Academic Appointments

Honors & Awards

  • The Award for Young Investigator, Japanese Society for Neuroscience (07/01/2020)
  • The Award for Young Investigator of Japanese Society for Neurochemistry, Japanese Society for Neurochemistry (7/26/2019)
  • Trainee Professional Development Awards, Society for Neuroscience (09/07/2017)

All Publications

  • Genetically encoded calcium indicators to probe complex brain circuit dynamics in vivo. Neuroscience research Inoue, M. n. 2020


    Over the past two decades, genetically encoded calcium indicators (GECIs) have been used extensively to report intracellular calcium (Ca2+) dynamics in order to readout neuronal and network activity in living tissue. Single wavelength GECIs, such as GCaMP, have been widely adapted due to advances in dynamic range, sensitivity, and kinetics. Additionally, recent efforts in protein engineering have expanded the GECI color palette to enable direct optical interrogation of more complex circuit dynamics. Here, I discuss the engineering, application, and future directions of the most recently developed GECIs for in vivo neuroscience research.

    View details for DOI 10.1016/j.neures.2020.05.013

    View details for PubMedID 32531233

  • Comprehensive Dual- and Triple-Feature Intersectional Single-Vector Delivery of Diverse Functional Payloads to Cells of Behaving Mammals. Neuron Fenno, L. E., Ramakrishnan, C. n., Kim, Y. S., Evans, K. E., Lo, M. n., Vesuna, S. n., Inoue, M. n., Cheung, K. Y., Yuen, E. n., Pichamoorthy, N. n., Hong, A. S., Deisseroth, K. n. 2020


    The resolution and dimensionality with which biologists can characterize cell types have expanded dramatically in recent years, and intersectional consideration of such features (e.g., multiple gene expression and anatomical parameters) is increasingly understood to be essential. At the same time, genetically targeted technology for writing in and reading out activity patterns for cells in living organisms has enabled causal investigation in physiology and behavior; however, cell-type-specific delivery of these tools (including microbial opsins for optogenetics and genetically encoded Ca2+ indicators) has thus far fallen short of versatile targeting to cells jointly defined by many individually selected features. Here, we develop a comprehensive intersectional targeting toolbox including 39 novel vectors for joint-feature-targeted delivery of 13 molecular payloads (including opsins, indicators, and fluorophores), systematic approaches for development and optimization of new intersectional tools, hardware for in vivo monitoring of expression dynamics, and the first versatile single-virus tools (Triplesect) that enable targeting of triply defined cell types.

    View details for DOI 10.1016/j.neuron.2020.06.003

    View details for PubMedID 32574559

  • Rational Engineering of XCaMPs, a Multicolor GECI Suite for In Vivo Imaging of Complex Brain Circuit Dynamics CELL Inoue, M., Takeuchi, A., Manita, S., Horigane, S., Sakamoto, M., Kawakami, R., Yamaguchi, K., Otomo, K., Yokoyama, H., Kim, R., Yokoyama, T., Takemoto-Kimura, S., Abe, M., Okamura, M., Kondo, Y., Quirin, S., Ramakrishnan, C., Imamura, T., Sakimura, K., Nemoto, T., Kano, M., Fujii, H., Deisseroth, K., Kitamura, K., Bito, H. 2019; 177 (5): 1346-+
  • Cortical layer-specific critical dynamics triggering perception. Science (New York, N.Y.) Marshel, J. H., Kim, Y. S., Machado, T. A., Quirin, S. n., Benson, B. n., Kadmon, J. n., Raja, C. n., Chibukhchyan, A. n., Ramakrishnan, C. n., Inoue, M. n., Shane, J. C., McKnight, D. J., Yoshizawa, S. n., Kato, H. E., Ganguli, S. n., Deisseroth, K. n. 2019


    Perceptual experiences may arise from neuronal activity patterns in mammalian neocortex. We probed mouse neocortex during visual discrimination using a red-shifted channelrhodopsin (ChRmine, discovered through structure-guided genome mining) alongside multiplexed multiphoton-holography (MultiSLM), achieving control of individually-specified neurons spanning large cortical volumes with millisecond precision. Stimulating a critical number of stimulus-orientation-selective neurons drove widespread recruitment of functionally-related neurons, a process enhanced by (but not requiring) orientation-discrimination task learning. Optogenetic targeting of orientation-selective ensembles elicited correct behavioral discrimination. Cortical layer specific-dynamics were apparent, as emergent neuronal activity asymmetrically propagated from layer-2/3 to layer-5, and smaller layer-5 ensembles were as effective as larger layer-2/3 ensembles in eliciting orientation discrimination behavior. Population dynamics emerging after optogenetic stimulation both correctly predicted behavior and resembled natural neural representations of visual stimuli.

