Molecular mechanism regulating expression of estrous behavior in ruminants

• Project/Area Number: 16H05014
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Animal production science
• Research Institution: Nagoya University
• Principal Investigator: Ohkura Satoshi 名古屋大学, 生命農学研究科, 教授 (20263163)
• Project Period (FY): 2016-04-01 – 2020-03-31
• Project Status: Completed (Fiscal Year 2020)
• Budget Amount: ¥17,420,000 (Direct Cost: ¥13,400,000、Indirect Cost: ¥4,020,000); Fiscal Year 2019: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000); Fiscal Year 2018: ¥2,600,000 (Direct Cost: ¥2,000,000、Indirect Cost: ¥600,000); Fiscal Year 2017: ¥3,120,000 (Direct Cost: ¥2,400,000、Indirect Cost: ¥720,000); Fiscal Year 2016: ¥9,490,000 (Direct Cost: ¥7,300,000、Indirect Cost: ¥2,190,000)
• Keywords: 応用動物 / 行動学 / 獣医学 / 畜産学 / 神経科学 / ヤギ / エストロジェン / 発情行動

Outline of Final Research Achievements

The present study aimed to elucidate the neural mechanisms controlling estrus behavior in ruminants using genetically modified goats that can conditionally knock out estrogen receptor (ER) expression, which plays a primary role in inducing estrus behavior. First, we constructed a model that induced estrus behavior by hormone treatment and found that the increase in the behavioral activity associated with estrus was a quantitative parameter to evaluate the estrus behavior in goats. Second, we identified the nucleotide sequence and deduced amino acid sequence of the goat ER gene mRNA, and optimized the goat genetic engineering technology to implement genome editing technology by “gene editing by electroporation of Cas9 protein (GEEP)” method in 1-cell stage embryo. Third, we developed a method to implant a microinjection cannula stereotaxically in the brain region where goat ER is highly expressed.

Academic Significance and Societal Importance of the Research Achievements

本研究において確立した、発情に伴う行動量増加を指標とした発情行動の定量的評価系は、発情行動発現のタイミングを正確に測ることが可能となる点で、反芻動物の発情行動を制御する神経機構の解明に大きく寄与する実験モデル系となる。また、本研究により確立したヤギの遺伝子工学基盤（過排卵処置、卵管内受精卵（1細胞期胚）採取法、GEEP法によるゲノム編集胚の作出など）は、ヤギをモデルとしたゲノム編集技術の確立に寄与し、遺伝子改変技術の対象動物としてのヤギの有用性を飛躍的に高めることができる。

