Biology subjects

Waters, J.

Publications and source records attributed to Waters, J..

35 records · Page 2Linked to original sources

bioRxiv2021

Hematopoietic stem cell requirement for macrophage regeneration is tissue-specific

Tissue-resident macrophages (TRM{Phi}) are important immune sentinels responsible for maintaining tissue and immune homeostasis within their specific niche. Recently, the origins of TRM{Phi} have undergone intense scrutiny where now most TRM{Phi} are thought to originate early during embryonic development independent of hematopoietic stem cells (HSCs). We previously characterized two distinct subsets of mouse peritoneal cavity macrophages (Large and Small Peritoneal Macrophages; LPM and SPM, respectively) whose origins and relationship to both fetal and adult long-term (LT)-HSCs have not been fully investigated. Here we employ highly purified LT-HSC transplantation and in vivo lineage tracing to show a dual ontogeny for LPM and SPM, where the initial wave of peritoneal macrophages is seeded from yolk sac-derived precursors, which later require LT-HSCs for regeneration. In contrast, transplanted fetal and adult LT-HSCs are not able to regenerate brain-resident microglia. Thus, we demonstrate that LT-HSCs retain the potential to develop into TRM{Phi}, but their requirement is tissue-specific.

immunology↗

bioRxiv2020

The effect of inclusion criteria on the functional properties reported in mouse visual cortex

Neurophysiology studies require the use of inclusion criteria to identify neurons responsive to the experimental stimuli. Five recent studies used calcium imaging to measure the preferred tuning properties of layer 2/3 pyramidal neurons in mouse visual areas. These five studies employed different inclusion criteria and report different, sometimes conflicting results. Here, we examine how different inclusion criteria can impact reported tuning properties, modifying inclusion criteria to select different sub-populations from the same dataset of almost 10,000 layer 2/3 neurons from the Allen Brain Observatory. The choice of inclusion criteria greatly affected the mean tuning properties of the resulting sub-populations; indeed, the differences in mean tuning due to inclusion criteria were often of comparable magnitude to the differences between studies. In particular, the mean preferred temporal frequencies of visual areas changed markedly with inclusion criteria, such that the rank ordering of visual areas based on their temporal frequency preferences changed with the percentage of neurons included. It has been suggested that differences in temporal frequency tuning support a hierarchy of mouse visual areas. These results demonstrate that our understanding of the functional organization of the mouse visual cortex obtained from previous experiments critically depends on the inclusion criteria used.

neuroscience↗

bioRxiv2019

The spatial structure of feedforward information in mouse primary visual cortex

Location-sensitive and motion-sensitive units are the two major functional types of feedforward projections from lateral genicular nucleus (LGN) to primary visual cortex (V1) in mouse. The distribution of these inputs in cortical depth remains under debate. By measuring the calcium activities of LGN axons in V1 of awake mice, we systematically mapped their functional and structural properties. Although both types distributed evenly across cortical depth, we found that they differ significantly across multiple modalities. Compared to the location-sensitive axons, which possessed confined spatial receptive fields, the motion-sensitive axons lacked spatial receptive fields, preferred lower temporal, higher spatial frequencies and had wider horizontal bouton spread. Furthermore, the motion-sensitive axons showed a strong depth-dependent motion direction bias while the location-sensitive axons showed a depth-independent OFF dominance. Overall, our results suggest a new model of receptive biases and laminar structure of thalamic inputs to V1. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=177 SRC="FIGDIR/small/888156v2_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@35c887org.highwire.dtl.DTLVardef@1001d54org.highwire.dtl.DTLVardef@8585b2org.highwire.dtl.DTLVardef@c8a35_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2019

On the correspondence of electrical and optical physiology in in vivo population-scale two-photon calcium imaging.

