Biology subjects

Kilborn, K.

Publications and source records attributed to Kilborn, K..

5 recordsLinked to original sources

bioRxiv2024

Opto2P-FCM: A MEMS based miniature two-photon microscope with two-photon patterned optogenetic stimulation

Multiphoton microscopy combined with optogenetic photostimulation is a powerful technique in neuroscience enabling precise control of cellular activity to determine the neural basis of behavior in a live animal. Two-photon patterned photostimulation has taken this further by allowing interrogation at the individual neuron level. However, it remains a challenge to implement imaging of neural activity with spatially patterned two-photon photostimulation in a freely moving animal. We developed a miniature microscope for high resolution two-photon fluorescence imaging with patterned two-photon optogenetic stimulation. The design incorporates a MEMS scanner for two-photon imaging and a second beam path for patterned two-photon excitation in a compact and lightweight design that can be head-attached to a freely moving animal. We demonstrate cell-specific optogenetics and high resolution MEMS based two-photon imaging in a freely moving mouse. The new capabilities of this miniature microscope design can enable cell-specific studies of behavior that can only be done in freely moving animals.

neuroscience↗

bioRxiv2024

CelFDrive: Artificial Intelligence assisted microscopy for automated detection of rare events

11.1 SummaryCelFDrive automates high-resolution 3D imaging cells of interest across a variety of fluorescence microscopes, integrating deep learning cell classification from auxiliary low resolution widefield images. CelFDrive enables efficient detection of rare events in large cell populations, such as the onset of cell division, and subsequent rapid switching to 3D imaging modes, increasing the speed for finding cells of interest by an order of magnitude. 1.2 Availability and ImplementationCelFDrive is available freely for academic purposes at the CelFDrive GitHub repository. and can be installed on Windows, macOS or Linux-based machines with relevant conda environments [1]. To interact with microscopy hardware requires additional software; we use SlideBook software from Intelligent Imaging Innovations (3i), but CelFDrive can be deployed with any microscope control software that can interact with a Python environment. Graphical Processing Units (GPUs) are recommended to increase the speed of application but are not required. On 3i systems the software can be deployed with a range of microscopes including their Lattice LightSheet microscope (LLSM) and spinning disk confocal (SDC). 1.3 Contacts.brooks.2@warwick.ac.uk

bioengineering↗

bioRxiv2023

Long-term in vivo three-photon imaging reveals region-specific differences in healthy and regenerative oligodendrogenesis

The generation of new myelin-forming oligodendrocytes in the adult CNS is critical for cognitive function and regeneration following injury. Oligodendrogenesis varies between gray and white matter regions suggesting that local cues drive regional differences in myelination and the capacity for regeneration. Yet, the determination of regional variability in oligodendrocyte cell behavior is limited by the inability to monitor the dynamics of oligodendrocytes and their transcriptional subpopulations in white matter of the living brain. Here, we harnessed the superior imaging depth of three-photon microscopy to permit long-term, longitudinal in vivo three-photon imaging of an entire cortical column and underlying subcortical white matter without cellular damage or reactivity. Using this approach, we found that the white matter generated substantially more new oligodendrocytes per volume compared to the gray matter, yet the rate of population growth was proportionally higher in the gray matter. Following demyelination, the white matter had an enhanced population growth that resulted in higher oligodendrocyte replacement compared to the gray matter. Finally, deep cortical layers had pronounced deficits in regenerative oligodendrogenesis and restoration of the MOL5/6-positive oligodendrocyte subpopulation following demyelinating injury. Together, our findings demonstrate that regional microenvironments regulate oligodendrocyte population dynamics and heterogeneity in the healthy and diseased brain.

neuroscience↗

bioRxiv2021

Characterization of red fluorescent reporters for dual-color in vivo three-photon microscopy

SignificanceThree-photon (3P) microscopy significantly increases the depth and resolution of in vivo imaging due to decreased scattering and nonlinear optical sectioning. Simultaneous excitation of multiple fluorescent proteins is essential to study multicellular interactions and dynamics in the intact brain. AimWe characterized the excitation laser pulses at a range of wavelengths for 3P microscopy, and then explored the application of tdTomato or mScarlet and EGFP for dual-color single-excitation structural 3P imaging deep in the living mouse brain. ApproachWe used frequency-resolved optical gating to measure the spectral intensity, phase, and retrieved pulse widths at a range of wavelengths. Then, we performed in vivo single wavelength-excitation 3P imaging in the 1225 - 1360 nm range deep in the mouse cerebral cortex to evaluate the performance of tdTomato or mScarlet in combination with EGFP. ResultsWe find that tdTomato and mScarlet, expressed in oligodendrocytes and neurons respectively, have high signal-to-background in the 1300-1360 nm range, consistent with enhanced 3P cross sections. ConclusionsThese results suggest that a single excitation wavelength source is advantageous for multiple applications of dual-color brain imaging and highlight the importance of empirical characterization of individual fluorophores for 3P microscopy.

neuroscience↗

bioRxiv2019

Brain BOLD MRI O2 and CO2 stress testing: Implications for perioperative neurocognitive disorder following surgery

Respiratory end-tidal (ET) gas control is fundamental to anaesthetic management. The range of ET O2 and CO2 during the conduct of anaesthesia can significantly deviate from values in the awake state. Recent work shows ET CO2 influences the incidence of perioperative neurocognitive disorder (POND). We examine the effects of controlled alterations in both ET O2 and CO2 on cerebral blood flow (CBF) in awake adults using BOLD MRI. Twelve healthy adults had BOLD and CBF responses measured to alterations in ET CO2 and O2 in various combinations commonly observed under anaesthesia. Dynamic alterations in regional BOLD and CBF were seen in all subjects with expected and inverse responses to both stimuli. These effects were incremental and rapid (within seconds). The most dramatic effects were seen with combined hyperoxia and hypocapnia. Inverse responses increased with age. Here we show that human brain CBF responds dramatically to alterations in ET respiratory gas tensions commonly seen during anaesthesia. Such alterations may impact the observed incidence of POND following surgery and intensive care, and is an important area for further investigation.

neuroscience↗