Systems Neuroscience Imaging Resource

National Institute Of Mental Health

Investigators

Linked publications & trials

Abstract

Summary The mission of the NIMH Systems Neuroscience Imaging Resource (SNIR) is to make advanced light microscopy related techniques available to Intramural Program investigators. This is its ninth year of operation. SNIR functions can be divided into three interacting domains: acquisition and maintenance of equipment and software, development and implementation of procedures, and training. During the COVID-19 pandemic the resource made many of its systems available through remote access. This has been continued and expanded and includes the ability to use all image processing workstations and software remotely. Major supported equipment includes: 1) Zeiss AxioscanZ1 slide scanning microscope (2016 acquisition). This is a high quality widefield microscope with transmitted brightfield and fluorescent epi-illumination capacity. Its most significant feature is the ability to program multichannel tiled acquisition of large areas from up to 100 microscope slides. It is being actively used by investigators from >10 intramural laboratories for projects that include whole brain mapping of gene expression profiles and projections of genetically tagged and fluorescently labeled neuron populations. The system is used to capacity. 2) Zeiss LSM780 microscope (2011 acquisition, now a secondary instrument). This is a high quality inverted confocal microscope with 405, 488, 514, 561, 594 and 633 nm lasers, a 32-channel GaAsP based spectral detector and 2 conventional PMTs. 3) Leica SP8 confocal/multiphoton system (2017 acquisition). This is an upright microscope equipped with both standard objectives and long working distance dipping objectives designed for work with thick cleared samples. It is equipped with 405, 488, 552 and 638 nm fixed lasers and an Insight X3 tunable IR laser, and both internal, and external non-descanned, PMT and HyD detectors. It has capacity to perform fluorescent lifetime imaging microscopy (FLIM). 4) Nikon A1HR confocal system (2020 acquisition). This is set up for both widefield epillumination and laser scanning confocal imaging and includes both galvanometer and resonant scanning ability. It is equipped with 405, 488, 561, 630 and 750 nm lasers. 6) Leica Stellaris confocal instrument (2020 acquisition). This is equipped with a 405nm diode and a tunable pulsed white light laser as well as 5 tunable high sensitivity detectors that support time gated acquisition, providing precise control of excitation wavelength and emission windows from 405 to 800 nm. Image acquisition can be done in galvanometer or resonant modes. Capability for FLIM was recently added. 5) Nikon Biopipeline Slide System (2020 acquisition). This is a slide scanning instrument that can provide customizable control over acquisition, including imaging of selected regions of interest in either widefield or confocal mode (using confocal components repurposed from a Nikon C2 system). 6) Intelligent Imaging Innovations (3i) Cleared Tissue Lightsheet Microscope (CTLS) (2022 acquisition; 2024 upgrade to Axl configuration). This is an advanced light sheet instrument designed for cleared tissue imaging. It was recently upgraded to the Axl configuration, which uses electronically tunable lenses for illumination so that the lighsheet waist can be swept across the field in synchrony with the cameras rolling shutter, creating a uniform sheet thickness across the field. It is used with (NA 0.25, 0.35) non-immersion objectives that allow essentially any clearing media to be imaged. It is equipped with 488, 561, 633 and 750 nm lasers. Whole mouse brains immunolabeled with the iDISCO technique and cleared with CUBIC, SHIELD and related procedures are being imaged routinely and pilot work with rat and marmoset is underway. Major supported software includes: Microbrightfield Brainmaker and Neurolucida 360. These packages facilitate reconstruction and analysis of the distribution and morphology of labeled neurons. Arivis Pro (Vision4D) is available for visualization of large - dimensional datasets and implementation of analysis pipelines including segmentation using explicit algorithms and machine learning. In addition, custom python-based code was developed within the group for denoising, deconvolution, dehazing, stitching, segmentation and registration of massive data sets, taking advantage of the NIH Biowulf computational cluster. Training provided this year included: 1) Initial use of each of the microscopes and the software packages described above. 2) Development of optimized acquisition and analysis strategies. 3) Ad hoc assistance during microscope and software use. 3) Use of iDISCO based labeling and clearing for whole mouse brain mapping of immediate early gene distribution. 4) Use of a custom pipeline for atlas registration and whole brain analysis of cells with immediate early gene activation. 5) Whole brain imaging using CUBIC and SHIELD for clearing. 6) Multiplex in situ hybridization. 7) Custom pipelines for multiple round in situ hybridization including subtraction of endogenous fluorescent material (lipofuchsin) and registration of material that has been stripped and reprobed. Recent publications that used SNIR resources are listed in the project bibliography and in addition include: Keary KM III, Sojka E, Gonzalez M, Li Z (2024) Dendritic Spine Quantification Using an Automatic Three-Dimensional Neuron Reconstruction Software. J Vis Exp. 2024 Sep 27. Ma, J., O’Malley, J.J., Kreiker, M. et al. Convergent direct and indirect cortical streams shape avoidance decisions in mice via the midline thalamus. Nat Commun 15, 6598 (2024). https://doi.org/10.1038/s41467-024-50941-6 Inácio AR, Lam KC, Zhao Y, Pereira F, Gerfen CR, Lee S. Brain-wide presynaptic networks of functionally distinct cortical neurons. Nature. 2025 May;641(8061):162-172. doi: 10.1038/s41586-025-08631-w. Epub 2025 Feb 26. PMID: 40011781; PMCID: PMC12043506. Ma J, O'Malley JJ, Kreiker M, Leng Y, Khan I, Kindel M, Penzo MA. Convergent direct and indirect cortical streams shape avoidance decisions in mice via the midline thalamus. Nat Commun. 2024 Aug 4;15(1):6598. doi: 10.1038/s41467-024-50941-6. PMID: 39097600; PMCID: PMC11297946. Beas S, Khan I, Gao C, Loewinger G, Macdonald E, Bashford A, Rodriguez-Gonzalez S, Pereira F, Penzo MA. Dissociable encoding of motivated behavior by parallel thalamo-striatal projections. Curr Biol. 2024 Apr 8;34(7):1549-1560.e3. doi: 10.1016/j.cub.2024.02.037. Epub 2024 Mar 7. PMID: 38458192; PMCID: PMC11003833. Nagarajan G, Matrov D, Pearson AC, Yen CC, Bradley SP, Chudasama Y. Cingulate cortex shapes early postnatal development of social vocalizations. Elife. 2025 Jul 25;13:RP97125. doi: 10.7554/eLife.97125. PMID: 40709916; PMCID: PMC12296256. Visocky V, Turner CJ, Lowrie MH, Alibro A, Messanvi F, Chudasama Y. Noradrenergic modulation of stress induced catecholamine release: Opposing influence of FG7142 and yohimbine. Prog Neuropsychopharmacol Biol Psychiatry. 2025 Apr 2;138:111314. doi: 10.1016/j.pnpbp.2025.111314. Epub 2025 Mar 5. PMID: 40054569.