Multiphoton calcium imaging is commonly used to monitor the spiking of large populations of neurons. Recovering action potentials from fluorescence necessitates calibration experiments, often with simultaneous imaging and cell-attached recording. Here we performed calibration for imaging conditions matching those of the Allen Brain Observatory. We developed a novel crowd-sourced, algorithmic approach to quality control. Our final data set was 50 recordings from 35 neurons in 3 mouse lines. Our calibration indicated that 3 or more spikes were required to produce consistent changes in fluorescence. Moreover, neither a simple linear model nor a more complex biophysical model accurately predicted fluorescence for small numbers of spikes (1-3). We observed increases in fluorescence corresponding to prolonged depolarizations, particularly in Emx1-IRES-Cre mouse line crosses. Our results indicate that deriving spike times from fluorescence measurements may be an intractable problem in some mouse lines.

neuroscience↗

bioRxiv2019

Relationship between spiking activity and simultaneously recorded fluorescence signals in transgenic mice expressing GCaMP6

Two-photon calcium imaging is often used with genetically encoded calcium indicators (GECIs) to investigate neural dynamics, but the relationship between fluorescence and action potentials (spikes) remains unclear. Pioneering work linked electrophysiology and calcium imaging in vivo with viral GECI expression, albeit in a small number of cells. Here we characterized the spikefluorescence transfer function in vivo of 91 layer 2/3 pyramidal neurons in primary visual cortex in four transgenic mouse lines expressing GCaMP6s or GCaMP6f. We found that GCaMP6s cells have spike-triggered fluorescence responses of larger amplitude, lower variability and greater single-spike detectability than GCaMP6f. Mean single-spike detection rates at high spatiotemporal resolution measured in our data was >70% for GCaMP6s and ~40-50% for GCaMP6f (at 5% false positive rate). These rates are estimated to decrease to 25-35% for GCaMP6f under generally used population imaging conditions. Our ground-truth dataset thus supports more refined inference of neuronal activity from calcium imaging.

neuroscience↗

bioRxiv2019

Dual-plane 3-photon microscopy with remote focusing

3-photon excitation enables in vivo fluorescence microscopy deep in densely labeled and highly scattering samples. To date, 3-photon excitation has been restricted to scanning a single focus, limiting the speed of volume acquisition. Here, for the first time to our knowledge, we implemented and characterized dual-plane 3-photon microscopy with temporal multiplexing and remote focusing, and performed simultaneous in vivo calcium imaging of two planes deep in the cortex of a pan-excitatory GCaMP6s transgenic mouse. This method is a straightforward and generalizable modification to single-focus 3PE systems, doubling the rate of volume (column) imaging with off-the-shelf components and minimal technical constraints.

neuroscience↗

bioRxiv2019

Interactions between the gut microbiome and mucosal immunoglobulins A, M and G in the developing infant gut

ObjectiveInteractions between the gut microbiome and immunoglobulin (Ig) A in infancy are important for future health. IgM and IgG are also present, however, their interactions with the microbiome in the developing infant are less understood.\n\nDesignWe employed stool samples sampled 15 times in infancy from 32 healthy subjects at 4 locations in 3 countries (from the TEDDY study). We characterized patterns of microbiome development in relation to levels of IgA, IgG and IgM. For 8 infants from a single location, we FACS-sorted microbial cells from stool by Ig status. We used 16S rRNA gene profiling on full and sorted microbiomes to assess patterns of antibody coating in relation to age and other factors.\n\nResultsAll antibodies decreased in concentration with age, but were augmented by breastmilk feeding regardless of infant age. Levels of IgA correlated with the relative abundances of OTUs belonging to the Bifidobacteria and Enterobacteriaceae, which dominated the early microbiome, and IgG levels correlated with Haemophilus. The diversity of Ig-coated microbiota was influenced by breastfeeding and age, but birth mode. IgA and IgM coated the same microbiota, while IgG targeted a different subset. Blautia generally evaded antibody coating, while members of the Bifidobacteria and Enterobacteriaceae were high in IgA/M.\n\nConclusionIgA/M have similar dynamics with respect to microbiome development with age, and their interactions with the microbiome are influenced by breastfeeding status. IgG generally does not coat the commensal microbiota.\n\nSummaryWhat is already known on this subject? O_LISecretory IgA coats ~50% of microbiota in the gut\nC_LIO_LIIgM and IgG are less prevalent and coat a lower fraction in the adult, dynamics in the infant gut are not well characterized.\nC_LIO_LIBreastmilk is a source of IgA to the infant gut and decreases with time.\nC_LIO_LIIgA coating of microbial cells in infant gut microbiome decreases over time.\nC_LI\n\nWhat are the new findings? O_LIBreastfeeding augments the IgA coating of the microbiome at all ages.\nC_LIO_LIIgA and IgM coat many of the same cells, whereas few are coated by IgG alone.\nC_LIO_LIBifidobacteria, Enterobacteriaceae, Ruminococcus gnavus are enriched in IgA/M-coated cell fraction, Blautia is enriched in uncoated fraction.\nC_LIO_LIIgG levels correlated with Haemophilus.\nC_LI\n\nHow might it impact on clinical practice in the foreseeable future? O_LIIg-coated fraction of the gut microbiome could serve as a useful tool for tracking development of the infant gut microbiome, and/or identifying aberrations to immune sensing of the microbiome.\nC_LI

