Time Series Tracking of Cerebral Microvascular Adaptation to Hypoxia and Hyperoxia Imaged with Repeated In Vivo Two-Photon Microscopy.

Abstract

The present study describes methodological aspects of image analysis for angiographic image data with long-term two-photon microscopy acquired for the investigation of dynamic changes in the three-dimensional (3D) network structure of the capillaries (less than 8μm in diameter) in the mouse cerebral cortex. Volume images of the identical capillaries over different periods of days up to 32days were compared for adaptation under either chronic hypoxia (8-9% O2) or hyperoxia (40-50% O2). We observed that the median diameters of measured capillaries were 5.8, 8.4, 9.0, and 8.4μm at 0, 1, 2, and 3weeks during exposure to hypoxia, respectively (N=1, n=2193 pairs at day 0), and 5.4, 5.7, 5.4, 6.0, and 6.1μm measured weekly up to 32days under hyperoxia (N=1, n=1025 pairs at day 0). In accordance with these changes in capillary diameters, tissue space was also observed to change in a depth-dependent manner under hypoxia, but not hyperoxia. The present methods provide us with a method to quantitatively determine three-dimensional vascular and tissue morphology with the aid of a computer-assisted graphical user interface, which facilitates morphometric analysis of the cerebral microvasculature and its correlation with the adaptation of brain cells imaged simultaneously with the microvasculature.