Two-Photon Excited Fluorescence Microscopy for the Non-Invasive Characterization of Brain Cell Metabolism and Structure
excited fluorescence (TPEF) imaging is a robust and versatile non-invasive,
non-destructive, high-resolution technique for studying cell structure and function in
2- and 3-dimensional in vitro systems. This thesis describes three applications of TPEF
for studying brain cell structure and function. In the first application, TPEF is used
to capture endogenous fluorescence ... read moreof NADH and FAD in 2D cultures of primary rat neurons
and astrocytes, as well as in cultures of adult human neural progenitor cells (AHNPs).
Analyzing distributions of pixel-wise optical redox ratios, defined as FAD/(FAD+NADH),
reveals differences in astrocyte and neuron metabolism consistent with their known
tendencies towards glycolysis and oxidative phosphorylation, respectively. Alterations
in astrocyte and neuron redox ratio distributions in response to manganese toxicity are
consistent with apoptosis and oxidative stress, and are recapitulated in a study with
Parkinson's Disease-derived AHNPs. The second application of TPEF utilizes an automated
imaging approach to quantify network density of 3D bioengineered cortical tissue. The
analysis successfully shows an increase in neurite density in cultures that incorporate
ECM derived from adult or fetal brains vs. a collagen control. The third application
describes efforts to calibrate a TPEF-based optical tweezer instrument to assess local
microrheometry of brain tissue models. In sum, these experiments demonstrate the broad
applicability of TPEF for characterizing brain cell and tissue structure and function in
Thesis (M.S.)--Tufts University, 2016.
Submitted to the Dept. of Biomedical Engineering.
Advisor: Irene Georgakoudi.
Committee: Irene Georgakoudi, David Kaplan, and Giuseppina Tesco.
Keyword: Biomedical engineering.read less
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