Photoacoustic microscopy (PAM) is a promising technique that relies on pulsed optical excitation and ultrasonic detection. Exquisite in vivo images of tissue microvasculature, including individual capillaries, have been obtained with excellent contrast based on optical absorption rather than reflectance. Physiological changes can alter the wavelength dependence of optical absorption, making functional PAM (fPAM) possible with optical excitation at multiple wavelengths. fPAM has received considerable interest for oxygenation studies of tumor microvasculature and monitoring brain activity in small animal models. A major limitation of fPAM is imaging speed. Although single-wavelength images can be acquired quickly, multi-wavelength images require excessively long acquisition times. Therefore, conventional fPAM cannot produce functional images of fast dynamical events (e.g. blood oxygenation in a developing embryonic heart). The slow wavelength tuning of conventional pulsed lasers makes it impossible to obtain video-rate fPAM images. Promising preliminary studies suggest that high speed fPAM is possible with a multicolor optical source based on stimulated Raman scattering. The fundamental hypothesis of this proposal is that wavelength agility can revolutionize the capabilities of fPAM. The PIs propose to develop this compact source to switch wavelengths in less than 1 millisecond, representing a 1000-fold improvement in switching speed over conventional lasers. This wavelength agile source provides the breakthrough necessary to realize video-rate fPAM. They further propose to combine our high speed fPAM system with their existing optical coherence tomography (OCT) system. This combined system will provide high resolution structural and functional images of both microvasculature and surrounding tissue at video frame rates.
