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Two-photon laser scanning microscopy (2PLSM) in living animals and brain tissue

2PLSM utilizes the ability of near-infrared lasers to penetrate deeply into highly scattering environments, permitting high resolution imaging at subcellular level in the intact brain.
Two 2PLSM set-ups are present in our department, one suitable for imaging in the living brain (in vivo) and the other adapted for use on freshly isolated brain slices (ex-vivo). Both set-ups combine the possibility of two-photon uncaging. In addition, the set-up for ex-vivo studies is equipped with a patch-clamp unit, while the one for in vivo studies with a fast 3D scanning piezo device enabling real-time imaging of large glial and neuronal networks.
The selective expression of fluorescent proteins in distinct brain cell populations (neurons, astrocytes, microglia) and the loading of specific dyes are exploited for morphological analysis as well as for functional studies, such as the detection of intracellular Ca2+ dynamics, in both physiological and pathological conditions. Physiological paradigms, such as sensory stimulation, as well as animal models of brain pathologies are utilized in these studies. See Volterra

 
Image of an astrocyte (green) contacting a bifurcating blood vessel (red) in layer 2 of the cerebral cortex in vivo


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