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Multiphoton Intravital Calcium Imaging.

ABSTRACT: Multiphoton intravital calcium imaging is a powerful technique that enables high-resolution longitudinal monitoring of cellular and subcellular activity hundreds of microns deep in the living organism. This unit addresses the application of 2-photon microscopy to imaging of genetically encoded calcium indicators (GECIs) in the mouse brain. The protocols in this unit enable real-time intravital imaging of intracellular calcium concentration simultaneously in hundreds of neurons, or at the resolution of single synapses, as mice respond to sensory stimuli or perform behavioral tasks. Protocols are presented for implantation of a cranial imaging window to provide optical access to the brain and for 2-photon image acquisition. Protocols for implantation of both open skull and thinned skull windows for single or multi-session imaging are described. © 2018 by John Wiley & Sons, Inc.

SUBMITTER: Cheetham CEJ

PROVIDER: S-EPMC6039231 | biostudies-literature | 2018 Jul

REPOSITORIES: biostudies-literature

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Multiphoton Intravital Calcium Imaging.

Cheetham Claire E J CEJ

Current protocols in cytometry 20180626 1

Multiphoton intravital calcium imaging is a powerful technique that enables high-resolution longitudinal monitoring of cellular and subcellular activity hundreds of microns deep in the living organism. This unit addresses the application of 2-photon microscopy to imaging of genetically encoded calcium indicators (GECIs) in the mouse brain. The protocols in this unit enable real-time intravital imaging of intracellular calcium concentration simultaneously in hundreds of neurons, or at the resolut ...[more]

PMID: 29944202

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Project description:BackgroundCancer is a highly complex disease which involves the co-operation of tumor cells with multiple types of host cells and the extracellular matrix. Cancer studies which rely solely on static measurements of individual cell types are insufficient to dissect this complexity. In the last two decades, intravital microscopy has established itself as a powerful technique that can significantly improve our understanding of cancer by revealing the dynamic interactions governing cancer initiation, progression and treatment effects, in living animals. This review focuses on intravital multiphoton microscopy (IV-MPM) applications in mouse models of cancer.Recent findingsIV-MPM studies have already enabled a deeper understanding of the complex events occurring in cancer, at the molecular, cellular and tissue levels. Multiple cells types, present in different tissues, influence cancer cell behavior via activation of distinct signaling pathways. As a result, the boundaries in the field of IV-MPM are continuously being pushed to provide an integrated comprehension of cancer. We propose that optics, informatics and cancer (cell) biology are co-evolving as a new field. We have identified four emerging themes in this new field. First, new microscopy systems and image processing algorithms are enabling the simultaneous identification of multiple interactions between the tumor cells and the components of the tumor microenvironment. Second, techniques from molecular biology are being exploited to visualize subcellular structures and protein activities within individual cells of interest, and relate those to phenotypic decisions, opening the door for "in vivo cell biology". Third, combining IV-MPM with additional imaging modalities, or omics studies, holds promise for linking the cell phenotype to its genotype, metabolic state or tissue location. Finally, the clinical use of IV-MPM for analyzing efficacy of anti-cancer treatments is steadily growing, suggesting a future role of IV-MPM for personalized medicine.ConclusionIV-MPM has revolutionized visualization of tumor-microenvironment interactions in real time. Moving forward, incorporation of novel optics, automated image processing, and omics technologies, in the study of cancer biology, will not only advance our understanding of the underlying complexities but will also leverage the unique aspects of IV-MPM for clinical use.

Dataset Information

Multiphoton Intravital Calcium Imaging.

Publications

Multiphoton Intravital Calcium Imaging.

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