Monitoring of reproductive cycle and clinic application of artificial breeding in endangered wildlife

• Project/Area Number: 15380215
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Clinical veterinary science
• Research Institution: Gifu University
• Principal Investigator: TSUBOTA Toshio Gifu University, Faculty of Applied Biological Science, Professor, 応用生物科学部, 教授 (10207441)
• Co-Investigator(Kenkyū-buntansha): MURASE Tetsuma Gifu University, Faculty of Applied Biological Science, Associate Professor, 応用生物科学部, 助教授 (30303514)
• Project Period (FY): 2003 – 2006
• Project Status: Completed (Fiscal Year 2006)
• Budget Amount: ¥13,800,000 (Direct Cost: ¥13,800,000); Fiscal Year 2006: ¥1,700,000 (Direct Cost: ¥1,700,000); Fiscal Year 2005: ¥1,700,000 (Direct Cost: ¥1,700,000); Fiscal Year 2004: ¥5,000,000 (Direct Cost: ¥5,000,000); Fiscal Year 2003: ¥5,400,000 (Direct Cost: ¥5,400,000)
• Keywords: endangered wildlife / artificial breeding / reproductive cycle / fecal sex steroid hormone / artificial insemination / in vitro maturation of ova / in vitro fertilization / conservation of species

Research Abstract

The purposes of this study were to monitor reproductive cycles by measuring sex steroid hormone concentrations non- invasively using molecular biology technique and to apply artificial breeding methods clinically under captive conditions in order to recover their populations in endangered wildlife. The following studies were conducted for 4 years in Tsushima wild cats and Japanese black bears.
1. Measurement of fecal sex steroid hormone concentrations in Tsushima wild cats and Japanese black bears
1) The method of measurement for fecal estradiol and progesterone concentrations using TR-FIA kit was established in domestic cats as a model for wild cats.
2) Reproductive cycles were monitored by measurement of fecal estradiol and progesterone concentrations using TR-FIA kit in Tsushima wild cats.
3) Regarding micro-measurement of fecal sex steroid hormone concentrations, preliminary studies of finding out ER Cell lines were only conducted.
2. Artificial insemination using cryopreserved semen in Japanese black bears
1) The semen was obtained from captive Japanese black bears by electroejaculation under immobilization.
2) The ejaculated semen was cryopreserved following examination of motility, concentrations and malformation rate of sperms.
3) The better method of cryopreservation of bear semen was established by examination for kinds of diluents and time of equilibration using glycerol.
4) Progesterone concentrations in blood during breeding season were measured in order to know ovulation manner of female bears.
5) Administrations of eCG and hCG for induction of artificial ovulations was experimented in female bears.

