visual scan of the identified domains
Calcium-dependent and independent calmodulin binding motifs. CaM, calmodulin; CaMBPs, CaM-binding proteins; CaMBDs, CaM-binding domains. Amino acids: A, Alanine; R, Arginine; Q, Glutamine; I, Isoleucine; L, Leucine; K, Lysine; F, Phenylalanine; W, Tryptophan; Y, Tyrosine; V, Valine; X, Any
Calcium-dependent calmodulin binding domains and motifs identified in human neuroinflammation proteins. ABCA7, ATP-binding cassette transporter subfamily A member A7; CD33, Myeloid cell surface antigen CD33; CH3L1/YKL-40, Chitinase-3-like protein 1; CLU, Clusterin; CR1, Complement receptor type 1; EPHA1, Ephrin type-A receptor 1; M4A4E, Membrane-spanning 4-domains subfamily A member 4E; M4A6A, Membrane-spanning 4-domains subfamily A member 6A; NLRP3, NACHT, LRR and PYD domains-containing protein 3; PILRA, Paired immunoglobulin-like type 2 receptor alpha; TREM2, Triggering receptor expressed on myeloid cells 2. ¹From O’Day, 2015; reanalyzed here. Green highlights: hydrophobic amino acids associated with binding motifs
Calmodulin binding proteins linked to neuroinflammation in specific neurodegenerative diseases. See text for details
Article
Calcium dysregulation (“Calcium Hypothesis”) is an early and critical event in Alzheimer’s and other neurodegenerative diseases. Calcium binds to and regulates the small regulatory protein calmodulin that in turn binds to and regulates several hundred calmodulin binding proteins. Initial and continued research has shown that many calmodulin binding proteins mediate multiple events during the onset and progression of Alzheimer’s disease, thus establishing the “Calmodulin Hypothesis”. To gain insight into the general applicability of this hypothesis, the involvement of calmodulin in neuroinflammation in Alzheimer’s, amyotrophic lateral sclerosis, Huntington’s disease, Parkinson’s disease, frontotemporal dementia, and other dementias was explored. After a literature search for calmodulin binding, 11 different neuroinflammatory proteins (TREM2, CD33, PILRA, CR1, MS4A, CLU, ABCA7, EPHA1, ABCA1, CH3L1/YKL-40 and NLRP3) were scanned for calmodulin binding domains using the Calmodulin Target Database. This analysis revealed the presence of at least one binding domain within which visual scanning demonstrated the presence of valid binding motifs. Coupled with previous research that identified 13 other neuroinflammation linked proteins (BACE1, BIN1, CaMKII, PP2B, PMCA, NOS, NMDAR, AchR, Ado A2AR, Aβ, APOE, SNCA, TMEM175), this work shows that at least 24 critical proteins involved in neuroinflammation are putative or proven calmodulin binding proteins. Many of these proteins are linked to multiple neurodegenerative diseases indicating that calmodulin binding proteins lie at the heart of neuroinflammatory events associated with multiple neurodegenerative diseases. Since many calmodulin-based pharmaceuticals have been successfully used to treat Huntington’s and other neurodegenerative diseases, these findings argue for their immediate therapeutic implementation.