    View details for DOI 10.1126/science.aaw5202

    View details for PubMedID 31320556

  • Kilohertz two-photon brain imaging in awake mice. Nature methods Zhang, T. n., Hernandez, O. n., Chrapkiewicz, R. n., Shai, A. n., Wagner, M. J., Zhang, Y. n., Wu, C. H., Li, J. Z., Inoue, M. n., Gong, Y. n., Ahanonu, B. n., Zeng, H. n., Bito, H. n., Schnitzer, M. J. 2019


    Two-photon microscopy is a mainstay technique for imaging in scattering media and normally provides frame-acquisition rates of ~10-30 Hz. To track high-speed phenomena, we created a two-photon microscope with 400 illumination beams that collectively sample 95,000-211,000 µm2 areas at rates up to 1 kHz. Using this microscope, we visualized microcirculatory flow, fast venous constrictions and neuronal Ca2+ spiking with millisecond-scale timing resolution in the brains of awake mice.

    View details for DOI 10.1038/s41592-019-0597-2

    View details for PubMedID 31659327

  • Functional emergence of a column-like architecture in layer 5 of mouse somatosensory cortex in vivo. The journal of physiological sciences : JPS Koizumi, K. n., Inoue, M. n., Chowdhury, S. n., Bito, H. n., Yamanaka, A. n., Ishizuka, T. n., Yawo, H. n. 2019; 69 (1): 65–77


    To investigate how the functional architecture is organized in layer 5 (L5) of the somatosensory cortex of a mouse in vivo, the input-output relationship was investigated using an all-optical approach. The neural activity in L5 was optically recorded using a Ca2+ sensor, R-CaMP2, through a microprism inserted in the cortex under two-photon microscopy, while the L5 was regionally excited using optogenetics. The excitability was spread around the blue-light irradiated region, but the horizontal propagation was limited to within a certain distance (λ < 130 μm from the center of the illumination spot). When two regions were photostimulated with a short interval, the excitability of each cluster was reduced. Therefore, a column-like architecture had functionally emerged with reciprocal inhibition through a minimal number of synaptic relays. This could generate a synchronous output from a region of L5 with simultaneous enhancement of the signal-to-noise ratio by silencing of the neighboring regions.

    View details for DOI 10.1007/s12576-018-0618-4

    View details for PubMedID 29761270

  • Rational Engineering of XCaMPs, a Multicolor GECI Suite for In Vivo Imaging of Complex Brain Circuit Dynamics. Cell Inoue, M. n., Takeuchi, A. n., Manita, S. n., Horigane, S. I., Sakamoto, M. n., Kawakami, R. n., Yamaguchi, K. n., Otomo, K. n., Yokoyama, H. n., Kim, R. n., Yokoyama, T. n., Takemoto-Kimura, S. n., Abe, M. n., Okamura, M. n., Kondo, Y. n., Quirin, S. n., Ramakrishnan, C. n., Imamura, T. n., Sakimura, K. n., Nemoto, T. n., Kano, M. n., Fujii, H. n., Deisseroth, K. n., Kitamura, K. n., Bito, H. n. 2019


    To decipher dynamic brain information processing, current genetically encoded calcium indicators (GECIs) are limited in single action potential (AP) detection speed, combinatorial spectral compatibility, and two-photon imaging depth. To address this, here, we rationally engineered a next-generation quadricolor GECI suite, XCaMPs. Single AP detection was achieved within 3-10 ms of spike onset, enabling measurements of fast-spike trains in parvalbumin (PV)-positive interneurons in the barrel cortex in vivo and recording three distinct (two inhibitory and one excitatory) ensembles during pre-motion activity in freely moving mice. In vivo paired recording of pre- and postsynaptic firing revealed spatiotemporal constraints of dendritic inhibition in layer 1 in vivo, between axons of somatostatin (SST)-positive interneurons and apical tufts dendrites of excitatory pyramidal neurons. Finally, non-invasive, subcortical imaging using red XCaMP-R uncovered somatosensation-evoked persistent activity in hippocampal CA1 neurons. Thus, the XCaMPs offer a critical enhancement of solution space in studies of complex neuronal circuit dynamics. VIDEO ABSTRACT.