Research Products

• [Int'l Joint Research] フランス国立農業研究所(フランス)
• [Journal Article] Estrogen increases <i>KISS1</i> expression in newly generated immortalized <i>KISS1</i>-expressing cell line derived from goat preoptic area (2021)
  • Author(s): OSHIMO Yukina、MUNETOMO Arisa、MAGATA Fumie、SUETOMI Yuta、SONODA Shuhei、TAKEUCHI Yukari、TSUKAMURA Hiroko、OHKURA Satoshi、MATSUDA Fuko
  • Journal Title: Journal of Reproduction and Development Volume: 67 Issue: 1 Pages: 15-23
  • DOI: 10.1262/jrd.2020-053
  • NAID: 130007986318
  • ISSN: 0916-8818, 1348-4400
  • Peer Reviewed / Open Access
• [Journal Article] Establishment of immortalised cell lines derived from female Shiba goat KNDy and GnRH neurones. (2020)
  • Author(s): Suetomi Y, Tatebayashi R, Sonoda S, Munetomo A, Matsuyama S, Inoue N, Uenoyama Y, Takeuchi Y,Tsukamura H, Ohkura S, Matsuda F.
  • Journal Title: Neuroendocrinology Volume: 32(6) Issue: 6 Pages: 12857-12857
  • DOI: 10.1111/jne.12857
  • Peer Reviewed
• [Journal Article] Facilitatory and inhibitory role of central amylin administration in the regulation of the gonadotropin-releasing hormone pulse generator activity in goats (2020)
  • Author(s): Kitagawa, Y., Sasaki, T., Suzumura, R., Morishima, A., Tatebayashi, R., Assadullah, Ieda, N., Morita, Y., Matsuyama, S., Inoue, N., Uenoyama, Y. Tsukamura, H. and Ohkura. S.
  • Journal Title: Neuroscience Letters Volume: 736 Pages: 135276-135276
  • DOI: 10.1016/j.neulet.2020.135276
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Peripheral administration of SB223412, a selective neurokinin-3 receptor antagonist, suppresses pulsatile luteinizing hormone secretion by acting on the gonadotropin-releasing hormone pulse generator in estrogen-treated ovariectomized female goats. (2020)
  • Author(s): Sasaki T, Sonoda T, Tatebayashi R, Kitagawa Y, Oishi S, Yamamoto K, Fujii N, Inoue N, Uenoyama Y, Tsukamura H, Maeda KI, Matsuda F, Morita Y, Matsuyama S, Ohkura S.
  • Journal Title: J Reprod Dev Volume: - Issue: 4 Pages: 351-357
  • DOI: 10.1262/jrd.2019-145
  • NAID: 130007888728
  • Peer Reviewed / Open Access
• [Journal Article] Role of kisspeptin neurons as a GnRH surge generator: Comparative aspects in rodents and non‐rodent mammals (2019)
  • Author(s): Matsuda Fuko、Ohkura Satoshi、Magata Fumie、Munetomo Arisa、Chen Jing、Sato Marimo、Inoue Naoko、Uenoyama Yoshihisa、Tsukamura Hiroko
  • Journal Title: Journal of Obstetrics and Gynaecology Research Volume: 45 Issue: 12 Pages: 2318-2329
  • DOI: 10.1111/jog.14124
  • Peer Reviewed
• [Journal Article] Peripheral administration of κ-opioid receptor antagonist stimulates gonadotropin-releasing hormone pulse generator activity in ovariectomized, estrogen-treated female goats (2019)
  • Author(s): Sasaki T.、Ito D.、Sonoda T.、Morita Y.、Wakabayashi Y.、Yamamura T.、Okamura H.、Oishi S.、Noguchi T.、Fujii N.、Uenoyama Y.、Tsukamura H.、Maeda K.I.、Matsuda F.、Ohkura S.
  • Journal Title: Domestic Animal Endocrinology Volume: 68 Pages: 83-91
  • DOI: 10.1016/j.domaniend.2018.12.011
  • Peer Reviewed / Open Access
• [Journal Article] A neurokinin 3 receptor-selective agonist accelerates pulsatile luteinizing hormone secretion in lactating cattle† (2017)
  • Author(s): Nakamura Sho、Wakabayashi Yoshihiro、Yamamura Takashi、Ohkura Satoshi、Matsuyama Shuichi
  • Journal Title: Biology of Reproduction Volume: 97 Issue: 1 Pages: 81-90
  • DOI: 10.1093/biolre/iox068
  • Peer Reviewed
• [Journal Article] The two populations of kisspeptin neurons are involved in the ram-induced LH pulsatile secretion and LH surge in anestrous ewes (2017)
  • Author(s): Fabre-Nys Claude、Cognie Juliette、Dufourny Laurence、Ghenim Meriem、Martinet Stephanie、Lasserre Olivier、Lomet Didier、Millar Robert P、Ohkura Satoshi、Suetomi Yuta
