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ELife Mar 2016Genetically encoded calcium indicators (GECIs) allow measurement of activity in large populations of neurons and in small neuronal compartments, over times of...
Genetically encoded calcium indicators (GECIs) allow measurement of activity in large populations of neurons and in small neuronal compartments, over times of milliseconds to months. Although GFP-based GECIs are widely used for in vivo neurophysiology, GECIs with red-shifted excitation and emission spectra have advantages for in vivo imaging because of reduced scattering and absorption in tissue, and a consequent reduction in phototoxicity. However, current red GECIs are inferior to the state-of-the-art GFP-based GCaMP6 indicators for detecting and quantifying neural activity. Here we present improved red GECIs based on mRuby (jRCaMP1a, b) and mApple (jRGECO1a), with sensitivity comparable to GCaMP6. We characterized the performance of the new red GECIs in cultured neurons and in mouse, Drosophila, zebrafish and C. elegans in vivo. Red GECIs facilitate deep-tissue imaging, dual-color imaging together with GFP-based reporters, and the use of optogenetics in combination with calcium imaging.
Topics: Animals; Biosensing Techniques; Caenorhabditis elegans; Calcium; Cells, Cultured; Drosophila; Intravital Microscopy; Luminescent Proteins; Mice; Neurons; Neurophysiology; Zebrafish; Red Fluorescent Protein
PubMed: 27011354
DOI: 10.7554/eLife.12727 -
Frontiers in Immunology 2020
Topics: Animals; Diagnostic Imaging; Humans; Inflammation; Intravital Microscopy; Leukocytes
PubMed: 33013902
DOI: 10.3389/fimmu.2020.02137 -
Methods in Molecular Biology (Clifton,... 2022Atherosclerosis is a lipid-driven inflammatory disorder that narrows the arterial lumen and can induce life-threatening complications from coronary artery disease,...
Atherosclerosis is a lipid-driven inflammatory disorder that narrows the arterial lumen and can induce life-threatening complications from coronary artery disease, cerebrovascular disease, and peripheral artery disease. On a mechanistic level, the development of novel cellular-resolution intravital microscopy imaging approaches has recently enabled in vivo studies of underlying biological processes governing disease onset and progress. In particular, multiphoton microscopy has emerged as a promising intravital imaging tool utilizing two-photon-excited fluorescence and second-harmonic generation that provides subcellular resolution and increased imaging depths beyond confocal and epifluorescence microscopy. In this chapter, we describe the state-of-the-art multiphoton microscopy applied to the study of murine atherosclerosis.
Topics: Animals; Atherosclerosis; Intravital Microscopy; Mice; Microscopy, Fluorescence, Multiphoton; Photons
PubMed: 35237994
DOI: 10.1007/978-1-0716-1924-7_40 -
Proceedings of the National Academy of... Nov 2015Imaging reveals complex structures and dynamic interactive processes, located deep inside the body, that are otherwise difficult to decipher. Numerous imaging modalities... (Review)
Review
Imaging reveals complex structures and dynamic interactive processes, located deep inside the body, that are otherwise difficult to decipher. Numerous imaging modalities harness every last inch of the energy spectrum. Clinical modalities include magnetic resonance imaging (MRI), X-ray computed tomography (CT), ultrasound, and light-based methods [endoscopy and optical coherence tomography (OCT)]. Research modalities include various light microscopy techniques (confocal, multiphoton, total internal reflection, superresolution fluorescence microscopy), electron microscopy, mass spectrometry imaging, fluorescence tomography, bioluminescence, variations of OCT, and optoacoustic imaging, among a few others. Although clinical imaging and research microscopy are often isolated from one another, we argue that their combination and integration is not only informative but also essential to discovering new biology and interpreting clinical datasets in which signals invariably originate from hundreds to thousands of cells per voxel.
Topics: Animals; Biomarkers; Diagnostic Imaging; Engineering; Humans; Intravital Microscopy; Neoplasms; Tomography, Optical Coherence
PubMed: 26598657
DOI: 10.1073/pnas.1508524112 -
Science (New York, N.Y.) Apr 2020Embryonic development is a complex process that is unamenable to direct observation. In this study, we implanted a window to the mouse uterus to visualize the developing...
Embryonic development is a complex process that is unamenable to direct observation. In this study, we implanted a window to the mouse uterus to visualize the developing embryo from embryonic day 9.5 to birth. This removable intravital window allowed manipulation and high-resolution imaging. In live mouse embryos, we observed transient neurotransmission and early vascularization of neural crest cell (NCC)-derived perivascular cells in the brain, autophagy in the retina, viral gene delivery, and chemical diffusion through the placenta. We combined the imaging window with in utero electroporation to label and track cell division and movement within embryos and observed that clusters of mouse NCC-derived cells expanded in interspecies chimeras, whereas adjacent human donor NCC-derived cells shrank. This technique can be combined with various tissue manipulation and microscopy methods to study the processes of development at unprecedented spatiotemporal resolution.