 
Representative profiles of TRH and TRH-like peptide responses in male rats to RF treatment. The response patterns in A and D are consistent with rapid and sustained increase in peptide release (reduced peptide level). The profile in B suggests rapid and sustained decrease in peptide release during and after RF exposure. C could be explained by RF stimulation of sustained peptide release which is compensated by increased peptide synthesis. Withdrawal of RF reduced peptide release but the compensatory increase in peptide synthesis results in a rebound increase in peptide content. The persistence of changes in TRH and TRH-like peptide levels in the WD group after RF has completely cleared from the GI tract is consistent with lingering effects of an altered microbiome
Article
Background The TRH/TRH-R1 receptor signaling pathway within the neurons of the dorsal vagal complex is an important mediator of the brain-gut axis. Mental health and protection from a variety of neuropathologies, such as autism, Attention Deficit Hyperactivity Disorder, Alzheimer’s and Parkinson’s disease, major depression, migraine and epilepsy are influenced by the gut microbiome and is mediated by the vagus nerve. The antibiotic rifaximin (RF) does not cross the gut-blood barrier. It changes the composition of the gut microbiome resulting in therapeutic benefits for traveler’s diarrhea, hepatic encephalopathy, and prostatitis. TRH and TRH-like peptides, with the structure pGlu-X-Pro-NH2, where “X” can be any amino acid residue, have reproduction-enhancing, caloric-restriction-like, anti-aging, pancreatic-β cell-, cardiovascular-, and neuroprotective effects. TRH and TRH-like peptides occur not only throughout the CNS but also in peripheral tissues. To elucidate the involvement of TRH-like peptides in brain-gut-reproductive system interactions 16 male Sprague–Dawley rats, 203 ± 6 g, were divided into 4 groups (n = 4/group): the control (CON) group remained on ad libitum Purina rodent chow and water for 10 days until decapitation, acute (AC) group receiving 150 mg RF/kg powdered rodent chow for 24 h providing 150 mg RF/kg body weight for 200 g rats, chronic (CHR) animals receiving RF for 10 days; withdrawal (WD) rats receiving RF for 8 days and then normal chow for 2 days. Results Significant changes in the levels of TRH and TRH-like peptides occurred throughout the brain and peripheral tissues in response to RF. The number of significant changes in TRH and TRH-like peptide levels in brain resulting from RF treatment, in descending order were: medulla (16), piriform cortex (8), nucleus accumbens (7), frontal cortex (5), striatum (3), amygdala (3), entorhinal cortex (3), anterior (2), and posterior cingulate (2), hippocampus (1), hypothalamus (0) and cerebellum (0). The corresponding ranking for peripheral tissues were: prostate (6), adrenals (4), pancreas (3), liver (2), testis (1), heart (0). Conclusions The sensitivity of TRH and TRH-like peptide expression to RF treatment, particularly in the medulla oblongata and prostate, is consistent with the participation of these peptides in the therapeutic effects of RF.
 
Article
Background Retinoic acid-related orphan receptor alpha (RORA) has been reported to be suppressed in autistic patients and is associated with autism spectrum disorders (ASD), although the potential role and mechanism of RORA on gastrointestinal (GI) symptoms in ASD patients is still not reported. In this study, we aim to investigate the contribution of RORA to GI symptoms through a maternal diabetes-mediated autism-like mouse model. Results Male offspring of diabetic dams were treated with either superoxide dismutase (SOD) mimetic MnTBAP or RORA agonist SR1078, or were crossbred with intestine epithelial cells (IEC)-specific RORA knockout (RORA −/− ) mouse. Gene expression, oxidative stress and inflammation were measured in brain tissues, peripheral blood mononuclear cells (PBMC) and IEC, and GI symptoms were evaluated. Our results showed that SOD mimetic MnTBAP completely, while RORA agonist SR1078 partly, reversed maternal diabetes-mediated oxidative stress and inflammation in the brain, PBMC and IEC, as well as GI symptoms, including intestine permeability and altered gut microbiota compositions. IEC-specific RORA deficiency either mimicked or worsened maternal diabetes-mediated GI symptoms as well as oxidative stress and inflammation in IEC, while there was little effect on maternal diabetes-mediated autism-like behaviors. Conclusions We conclude that RORA suppression contributes to maternal diabetes-mediated GI symptoms in autism-like mouse offspring, this study provides a potential therapeutical target for maternal diabetes-mediated GI symptoms in offspring through RORA activation.