microbiology↗

bioRxiv2019

Separation of hemodynamic signals from GCaMP fluorescence measured with widefield imaging

Widefield calcium imaging is often used to measure brain dynamics in behaving mice. With a large field of view and a high sampling rate, widefield imaging can monitor activity from several distant cortical areas simultaneously, revealing cortical interactions. Interpretation of widefield images is complicated, however, by the absorption of light by hemoglobin, which can substantially affect the measured fluorescence. One approach to separating hemodynamics and calcium signals is to use multi-wavelength backscatter recordings to measure light absorption by hemoglobin. Following this approach, we develop a spatially-detailed regression-based method to estimate hemodynamics. The spatially-detailed model is based on a linear form of the Beer-Lambert relationship, but is fit at every pixel in the image and does not rely on the estimation of physical parameters. In awake mice of three transgenic lines, the Spatial Model offers improved separation of hemodynamics and changes in GCaMP fluorescence. The improvement is pronounced near blood vessels and, in contrast with other models based on regression or the Beer-Lambert law, can remove vascular artifacts along the sagittal midline. Compared to other separation approaches, the spatially-detailed model permits more accurate fluorescence-based determination of neuronal activity across the cortex.\n\nNEW & NOTEWORTHYThis manuscript addresses a well-known and strong source of contamination in widefield calcium imaging data: hemodynamics. To guide researchers towards the best method to separate calcium signals from hemodynamics, we compare the performance of several commonly used methods in three commonly-used Cre-driver lines, and we present a novel regression model that out-performs the other techniques we consider.

neuroscience↗

bioRxiv2019

Superficial bound of the depth limit of 2-photon imaging in mouse brain

2-photon fluorescence microscopy has been used extensively to probe the structure and functions of cells in living biological tissue. 2-photon excitation generates fluorescence from the focal plane, but also from outside the focal plane, with out-of-focus fluorescence increasing as the focus is pushed deeper into tissue. It has been suggested that the 2-photon depth limit, beyond which results become inaccurate, is where in- and out-of-focus fluorescence are equal. We found the depth limit of 2-photon excitation in mice with GCaMP6 indicator expression in all layers of visual cortex, by comparing near-simultaneous 2- and 3-photon excitation. 2-photon results were accurate only superficial to 450 m, matching the depth at which in-and out-of-focus fluorescence were equal. The expected depth limit is deeper in tissue with fewer fluorophores outside the plane of interest. Our results, from tissue with a largely homogenous distribution of fluorophores, establish a superficial bound on the 2-photon depth limit in the mouse visual cortex.

neuroscience↗

bioRxiv2019

Identification of genetic markers for cortical areas using a Random Forest classification routine and the Allen Mouse Brain Atlas

The mammalian neocortex is subdivided into a series of cortical areas that are functionally and anatomically distinct, and are often distinguished in brain sections using histochemical stains and other markers of protein expression. We searched the Allen Mouse Brain Atlas, a database of gene expression, for novel markers of cortical areas. We employed a random forest algorithm to screen for genes that change expression at area borders. We found novel genetic markers for 19 of 39 areas and provide code that quickly and efficiently searches the Allen Mouse Brain Atlas.