  • Journal Title: Endocrinology Volume: 158 Issue: 11 Pages: 3914-3928
  • DOI: 10.1210/en.2017-00429
  • Peer Reviewed
• [Journal Article] Immunohistochemical characterization of the arcuate kisspeptin/neurokinin B/dynorphin (KNDy) and preoptic kisspeptin neuronal populations in the hypothalamus during the estrous cycle in heifers. (2016)
  • Author(s): Hassaneen, A.S.A., Naniwa, Y., Suetomi, Y., Matsuyama, S., Kimura, K., Ieda, N., Inoue, N., Uenoyama, Y., Tsukamura, H., Maeda, K.-I., Matsuda, F. and Ohkura, S.
  • Journal Title: Journal of Reproduction and Development Volume: 62 Issue: 5 Pages: 471-477
  • DOI: 10.1262/jrd.2016-075
  • NAID: 130005431286
  • Peer Reviewed / Open Access
• [Journal Article] Molecular and Epigenetic Mechanism Regulating Hypothalamic Kiss1 Gene Expression in Mammals. (2016)
  • Author(s): Uenoyama Y, Tomikawa J, Inoue N, Goto T, Minabe S, Ieda N, Nakamura S, Watanabe Y, Ikegami K, Matsuda F, Ohkura S, Maeda KI, Tsukamura H.
  • Journal Title: Neuroendocrinology Volume: 印刷中 Issue: 6 Pages: 640-649
  • DOI: 10.1159/000445207
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Presentation] シバヤギにおけるGnRHパルス発生中枢の制御に関わるセロトニン受容体サブタイプの検討 (2020)
  • Author(s): 鈴村玲香，森田康広，松山秀一，佐々木拓弥，北川悠梨，井上直子，上野山賀久，束村博子，大蔵 聡
  • Organizer: 第113回日本繁殖生物学会大会
• [Presentation] 雌ヤギ由来KNDyニューロンおよびGnRHニューロン不死化細胞株の樹立 (2020)
  • Author(s): 松田二子，末富祐太，舘林亮輝，園田修平，棟朝亜理紗，松山秀一，井上直子，上野山賀久，武内ゆかり，束村博子，大蔵 聡
  • Organizer: 第25回日本生殖内分泌学会学術集会
• [Presentation] GnRHパルス発生中枢制御機構におけるカルシトニン受容体の役割 (2019)
  • Author(s): 北川悠梨，佐々木拓弥，森島 愛，舘林亮輝，森田康広，松山秀一，井上直子，上野山賀久，束村博子，大蔵 聡
  • Organizer: 第37回内分泌代謝学サマーセミナー
• [Presentation] 生殖内分泌の基礎 －GnRH分泌制御の神経内分泌メカニズム－ (2019)
  • Author(s): 大蔵 聡
  • Organizer: 第1 回日本生殖発生毒性フォーラム
  • Invited
• [Presentation] Neuroendocrine control of reproductive functions in domestic ruminants (2019)
  • Author(s): Ohkura, S.
  • Organizer: The 2nd International Conference on Mathematics and Natural Sciences (IConMNS) 2019
  • Int'l Joint Research / Invited
• [Presentation] パルス状GnRH分泌制御機構におけるカルシトニン受容体の役割 (2019)
  • Author(s): 北川悠梨，佐々木拓弥，森島 愛，舘林亮輝，森田康広，松山秀一，井上直子，上野山賀久，束村博子，大蔵 聡
  • Organizer: 第112回日本繁殖生物学会大会
• [Presentation] シバヤギのパルス状GnRH分泌制御メカニズムにおけるセロトニンの役割 (2019)
  • Author(s): 佐々木拓弥，森島 愛，中西真莉菜，鈴村玲香，舘林亮輝，北川悠梨，森田康広，松山秀一，井上直子，上野山賀久，束村博子，大蔵 聡
  • Organizer: 第112回日本繁殖生物学会大会
• [Presentation] ヤギ視索前野由来キスペプチンニューロン不死化細胞株の樹立 (2019)
  • Author(s): 大下雪奈，棟朝亜理紗，末富祐太，真方文絵，束村博子，大蔵 聡，松田二子
  • Organizer: 第112回日本繁殖生物学会大会
• [Presentation] 家畜における繁殖の神経内分泌学－基礎的知見と臨床応用の可能性－ (2019)
  • Author(s): 松田二子，大蔵 聡
  • Organizer: （公社）日本畜産学会主催日本家畜臨床学会共催公開シンポジウム「畜産研究の成果を獣医臨床フィールドへ」
  • Invited
• [Presentation] The role of calcitonin receptor signaling in the regulation of GnRH pulse generator activity in goats (2019)
  • Author(s): Kitagawa, Y., Sasaki, T., Morishima, A., Tatebayashi, R., Morita, Y., Matsuyama, S., Inoue, N., Uenoyama, Y., Tsukamura, H., Ohkura, S.
  • Organizer: 日本畜産学会第125回大会
• [Presentation] シバヤギのGnRH パルス発生メカニズムにおけるセロトニンの役割 (2018)
  • Author(s): 森島 愛，佐々木拓弥，舘林亮輝，北川悠梨，森田康広，松山秀一，井上直子，上野山賀久，束村博子，大蔵 聡
  • Organizer: 日本下垂体研究会第33回学術集会
• [Presentation] シバヤギにおけるパルス状GnRH 分泌調節メカニズムに対するアミリンの作用 (2018)
  • Author(s): 北川悠梨，佐々木拓弥，森島 愛，舘林亮輝，森田康広，松山秀一，井上直子，上野山賀久，束村博子，大蔵 聡
  • Organizer: 日本下垂体研究会第33回学術集会
• [Presentation] ニューロキニンB受容体拮抗剤の経口投与はパルス状LH分泌を抑制する (2018)
  • Author(s): 佐々木拓弥，園田朋也，大石真也，藤井信孝，森田康広，松山秀一，井上直子，上野山賀久，束村博子，前多敬一郎，松田二子，大蔵 聡
  • Organizer: 第111回日本繁殖生物学会大会
• [Presentation] 私が出会った生殖神経内分泌学－農学分野における基礎研究の意義－ (2018)
  • Author(s): 大蔵 聡
  • Organizer: 第43回日本比較内分泌学会
  • Invited
• [Presentation] シバヤギにおける発情に伴う行動量の増加とLHサージの時間的関係 (2017)
  • Author(s): 今岡麻子・末富祐太・佐々木拓弥・大蔵 聡
  • Organizer: 日本畜産学会第122回大会
  • Place of Presentation: 兵庫県神戸市
  • Year and Date: 2017-03-28