 
The chemical structure of Q808
Effects of Q808 on PTZ kindling-induced A seizure stage and B body weight. Q808 effectively alleviates seizures in PTZ-kindling model rats (p = 0.003). The body weight of rats in each group have no difference. These data were shown as mean ± SEM and seizure score was analyzed with Mann–Whitney U test. **p < 0.01
A Venn diagram and B α-diversity analysis for control, model, and Q808 groups (n = 6). Venn diagram illustrates the average unique and common OTU numbers in three groups. The α-diversity (Chao 1 index) showed that the diversity of the gut microbial community was significantly decreased in model group compared with control (p = 0.021) and significantly increased in Q808 group compared with model group (p = 0.001). Differences in relative abundances of OTUs were calculated using Tukey’s honest significance test by R package. Indexes of α-diversity (Chao 1 index) were analyzed by the Kruskal–Wallis test. *p < 0.05, **p < 0.01
Alterations in the gut microbiota of PTZ-induced seizure model rats depending on Q808 treatment. A Two-dimensional principal coordinate analysis (PCoA) of the control, model, and Q808 groups. Each sample is marked by a dot. Samples that are more similar to one another are closer together. B Distribution of gut microbiota in the control, model, and Q808 groups at the phylum level. C Distribution of gut microbiota in the control, model, and Q808 groups at the genus level (n = 6). β-diversity was calculated by the pMANOVA analysis based on Bray Curtis distances
Normalized intensity of neurotransmitters in the hippocampi of the rats in three groups (n = 6). In the seizure model group, the content of GABA was drastically decreased compared with that in control group (p = 0.011). After Q808 treatment, the content of acetylcholine increased (p = 0.002 compared with the PTZ group), whereas the levels of 3-methoxytyramine, glutamine, and 5-HIAA decreased (p = 0.003, p = 0.001, and p = 0.0004, separately, compared with the control group). No significant differences were observed in the levels of normetanephrine, norepinephrine, glutamate, dopamine, histamine, and serotonin among the three groups. Data were means ± SEM and analyzed using the Kruskal–Wallis and Dunn’s non-parametric test. *p < 0.05, **p < 0.01, ***p < 0.001
Article
Background The gut microbiota can modulate brain function and behavior and is increasingly recognized as an important factor in mediating the risk of epilepsy and the effects of seizure interventions. Drug therapy is one of the factors that influence the composition of the intestinal microbiota. Q808 is an innovative chemical with strong anticonvulsant activity and low neurotoxicity. However, studies evaluating the effect of Q808 on gut microbial communities are lacking. In this study, we aimed to evaluate the anticonvulsant activity of Q808 on a pentylenetetrazol (PTZ)—induced seizure model and analyze and compare the intestinal microbiota composition of non-PTZ vehicle control group, the PTZ-induced seizure model rats with and without Q808, through 16S rDNA sequencing. Neurotransmitter levels in the hippocampus were quantitatively estimated using HPLC–MS. Results The results suggest that Q808 effectively alleviates seizures in chronic PTZ-kindled model rats. Additionally, based on the analyzed abundance of the gut microbiota, dysbacteriosis of model rats was found to be corrected after Q808 treatment at the phylum level. The unique bacterial taxa (e.g., Lactobacillus ) that are associated with acetylcholine production, were significantly increased. Several short-chain fatty acids (SCFAs)-producing bacteria, including Roseburia, Alloprevptella , Prevotellaceae_NK3B31_group , Prevotellaceae_UCG-001 , and Prevotella_9 , were enriched. In the hippocampus, the contents of acetylcholine increased, whereas the levels of 3-methoxytyramine, glutamine, and 5-hydroxyindole acetic acid (5-HIAA) decreased after Q808 treatment. Conclusions This study demonstrates that Q808 can be used to remodel the dysbiosis of the gut microbiome and influence neurotransmitter levels in the hippocampus of PTZ-induced seizure model rats. We hope that these novel findings prompt further research on the interaction between gut microbiota and seizures and the mechanism of Q808.
 
Article
Background In the horizon of therapeutic restrictions in intracerebral hemorrhage (ICH), recently, non-invasive transcranial electrical stimulation (tES) has achieved considerable prosperities. Translational studies have postulated that transcranial direct current stimulation (tDCS) and the other types of tES remain potentially a novel therapeutic option to reverse or stabilize cognitive and motor impairments. Objective The aim of this study was to comparatively evaluate the effects of the four main paradigms of tES, including tDCS, transcranial alternating (tACS), pulsed (tPCS), and random noise (tRNS) stimulations on collagenase-induced sensorimotor impairments and striatum tissue damage in male rats. Methods To induce ICH, 0.5 μl of collagenase was injected into the right striatum of male Sprague Dawley rats. One day after surgery, tES, was applied to the animals for seven consecutive days. Motor functions were appraised by neurological deficit score, rotarod, and wire hanging tests on the day before surgery and postoperative days 3, 7, and 14. After behavioral tests, brain tissue was prepared appropriately to perform the stereological evaluations. Results The results indicated that the application of the four tES paradigms (tDCS, tACS, tRNS, and tPCS) significantly reversed motor disorders in collagenase-induced ICH groups. Further, the motor function improvement of tACS and tRNS receiving rats in wire-hanging and rotarod tests were higher than the other two tES receiving groups. Structural changes and stereological assessments also confirmed the results of behavioral functions. Conclusion Our findings suggest that in addition to tDCS application in the treatment of ICH, other tES paradigms, especially tACS and tRNS may be considered as add-on therapeutic strategies in stroke.
