Biology subjects

Wallace, D. J.

Publications and source records attributed to Wallace, D. J..

7 recordsLinked to original sources

bioRxiv2025

Simultaneous 2- and 3-photon multiplane imaging across cortical layers in freely moving mice

Head-mounted multiphoton microscopes enable imaging of activity from neuronal populations spread throughout the cortical layers in freely moving mice, but so far have been restricted to recording from one cortical layer at a time. Here, combining 2- and 3-photon based excitation delivered through multiple fibers, we built a head-mounted multiplane microscope enabling near simultaneous imaging (8ns between planes) of neuronal activity from five vertically separated planes, spread across multiple cortical layers. Both excitation pathways had remote focusing mechanisms for fine axial adjustments enabling activity recordings from the same neuronal populations over weeks in freely behaving mice. The lightweight microscope utilized an onboard, 2-channel detection system designed to enable activity recordings from neuronal populations spread across visual-cortex layers in both lit and dark conditions as well as imaging activity across posterior parietal cortex layers during complex gap-crossing behaviors. We show that during gap- crossing tasks, layer 5 and 2/3 neuronal subpopulations in posterior parietal cortex have differential pattern sequences during free decision making. Editorial summaryA light-weight head-mounted multiplane microscope allows simultaneous imaging from >1800 neurons spread across the cortical layers in freely moving mice performing complex behavioral tasks, sampled over weeks.

neuroscience↗

bioRxiv2025

Acute obstructive uropathy in male C57Bl/6 mice after general anesthesia with fentanyl, midazolam and medetomidine

The selective alpha-2 adrenoreceptor agonist medetomidine is commonly used as a component of combination solutions for general anesthesia in rodents and small animals. However, it has also been reported to increase the risk of obstructive uropathy in male mice. Here we report that in a recent cohort of mice undergoing two sequential surgical procedures using the anesthetic combination of fentanyl, midazolam and medetomidine (FMM) for general anesthesia, 60% of the male mice developed severe obstructive uropathy within 48 hours of the second surgery. We also report one case of an older male mouse developing severe obstructive uropathy after a single general anesthesia with FMM. In a subsequent pilot study involving five male mice undergoing similar surgical procedures, also with two sequential surgeries, but using xylazine as the alpha-2 adrenoreceptor agonist in the narcotic combination, none of the animals developed obstructive uropathy in the post-operative period or thereafter. We suggest that replacement of medetomidine with xylazine should be considered for general anesthesia in male mice undergoing multiple successive anesthesia events or in for procedures in older males.

pathology↗

bioRxiv2022

A three-photon head-mounted microscope for imaging all layers of visual cortex in freely moving mice.

Recent advances in head-mounted microscopes have enabled imaging of neuronal activity using genetic-tools in freely moving mice but these microscopes are restricted to recording in minimally lit arenas and imaging upper cortical layers. Here we built a 2 gram, three-photon excitation-based microscope, containing a z-drive that enabled access to all cortical layers while mice freely behaved in a fully lit environment. We show that neuronal population activity in cortical layer-4 and layer-6 was differentially modulated by lit and dark conditions during free exploration.

neuroscience↗

bioRxiv2021

Anatomically-based skeleton kinetics and pose estimation in freely-moving rodents

Forming a complete picture of the relationship between neural activity and body kinetics requires quantification of skeletal joint biomechanics during behavior. However, without detailed knowledge of the underlying skeletal motion, inferring joint kinetics from surface tracking approaches is difficult, especially for animals where the relationship between surface anatomy and skeleton changes during motion. Here we developed a videography-based method enabling detailed three-dimensional kinetic quantification of an anatomically defined skeleton in untethered freely-behaving animals. This skeleton-based model has been constrained by anatomical principles and joint motion limits and provided skeletal pose estimates for a range of rodent sizes, even when limbs were occluded. Model-inferred joint kinetics for both gait and gap-crossing behaviors were verified by direct measurement of limb placement, showing that complex decision-making behaviors can be accurately reconstructed at the level of skeletal kinetics using our anatomically constrained model.

