Minimally invasive, wide-field two-photon imaging of the brainstem at cellular resolution

Masakazu Agetsuma; Azumi Hatakeyama; Daisuke Yamada; Hiroshi Kuniishi; Chihiro Ito; Eri Takeuchi; Shinji Tsuji; Motosuke Tsutsumi; Takako Ichiki; Kohei Otomo; Toshinori Yoshioka; Tomoko Kobayashi; Atsushi Noritake; Yoshitsugu Aoki; Tomomi Nemoto; Hiroshi Yukawa; Akiyoshi Saitoh; Junichi Nabekura; Masayuki Sekiguchi

doi:10.1016/j.crmeth.2025.101010

Minimally invasive, wide-field two-photon imaging of the brainstem at cellular resolution

Masakazu Agetsuma^*, Azumi Hatakeyama, Daisuke Yamada^*, Hiroshi Kuniishi, Chihiro Ito, Eri Takeuchi, Shinji Tsuji, Motosuke Tsutsumi, Takako Ichiki, Kohei Otomo, Toshinori Yoshioka, Tomoko Kobayashi, Atsushi Noritake, Yoshitsugu Aoki, Tomomi Nemoto, Hiroshi Yukawa, Akiyoshi Saitoh, Junichi Nabekura, Masayuki Sekiguchi

^*Corresponding author for this work

Integrative Neuroscience

Research output: Contribution to journal › Article › peer-review

Abstract

Brain-viscera communication is crucial for regulating mental health, with the vagus nerve being a key structure mediating this interaction. Clinically, artificial vagus nerve stimulation (VNS) is used to treat various neuropsychiatric disorders, highlighting the importance of vagal afferent fibers in emotion regulation. The nucleus tractus solitarii (NTS) is a brainstem structure proposed to receive signals from vagal afferents and relay them to brain networks for emotion regulation. However, due to the anatomical complexity and difficulty in accessing the deep-brain NTS region in vivo, its underlying mechanisms remain unclear. Here, we developed a wide-field and deep-brain two-photon imaging method using a double-prism optical interface. This approach enables cellular-resolution imaging to specifically detect NTS neural activity while largely preserving the overlying cerebellum, a region also implicated in emotion regulation. We evaluated NTS neuronal responses to VNS and a gastrointestinal hormone, demonstrating the method's utility for investigating the vagus-NTS pathway in vivo.

Original language	English
Article number	101010
Journal	Cell Reports Methods
Volume	5
Issue number	4
DOIs	https://doi.org/10.1016/j.crmeth.2025.101010
State	Published - 2025/04/21

Keywords

CP: Imaging
CP: Neuroscience
brain-viscera communication
brainstem
emotion
in vivo two-photon calcium imaging
nucleus tractus solitarii
vagus nerve stimulation

ASJC Scopus subject areas

Biotechnology
Biochemistry
Biochemistry, Genetics and Molecular Biology (miscellaneous)
Genetics
Radiology Nuclear Medicine and imaging
Computer Science Applications

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

Access to Document

10.1016/j.crmeth.2025.101010

Cite this

Agetsuma, M., Hatakeyama, A., Yamada, D., Kuniishi, H., Ito, C., Takeuchi, E., Tsuji, S., Tsutsumi, M., Ichiki, T., Otomo, K., Yoshioka, T., Kobayashi, T., Noritake, A., Aoki, Y., Nemoto, T., Yukawa, H., Saitoh, A., Nabekura, J., & Sekiguchi, M. (2025). Minimally invasive, wide-field two-photon imaging of the brainstem at cellular resolution. Cell Reports Methods, 5(4), Article 101010. https://doi.org/10.1016/j.crmeth.2025.101010

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title = "Minimally invasive, wide-field two-photon imaging of the brainstem at cellular resolution",

abstract = "Brain-viscera communication is crucial for regulating mental health, with the vagus nerve being a key structure mediating this interaction. Clinically, artificial vagus nerve stimulation (VNS) is used to treat various neuropsychiatric disorders, highlighting the importance of vagal afferent fibers in emotion regulation. The nucleus tractus solitarii (NTS) is a brainstem structure proposed to receive signals from vagal afferents and relay them to brain networks for emotion regulation. However, due to the anatomical complexity and difficulty in accessing the deep-brain NTS region in vivo, its underlying mechanisms remain unclear. Here, we developed a wide-field and deep-brain two-photon imaging method using a double-prism optical interface. This approach enables cellular-resolution imaging to specifically detect NTS neural activity while largely preserving the overlying cerebellum, a region also implicated in emotion regulation. We evaluated NTS neuronal responses to VNS and a gastrointestinal hormone, demonstrating the method's utility for investigating the vagus-NTS pathway in vivo.",

