To solve the problem that conventional selective activation methods cannot share the scanning path in laser scanning confocal microscopy and is difficult to be coaxial with the imaging path, we design a coaxial scanning real-time light stimulus system. The field-programmable gate array controls the acousto-optic modulator to modulate the light source of the laser scanning confocal microscope. After the light path scanning, the output of stimulus can be synchronized with the imaging scanning. The system uses the peripheral component interconnect express hard core integrated by the field-programmable gate array in Xilinx KC705 development board, and uses the peripheral component interconnect express interface to transmit the stimulus image of the host computer to the field-programmable gate array. The test results show that the proposed system can satisfy the transmission requirement and adjust the light stimulus area effectively.