Two-photon laser scanning microscopy as a tool to study cortical vasodynamics under normal and ischemic conditions

This review focuses on the application of two-photon laser scanning microscopy (TPLSM) for in vivo imaging of vascular reactivity, neurovascular communication and as interventional tool for altering vascular and neuronal networks. A mechanistic understanding of hemodynamics requires a systematic analysis of microvascular responses to changes in underlying neuronal activity. This analysis needs to be carried at the single vessel level in the context of the angioarchitecture of vascular networks (Scremin 1995). Moreover, interventional strategies are needed to manipulate specific neuronal and vascular structures to test the role in neurovascular ensemble responses. In this context, recent advances in TPLSM (Denk et al. 1990) allows one to measure the activity of single, visually-identified blood vessels with the potential for simultaneous measurements of neuronal and glial activity using calcium indicators (Stosiek et al. 2003; Ohki et al. 2005; Garaschuk et al. 2006), and targeted occlusion of single blood vessels (Nishimura et al. 2006; Schaffer et al. 2006; Nishimura et al. 2007). While technical aspects of TPLSM have been recently extensively reviewed (Denk et al. 1990; Denk and Svoboda 1997; Svoboda and Yasuda 2006), here we focus on the application of TPLSM to study cortical vascular dynamics under normal conditions and following experimentally-induced ischemic stroke.