Essential physiologic processes are also evident in Figure 2. These processes can be quantified and the alteration of these processes can serve as a marker of drug efficacy or toxicity. For instance, the unfiltered large molecular weight probe (green) is retained within the vasculature and effectively defines the vascular space (arrow). Microvascular perfusion rates can be determined by the geometry of red blood cell “shadows,” which form voids in the signal of the large probe coursing through the capillaries.
Integrity of the microvascular endothelial barrier is reflected by the complete retention of the large probe in the vasculature and the absence of leakage into the interstitial space. Furthermore, the absence of signal voids from white blood cells adherent to or rolling along the microvascular wall demonstrates the lack of leukocyte binding to the endothelium or leukocyte activation characteristic of an inflammatory process.
Finally, the nuclear label serves as a monitor for processes such as nuclear fragmentation that would herald apoptosis and cell death. As mentioned above, these quantifiable physiological readouts of organ function, vascular function, inflammation, and cell injury can be used to detect highly localized drug toxicities in a variety of organs that are otherwise undetectable in conventional assays. More importantly, the ability to spatially correlate physiological readouts with cellular and subcellular distributions of drugs gives investigators the ability to detect drug efficacy and/or toxicity at the level of individual cells. This level of resolution can provide drug developers with vital insights into drug actions.
Intravital multiphoton microscopy offers the advantages of both microscopic tissue analysis and in vivo imaging. As with microscopic analysis of histological tissue samples, intravital multiphoton microscopy provides subcellular resolution of multiple parameters. As with other forms of in vivo imaging, intravital multiphoton microscopy is free from fixation artifacts and supports dynamic studies and longitudinal studies of single animals over time. The synergy of these capabilities results in an investigational tool with capabilities that can complement and extend traditional ADMET approaches in drug development.
Timothy Alan Sutton, M.D., Ph.D., is CSO, and Kenneth William Dunn, Ph.D., is vp, scientific development, at INphoton. Web: www.inphoton.com. E-mail: [email protected]