SmartEM: machine-learning guided electron microscopy.

Connectomics provides essential nanometer-resolution, synapse-level maps of neural circuits to understand brain activity and behavior. However, few researchers have access to the high-throughput electron microscopes necessary to generate enough data for whole circuit or brain reconstruction. To date, machine-learning methods have been used after the collection of images by electron microscopy (EM) to accelerate and improve neuronal segmentation, synapse reconstruction and other data analysis. With the computational improvements in processing EM images, acquiring EM images has now become the rate-limiting step. Here, in order to speed up EM imaging, we integrate machine-learning into real-time image acquisition in a singlebeam scanning electron microscope. This SmartEM approach allows an electron microscope to perform intelligent, data-aware imaging of specimens. SmartEM allocates the proper imaging time for each region of interest - scanning all pixels equally rapidly, then re-scanning small subareas more slowly where a higher quality signal is required to achieve accurate segmentability, in significantly less time. We demonstrate that this pipeline achieves a 7-fold acceleration of image acquisition time for connectomics using a commercial single-beam SEM. We apply SmartEM to reconstruct a portion of mouse cortex with the same accuracy as traditional microscopy but in less time.

A petavoxel fragment of human cerebral cortex reconstructed at nanoscale resolution.

To fully understand how the human brain works, knowledge of its structure at high resolution is needed. Presented here is a computationally intensive reconstruction of the ultrastructure of a cubic millimeter of human temporal cortex that was surgically removed to gain access to an underlying epileptic focus. It contains about 57,000 cells, about 230 millimeters of blood vessels, and about 150 million synapses and comprises 1.4 petabytes. Our analysis showed that glia outnumber neurons 2:1, oligodendrocytes were the most common cell, deep layer excitatory neurons could be classified on the basis of dendritic orientation, and among thousands of weak connections to each neuron, there exist rare powerful axonal inputs of up to 50 synapses. Further studies using this resource may bring valuable insights into the mysteries of the human brain.

Immersion Fixation and Staining of Multicubic Millimeter Volumes for Electron Microscopy-Based Connectomics of Human Brain Biopsies.

Connectomics allows mapping of cells and their circuits at the nanometer scale in volumes of approximately 1 mm3. Given that the human cerebral cortex can be 3 mm in thickness, larger volumes are required. Larger-volume circuit reconstructions of human brain are limited by 1) the availability of fresh biopsies; 2) the need for excellent preservation of ultrastructure, including extracellular space; and 3) the requirement of uniform staining throughout the sample, among other technical challenges. Cerebral cortical samples from neurosurgical patients are available owing to lead placement for deep brain stimulation. Described here is an immersion fixation, heavy metal staining, and tissue processing method that consistently provides excellent ultrastructure throughout human and rodent surgical brain samples of volumes 2 × 2 × 2 mm3 and up to 37 mm3 with one dimension ≤2 mm. This method should allow synapse-level circuit analysis in samples from patients with psychiatric and neurologic disorders.

Intraarterial transplantation of human umbilical cord blood mononuclear cells is more efficacious and safer compared with umbilical cord mesenchymal stromal cells in a rodent stroke model.

INTRODUCTION: Stroke is the second leading cause of death worldwide, claims six lives every 60 seconds, and is a leading cause of adult disability across the globe. Tissue plasminogen activator, the only United States Food and Drug Administration (FDA)-approved drug currently available, has a narrow therapeutic time window of less than 5 hours. In the past decade, cells derived from the human umbilical cord (HUC) have emerged as a potential therapeutic alternative for stroke; however, the most effective HUC-derived cell population remains unknown. METHODS: We compared three cell populations derived from the human umbilical cord: cord blood mononuclear cells (cbMNCs); cord blood mesenchymal stromal cells (cbMSCs), a subpopulation of cbMNCs; and cord matrix MSCs (cmMSCs). We characterized these cells in vitro with flow cytometry and assessed the cells' in vivo efficacy in a 2-hour transient middle cerebral artery occlusion (MCAo) rat model of stroke. cbMNCs, cbMSCs, and cmMSCs were each transplanted intraarterially at 24 hours after stroke. RESULTS: A reduction in neurologic deficit and infarct area was observed in all three cell groups; however, this reduction was significantly enhanced in the cbMNC group compared with the cmMSC group. At 2 weeks after stroke, human nuclei-positive cells were present in the ischemic hemispheres of immunocompetent stroke rats in all three cell groups. Significantly decreased expression of rat brain-derived neurotrophic factor mRNA was observed in the ischemic hemispheres of all three cell-treated and phosphate-buffered saline (PBS) group animals compared with sham animals, although the decrease was least in cbMNC-treated animals. Significantly decreased expression of rat interleukin (IL)-2 mRNA and IL-6 mRNA was seen only in the cbMSC group. Notably, more severe complications (death, eye inflammation) were observed in the cmMSC group compared with the cbMNC and cbMSC groups. CONCLUSIONS: All three tested cell types promoted recovery after stroke, but cbMNCs showed enhanced recovery and fewer complications compared with cmMSCs.