Research Products

• [Journal Article] Ovulation induction by administration of equine and human chorionic gonadotropin in captive Japanese black bears (Ursus thibetanus japonicus). (2007)
  • Author(s): Okano, T., Nakamura, S., Komatsu, T., Murase, T., Asano, M., Tsubota, T.
  • Journal Title: Jpn. J. Zoo Wildl. Med. 13(印刷中)
  • NAID: 110006277189
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Effects of final dulution rate, sperm concentration and times for cooling and glycerol equibration on post-thaw characteristics of canine spermatozoa. (2006)
  • Author(s): Okano, T., Murase, T., Asano, M., Tsubota, T.
  • Journal Title: J. Vet. Med. Sci. 68 Pages: 1133-1137
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Characteristics of captive Japanese black bears (Ursus thibetanus japonicus) semen collected by electroejaculation with different voltages for stimulation and frozen-thawed under different conditions (2006)
  • Author(s): Okano, T., Murase, T., Yayota, C., Komatsu, T., Miyazawa, K., Asano, M., Tsubota, T.
  • Journal Title: Anim. Reprod. Sci. 95 Pages: 134-143
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Characteristics of frozen-thawed spermatozoa cryopreserved with different concentrations of glycerol in captive Japanese black bears (Ursus thibetanus japonicus) (2006)
  • Author(s): Okano, T., Nakamura, S., Komatsu, T., Murase, T., Miyazawa, K., Asano, M., Tsubota, T.
  • Journal Title: J. Vet. Med. Sci. 68 Pages: 1101-1104
  • NAID: 110004820616
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Incidence of ovulation without coital stimuli in captive Japanese black bears (Ursus thibetanus japonicus) based on serum progesterone profiles (2006)
  • Author(s): Okano, T., Nakamura, S., Nakashita, R., Komatsu, T., Murase, T., Asano, M., Tsubota.T.
  • Journal Title: J. Vet. Med. Sci. 68 Pages: 1133-1137
  • NAID: 110004820624
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Characteristics of captive Japanese black bears (Ursus thibetanus japonicus) semen collected by electroejaculation with different voltages for stimulation and frozen-thawed under different conditions. (2006)
  • Author(s): Okano, T., Murase, T., Yayota, C., Komatsu, T., Miyazawa, K., Asano, M., Tsubota, T.
  • Journal Title: Anim. Reprod. Sci. 95 Pages: 134-143
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Characteristics of frozen-thawed spermatozoa cryopreserved with different concentrations of glycerol in captive Japanese black bears (Ursus thibetanus japonicus). (2006)
  • Author(s): Okano, T., Nakamura, S., Komatsu, T., Murase, T., Miyazawa, K., Asano, M., Tsubota, T.
  • Journal Title: J. Vet. Med. Sci. 68 Pages: 1101-1104
  • NAID: 110004820616
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Incidence of ovulation without coital stimuli in captive Japanese black bears (Ursus thibetanus japonicus) based on serum progesterone profiles. (2006)
  • Author(s): Okano, T., Nakamura, S., Nakashita, R., Komatsu, T., Murase, T., Asano, M., Tsubota, T.
  • Journal Title: J. Vet. Med. Sci. 68 Pages: 1133-1137
  • NAID: 110004820624
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Characteristics of captive Japanese black bears ( Ursus thibetanus japonicus ) semen Collected by electroejaculation with different voltages for stimulation and frozen-thawed under different conditions. (2006)
  • Author(s): Okano, T., Murase, T., Yayota, C., Komatsu, T., Miyazawa, K., Asano, M., Tsubota, T.
  • Journal Title: Anim. Reprod. Sci. 95 Pages: 134-143
• [Journal Article] Electroejaculation and semen cryopreservation of free-ranging Japanese black bears (Ursus thibetanus japonicus) (2004)
  • Author(s): Okano, T., Murase, T., Tsubota, T.
  • Journal Title: J. Vet. Med. Sci. 66 Pages: 1371-1376
  • NAID: 110003963617
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Effects of final dilution rate, sperm concentration and times for cooling and glycerol equilibration on post-thaw characteristics of canine spermatozoa. (2004)
  • Author(s): Okano, T., Murase, T., Asano, M., Tsubota, T.
  • Journal Title: J. Vet. Med. Sci. 66 Pages: 1359-1364
  • NAID: 110003963615
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Electroejaculation and semen cryopreservation of free-ranging Japanese black bears (Ursus thibetanus japonicus). (2004)
  • Author(s): Okano, T., Murase, T., Tsubota, T.
  • Journal Title: J. Vet. Med. Sci. 66 Pages: 1371-1376
  • NAID: 110003963617
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Effects of final dilution rate, sperm concentration and times for cooling and glycerol equilibration on post-thaw characteristics of canine spermatozoa. (2004)
  • Author(s): Okano, T., Murase, T., Asano, M., Tsubota, T.
  • Journal Title: J.Vet.Med.Sci. 66(11) Pages: 1359-1364
  • NAID: 110003963615
• [Journal Article] Electroejaculation and semen cryopreservation of free-ranging Japanese black bears (Ursus thibetanus japonicus). (2004)
  • Author(s): Okano, T., Murase, T., Tsubota, T.
  • Journal Title: J.Vet.Med.Sci. 66(11) Pages: 1371-1376
  • NAID: 110003963617
• [Journal Article] Spermatogenesis, serum testosterone levels and immunolocalization of stcroidogenic enzymes in the wild male Japanese black bear (Ursus thibelanus japonicus) (2003)
  • Author(s): Okano, T., Murase, T., Tsubota, T.
  • Journal Title: J. Vet. Med. Sci. 65 Pages: 1093-1099
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Spermatogenesis, serum testosterone levels and immunolocalization of steroidogenic enzymes in the wild male Japanese black bear (Ursus thibetanus japonicus). (2003)
  • Author(s): Okano, T., Murase, T., Tsubota, T.
  • Journal Title: J. Vet. Med. Sci. 65 Pages: 1093-1099
  • NAID: 110003886132
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Ovulation Induction by Administration of Equine and Human Chorionic Gonadotropin in Captive Japanese Black Bears (Ursus thibetanus japonicus).
  • Author(s): Okano, T., Nakamura, S., Komatsu, T., Murase, T., Asano, M., Tsubota, T.
  • Journal Title: Jpn J. Zoo Wildl. Med. (in press)
  • NAID: 110006277189
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Ovulation induction by administration of equine and human chorionic gonadotropin in captive Japanese black bears (Ursus thibetanus japonicus).
  • Author(s): Okano, T., Nakamura, S., Komatsu, T., Murase, T., Asano, M., Tsubota, T.
  • Journal Title: Jpn. J. Zoo Wildl. Med. 12(印刷中)
  • NAID: 110006277189
• [Journal Article] Characteristics of captive Japanese black bears (Ursus thibetanus japonicus) semen collected by electroejaculation with different voltages for stimulation and frozen-thawed under different conditions.
  • Author(s): Okano, T., Murase, T., Yayota, C., Komatsu, T., Miyazawa, K., Asano, M., Tsubota, T.
  • Journal Title: Anim.Reprod.Sci. (印刷中)
• [Book] 獣医繁殖学第3版 (2006)
  • Author(s): 坪田敏男
  • Total Pages: 22
  • Publisher: 文永堂出版
  • Description: 「研究成果報告書概要(和文)」より
• [Book] Thriogenology (2006)
  • Author(s): Tsubota, T.
  • Total Pages: 22
  • Publisher: Japan Forest Technology Association
  • Description: 「研究成果報告書概要(欧文)」より
• [Book] 森の野生動物に学ぶ101のヒント (2003)
  • Author(s): 坪田敏男
  • Total Pages: 2
  • Publisher: (社)日本林業技術協会
  • Description: 「研究成果報告書概要(和文)」より
• [Book] One hundred and one hints obtained from wildlife in forests (2003)
  • Author(s): Tsubota, T.
  • Total Pages: 2
  • Publisher: Buneido Publishing Co.,Ltd.
  • Description: 「研究成果報告書概要(欧文)」より