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5 Janelia Publications

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    Ji LabGENIE
    08/20/18 | In vivo measurement of afferent activity with axon-specific calcium imaging.
    Broussard GJ, Liang Y, Fridman M, Unger EK, Meng G, Xiao X, Ji N, Petreanu L, Tian L
    Nature Neuroscience. 2018 Aug 20:. doi: 10.1038/s41593-018-0211-4

    In vivo calcium imaging from axons provides direct interrogation of afferent neural activity, informing the neural representations that a local circuit receives. Unlike in somata and dendrites, axonal recording of neural activity-both electrically and optically-has been difficult to achieve, thus preventing comprehensive understanding of neuronal circuit function. Here we developed an active transportation strategy to enrich GCaMP6, a genetically encoded calcium indicator, uniformly in axons with sufficient brightness, signal-to-noise ratio, and photostability to allow robust, structure-specific imaging of presynaptic activity in awake mice. Axon-targeted GCaMP6 enables frame-to-frame correlation for motion correction in axons and permits subcellular-resolution recording of axonal activity in previously inaccessible deep-brain areas. We used axon-targeted GCaMP6 to record layer-specific local afferents without contamination from somata or from intermingled dendrites in the cortex. We expect that axon-targeted GCaMP6 will facilitate new applications in investigating afferent signals relayed by genetically defined neuronal populations within and across specific brain regions.

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    Ji Lab
    08/01/18 | Optical alignment device for two-photon microscopy.
    Galiñanes GL, Marchand PJ, Turcotte R, Pellat S, Ji N, Huber D
    Biomedical Optics Express. 2018 Aug 1;9(8):3624-9. doi: 10.1364/BOE.9.003624

    Two-photon excitation fluorescence microscopy has revolutionized our understanding of brain structure and function through the high resolution and large penetration depth it offers. Investigating neural structures in vivo requires gaining optical access to the brain, which is typically achieved by replacing a part of the skull with one or several layers of cover glass windows. To compensate for the spherical aberrations caused by the presence of these layers of glass, collar-correction objectives are typically used. However, the efficiency of this correction has been shown to depend significantly on the tilt angle between the glass window surface and the optical axis of the imaging system. Here, we first expand these observations and characterize the effect of the tilt angle on the collected fluorescence signal with thicker windows (double cover slide) and compare these results with an objective devoid of collar-correction. Second, we present a simple optical alignment device designed to rapidly minimize the tilt angle in vivo and align the optical axis of the microscope perpendicularly to the glass window to an angle below 0.25°, thereby significantly improving the imaging quality. Finally, we describe a tilt-correction procedure for users in an in vivo setting, enabling the accurate alignment with a resolution of <0.2° in only few iterations.

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    Ji Lab
    06/01/18 | Adaptive optical microscopy for neurobiology.
    Rodriguez C, Ji N
    Current Opinion in Neurobiology. 2018 Jun;50:83-91. doi: 10.1016/j.conb.2018.01.011


    • Biological specimens introduce wavefront aberrations and deteriorate the image quality of optical microscopy.
    • Adaptive optics is used in optical microscopy to recover ideal imaging performance.
    • Adaptive optical imaging improves structural imaging of neurons, allowing for synaptic-level resolution at depth.
    • Adaptive optical imaging leads to a more accurate characterization of the functional properties of neurons.

    With the ability to correct for the aberrations introduced by biological specimens, adaptive optics—a method originally developed for astronomical telescopes—has been applied to optical microscopy to recover diffraction-limited imaging performance deep within living tissue. In particular, this technology has been used to improve image quality and provide a more accurate characterization of both structure and function of neurons in a variety of living organisms. Among its many highlights, adaptive optical microscopy has made it possible to image large volumes with diffraction-limited resolution in zebrafish larval brains, to resolve dendritic spines over 600μm deep in the mouse brain, and to more accurately characterize the orientation tuning properties of thalamic boutons in the primary visual cortex of awake mice.

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    Ji Lab
    04/15/18 | Three-photon fluorescence microscopy with an axially elongated Bessel focus.
    Rodriguez C, Liang Y, Lu R, Ji N
    Optics Letters. 2018 Apr 15;43(8):1914-1917. doi: 10.1364/OL.43.001914

    Volumetric imaging tools that are simple to adopt, flexible, and robust are in high demand in the field of neuroscience, where the ability to image neurons and their networks with high spatiotemporal resolution is essential. Using an axially elongated focus approximating a Bessel beam, in combination with two-photon fluorescence microscopy, has proven successful at such an endeavor. Here, we demonstrate three-photon fluorescence imaging with an axially extended Bessel focus. We use an axicon-based module that allowed for the generation of Bessel foci of varying numerical apertures and axial lengths, and apply this volumetric imaging tool to image mouse brain slices and for in vivo imaging of the mouse brain.

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    Ji Lab
    04/01/18 | 50 Hz volumetric functional imaging with continuously adjustable depth of focus.
    Lu R, Tanimoto M, Koyama M, Na J
    Biomedical Optics Express. 2018 Apr;9(4):1964-76. doi: 10.1364/BOE.9.001964

    Understanding how neural circuits control behavior requires monitoring a large population of neurons with high spatial resolution and volume rate. Here we report an axicon-based Bessel beam module with continuously adjustable depth of focus (CADoF), that turns frame rate into volume rate by extending the excitation focus in the axial direction while maintaining high lateral resolutions. Cost-effective and compact, this CADoF Bessel module can be easily integrated into existing two-photon fluorescence microscopes. Simply translating one of the relay lenses along its optical axis enabled continuous adjustment of the axial length of the Bessel focus. We used this module to simultaneously monitor activity of spinal projection neurons extending over 60 µm depth in larval zebrafish at 50 Hz volume rate with adjustable axial extent of the imaged volume.

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