• [Presentation] Microimplants of estradiol in the medial preoptic area induce a surge-like secretion of luteinizing hormone in male Shiba goats. (2017)
  • Author(s): Watanabe, M., Sasaki, T., Tatebayashi, R., Suetomi, Y. and Ohkura, S.
  • Organizer: The 4th World Congress on Reproductive Biology
  • Int'l Joint Research
• [Presentation] TALEN-mediated Cre recombinase knockin in KISS1 locus of goat embryonic fibroblasts. (2017)
  • Author(s): Tatebayashi, R., Sakuma, T., Yamamoto, T., Ohkura, S. and Matsuda, F.
  • Organizer: The 4th World Congress on Reproductive Biology
  • Int'l Joint Research
• [Presentation] A neurokinin 3 receptor-selective agonist, senktide, stimulates pulsatile luteinizing hormone secretion in lactating cattle. (2017)
  • Author(s): Nakamura, S., Wakabayashi, Y., Yamamura, T., Ohkura, S. and Matsuyama, S.
  • Organizer: The 4th World Congress on Reproductive Biology
  • Int'l Joint Research
• [Presentation] Neurokinin B/neurokinin-3 receptor signaling is involved in the regulation of GnRH pulse generation in male goats. (2017)
  • Author(s): Yamamura, T, Ohkura, S. and Wakabayashi, Y.
  • Organizer: The 4th World Congress on Reproductive Biology
  • Int'l Joint Research
• [Presentation] Establishment of immortalized GnRH neuronal cell lines derived from the cattle brain. (2017)
  • Author(s): Ozaki, R., Uenoyama, Y., Suetomi, Y., Matsuyama, S., Kimura, K., Ieda, N., Inoue, N., Tsukamura, H., Ohkura, S. and Matsuda, F.
  • Organizer: The 4th World Congress on Reproductive Biology
  • Int'l Joint Research
• [Presentation] Introduction of DREADD in goat arcuate nucleus using adeno-associated virus. (2017)
  • Author(s): Tatebayashi, R., Suetomi, Y., Kobayashi, K., Ohkura, S. and Matsuda, F.
  • Organizer: Society for Neuroscience 47th Annual Meeting
  • Int'l Joint Research
• [Presentation] 泌乳期ウシにおけるニューロキニンB受容体作動薬が黄体形成ホルモンのパルス状分泌に及ぼす影響. (2017)
  • Author(s): 中村 翔・若林嘉浩・山村 崇・大蔵 聡・松山秀一
  • Organizer: 日本下垂体研究会第32回学術集会
• [Presentation] シバヤギKNDyニューロン不死化細胞株の樹立 (2016)
  • Author(s): 末富祐太・舘林亮輝・束村博子・大蔵 聡・松田二子
  • Organizer: 第109回日本繁殖生物学会大会
  • Place of Presentation: 神奈川県相模原市
  • Year and Date: 2016-09-11
• [Presentation] Modification of KISS1 gene in goat embryonic fibroblasts using TALEN. (2016)
  • Author(s): Tatebayashi, R., Sakuma, T., Yamamoto, T., Ohkura, S. and Matsuda, F.
  • Organizer: International Symposium on Pituitary Gland and Its Related Systems
  • Place of Presentation: Honolulu, USA
  • Year and Date: 2016-09-01
  • Int'l Joint Research
• [Presentation] Establishment of a neuronal cell line derived from KNDy neuron in a goat (2016)
  • Author(s): Suetomi, Y., Tatebayashi, R., Tsukamura, H., Ohkura, S. and Matsuda, F.
  • Organizer: The 49th Annual Meeting of the Society for the Study of Reproduction
  • Place of Presentation: San Diego, USA
  • Year and Date: 2016-07-16
  • Int'l Joint Research
• [Book] 視床下部および下垂体ホルモン（「繁殖生物学 改訂版」） (2020)
  • Author(s): 大蔵 聡
  • Total Pages: 351
  • Publisher: インターズー
  • ISBN: 9784866711102
• [Book] The GnRH Neuron and its Control (Wiley-INF Masterclass in Neuroendocrinology Series) (2018)
  • Author(s): Goodman, R.L., Ohkura, S., Okamura, H., Coolen, L.M., Lehman, M.N.
  • Total Pages: 536
  • Publisher: Wiley-Blackwell
  • ISBN: 9781119233244
• [Book] Masterclass in Neuroendocrinology (2017)
  • Author(s): Goodman, R.L., Okhura, S., Okamura, H., Coolen, L.M. and Lehman, M.N.
  • Publisher: John Wiley & Sons, Inc.
• [Remarks] 名古屋大学大学院生命農学研究科 動物生産科学研究室
  • URL: https://www.agr.nagoya-u.ac.jp/~laps/
• [Remarks] 名古屋大学大学院生命農学研究科 動物生産科学研究室
  • URL: http://www.agr.nagoya-u.ac.jp/~laps/index-j.html
• [Remarks] 名古屋大学大学院生命農学研究科 動物生産科学第1研究分野
  • URL: http://www.agr.nagoya-u.ac.jp/~laps/index-j.html