keywords = "CP: Imaging, CP: Neuroscience, brain-viscera communication, brainstem, emotion, in vivo two-photon calcium imaging, nucleus tractus solitarii, vagus nerve stimulation",

author = "Masakazu Agetsuma and Azumi Hatakeyama and Daisuke Yamada and Hiroshi Kuniishi and Chihiro Ito and Eri Takeuchi and Shinji Tsuji and Motosuke Tsutsumi and Takako Ichiki and Kohei Otomo and Toshinori Yoshioka and Tomoko Kobayashi and Atsushi Noritake and Yoshitsugu Aoki and Tomomi Nemoto and Hiroshi Yukawa and Akiyoshi Saitoh and Junichi Nabekura and Masayuki Sekiguchi",

note = "Publisher Copyright: {\textcopyright} 2025 The Author(s)",

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Agetsuma, M, Hatakeyama, A, Yamada, D, Kuniishi, H, Ito, C, Takeuchi, E, Tsuji, S, Tsutsumi, M, Ichiki, T, Otomo, K, Yoshioka, T, Kobayashi, T, Noritake, A, Aoki, Y, Nemoto, T, Yukawa, H, Saitoh, A, Nabekura, J & Sekiguchi, M 2025, 'Minimally invasive, wide-field two-photon imaging of the brainstem at cellular resolution', Cell Reports Methods, vol. 5, no. 4, 101010. https://doi.org/10.1016/j.crmeth.2025.101010

TY - JOUR

T1 - Minimally invasive, wide-field two-photon imaging of the brainstem at cellular resolution

AU - Agetsuma, Masakazu

AU - Hatakeyama, Azumi

AU - Yamada, Daisuke

AU - Kuniishi, Hiroshi

AU - Ito, Chihiro

AU - Takeuchi, Eri

AU - Tsuji, Shinji

AU - Tsutsumi, Motosuke

AU - Ichiki, Takako

AU - Otomo, Kohei

AU - Yoshioka, Toshinori

AU - Kobayashi, Tomoko

AU - Noritake, Atsushi

AU - Aoki, Yoshitsugu

AU - Nemoto, Tomomi

AU - Yukawa, Hiroshi

AU - Saitoh, Akiyoshi

AU - Nabekura, Junichi

AU - Sekiguchi, Masayuki

PY - 2025/4/21

Y1 - 2025/4/21

N2 - Brain-viscera communication is crucial for regulating mental health, with the vagus nerve being a key structure mediating this interaction. Clinically, artificial vagus nerve stimulation (VNS) is used to treat various neuropsychiatric disorders, highlighting the importance of vagal afferent fibers in emotion regulation. The nucleus tractus solitarii (NTS) is a brainstem structure proposed to receive signals from vagal afferents and relay them to brain networks for emotion regulation. However, due to the anatomical complexity and difficulty in accessing the deep-brain NTS region in vivo, its underlying mechanisms remain unclear. Here, we developed a wide-field and deep-brain two-photon imaging method using a double-prism optical interface. This approach enables cellular-resolution imaging to specifically detect NTS neural activity while largely preserving the overlying cerebellum, a region also implicated in emotion regulation. We evaluated NTS neuronal responses to VNS and a gastrointestinal hormone, demonstrating the method's utility for investigating the vagus-NTS pathway in vivo.

AB - Brain-viscera communication is crucial for regulating mental health, with the vagus nerve being a key structure mediating this interaction. Clinically, artificial vagus nerve stimulation (VNS) is used to treat various neuropsychiatric disorders, highlighting the importance of vagal afferent fibers in emotion regulation. The nucleus tractus solitarii (NTS) is a brainstem structure proposed to receive signals from vagal afferents and relay them to brain networks for emotion regulation. However, due to the anatomical complexity and difficulty in accessing the deep-brain NTS region in vivo, its underlying mechanisms remain unclear. Here, we developed a wide-field and deep-brain two-photon imaging method using a double-prism optical interface. This approach enables cellular-resolution imaging to specifically detect NTS neural activity while largely preserving the overlying cerebellum, a region also implicated in emotion regulation. We evaluated NTS neuronal responses to VNS and a gastrointestinal hormone, demonstrating the method's utility for investigating the vagus-NTS pathway in vivo.

KW - CP: Imaging

KW - CP: Neuroscience

KW - brain-viscera communication

KW - brainstem

KW - emotion

KW - in vivo two-photon calcium imaging

KW - nucleus tractus solitarii

KW - vagus nerve stimulation

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U2 - 10.1016/j.crmeth.2025.101010

DO - 10.1016/j.crmeth.2025.101010

M3 - 学術論文

C2 - 40187350

AN - SCOPUS:105001822426

SN - 2667-2375

VL - 5

JO - Cell Reports Methods

JF - Cell Reports Methods

IS - 4

M1 - 101010

ER -