title: Microfiberoptic measurement of extracellular space volume in brain and tumor slices based on fluorescent dye partitioning.
abstract: The fractional volume occupied by extracellular space in tissues, termed alpha, is an important parameter of tissue architecture that affects cellular functions and drug delivery. We report a technically simple fluorescent dye partitioning method to measure alpha in tissue slices based on microfiberoptic detection of dye fluorescence in tissue versus overlying solution. Microfiberoptic tip geometry and dyes were selected for alpha determination from fluorescence intensity ratios, without the need to correct for illumination profile, light scattering/absorption, or dye binding. The method was validated experimentally using cell-embedded gels of specified alpha-values and optical properties. In mouse brain slices, alpha was strongly location-dependent, ranging from 0.16 in thalamus to 0.22 in brainstem, and was sensitive to cell volume changes. Aquaporin-4 water channel gene deletion caused significant extracellular space expansion, with alpha = 0.181 +/- 0.002 in cortex in wild-type mice and 0.211 +/- 0.003 in Aquaporin-4 knockout mice. In slices of LLC1 cell tumors grown in mice to approximately 5 mm diameter, alpha decreased remarkably from approximately 0.45 in superficial tumor to <0.25 in deeper (>100 mum) tumor. Fluorescent dye partitioning with microfiberoptic detection permits rapid, accurate, and anisotropy-insensitive determination of alpha-values in tissue slices.
authors: Department of Medicine and Department of Physiology, University of California, San Francisco, California, USA.
authors: Hua
authors: H
authors: Zhang
authors: A S
authors: AS
authors: Verkman
mesh_terms: D000818:Animals; D051401:Aquaporin 4 / Q000378:metabolism; D001921:Brain / Q000378:metabolism* / Q000473:pathology; D001932:Brain Neoplasms / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D005110:Extracellular Space / Q000378:metabolism*; D005336:Fiber Optic Technology / Q000379:methods*; D005456:Fluorescent Dyes / Q000378:metabolism*; D017353:Gene Deletion; D051379:Mice; D015203:Reproducibility of Results
chemical_list: D051401:Aquaporin 4; D005456:Fluorescent Dyes