Optical Precise Ablation of Targeted Individual Neurons In Vivo.

Abstract

Targeted cell ablation is a powerful strategy for investigating the function of individual neurons within neuronal networks. Multiphoton ablation technology by a tightly focused femtosecond laser, with its significant advantages of noninvasiveness, high efficiency, and single-cell resolution, has been widely used in the study of neuroscience. However, the firing activity of the ablated neuron and its impact on the surrounding neurons and entire neuronal ensembles are still unclear. In this study, we describe the depolarization process of targeted neuron ablation by a femtosecond laser based on a standard two-photon microscope in vitro and in vivo. The photoporation damages the cell membrane, depolarizes the membrane potential, and thus disables the neuron's ability to fire action potentials. The dysfunctional neuron after laser ablation affects both the responses of surrounding neighbors and the functions of ensemble neurons in vivo. Although abnormal Ca²⁺ responses in spatially surrounding neurons are observed, the damage effect is confined to the focal volume. The function of the neuronal ensembles that associate with a specific visual stimulation is not influenced by the ablation of an individual member of the ensemble, indicating the redundancy of the ensemble organization. This study thus provides an insight into the targeted neuron ablation as well as the role of an individual neuron in an ensemble.