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Targeted cell ablation is an important strategy for dissecting the function of individual cells within biological tissues. Here we developed an amplified femtosecond laser-coupled two-photon microscopy (AFL-TPM) system that allows instantaneous and targeted ablation of individual cells and real-time monitoring of neuronal network changes in the living mouse cortex. Through precise and iterative control of the laser power and position, individual cells could be ablated by a single femtosecond light pulse with minimum collateral damage. We further show that ablation of individual somatostatin-expressing interneuron increases the activity of nearby neurons in the primary motor cortex during motor learning. Through precise dendrotomy, we reveal that different dendritic branches of layer 5 pyramidal neurons are structurally and functionally independent. By ablating individual cells and their processes in a spatiotemporally specific manner, the AFL-TPM system could serve as an important means for understanding the functions of cells within the complicated neuronal network.

To understand neural circuit mechanisms underlying behavior, it is crucial to observe the dynamics of neuronal structure and function in different regions of the brain. Since current noninvasive imaging technologies allow cellular-resolution imaging of neurons only within ~1 mm below the cortical surface, the majority of mouse brain tissue remains inaccessible. While miniature optical imaging probes allow access to deep brain regions, cellular-resolution imaging is typically restricted to a small tissue volume. To increase the tissue access volume, we developed a clear optically matched panoramic access channel technique (COMPACT). With probe dimensions comparable to those of common gradient-index lenses, COMPACT enables a two to three orders of magnitude greater tissue access volume. We demonstrated the capabilities of COMPACT by multiregional calcium imaging in mice during sleep. We believe that large-volume in vivo imaging with COMPACT will be valuable to a variety of deep tissue imaging applications.