Neurocircuitry underlying the antidepressant effect of retrograde facial botulinum toxin in mice.

BACKGROUNDS: Botulinum toxin type A (BoNT/A) is extensively applied in spasticity and dystonia as it cleaves synaptosome-associated protein 25 (SNAP25) in the presynaptic terminals, thereby inhibiting neurotransmission. An increasing number of randomized clinical trials have suggested that glabellar BoNT/A injection improves depressive symptoms in patients with major depressive disorder (MDD). However, the underlying neuronal circuitry of BoNT/A-regulated depression remains largely uncharacterized. RESULTS: Here, we modeled MDD using mice subjected to chronic restraint stress (CRS). By pre-injecting BoNT/A into the unilateral whisker intrinsic musculature (WIM), and performing behavioral testing, we showed that pre-injection of BoNT/A attenuated despair- and anhedonia-like phenotypes in CRS mice. By applying immunostaining of BoNT/A-cleaved SNAP25 (cl.SNAP25197), subcellular spatial localization of SNAP25 with markers of cholinergic neurons (ChAT) and post-synaptic membrane (PSD95), and injection of monosynaptic retrograde tracer CTB-488-mixed BoNT/A to label the primary nucleus of the WIM, we demonstrated that BoNT/A axonal retrograde transported to the soma of whisker-innervating facial motoneurons (wFMNs) and subsequent transcytosis to synaptic terminals of second-order neurons induced central effects. Furthermore, using transsynaptic retrograde and monosynaptic antegrade viral neural circuit tracing with c-Fos brain mapping and co-staining of neural markers, we observed that the CRS-induced expression of c-Fos and CaMKII double-positive neurons in the ventrolateral periaqueductal grey (vlPAG), which sent afferents to wFMNs, was down-regulated 3 weeks after BoNT/A facial pre-administration. Strikingly, the repeated and targeted silencing of the wFMNs-projecting CaMKII-positive neurons in vlPAG with a chemogenetic approach via stereotactic injection of recombinant adeno-associated virus into specific brain regions of CRS mice mimicked the antidepressant-like action of BoNT/A pre-treatment. Conversely, repeated chemogenetic activation of this potential subpopulation counteracted the BoNT/A-improved significant antidepressant behavior. CONCLUSION: We reported for the first time that BoNT/A inhibited the wFMNs-projecting vlPAG excitatory neurons through axonal retrograde transport and cell-to-cell transcytosis from the injected location of the WIM to regulate depressive-like phenotypes of CRS mice. For the limited and the reversibility of side effects, BoNT/A has substantial advantages and potential application in MDD.

(+)-Borneol inhibits the generation of reactive oxygen species and neutrophil extracellular traps induced by phorbol-12-myristate-13-acetate.

Background and purpose: Neutrophil extracellular traps (NETs) are special web-like structures that can be generated in both infectious and noninfectious diseases. Previous studies showed that reactive oxygen species (ROS) were crucial in the formation of NETs (NETosis). The purpose of this study is to evaluate the effect of (+)-borneol, an antioxidant, on NETosis. Methods: Human neutrophils were stimulated with phorbol-12-myristate-13-acetate (PMA) to induce NETosis in vitro. Neutrophils treated with (+)-borneol at three different time points (-30 min, 0, and 30 min) associated with PMA stimulation were used to examine the effect of (+)-borneol on the formation of NETs. The ROS generation of neutrophils was also measured to explore the potential mechanism of the inhibitory effect of (+)-borneol on NETosis. Results: (+)-Borneol pretreatment inhibited NETosis induced by PMA. Immunofluorescence staining visualized and confirmed the inhibitory effect. (+)-Borneol inhibited the burst of ROS in neutrophils caused by PMA. Suppressing NADPH oxidase or protein kinase C (PKC) eliminated the effect of (+)-borneol on NETosis. Moreover, inhibiting Toll-like receptor 2 (TLR2) led to increased NETosis which can be inhibited by (+)-borneol. Conclusion: (+)-Borneol decreases the ROS level in activated neutrophils and inhibits NETosis triggered by PMA stimulation in vitro. (+)-Borneol therapy may be effective in some NET-dependent conditions.

Pathological Gait Signatures of Post-stroke Dementia With Toe-Off and Heel-to-Ground Angles Discriminate From Alzheimer's Disease.

