TY - JOUR T1 - <em>In vivo</em> two-photon imaging of dendritic spines in marmoset neocortex JF - eneuro JO - eneuro DO - 10.1523/ENEURO.0019-15.2015 SP - ENEURO.0019-15.2015 AU - Osamu Sadakane AU - Akiya Watakabe AU - Masanari Ohtsuka AU - Masafumi Takaji AU - Tetsuya Sasaki AU - Masatoshi Kasai AU - Tadashi Isa AU - Go Kato AU - Junichi Nabekura AU - Hiroaki Mizukami AU - Keiya Ozawa AU - Hiroshi Kawasaki AU - Tetsuo Yamamori Y1 - 2015/08/27 UR - http://www.eneuro.org/content/early/2015/08/27/ENEURO.0019-15.2015.abstract N2 - Two-photon microscopy in combination with a technique involving the artificial expression of fluorescent protein has enabled the direct observation of dendritic spines in living brains. However, the application of this method to primate brains has been hindered by the lack of appropriate labeling techniques for visualizing dendritic spines. Here, we developed an AAV-vector-based fluorescent protein expression system for visualizing dendritic spines in vivo in the marmoset neocortex. For the clear visualization of each spine, the expression of reporter fluorescent protein should be both sparse and strong. To fulfill these requirements, we amplified fluorescent signals using the transactivator (tTA)-TET responsive element (TRE) system and by titrating down the amount of Thy1S-promoter-driven tTA for sparse expression. By this method, we were able to visualize dendritic spines in the marmoset cortex by two-photon microscopy in vivo and analyze the turnover of spines in the prefrontal cortex. Our results demonstrated that short spines in the marmoset cortex tend to change more frequently than long spines. The comparison of in vivo samples with fixed samples showed that we did not detect all existing spines by our method. Although we found glial cell proliferation, the damage of tissues caused by window construction was relatively small judging from the comparison of spine length between samples with or without window construction. Our new labeling technique for two-photon imaging to visualize in vivo dendritic spines of the marmoset neocortex can be applicable to examining circuit reorganization and synaptic plasticity in primates.Significance Statement: Investigation of non-human primate brains is important for the understanding of the human brain. However, because of technical difficulties, several important methods that have been used in rodent studies are not available for primate studies. Two-photon imaging of dendritic spines has been used in rodent studies, which clarified the basis of neural circuit plasticity, but there has been no report of the application of this imaging method to primate brains. Therefore, in this study, we developed an AAV-vector-based fluorescent protein expression system for use in the studies of the marmoset neocortex. Our approach enabled the sparse yet strong expression of fluorescent protein in neurons. This labeling technique will be applicable to the research of circuit reorganization of primate brains. ER -