neuroscience↗

bioRxiv2019

Gut microbiota influence B cell function in a TLR5-dependent manner

Toll-like receptor (TLR) 5-deficient mice display aberrantly low levels of flagellin-specific antibodies (Flic-IgA) secreted into the gut, combined with excess bacterial flagellin in the gut, and together these attributes define microbiome dysbiosis (T5-dysbiosis). How TLR5 signaling deficiency results in T5-dysbiosis is unclear. Here, we address the role of B cells in T-dysbiosis. We observed that B cells do not express TLR5, and that B cell transplantation from TLR5-/- mouse donors into B-cell deficient mice resulted in a slight reduction in Flic-IgA levels compared to B-cells from WT donors. Bone marrow transplants from WT and TLR5-/- donors into recipients of both genotypes confirmed that TLR5 signaling by non-hematopoietic cells is required for T5-dysbiosis. We observed TLR5 deficiency was associated with an expanded population of IgA+ B cells. TLR5-/- mice tended to have higher richness for the IgA gene hypervariable region (CDR3 gene) variants. Transplantation of microbiomes from TLR5-/- and WT microbiomes donors into germfree mice resulted in a higher proportion of IgA-secreting B cells, and higher overall fecal IgA and anti-Flic IgA for TLR5-/- microbiome recipients. This observation indicated that the TLR5-/- mouse microbiome elicits an anti-flagellin antibody response that requires TLR5 signaling. Together these results indicate that TLR5 signaling on epithelial cells influences B cell populations and antibody repertoire.

immunology↗

bioRxiv2018

Activation of neuromodulatory axon projections in primary visual cortex during periods of locomotion and pupil dilation

Neuromodulators such as acetylcholine, noradrenaline (norepinephrine), and serotonin are released into the cortex by axons ascending from subcortical nuclei. These neuromodulators have been hypothesized to influence cortical function during behavioral periods such as arousal, locomotion, exploration, and attention. To determine when these neuromodulatory projections were active, we expressed the genetically-encoded calcium sensor GCaMP6 in neuromodulatory axons which project to the mouse primary visual cortex and performed two-photon microscopy to monitor their activity in vivo. We observed that the fluorescence of both cholinergic and noradrenergic axons increased during periods of pupil dilation, with the fluorescence of the axons rising less than one second before eye pupil dilation. We also observed increases in cholinergic and noradrenergic axon fluorescence periods of locomotion, which was accompanied by pupil dilation and nasal (forward) movement of both pupils. Locomotion was preceded by a rise in axonal fluorescence with a timing and amplitude that matched the subsequent pupil dilation, but axon fluorescence was more sustained than expected from the pupil dilation, suggesting that there is an additional physiological factor that affects cholinergic and noradrenergic axon activity in primary visual cortex during locomotion.

neuroscience↗

bioRxiv2018

Biological variation in the sizes, shapes and locations of visual cortical areas in the mouse

Visual cortex is organized into discrete sub-regions or areas that are arranged into a hierarchy and serve different functions in the processing of visual information. In our previous work, we noted that retinotopic maps of cortical visual areas differed between mice, but did not quantify these differences or determine the relative contributions of biological variation and measurement noise. Here we quantify the biological variation in the size, shape and locations of 11 visual areas in the mouse. We find that there is substantial biological variation in the sizes of visual areas, with some visual areas varying in size by two-fold across the population of mice.

neuroscience↗

bioRxiv2018

A large-scale, standardized physiological survey reveals higher order coding throughout the mouse visual cortex

To understand how the brain processes sensory information to guide behavior, we must know how stimulus representations are transformed throughout the visual cortex. Here we report an open, large-scale physiological survey of neural activity in the awake mouse visual cortex: the Allen Brain Observatory Visual Coding dataset. This publicly available dataset includes cortical activity from nearly 60,000 neurons collected from 6 visual areas, 4 layers, and 12 transgenic mouse lines from 221 adult mice, in response to a systematic set of visual stimuli. Using this dataset, we reveal functional differences across these dimensions and show that visual cortical responses are sparse but correlated. Surprisingly, responses to different stimuli are largely independent, e.g. whether a neuron responds to natural scenes provides no information about whether it responds to natural movies or to gratings. We show that these phenomena cannot be explained by standard local filter-based models, but are consistent with multi-layer hierarchical computation, as found in deeper layers of standard convolutional neural networks.