Given the limited power of neuropsychological tests, there is a need for a simple, reliable means, such as gait, to identify mild dementia and its subtypes. However, gait characteristics of patients with post-stroke dementia (PSD) and Alzheimer's disease (AD) are unclear. We sought to describe their gait signatures and to explore gait parameters distinguishing PSD from post-stroke non-dementia (PSND) and patients with AD. We divided 3-month post-stroke patients into PSND and PSD groups based on the Mini-Mental State Examination (MMSE), Montreal Cognitive Assessment (MoCA), and the activity of daily living (ADL). Thirty-one patients with AD and thirty-two healthy controls (HCs) were also recruited. Ten gait parameters in one single and two dual-task gait tests (counting-backward or naming-animals while walking) were compared among the groups, with adjustment for baseline demographic covariates and the MMSE score. The area under the receiver operating characteristic curve (AUC) was used to identify parameters discriminating PSD from individuals with PSND and AD. Patients with PSD and patients with AD showed impaired stride length, velocity, stride time, and cadence while patients with PSD had altered stance and swing phase proportions (all p ≤ 0.01, post hoc). Patients with AD had smaller toe-off (ToA) and heel-to-ground angles (HtA) (p ≤ 0.01) than HCs in dual-task gait tests. Individuals with PSD had a shorter stride length, slower velocity, and altered stance and swing phase percentages in all tests (p ≤ 0.01), but a higher coefficient of variation of stride length (CoVSL) and time (CoVST) only in the naming animals-task gait test (p ≤ 0.001) than individuals with PSND. ToA and HtA in the naming animals-task gait test were smaller in individuals with AD than those with PSD (p ≤ 0.01). Statistical significance persisted after adjusting for demographic covariates, but not for MMSE. The pace and the percentage of stance or swing phase in all tests, CoVST in the dual-task paradigm, and CoVSL only in the naming animals-task gait test (moderate accuracy, AUC > 0.700, p ≤ 0.01) could distinguish PSD from PSND. Furthermore, the ToA and HtA in the naming animals-task gait paradigm discriminated AD from PSD (moderate accuracy, AUC > 0.700, p ≤ 0.01). Thus, specific gait characteristics could allow early identification of PSD and may allow non-invasive discrimination between PSD and AD, or even other subtypes of dementia.

Structural and Functional Deficits in Patients with Poststroke Dementia: A Multimodal MRI Study.

Although many neuroimaging studies have reported structural and functional abnormalities in the brains of patients with cognitive impairments following stroke, little is known about the pattern of such brain reorganization in poststroke dementia (PSD). The present study was aimed at investigating alterations in spontaneous brain activity and gray matter volume (GMV) in PSD patients. We collected T1-weighted and resting-state functional magnetic resonance imaging data from 20 PSD patients, 24 poststroke nondementia (PSND) patients, and 21 well-matched normal controls (NCs). We compared the differences among the groups in GMV and the fractional amplitude of low-frequency fluctuations (fALFF). Then, we evaluated the relationship between these brain measures and cognitive assessments and explored the possible distinguisher for PSD by receiver operating characteristic (ROC) curve analysis. PSD patients showed smaller GMV in the right superior temporal gyrus and lower fALFF values in the right inferior frontal gyrus than both PSND patients and NCs, but such differences were not observed between PSND patients and NCs. Moreover, GMV in the left medial prefrontal cortex showed a significant positive correlation with the Mini-Cog assessment in PSD patients, and GMV in the left CPL displayed the highest area under the ROC curve among all the features for classifying PSD versus PSND patients. Our findings suggest that PSD patients show dementia-specific structural and functional alteration patterns, which may help elucidate the pathophysiological mechanisms underlying PSD.

Inhibition of endoplasmic reticulum stress reverses synaptic plasticity deficits in striatum of DYT1 dystonia mice.

BACKGROUND AND OBJECTIVE: Striatal plasticity alterations caused by endoplasmic reticulum (ER) stress is supposed to be critically involved in the mechanism of DYT1 dystonia. In the current study, we expanded this research field by investigating the critical role of ER stress underlying synaptic plasticity impairment imposed by mutant heterozygous Tor1a+/- in a DYT1 dystonia mouse model. METHODS: Heterozygous Tor1a+/- mouse model for DYT1 dystonia was established. Wild-type (Tor1a+/+, N=10) and mutant (Tor1a+/-, N=10) mice from post-natal day P25 to P35 were randomly distributed to experimental and control groups. Patch-clamp and current-clamp recordings of SPNs were conducted with intracellular electrodes for electrophysiological analyses. Striatal changes of the direct and indirect pathways were investigated via immunofluorescence. Golgi-Cox staining was conducted to observe spine morphology of SPNs. To quantify postsynaptic signaling proteins in striatum, RNA-Seq, qRT-PCR and WB were performed in striatal tissues. RESULTS: Long-term depression (LTD) was failed to be induced, while long-term potentiation (LTP) was further strengthened in striatal spiny projection neurons (SPNs) from the Tor1a+/- DYT1 dystonia mice. Spine morphology analyses revealed a significant increase of both number of mushroom type spines and spine width in Tor1a+/- SPNs. In addition, increased AMPA receptor function and the reduction of NMDA/AMPA ratio in the postsynaptic of Tor1a+/- SPNs was observed, along with increased ER stress protein levels in striatum of Tor1a+/- DYT1 dystonia mice. Notably, ER stress inhibitors, tauroursodeoxycholic acid (TUDCA), could rescue LTD as well as AMPA currents. CONCLUSION: The current study illustrated the role of ER stress in mediating structural and functional plasticity alterations in Tor1a+/- SPNs. Inhibition of the ER stress by TUDCA is beneficial in reversing the deficits at the cellular and molecular levels. Remedy of dystonia associated neurological and motor functional impairment by ER stress inhibitors could be a recommendable therapeutic agent in clinical practice.

