We introduce a web-based tool, Peak Annotation and Visualization (PAVIS), for annotating and visualizing ChIP-seq peak data. PAVIS is designed with non-bioinformaticians in mind and presents a straightforward user interface to facilitate biological interpretation of ChIP-seq peak or other genomic enrichment data. PAVIS, through association with annotation, provides relevant genomic context for each peak, such as peak location relative to genomic features including transcription start site, intron, exon or 5'/3'-untranslated region. PAVIS reports the relative enrichment P-values of peaks in these functionally distinct categories, and provides a summary plot of the relative proportion of peaks in each category. PAVIS, unlike many other resources, provides a peak-oriented annotation and visualization system, allowing dynamic visualization of tens to hundreds of loci from one or more ChIP-seq experiments, simultaneously. PAVIS enables rapid, and easy examination and cross-comparison of the genomic context and potential functions of the underlying genomic elements, thus supporting downstream hypothesis generation.
The continuous addition of new dentate granule cells (DGCs), which is regulated exquisitely by brain activity, renders the hippocampus plastic. However, how neural circuits encode experiences to affect the addition of adult-born neurons remains unknown. Here, we used endoscopic Ca 2ϩ imaging to track the real-time activity of individual DGCs in freely behaving mice. For the first time, we found that active DGCs responded to a novel experience by increasing their Ca 2ϩ event frequency preferentially. This elevated activity, which we found to be associated with object exploration, returned to baseline by 1 h in the same environment, but could be dishabituated via introduction to a novel environment. To transition seamlessly between environments, we next established a freely controllable virtual reality system for unrestrained mice. We again observed increased firing of active neurons in a virtual enriched environment. Interestingly, multiple novel virtual experiences increased the number of newborn neurons accumulatively compared with a single experience. Finally, optogenetic silencing of existing DGCs during novel environmental exploration perturbed experience-induced neuronal addition. Our study shows that the adult brain conveys novel, enriched experiences to increase the addition of adult-born hippocampal neurons by increasing the firing of active DGCs.
Visualizing diverse anatomical and functional traits that span many spatial scales with high spatio-temporal resolution provides insights into the fundamentals of living organisms. Light-field microscopy (LFM) has recently emerged as a scanning-free, scalable method that allows for high-speed, volumetric functional brain imaging. Given those promising applications at the tissue level, at its other extreme, this highly-scalable approach holds great potential for observing structures and dynamics in single-cell specimens. However, the challenge remains for current LFM to achieve a subcellular level, neardiffraction-limited 3D spatial resolution. Here, we report high-resolution LFM (HR-LFM) for live-cell imaging with a resolution of 300-700 nm in all three dimensions, an imaging depth of several micrometers, and a volume acquisition time of milliseconds. We demonstrate the technique by imaging various cellular dynamics and structures and tracking single particles. The method may advance LFM as a particularly useful tool for understanding biological systems at multiple spatio-temporal levels.
Visualizing diverse anatomical and functional traits that span many spatial scales with high spatio-temporal resolution provides insights into the fundamentals of living organisms. Light-field microscopy (LFM) has recently emerged as a scanning-free, scalable method that allows for high-speed, volumetric functional brain imaging. Given those promising applications at the tissue level, at its other extreme, this highly-scalable approach holds great potential for observing structures and dynamics in single-cell specimens. However, the challenge remains for current LFM to achieve subcellular level, near-diffraction-limited 3D spatial resolution. Here, we report high-resolution LFM (HR-LFM) for live-cell imaging with a resolution of 300-700 nm in all three dimensions, an imaging depth of several micrometers, and a volume acquisition time of milliseconds. We demonstrate the technique by imaging various cellular dynamics and structures and tracking single particles. The method may advance LFM as a particularly useful tool for understanding biological systems at multiple spatiotemporal levels. Fig. 13. Imaging Golgi-derived membrane vesicles in living COS-7 cells at a volume acquisition time of 5 ms using HR-LFM. (a) Reconstructed 3D images of vesicles in living COS-7 cells labeled with mEmerald-Golgi-7 acquired at a volume acquisition time of 5 ms. The left and right are reconstructed 3D images at t = 0.005s and 5.000s of a 1000-time-point series, respectively. The depth-information is color-coded according to the color scale bar. (b) Zoomedin images of the corresponding boxed region in (a) at t = 0.005s, 3.000s, 3.500s, 4.000s, 4.500s and 5.000s, respectively. White arrows indicate vesicles moving during time-points. (c) Zoomed-in image (leftmost) of the boxed region in (b) at t = 0.005s and its z-stack images from z = 0.2 µm to +1.3 µm at an axial step size of 100 nm, resolving several nearby vesicles. (d) Zoomed-in images of the corresponding boxed region in (a) at t = 0.005s, 0.500s, 1.000s, 1.500s, 2.000s, 2.500s, 3.000s and 3.500s, respectively. Scale bars: 10 µm (a), 1 µm (b-d).
We report a depth-extended, high-resolution fluorescence microscopy system based on interfering Bessel beams generated with double-ring phase (DRiP) modulation. The DRiP method effectively suppresses the Bessel side lobes, exhibiting a high resolution of the main lobe throughout a four-to five-fold improved depth of focus (DOF), compared to conventional wide-field microscopy. We showed both theoretically and experimentally the generation and propagation of a DRiP point-spread function (DRiP-PSF) of the imaging system. We further developed an approach for creating an axially-uniform DRiP-PSF and successfully demonstrated diffraction-limited, depth-extended imaging of cellular structures. We expect the DRiP method to contribute to the fast-developing field of non-diffractingbeam-enabled optical microscopy and be useful for various types of imaging modalities.
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