Topics: Animals; Brain; Cell Division; Cell Movement; Chimera; Electroporation; Embryo, Mammalian; Embryonic Development; Female; Gene Transfer Techniques; Intravital Microscopy; Mice; Mice, Transgenic; Neovascularization, Physiologic; Neural Crest; Placenta; Pregnancy; Retina; Synaptic Transmission; Uterus
PubMed: 32273467
DOI: 10.1126/science.aba0210 -
Immunology and Cell Biology Jul 2017In this review, we discuss intravital microscopy of immune cells, starting from its historic origins to current applications in diverse organs. It is clear from a... (Review)
Review
In this review, we discuss intravital microscopy of immune cells, starting from its historic origins to current applications in diverse organs. It is clear from a quantitative review of the literature that intravital microscopy is a key tool in both historic and contemporary immunological research, providing unique advances in our understanding of immune responses. We have chosen to focus this review on how intravital microscopy methodologies are used to image specific organs or systems and we present recent descriptions of fundamental immunological processes that could not have been achieved by other methods. The following target organs/systems are discussed in more detail: cremaster muscle, skin (ear and dorsal skin fold chamber), lymph node, liver, lung, mesenteric vessels, carotid artery, bone marrow, brain, spleen, foetus and lastly vessels of the knee joint.
Topics: Animals; Histological Techniques; Humans; Immunologic Techniques; Intravital Microscopy
PubMed: 28366932
DOI: 10.1038/icb.2017.25 -
Essays in Biochemistry Oct 2019In this review, we present recent findings on the dynamic nature of the tumour microenvironment (TME) and how intravital microscopy studies have defined TME components... (Review)
Review
In this review, we present recent findings on the dynamic nature of the tumour microenvironment (TME) and how intravital microscopy studies have defined TME components in a spatiotemporal manner. Intravital microscopy has shed light into the nature of the TME, revealing structural details of both tumour cells and other TME co-habitants in vivo, how these cells communicate with each other, and how they are organized in three-dimensional space to orchestrate tumour growth, invasion, dissemination and metastasis. We will review different imaging tools, imaging reporters and fate-mapping strategies that have begun to uncover the complexity of the TME in vivo.
Topics: Animals; Extracellular Matrix; Humans; Intravital Microscopy; Neoplasm Metastasis; Neoplasms; Signal Transduction; Tumor Microenvironment
PubMed: 31654075
DOI: 10.1042/EBC20190019 -
Trends in Cell Biology May 2022Highly motile and functionally diverse immune cells orchestrate effective immune responses through complex and dynamic cooperative behavior. Multiphoton intravital... (Review)
Review
Highly motile and functionally diverse immune cells orchestrate effective immune responses through complex and dynamic cooperative behavior. Multiphoton intravital microscopy (MP-IVM) presents a unique and powerful tool to study the coordinated action of immune cell interactions in situ. Here, we review the current state of intravital microscopy in deepening our understanding of the immune system and discuss its fundamental limitations. In addition, we draw insights from recent technical advances in multiplex static tissue-imaging methods and propose an approach that could enable simultaneous visualization of cellular dynamics, deep phenotyping, and transcriptional states through a new type of correlative microscopy that combines these imaging technologies with advances in complex data analysis.
Topics: Cell Communication; Humans; Immune System; Intravital Microscopy; Microscopy, Fluorescence, Multiphoton
PubMed: 34920936
DOI: 10.1016/j.tcb.2021.11.007 -
Genesis (New York, N.Y. : 2000) Sep 2022Craniofacial and appendicular bone homeostasis is dynamically regulated by a balance between bone formation and resorption by osteoblasts and osteoclasts, respectively.... (Review)
Review
Craniofacial and appendicular bone homeostasis is dynamically regulated by a balance between bone formation and resorption by osteoblasts and osteoclasts, respectively. Despite the developments in multiple imaging techniques in bone biology, there are still technical challenges and limitations in the investigation of spatial/anatomical location of rare stem/progenitor cells and their molecular regulation in tooth and craniofacial bones of living animals. Recent advances in live animal imaging techniques for the craniofacial and dental apparatus can provide new insights in real time into bone stem/progenitor cell dynamics and function in vivo. Here, we review the current inventions and applications of the noninvasive intravital imaging technique and its practical uses and limitations in the analysis of stem/progenitor cells in craniofacial and dental apparatus in vivo. Furthermore, we also explore the potential applications of intravital microscopy in the dental field.
Topics: Animals; Bone and Bones; Intravital Microscopy; Molecular Imaging; Osteoclasts; Stem Cells
PubMed: 35980285
DOI: 10.1002/dvg.23498 -
Frontiers in Endocrinology 2022Human islet transplantations into rodent models are an essential tool to aid in the development and testing of islet and cellular-based therapies for diabetes prevention... (Review)
Review
Human islet transplantations into rodent models are an essential tool to aid in the development and testing of islet and cellular-based therapies for diabetes prevention and treatment. Through the ability to evaluate human islets in an setting, these studies allow for experimental approaches to answer questions surrounding normal and disease pathophysiology that cannot be answered using other and techniques alone. Intravital microscopy enables imaging of tissues in living organisms with dynamic temporal resolution and can be employed to measure biological processes in transplanted human islets revealing how experimental variables can influence engraftment, and transplant survival and function. A key consideration in experimental design for transplant imaging is the surgical placement site, which is guided by the presence of vasculature to aid in functional engraftment of the islets and promote their survival. Here, we review transplantation sites and mouse models used to study beta cell biology using intravital microscopy and we highlight fundamental observations made possible using this methodology.
Topics: Mice; Animals; Humans; Islets of Langerhans Transplantation; Islets of Langerhans; Graft Survival; Intravital Microscopy; Insulin-Secreting Cells; Disease Models, Animal
PubMed: 36277698
DOI: 10.3389/fendo.2022.992540