Altered Functional Connectivity of Hippocampal Subfields in Poststroke Dementia.

BACKGROUND: The hippocampus (HP) plays a critical role in memory and orientational functions and is functionally heterogeneous along the longitudinal anterior-posterior axis. Although the previous study has reported volumetric atrophy in hippocampal subfields of patients with poststroke dementia (PSD), how the functional connectivity (FC) is altered in these subfields remains unclear. PURPOSE: To examine the FC changes of the HP subfields in patients with PSD. STUDY TYPE: Prospective. POPULATION: Seventeen normal controls, 20 PSD, and 24 nondemented poststroke (PSND) patients. FIELD STRENGTH/SEQUENCE: A 3.0 T/ T1-weighted imaging, resting-state functional and diffusion tensor imaging. ASSESSMENT: We first segmented the HP using independent component analysis, and then used granger causality analysis to calculate the directed FCs (dFCs) between the subfields and the whole brain, and compared the dFCs among PSD, PSND, and controls. STATISTICAL TESTS: Student's t-test, chi-square test, one-way ANCOVA, multiple regression, support vector machine, multiple comparison correction, and reproducibility analysis. A P value < 0.05 was considered statistically significant. RESULTS: Our results showed HP was functionally divided into HPhead , HPbody , and HPtail bilaterally along the longitudinal axis. PSD patients showed significant dementia-specific decreases in the inward information flow and increases in the outward information flow associated with the bilateral entire HP/HPhead and left HPbody (P < 0.05). Moreover, we observed significant correlations (P < 0.05) between the cognition score and the dFCs related to the bilateral entire HP and left HPhead in the PSD group. Furthermore, dFCs of the HP and its subfields improved the classification between the PSD and PSND patients (accuracy/sensitivity/specificity: 94%/95%/93%) compared to the clinical and demographic parameters alone. DATA CONCLUSION: These findings suggest that altered transmission and reception of information in the HP. These alternations were specific to individual subfields in PSD patients and may offer insight into the neurophysiological mechanisms underlying PSD. EVIDENCE LEVEL: 1 TECHNICAL EFFICACY: Stage 2.

Gait characteristics and clinical relevance of hereditary spinocerebellar ataxia on deep learning.

BACKGROUND: Deep learning has always been at the forefront of scientific research. It has also been applied to medical research. Hereditary spinocerebellar ataxia (SCA) is characterized by gait abnormalities and is usually evaluated semi-quantitatively by scales. However, more detailed gait characteristics of SCA and related objective methods have not yet been established. Therefore, the purpose of this study was to evaluate the gait characteristics of SCA patients, as well as to analyze the correlation between gait parameters, clinical scales, and imaging on deep learning. METHODS: Twenty SCA patients diagnosed by genetic detection were included in the study. Ten patients who were tested via functional magnetic resonance imaging (fMRI) were included in the SCA imaging subgroup. All SCA patients were evaluated with the International Cooperative Ataxia Rating Scale (ICARS) and Scale for the Assessment and Rating of Ataxia (SARA) clinical scales. The gait control group included 16 healthy subjects, and the imaging control group included seven healthy subjects. Gait data consisting of 10 m of free walking of each individual in the SCA group and the gait control group were detected by wearable gait-detection equipment. Stride length, stride time, velocity, supporting-phase percentage, and swinging-phase percentage were extracted as gait parameters. Cerebellar volume and the midsagittal cerebellar proportion in the posterior fossa (MRVD) were calculated according to MR. RESULTS: There were significant differences in stride length, velocity, supporting-phase percentage, and swinging-phase percentage between the SCA group and the gait control group. The stride length and stride velocity of SCA groups were lower while supporting phase was longer than those of the gait control group. SCA group's velocity was negatively correlated with both the ICARS and SARA scores. The cerebellar volume and MRVD of the SCA imaging subgroup were significantly smaller than those of the imaging control group. MRVD was significantly correlated with ICARS and SARA scores, as well as stride velocity variability. CONCLUSION: SCA gait parameters were characterized by a reduced stride length, slower walking velocity, and longer supporting phase. Additionally, a smaller cerebellar volume correlated with an increased irregularity in gait. Gait characteristics exhibited considerable clinical relevance to hereditary SCA. We conclude that a combination of gait parameters, ataxia scales, and MRVD may represent more objective markers for clinical evaluations of SCA.

