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      Dense EM-based reconstruction of the interglomerular projectome in the zebrafish olfactory bulb.

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          Abstract

          The dense reconstruction of neuronal circuits from volumetric electron microscopy (EM) data has the potential to uncover fundamental structure-function relationships in the brain. To address bottlenecks in the workflow of this emerging methodology, we developed a procedure for conductive sample embedding and a pipeline for neuron reconstruction. We reconstructed ∼98% of all neurons (>1,000) in the olfactory bulb of a zebrafish larva with high accuracy and annotated all synapses on subsets of neurons representing different types. The organization of the larval olfactory bulb showed marked differences from that of the adult but similarities to that of the insect antennal lobe. Interneurons comprised multiple types but granule cells were rare. Interglomerular projections of interneurons were complex and bidirectional. Projections were not random but biased toward glomerular groups receiving input from common types of sensory neurons. Hence, the interneuron network in the olfactory bulb exhibits a specific topological organization that is governed by glomerular identity.

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          Most cited references31

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          Optimization of a GCaMP calcium indicator for neural activity imaging.

          Genetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Recent efforts in protein engineering have significantly increased the performance of GECIs. The state-of-the art single-wavelength GECI, GCaMP3, has been deployed in a number of model organisms and can reliably detect three or more action potentials in short bursts in several systems in vivo. Through protein structure determination, targeted mutagenesis, high-throughput screening, and a battery of in vitro assays, we have increased the dynamic range of GCaMP3 by severalfold, creating a family of "GCaMP5" sensors. We tested GCaMP5s in several systems: cultured neurons and astrocytes, mouse retina, and in vivo in Caenorhabditis chemosensory neurons, Drosophila larval neuromuscular junction and adult antennal lobe, zebrafish retina and tectum, and mouse visual cortex. Signal-to-noise ratio was improved by at least 2- to 3-fold. In the visual cortex, two GCaMP5 variants detected twice as many visual stimulus-responsive cells as GCaMP3. By combining in vivo imaging with electrophysiology we show that GCaMP5 fluorescence provides a more reliable measure of neuronal activity than its predecessor GCaMP3. GCaMP5 allows more sensitive detection of neural activity in vivo and may find widespread applications for cellular imaging in general.
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            Radiation damage in the TEM and SEM.

            We review the various ways in which an electron beam can adversely affect an organic or inorganic sample during examination in an electron microscope. The effects considered are: heating, electrostatic charging, ionization damage (radiolysis), displacement damage, sputtering and hydrocarbon contamination. In each case, strategies to minimise the damage are identified. In the light of recent experimental evidence, we re-examine two common assumptions: that the amount of radiation damage is proportional to the electron dose and is independent of beam diameter; and that the extent of the damage is proportional to the amount of energy deposited in the specimen.
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              Divisive normalization in olfactory population codes.

              In many regions of the visual system, the activity of a neuron is normalized by the activity of other neurons in the same region. Here we show that a similar normalization occurs during olfactory processing in the Drosophila antennal lobe. We exploit the orderly anatomy of this circuit to independently manipulate feedforward and lateral input to second-order projection neurons (PNs). Lateral inhibition increases the level of feedforward input needed to drive PNs to saturation, and this normalization scales with the total activity of the olfactory receptor neuron (ORN) population. Increasing total ORN activity also makes PN responses more transient. Strikingly, a model with just two variables (feedforward and total ORN activity) accurately predicts PN odor responses. Finally, we show that discrimination by a linear decoder is facilitated by two complementary transformations: the saturating transformation intrinsic to each processing channel boosts weak signals, while normalization helps equalize responses to different stimuli. Copyright 2010 Elsevier Inc. All rights reserved.
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                Author and article information

                Journal
                Nat. Neurosci.
                Nature neuroscience
                1546-1726
                1097-6256
                Jun 2016
                : 19
                : 6
                Affiliations
                [1 ] Friedrich Miescher Institute for Biomedical Research, Basel, Switzerland.
                [2 ] University of Basel, Basel, Switzerland.
                Article
                nn.4290
                10.1038/nn.4290
                27089019
                aaf6ee43-110f-4876-83ef-2de917e512dd
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