Structural Physiology

Functional imaging is a central theme in modern biology and medicine. All biological functions involve a multitude of interactions at the molecular, cellular, and system levels, and it is ultimately desirable to perform molecular and cellular imaging in intact preparations in which the original in vivo functions are preserved. We have been exploring two-photon excitation microscopy with a new type of laser, an infrared femtosecond-pulse laser, as a means to achieve this goal. The two-photon microscope has the ability to penetrate deep into tissues and is the only imaging instrument that allows investigations of intact tissues at the cellular and molecular levels. Two-photon microscopy can also be readily combined with molecular biological and other physiological methods, and it promises to provide important insight into various biological processes in the coming years. Our research interests have two main focuses: (1) the dynamics of synapses in the cerebral cortex and (2) exocytosis in both neurons and secretory cells. We welcome multidisciplinary collaborations to promote our research goals and to help to adapt the new microscopic techniques and lasers to a wide range of biomedical applications.

1. Dynamics of synapses in the cerebral cortex
   We have developed a method to stimulate and control single synapses in the cerebral cortex with the use of two-photon excitation of photoactive glutamate analogs. Our investigations have revealed that the major functions of synapses depend on their structure. We have thus shown that small synapses are ready to learn, enlarging rapidly (within 10 s) after stimulation, whereas large synapses are structurally stable and act as long-term memory traces. These observations suggest that learning, memory, and other activities of the brain are mediated by changes in synaptic structure, and that they can be directly visualized. Moreover, we are now in a position to manipulate synaptic plasticity with a laser beam at the level of the individual synapse　(refs. 1,2,6,9). Such notions and methodology will be further exploited to understand brain functions and disorders.
2. Exocytosis
   Exocytosis is the most essential function of synaptic terminals and secretory cells. Knowledge of the mechanisms of and the ability to control exocytosis artificially have been limited, however. With the use of two-photon excitation–based simultaneous multicolor imaging of various tracers, we have, f or the first time, visualized exocytosis in intact islets of Langerhans, pancreatic acini, the adrenal medulla, and synaptic preparations (refs. 3-5,7,8,10). By further extending our approaches, we aim to develop new methods for imaging and control of secretory functions and their molecular processes in the cerebral cortex and secretory tissues.

1. Iino, Y., Sawada, T., Yamaguchi, Tajiri, M., K., Ishii, S., Kasai, H.^* & Yagishita, S.^* (2020) Dopamine D2 receptors in discrimination learning and spine enlargement. Nature 579: 555-560.
2. Moda-Sava, R.N., Murdock, M.H., et al., Kasai, H. & Liston, C. (2019). Sustained rescue of prefrontal circuit dysfunction by antidepressant-induced postsynaptic spine formation. Science, 364:
3. Hayashi-Takagi, A., Yagishita, S., Nakamura, M. Shirai, F., Wu, Y., Loshbaugh, A.L., Kuhlman, B., Hahn, K.M. and Kasai, H. (2015). Labelling and optical erasure of synaptic memory traces in the motor cortex. Nature (Article), 525:333-338. 
4. Takahashi, N., Sawada, W., Noguchi, J., Watanabe, S., Ucar, H., Hayashi-Takagi, A., Yagishita, S., Ohno, M., Tokumaru, H. & Kasai, H. (2015). Two-photon fluorescence lifetime imaging of primed SNARE complexes in presynaptic terminals and b Nature Communications 6:8531.
5. Yagishita, S., Hayashi-Takagi, A., Ellis-Davies, G.C.R., Urakubo, H., Ishii, S. & Kasai, H. (2014). A critical time window for dopamine action on the structural plasticity of dendritic spines. Science, 345:1616-1620.
6. Hayama, T., Noguchi, J., Watanabe, S., Ellis-Davies, G.C.R., Hayashi, A., Takahashi, N., Matsuzaki, M. & Kasai, H. (2013). GABA promotes the competitive selection of dendritic spines by controlling local Ca²⁺ Nature Neurosci. 16:1409-1416.