Treatment of bacterial meningitis: an overview of Cochrane systematic reviews.

BACKGROUND: Acute bacterial meningitis (ABM) is a rapidly developing acute inflammation of leptomeninges and underlying subarachnoid cerebrospinal fluid (CSF). ABM is caused by bacteria and has a case fatality rate of 20-30%. Most prevalent causes of ABM are Neisseria meningitis, Streptococcus pneumoniae and Haemophilus influenzae. The aim of this paper is to summarize the main findings from Cochrane systematic reviews that have considered the evidence for treatments of ABM. METHODS: We searched the Cochrane Library (issue 1, 2007) for relevant reviews using 'meningitis' as a search term. The titles of all the search results were examined to select reviews on treatment of ABM. The full text of each of the selected reviews was studied to summarize the evidence available in Cochrane systematic reviews. RESULTS: We found three Cochrane reviews that focused specifically on the treatment of ABM, addressing empiric antibiotic therapy, fluid therapy and effects of adjuvant corticosteroids respectively. No statistically significant difference was found between third generation cephalosporins and conventional antibiotics in the combined endpoint of death or deafness (risk difference (RD) -1%, 95% CI -4% to +2%). However, culture positivity of CSF at 10-48 h was significantly higher in the conventional antibiotic group and diarrhoea was significantly more common in the cephalosporin group. When third generation cephalosporins are not available, ampicillin-chloramphenicol combination may be used as an alternative empiric treatment, however both resistance pattern as well as availability should be considered while prescribing empiric therapy of community acquired ABM. The fluid therapy review found too few studies to provide any robust conclusion. In settings with high mortality rates and where patients present late, use of intravenous maintenance fluids seems preferable to a restricted fluid intake. The efficacy of adjuvant corticosteroids varied between high- and low-income countries suggesting greater mortality reduction in high-income countries (RR 0.74, 95% CI 0.52-1.05) than in low-income countries (RR 0.87, 95% CI 0.72-1.05) and a beneficial effect on severe hearing loss in high-income countries (RR 0.32, 95% CI 0.18-0.57), whereas, sparse data in low-income countries (RR 1.04, 95% CI 0.66-1.63). A four-day regimen of dexamethasone should be given preferably before or with the first dose of antibiotics for cases of ABM from high-income countries. CONCLUSION: In presence of sensitive organisms, third generation cephalosporins and conventional antibiotics lead to similar outcomes. More studies are needed to determine the antimicrobial resistance pattern against various antibiotics in rural and remote areas of developing as well as developed countries. To assess the effectiveness of either restricting or maintenance fluids in populations where patients present early and on death and disability when mortality rates are low, large trials should be conducted. More trials are needed to assess the use of adjuvant dexamethasone for ABM in low-income countries.

Prevention of bacterial meningitis: an overview of Cochrane systematic reviews.

Acute bacterial meningitis (ABM) is an acute inflammation of leptomeninges caused by bacteria, and has a case fatality rate of 10-30%. Prevention strategies, such as vaccination and prophylactic antibiotics, can prevent ABM and have substantial public health impact by reducing the disease burden associated with it. The aim of this paper is to summarize the main findings from Cochrane systematic reviews that have considered the evidence for measures to prevent ABM. We assessed the evidence available in the Cochrane Library. We found five Cochrane reviews focused on the prevention of ABM; three with use of vaccination and two with prophylactic antibiotics. Polysaccharide serogroup A vaccine is strongly protective for the first year, against serogroup A meningococcal meningitis in adults and children over 5 years of age. Meningococcal serogroup C conjugate (MCC) vaccine is safe and effective in infants. Haemophilus influenzae type b (Hib) vaccine is safe and effective against Hib-invasive disease at all ages. Ceftriaxone, rifampicin and ciprofloxacin are the most effective prophylactic antibiotics against Neisseria meningitidis. There is sufficient evidence to use polysaccharide serogroup A vaccine to prevent serogroup A meningococcal meningitis, MCC conjugate vaccines to prevent meningococcal C meningitis and Hib conjugate vaccine to prevent Hib infections. More studies are needed to evaluate the effects of Hib conjugate vaccine on mortality. Further, studies are required to compare the relative effectiveness of ceftriaxone, ciprofloxacin and rifampicin in chemoprophylaxis against meningococcal infection.