neuroscience↗

bioRxiv2021

Neutrophils contribute to ER stress in lung epithelial cells in the pristane-induced diffuse alveolar hemorrhage mouse model

Diffuse alveolar hemorrhage (DAH), although rare, is a life-threatening complication of systemic lupus erythematosus (SLE). Little is known about the pathophysiology of DAH in humans, although increasingly neutrophils, NETosis and inflammatory monocytes have been shown to play an important role in the pristane-induced model of SLE which develops lung hemorrhage and recapitulates many of the pathologic features of human DAH. Using this experimental model, we asked whether endoplasmic reticulum (ER) stress played a role in driving the pathology of pulmonary hemorrhage and what role infiltrating neutrophils had in this process. Analysis of lung tissue from pristane-treated mice showed genes associated with ER stress and NETosis were increased in a time-dependent manner and reflected the timing of CD11b+Ly6G+ neutrophil accumulation in the lung. Using precision cut lung slices from untreated mice we observed that neutrophils isolated from the peritoneal cavity of pristane-treated mice could directly induce the expression of genes associated with ER stress, namely Chop and Bip. Mice which had myeloid-specific deletion of PAD4 were generated and treated with pristane to assess the involvement of PAD4 and PAD4-dependent NET formation in pristane-induced lung inflammation. Specific deletion of PAD4 in myeloid cells resulted in decreased expression of ER stress genes in the pristane model, with accompanying reduction in IFN-driven genes and pathology. Lastly, coculture experiments of human neutrophils and human lung epithelial cell line (BEAS-2b) showed neutrophils from SLE patients induced significantly more ER stress and interferon-stimulated genes in epithelial cells compared to healthy control neutrophils. These results support a pathogenic role of neutrophils and NETs in lung injury during pristane-induced DAH through the induction of ER stress response and suggest that overactivation of neutrophils in SLE and NETosis may underlie development of DAH.

immunology↗

bioRxiv2021

Freely-moving mice visually pursue prey using a retinal area with least optic flow

Mice have a large visual field that is constantly stabilized by vestibular ocular reflex driven eye rotations that counter head-rotations. While maintaining their extensive visual coverage is advantageous for predator detection, mice also track and capture prey using vision. However, in the freely moving animal quantifying object location in the field of view is challenging. Here, we developed a method to digitally reconstruct and quantify the visual scene of freely moving mice performing a visually based prey capture task. By isolating the visual sense and combining amouse eye optic model with the head and eye rotations, the detailed reconstruction of the digital environment and retinal features were projected onto the corneal surface for comparison, and updated throughout the behavior. By quantifying the spatial location of objects in the visual scene and their motion throughout the behavior, we show that the image of the prey is maintained within a small area, the functional focus, in the upper-temporal part of the retina. This functional focus coincides with a region of minimal optic flow in the visual field and consequently minimal motion-induced image blur during pursuit, as well as the reported high density-region of Alpha-ON sustained retinal ganglion cells.

neuroscience↗

bioRxiv2018

Accurate action potential inference from a calcium sensor protein through biophysical modeling

Multiphoton imaging of genetically encoded calcium indicators is routinely used to report activity from populations of spatially resolved neurons in vivo. However, since the relationship between fluorescence and action potentials (APs) is nonlinear and varies over neurons, quantitatively inferring AP discharge is problematic. To address this we developed a biophysical model of calcium binding kinetics for the indicator GCaMP6s that accurately describes AP-evoked fluorescence changes in vivo. The models physical interpretation allowed the same parameters to describe GCaMP6s binding kinetics for both in vitro binding assays and in vivo imaging. Using this model, we developed an algorithm to infer APs from fluorescence and measured its accuracy with cell-attached electrical recordings. This approach consistently inferred more accurate AP counts and times than alternative methods for firing rates from 0 to >20 Hz, while requiring less training data. These results demonstrate the utility of quantitative, biophysically grounded models for complex biological data.

neuroscience↗