Meta-Regression Analysis of the Long-Term Effects of Pallidal and Subthalamic Deep Brain Stimulation for the Treatment of Isolated Dystonia.

OBJECTIVE: The globus pallidus internus (GPi) and subthalamic nucleus (STN) are therapeutic targets for deep brain stimulation (DBS) in the treatment of isolated dystonia. We conducted a meta-regression analysis on long-term studies of bilateral DBS in the GPi and STN to compare the relative effects of the 2 approaches. METHODS: We systematically searched the PubMed, Embase, and Cochrane Controlled Register of Trials databases to identify studies reporting the treatment outcomes of GPi DBS and STN DBS for isolated dystonia. The primary outcome measure was the change in the Burke-Fahn-Marsden dystonia rating scale movement score between the baseline and follow-up evaluations. We performed a regression analysis using a random effects model. RESULTS: A total of 42 follow-up evaluations (30 for GPi and 12 for STN) nested in 19 studies (16 of GPi and 3 of STN) were included in our analysis. The results from univariate regression analysis suggested that shorter disease duration and STN stimulation were associated with a greater standardized change in the Burke-Fahn-Marsden dystonia rating scale movement score. On combining the factors into 1 model, only the disease duration remained significant. The regression analysis results of the GPi and STN subgroups revealed more persistent improvement after STN stimulation. CONCLUSIONS: A shorter disease duration correlated positively with better DBS outcomes. The STN appeared to be an optimized stimulation target for the treatment of isolated dystonia, although randomized controlled trials are needed to compare the treatment efficacy of GPi DBS and STN DBS.

Unilateral whisker pad injection of botulinum toxin type a enhances spatial learning in mice.

The central cholinergic nervous system plays an important role in cognition, with acetylcholine hypofunction considered to be a major factor of dementia. Botulinum toxin type A (BoNT/A), a potent poison secreted by Clostridium botulinum, is used widely for dystonia treatment and facial cosmesis. BoNT/A injection inhibits acetylcholine release in the neuromuscular junction through cleavage of synaptosomal-associated protein of 25 kDa in cholinergic terminals. Furthermore, beyond the injection site, BoNT/A undergoes retrograde transport and transcytosis to the central nervous system from peripheral cholinergic terminals. However, whether peripheral BoNT/A injection affects the function of the central nervous system and induces learning deficits remains unclear. We injected mice with different doses of BoNT/A (2, 10, and 50 U/kg) or sterile saline (control) into the left whisker pad to test spatial learning performance at different times after injection using the Morris water maze. At 3 days and 4 weeks after injection, the spatial learning ability of the control and BoNT/A-treated mice showed no significant differences. Surprisingly, however, rather than spatial learning impairment at 6 weeks after injection, BoNT/A-treated mice spent less time than control mice in locating the experimental platform, indicating that BoNT/A facial injection might promote spatial learning. Furthermore, our study suggests that facial application of BoNT/A is safe and could play a positive role in ameliorating the spatial learning deficits associated with neurodegenerative diseases.

Antinociceptive effect of botulinum toxin A involves alterations in AMPA receptor expression and glutamate release in spinal dorsal horn neurons.

The use of botulinum toxin A (BTX-A) for various clinical therapeutic applications is increasing. It is widely believed that peripheral therapeutic or toxic effects of BTX-A are exclusively mediated by SNAP-25 cleavage. There is growing evidence of long-distance retrograde axonal transport of BTX-A on entering the central nervous system, subsequent to a local injection of the toxin. However, the prevalence of central antinociceptive effects after BTX-A peripheral application and its underlying mechanisms are unclear. Our results show that (1) BTX-A can undergo retrograde axonal transport to the dorsal horn after peripheral application; (2) Peripheral pretreatment with BTX-A decreases the expression and function of AMPA receptors in the spinal cord dorsal horn neurons; (3) Peripheral pretreatment with BTX-A does not change basal glutamate release, but decreases the effect of formalin-evoked release of glutamate in spinal cord dorsal horn neurons. These results suggest that peripheral application of BTX-A can change AMPA receptor expression in, and glutamate release from, spinal dorsal horn neurons, which may have significance in its central antinociceptive effects.