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内容記述 |
Two-photon optogenetics and simultaneous calcium imaging can be used to assess brain neuronal connectivity. This method enables visualization of the response with respect to the activity of the optically stimulated neuron, providing a direct assessment of the connectivity between neurons in the mouse brain in vivo. Resting-state neuronal synchrony, an indirect evaluation method, has been frequently employed to assess brain neuronal connectivity. The utility of two-photon optogenetics-based assessment of neuronal connectivity in pathophysiological research remains to be determined. In this study, we aimed to achieve the following two goals: to examine whether indirect resting-state neuronal synchrony reflects direct neural connectivity revealed by the two-photon optogenetics-based method, and to provide direct evidence for impaired neural connectivity caused by brain disease. In these experiments, C1V1-mScarlet, a type of red-shifted channelrhodopsin, was introduced into GCaMP6s-expressing transgenic mice with an adeno-associated virus. Two-photon optical stimulation (1064 nm) of a single neuron and simultaneous calcium imaging (920 nm) of the target and surrounding cells were performed. Neuronal connectivity was evaluated based on the correlation coefficients between the normalized percent change in the neuronal activity of the target and surrounding cells. The results showed that resting-state synchrony partially approximated two-photon optogenetics-based connectivity but was insufficient for assessing one-to-one connectivity between neurons. Furthermore, comparisons between two-photon optogenetics-based connectivity, resting-state synchrony, and synchrony induced by sensory stimulation suggest that the disagreement is due to inherent neural circuits that affect one-to-one connectivity. Our results showed the advantage of direct assessment using the two-photon optogenetics-based method. We then evaluated two-photon optogenetics-based connectivity using the common carotid artery occlusion (CCAO) model. The mean correlation coefficient in each neuron significantly decreased after CCAO, whereas it remained unchanged in the control group. Additionally, there was no significant decrease in resting-state neuronal synchrony in the CCAO group. These results suggest that the two-photon optogenetics-based method is a sensitive detector of neuronal connectivity impairments in brain diseases. |