 
Quantitative performance comparison of the BrainNet-GA CNN with competing methods: a ROC curves and b box plot graph
†Discriminative connections between brain regions for the classification of SSDs vs. HCs: a results of partial derivatives on a target class of SSDs and b HCs, and c§Circular plot showing increased (red color) or decreased (blue color) functional connectivity in patients compared to controls. †Green line and bar represent connectivity strength. The brighter color is, the greater its importance in the classification; Small circle in sky blue represents nodal strength. The more circle is filled, the greater its importance in the classification; §Red and blue lines represent hyperconnectivity and hypoconnectivity, respectively and darker line means more higher value
a Overview of Net-Global Covariance Pooling-Attention (Net-GA) block and b architecture of the BrainNet-GA CNN for the classification of SSDs vs. HCs. The Net-GA block consists of the BrainNetCNN (E2E, E2N and N2G filters) combined with global covariance pooling (2nd order pooling) and squeeze-excitation network (attention model). The BrainNet-GA CNN consists of typical convolutional layer plus Net-GA block
Article
Previous deep learning methods have not captured graph or network representations of brain structural or functional connectome data. To address this, we developed the BrainNet-Global Covariance Pooling-Attention Convolutional Neural Network (BrainNet-GA CNN) by incorporating BrainNetCNN and global covariance pooling into the self-attention mechanism. Resting-state functional magnetic resonance imaging data were obtained from 171 patients with schizophrenia spectrum disorders (SSDs) and 161 healthy controls (HCs). We conducted an ablation analysis of the proposed BrainNet-GA CNN and quantitative performance comparisons with competing methods using the nested tenfold cross validation strategy. The performance of our model was compared with competing methods. Discriminative connections were visualized using the gradient-based explanation method and compared with the results obtained using functional connectivity analysis. The BrainNet-GA CNN showed an accuracy of 83.13%, outperforming other competing methods. Among the top 10 discriminative connections, some were associated with the default mode network and auditory network. Interestingly, these regions were also significant in the functional connectivity analysis. Our findings suggest that the proposed BrainNet-GA CNN can classify patients with SSDs and HCs with higher accuracy than other models. Visualization of salient regions provides important clinical information. These results highlight the potential use of the BrainNet-GA CNN in the diagnosis of schizophrenia.