neuroscience↗

bioRxiv2018

Visual physiology of the Layer 4 cortical circuit in silico

Despite advances in experimental techniques and accumulation of large datasets concerning the composition and properties of the cortex, quantitative modeling of cortical circuits under in-vivo-like conditions remains challenging. Here we report and publicly release a biophysically detailed circuit model of layer 4 in the mouse primary visual cortex, receiving thalamo-cortical visual inputs. The 45,000-neuron model was subjected to a battery of visual stimuli, and results were compared to published work and new in vivo experiments. Simulations reproduced a variety of observations, including effects of optogenetic perturbations. Critical to the agreement between responses in silico and in vivo were the rules of functional synaptic connectivity between neurons. Interestingly, after extreme simplification the model still performed satisfactorily on many measurements, although quantitative agreement with experiments suffered. These results emphasize the importance of functional rules of cortical wiring and enable a next generation of data-driven models of in vivo neural activity and computations.\n\nAUTHOR SUMMARYHow can we capture the incredible complexity of brain circuits in quantitative models, and what can such models teach us about mechanisms underlying brain activity? To answer these questions, we set out to build extensive, bio-realistic models of brain circuitry employing systematic datasets on brain structure and function. Here we report the first modeling results of this project, focusing on the layer 4 of the primary visual cortex (V1) of the mouse. Our simulations reproduced a variety of experimental observations in a large battery of visual stimuli. The results elucidated circuit mechanisms determining patters of neuronal activity in layer 4 - in particular, the roles of feedforward thalamic inputs and specific patterns of intracortical connectivity in producing tuning of neuronal responses to the orientation of motion. Simplification of neuronal models led to specific deficiencies in reproducing experimental data, giving insights into how biological details contribute to various aspects of brain activity. To enable future development of more sophisticated models, we make the software code, the model, and simulation results publicly available.

neuroscience↗

bioRxiv2017

A suite of transgenic driver and reporter mouse lines with enhanced brain cell type targeting and functionality

Modern genetic approaches are powerful in providing access to diverse types of neurons within the mammalian brain and greatly facilitating the study of their function. We here report a large set of driver and reporter transgenic mouse lines, including 23 new driver lines targeting a variety of cortical and subcortical cell populations and 26 new reporter lines expressing an array of molecular tools. In particular, we describe the TIGRE2.0 transgenic platform and introduce Cre-dependent reporter lines that enable optical physiology, optogenetics, and sparse labeling of genetically-defined cell populations. TIGRE2.0 reporters broke the barrier in transgene expression level of single-copy targeted-insertion transgenesis in a wide range of neuronal types, along with additional advantage of a simplified breeding strategy compared to our first-generation TIGRE lines. These novel transgenic lines greatly expand the repertoire of high-precision genetic tools available to effectively identify, monitor, and manipulate distinct cell types in the mouse brain.

neuroscience↗

bioRxiv2017

Aberrant Cortical Activity In Multiple GCaMP6-Expressing Transgenic Mouse Lines

Transgenic mouse lines are invaluable tools for neuroscience but as with any technique, care must be taken to ensure that the tool itself does not unduly affect the system under study. Here we report aberrant electrical activity, similar to interictal spikes, and accompanying fluorescence events in some genotypes of transgenic mice expressing GCaMP6 genetically-encoded calcium sensors. These epileptiform events have been observed particularly, but not exclusively, in mice with Emx1-Cre and Ai93 transgenes, across multiple laboratories. The events occur at >0.1 Hz, are very large in amplitude (>1.0 mV local field potentials, >10% df/f widefield imaging signals), and typically cover large regions of cortex. Many properties of neuronal responses and behavior seem normal despite these events, though rare subjects exhibit overt generalized seizures. The underlying mechanisms of this phenomenon remain unclear, but we speculate about possible causes on the basis of diverse observations. We encourage researchers to be aware of these activity patterns while interpreting neuronal recordings from affected mouse lines and when considering which lines to study.

neuroscience↗