Random access two-photon microscope based on acousto-optic deflector

Two-photon microscopy has grown up to be an important technique in biology research, particularly in exploring the neuronal functions of the neurons. With large penetration depth and three-dimensional selectivity, this technique has been able to address the neuro-computing in brain slice or even in live animals. However, its imaging rate is limited by the mechanic scanning mechanism and cannot satisfy the requirement for imaging the encoding pattern of the neuron populations or integrating sites such as the spines. Laser beam steering with acousto-optic deflector (AOD) provides a fast scanning rate, as well as high precision, and high stability due to its inertial-free scanning mechanism. Moreover, 2D AOD scanning allows fast random access to each site of interest, and can thus devote dwell time to pixels of interest and increase both the signal-to-noise ratio and the frame-capture rate. However, scanning femtosecond laser beam with AOD is frustrated by the dispersive nature of the acousto-optic effect and crystal material. This presentation first shows a novel method to solve the problem of dispersion compensation. Based on this dispersion compensated AOD scanner, a random scanning two-photon microscope has been implemented to provide fast and flexible imaging rate with higher signal to noise ratio. A theoretical analysis is presented to explain the evolution of the femtosecond laser pulse in this kind of microscope. Finally, biological experiment is demonstrated to show the potential of recording fast neuronal activities with this technique.