Article
Epilepsy in the laser spotlight – multiphoton microscopy of hippocampal tissue
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Authors
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Ortrud Uckermann
- Klinik und Poliklinik für Neurochirurgie, Universitätsklinikum Carl Gustav Carus an der Technischen Universität Dresden
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Roberta Galli
- Klinisches Sensoring und Monitoring, Medizinische Fakultät, Technische Universität Dresden
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Susann Leupold
- Klinik und Poliklinik für Neurochirurgie, Universitätsklinikum Carl Gustav Carus an der Technischen Universität Dresden
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Katrin D. Geiger
- Abteilung Neuropathologie, Institut für Pathologie, Universitätsklinikum Carl Gustav Carus an der Technischen Universität Dresden
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Gabriele Schackert
- Klinik und Poliklinik für Neurochirurgie, Universitätsklinikum Carl Gustav Carus an der Technischen Universität Dresden
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Gerald Steiner
- Klinisches Sensoring und Monitoring, Medizinische Fakultät, Technische Universität Dresden
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Matthias Kirsch
- Klinik und Poliklinik für Neurochirurgie, Universitätsklinikum Carl Gustav Carus an der Technischen Universität Dresden
| Published: | June 2, 2015 |
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Outline
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Objective: Multiphoton microscopy enables the visualization of tissue properties and structure without any labeling or tissue preparation and can be applied on native tissue. We investigated the ability of this technique to detect cytological changes in epileptic brain tissue.
Method: Human brain tissue samples of ammon's horn sclerosis (n=11) were obtained during routine surgeries for treatment of epilepsy. Multiphoton microscopy was used to investigate fresh, bulk samples and tissue cryosections. Second harmonic generation (SHG) addressing collagen, two photon excited fluorescence (TPEF) visualizing endogenous fluorophores and coherent anti Stokes Raman scattering (CARS) addressing mainly tissue lipid content were combined in multimodal images. For reference, cryosections were subjected to standard H&E staining, and immunohistochemistry for GFAP, Ki67, NeuN, and CD34.
Results: Multimodal multiphoton microscopy of the unstained tissue displayed the hippocampal layering and micromorphological details in both bulk samples and cryosections: CARS identified white and gray matter layers and the high lateral resolution of the technique allowed the assessment of axonal structures. SHG revealed the size and location of blood vessels based on adventitial collagen. In addition, SHG-active corpora amylacea (CoA) were found in seven samples with varying densities up to 2181/ mm2. They were always located in close relationship to blood vessels and in areas with marked neuronal loss. Pyramidal neuronal cell bodies, but not granular cells were characterized by intense cytoplasmatic endogenous TPEF.
Conclusions: We developed an optical technique that analyses pathological aspects of ammon's horn sclerosis on native tissue without any labels or stainings. Most importantly, pyramidal cells and axonal tracts as factors of normal cortical layers as well as corpora amylacea indicative of sclerosis can be identified. In conclusion, multimodal multiphoton microscopy holds prospect for fast, intraoperative analysis of the extent of altered tissue composition in epilepsy surgery.
