Visualizing inhibitory and excitatory synapse dynamics In vivo

Abstract

Structural plasticity is one of the physical manifestations of circuit rewiring in the brain. Once thought to be relegated solely to developmental time periods, we now know that even in the mature brain inhibitory or excitatory connections can be made and broken, modifying the information flow within a circuit by enabling or removing specific information channels. However, the properties of inhibitory and excitatory synapse dynamics are not well understood. To address this issue, we utilized triple-color two photon microscopy to examine inhibitory and excitatory synapses across time with daily imaging. We found that the majority of dynamic spines at these intervals lacked a mature excitatory synapse as indicated by the absence of PSD-95. Inhibitory synapses were also highly dynamic during daily imaging, much more so than expected from previous results imaging at longer intervals, especially those located on spines which also contain an excitatory synapse. Surprisingly, we found that many inhibitory synapses, on the dendritic shaft and on spines, were also repeatedly removed and then reformed again at the same locations on the dendritic arbor. These recurrent inhibitory dynamic events at persistent locations represent a novel role for synapse dynamics, modulating local excitatory activity via their addition or removal. The rate of inhibitory synapse turnover was also modified by experience, as shown through their responses following monocular deprivation. We further sought to investigate these events on even shorter time scales by developing a dual color labeling strategy in combination with a newly developed line scanning temporal focusing two photon microscope, enabling imaging of the entire dendritic arbor and its inhibitory synapses in just a few minutes. This system allows for examination of synapse dynamics on the hourly time scale in vivo and can be expanded to study other molecular events that occur too fast for conventional two photon imaging.