Dual-wavelength high-speed functional photoacoustic microscopy of mouse brain with a Raman laser at 1-MHz A-line rate (Conference Presentation)

Label-free functional photoacoustic microscopy (fPAM) has become a popular technology in small-animal hemodynamic studies. Here we report a stimulated-Raman-scattering-based (SRS) dual-wavelength high-speed fPAM that has achieved volumetric imaging at a 1 MHz A-line rate with capillary-level resolution. Potassium gadolinium tungstate (KGd(WO4)2) crystal is used as a Raman shifter to convert the pump 532 nm picosecond-pulsed laser to the first order Stokes line at 558 nm through the SRS effect with ~40% efficiency and a much narrower line width compared with previous fiber-based SRS PAMs. We also developed a water-immersible micro-electro-mechanical system scanner for scanning a ~4-mm range at a 500 Hz B-scan rate, while maintaining the optic-acoustic confocal alignment. This scanner is assembled entirely from commercially available components, facilitating replication. The detection sensitivity of our fPAM is also improved by employing a high numerical aperture polyvinylidene fluoride ultrasonic transducer, whose acoustic impedance matches better with tissue coupling medium than traditional ceramic transducers. The high sensitivity combined with ~2.4 µm resolution enabled our fPAM to image single red blood cells with a signal-to-noise ratio of ~27 dB. Compared with our previous laser-pulse-width based fPAM, we achieved simultaneous imaging of hemoglobin concentration and oxygenation with a 5-fold increase in imaging speed. Moreover, our system works in a convenient free-space manner compared to previous SRS-based PAMs. We applied it to imaging vasculature and blood oxygen saturation on mouse brains in both resting and stimulated states.