 
Article
Background Methamphetamine (MA) is a non-selective monoamine releaser and thus releases serotonin (5-HT), norepinephrine (NE) and dopamine (DA) from corresponding nerve terminals into synapses. DOI ((±)-2, 5-dimethoxy-4-iodoamphetamine) is a direct-acting serotonergic 5-HT2A/C receptor agonist and induces the head-twitch response (HTR) via stimulation of 5-HT2A receptor in mice. While more selective serotonin releasers such as d-fenfluramine evoke the HTR, monoamine reuptake blockers (e.g., cocaine) suppress the DOI-evoked HTR via indirect stimulation of serotonergic 5-HT1A- and adrenergic ɑ2-receptors. Since the induction of HTR by DOI is age-dependent, we investigated whether: (1) during development MA can evoke the HTR by itself, and (2) acute pretreatment with either the selective 5-HT2A receptor antagonist EMD 281014 or low-doses of MA can: (i) modulate the DOI-induced HTR in mice across postnatal days 20, 30 and 60, and (ii) alter the DOI-induced c-fos expression in mice prefrontal cortex (PFC). To further explore the possible modulatory effect of MA on DOI-induced HTR, we investigated whether blockade of inhibitory serotonergic 5-HT1A- or adrenergic ɑ2-receptors by corresponding selective antagonists (WAY 100635 or RS 79948, respectively), can prevent the effect of MA on DOI-induced HTR during aging. Results Although neither EMD 281014 nor MA by themselves could evoke the HTR, acute pretreatment with either EMD 281014 (0.01, 0.05 and 0.1 mg/kg, i.p.) or MA (1, 2.5, 5 mg/kg, i.p.), dose-dependently suppressed the DOI-induced HTR across ages. While WAY 100635 significantly reversed the inhibitory effect of MA in 20- and 30-day old mice, RS 79948 failed to significantly counter MA’s inhibitory effect. Moreover, DOI significantly increased c-fos expressions in several PFC regions. EMD 281014 prevented the DOI-induced increases in c-fos expression. Despite the inhibitory effect of MA on DOI-induced HTR, MA alone or in combination with DOI, significantly increased c-fos expression in several regions of the PFC. Conclusion The suppressive effect of MA on the DOI-evoked HTR appears to be mainly due to functional interactions between the HTR-inducing 5-HT2A receptor and the inhibitory 5-HT1A receptor. The MA-induced increase in c-fos expression in different PFC regions may be due to MA-evoked increases in synaptic concentrations of 5-HT, NE and/or DA.
 
Increased Aβ42 in PDD and decreased p-TauS199/202 levels in DLB. Multivariate analysis of post-mortem samples from the TC and CP regions of controls and individuals with PDD and DLB. a Protein levels of Aβ42 in the TC by ELISA assay; 100 µg of each protein sample was used per ELISA reaction. N  = 5. b p-APPT668 and APP protein levels were measured in the TC samples by western blot. Twenty micrograms of protein were used per well for SDS-PAGE. Protein values are normalized to β-actin levels and control samples. Blots were cropped from different gels. Representative western blot images were shown. All blot images with significant changes were included in the Additional file 1: Fig. S1. c Statistical results of relative p-APPT668 protein levels normalized to APP levels in the TC. d p-TauS199/202 protein levels were measured in CP by western blot. Blots were cropped from different gels. Representative western blot images were shown. Protein values are normalized to β-actin levels and control samples. e Statistical results of relative p-TauS199/202 levels in the CP samples. Data are presented as the mean  ±  SEM. Kruskal–Wallis test was used to measure the differences between Control, PDD, and DLB samples. *p  < 0.05, **p  < 0.01, ***p  < 0.001. Sample numbers, Control  = 9, PDD  = 10, and DLB  = 9
Decreased TH in both PDD and DLB but increased α-synuclein in DLB in CP. a Multivariate analysis of post-mortem samples from TC by western blot. Twenty micrograms of protein were used per well for SDS-PAGE. Blots were cropped from different gels. Representative western blot images were shown. Statistical results of relative protein levels for DAT (b), DD2R (c), and TH (d). e Multivariate analysis of post-mortem samples from CP by western blot. Twenty micrograms of protein were used per well for SDS-PAGE. Blots were cropped from different gels. Representative western blot images were shown. All blot images with significant changes were included in the Additional file 1: Fig. S2. Statistical results of relative protein levels for TH (f) and α-synuclein (α-syn) (g). Protein values are normalized to β-actin levels and control samples. Data are presented as the mean  ±  SEM. Kruskal–Wallis test was used to measure the differences between Control, PDD, and DLB samples. *p  < 0.05, **p  < 0.01. Sample numbers, Control  = 9, PDD  = 9–10, and DLB  = 9