    View details for PubMedID 31080068

  • Modulation of prefrontal cortex excitation/inhibition balance rescues social behavior in CNTNAP2-deficient mice. Science translational medicine Selimbeyoglu, A. n., Kim, C. K., Inoue, M. n., Lee, S. Y., Hong, A. S., Kauvar, I. n., Ramakrishnan, C. n., Fenno, L. E., Davidson, T. J., Wright, M. n., Deisseroth, K. n. 2017; 9 (401)


    Alterations in the balance between neuronal excitation and inhibition (E:I balance) have been implicated in the neural circuit activity-based processes that contribute to autism phenotypes. We investigated whether acutely reducing E:I balance in mouse brain could correct deficits in social behavior. We used mice lacking the CNTNAP2 gene, which has been implicated in autism, and achieved a temporally precise reduction in E:I balance in the medial prefrontal cortex (mPFC) either by optogenetically increasing the excitability of inhibitory parvalbumin (PV) neurons or decreasing the excitability of excitatory pyramidal neurons. Surprisingly, both of these distinct, real-time, and reversible optogenetic modulations acutely rescued deficits in social behavior and hyperactivity in adult mice lacking CNTNAP2 Using fiber photometry, we discovered that native mPFC PV neuronal activity differed between CNTNAP2 knockout and wild-type mice. During social interactions with other mice, PV neuron activity increased in wild-type mice compared to interactions with a novel object, whereas this difference was not observed in CNTNAP2 knockout mice. Together, these results suggest that real-time modulation of E:I balance in the mouse prefrontal cortex can rescue social behavior deficits reminiscent of autism phenotypes.

    View details for PubMedID 28768803

  • Simultaneous fast measurement of circuit dynamics at multiple sites across the mammalian brain. Nature methods Kim, C. K., Yang, S. J., Pichamoorthy, N., Young, N. P., Kauvar, I., Jennings, J. H., Lerner, T. N., Berndt, A., Lee, S. Y., Ramakrishnan, C., Davidson, T. J., Inoue, M., Bito, H., Deisseroth, K. 2016; 13 (4): 325-328


    Real-time activity measurements from multiple specific cell populations and projections are likely to be important for understanding the brain as a dynamical system. Here we developed frame-projected independent-fiber photometry (FIP), which we used to record fluorescence activity signals from many brain regions simultaneously in freely behaving mice. We explored the versatility of the FIP microscope by quantifying real-time activity relationships among many brain regions during social behavior, simultaneously recording activity along multiple axonal pathways during sensory experience, performing simultaneous two-color activity recording, and applying optical perturbation tuned to elicit dynamics that match naturally occurring patterns observed during behavior.

    View details for DOI 10.1038/nmeth.3770

    View details for PubMedID 26878381

  • Rational design of a high-affinity, fast, red calcium indicator R-CaMP2 NATURE METHODS Inoue, M., Takeuchi, A., Horigane, S., Ohkura, M., Gengyo-Ando, K., Fujii, H., Kamijo, S., Takemoto-Kimura, S., Kano, M., Nakai, J., Kitamura, K., Bito, H. 2015; 12 (1): 64-70


    Fluorescent Ca(2+) reporters are widely used as readouts of neuronal activities. Here we designed R-CaMP2, a high-affinity red genetically encoded calcium indicator (GECI) with a Hill coefficient near 1. Use of the calmodulin-binding sequence of CaMKK-α and CaMKK-β in lieu of an M13 sequence resulted in threefold faster rise and decay times of Ca(2+) transients than R-CaMP1.07. These features allowed resolving single action potentials (APs) and recording fast AP trains up to 20-40 Hz in cortical slices. Somatic and synaptic activities of a cortical neuronal ensemble in vivo were imaged with similar efficacy as with previously reported sensitive green GECIs. Combining green and red GECIs, we successfully achieved dual-color monitoring of neuronal activities of distinct cell types, both in the mouse cortex and in freely moving Caenorhabditis elegans. Dual imaging using R-CaMP2 and green GECIs provides a powerful means to interrogate orthogonal and hierarchical neuronal ensembles in vivo.

    View details for DOI 10.1038/NMETH.3185

    View details for Web of Science ID 000347668600021

    View details for PubMedID 25419959