MATRIEX imaging: multiarea two-photon real-time in vivo explorer

Two-photon laser scanning microscopy has been extensively applied to study in vivo neuronal activity at cellular and subcellular resolutions in mammalian brains. However, the extent of such studies is typically confined to a single functional region of the brain. Here, we demonstrate a novel technique, termed the multiarea two-photon real-time in vivo explorer (MATRIEX), that allows the user to target multiple functional brain regions distributed within a zone of up to 12 mm in diameter, each with a field of view (FOV) of ~200 μm in diameter, thus performing two-photon Ca2+ imaging with single-cell resolution in all of the regions simultaneously. For example, we demonstrate real-time functional imaging of single-neuron activities in the primary visual cortex, primary motor cortex and hippocampal CA1 region of mice in both anesthetized and awake states. A unique advantage of the MATRIEX technique is the configuration of multiple microscopic FOVs that are distributed in three-dimensional space over macroscopic distances (>1 mm) both laterally and axially but that are imaged by a single conventional laser scanning device. In particular, the MATRIEX technique can be effectively implemented as an add-on optical module for an existing conventional single-beam-scanning two-photon microscope without requiring any modification to the microscope itself. Thus, the MATRIEX technique can be readily applied to substantially facilitate the exploration of multiarea neuronal activity in vivo for studies of brain-wide neural circuit function with single-cell resolution. The two-photon scanning microscopy imaging technique, commonly applied to study neuronal activity in single regions of mammalian brains, has now been adapted to observe differently functioning regions simultaneously. The innovation, called multiarea two-photon real-time in vivo explorer (MATRIEX), has been developed by Hongbo Jia and colleagues at the Suzhou Institute of Biomedical Engineering and Technology, Chinese Academy of Sciences, with co-workers in other institutions in China and Europe. Any conventional two-photon microscope can be transformed into a MATRIEX microscope by attaching an additional component. The different regions examined simultaneously, each around 200 μm in diameter, can be within a study zone of up to 12 mm in diameter and at depth intervals of more than 1 mm. The technique has been demonstrated by visualizing the transformation of both intraregional and interregional neuronal pairwise activity correlations from the anesthetized state to the awake state in the primary visual cortex, the primary motor cortex and the hippocampal CA1 region of mice. It will facilitate understanding of the functions of three-dimensional brain-wide neural circuits at single-cell resolution in vivo.