Increased neuroinflammation, mitophagy activity, and mitochondrial dynamics in PDD. a Multivariate analysis of post-mortem samples from TC by western blot. Twenty micrograms of protein were used per well for SDS-PAGE. Blots were cropped from different gels. Representative western blot images were shown. Statistical results of the relative protein levels of GFAP (b), IBA1 (c), and DRP1 (d). e Multivariate analysis of post-mortem samples from CP by western blot. Blots were cropped from different gels. Representative western blot images were shown. All blot images with significant changes were included in the Additional file 1: Fig. S3. Statistical results of the relative protein levels of LC3II (f), and MFN2 (g). Protein values are normalized to β-actin levels and control samples. Data are presented as the mean  ±  SEM. Kruskal–Wallis test was used to measure the differences between Control, PDD, and DLB samples. *p  < 0.05. Sample numbers, Control  = 9, PDD  =  10, and DLB  = 9
Increased p-LRRK2S935 in PDD and increased VPS35 in PDD compared to DLB in TC. a Multivariate analysis of post-mortem samples from the TC by western blot. Twenty micrograms of protein were used per well for SDS-PAGE. Blots were cropped from different gels. Representative western blot images were shown. All blot images with significant changes were included in the Additional file 1: Fig. S4. Statistical results of the relative protein levels of p-LRRK2S935 (b), p-Rab10T73/Rab10 (c), and VPS35 (d). Data are presented as the mean  ±  SEM. Kruskal–Wallis test was used to measure the differences between Control, PDD, and DLB samples. *p  < 0.05, **p  < 0.01. Sample numbers, Control  = 9, PDD  = 10, and DLB  = 9
Article
Background Parkinson’s disease (PD) and dementia with Lewy bodies (DLB) are common age-related neurodegenerative diseases comprising Lewy body spectrum disorders associated with cortical and subcortical Lewy body pathology. Over 30% of PD patients develop PD dementia (PDD), which describes dementia arising in the context of established idiopathic PD. Furthermore, Lewy bodies frequently accompany the amyloid plaque and neurofibrillary tangle pathology of Alzheimer’s disease (AD), where they are observed in the amygdala of approximately 60% of sporadic and familial AD. While PDD and DLB share similar pathological substrates, they differ in the temporal onset of motor and cognitive symptoms; however, protein markers to distinguish them are still lacking. Methods Here, we systematically studied a series of AD and PD pathogenesis markers, as well as mitochondria, mitophagy, and neuroinflammation-related indicators, in the substantia nigra (SN), temporal cortex (TC), and caudate and putamen (CP) regions of human post-mortem brain samples from individuals with PDD and DLB and condition-matched controls. Results We found that p-APPT668 (TC), α-synuclein (CP), and LC3II (CP) are all increased while the tyrosine hydroxylase (TH) (CP) is decreased in both PDD and DLB compared to control. Also, the levels of Aβ42 and DD2R, IBA1, and p-LRRK2S935 are all elevated in PDD compared to control. Interestingly, protein levels of p-TauS199/202 in CP and DD2R, DRP1, and VPS35 in TC are all increased in PDD compared to DLB. Conclusions Together, our comprehensive and systematic study identified a set of signature proteins that will help to understand the pathology and etiology of PDD and DLB at the molecular level.
 
Article
Background Imitative learning is highly effective from infancy to old age; however, little is known about the effects of observing errors during imitative learning. This study aimed to examine how observing errors affected imitative learning performance to maximize its effect. Methods In the pre-training session, participants were instructed to pinch at a target force (8 N) with auditory feedback regarding generated force while they watched videos of someone pinching a sponge at the target force. In the pre-test, participants pinched at the target force and did not view a model or receive auditory feedback. In Experiment 1, in the main training session, participants imitated models while they watched videos of pinching at either the incorrect force (error-mixed condition) or target force (correct condition). Then, the exact force generated was measured without receiving auditory feedback or viewing a model. In Experiment 2, using the same procedures, newly recruited participants watched videos of pinching at incorrect forces (4 and 24 N) as the error condition and the correct force as the correct condition. Results In Experiment 1, the average force was closer to the target force in the error-mixed condition than in the correct condition. In Experiment 2, the average force in the correct condition was closer to the target force than in the error condition. Conclusion Our findings indicated that observing error actions combined with correct actions affected imitation motor learning positively as error actions contained information on things to avoid in the target action. It provides further information to enhance imitative learning in mixed conditions compared to that with correct action alone.
 
Top-cited authors
Giulio Tononi
  • University of Wisconsin–Madison
Lutz Jäncke
  • University of Zurich
Michael Posner
  • University of Oregon
Yanhong Wu
  • Peking University
Jin Fan
  • City University of New York - Queens College