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Light activation and inactivation of neurons by optogenetic techniques has emerged as an important tool for studying neural circuit function and connectivity.To achieve a rapid and precise manipulation of neuronal or circuitry activity at sub-millisecond time resolution, we have designed an acousto-optic device (AOD)-assisted blue-laser stimulation system for exciting neurons with an optogenetic probe light-activated Channelrhdopsin-2 (ChR2) channels.We further evaluated the performance of this system for activating ChR2-expressing neurons in the neocortical slices acutely isolated from the Thy-1-ChR2-YFP transgenic or AAV vector injected mice.Our results demonstrate that the AOD-assisted light activation of ChR2 channels in neurons is capable of efficiently and precisely controlling the rate and temporal sequence of neuronal spikes of different types of neocortical neurons in brain slices, and this system is promising for the studies on the mapping of functional connectivity in a local circuit as well as on the synaptic integration in a single principle cell in brain slices.