272 OVUM RECOVERY AND BLASTOCYST DEVELOPMENT FOLLOWING INTRACYTOPLASMIC SPERM INJECTION IN CHIMPANZEES
I. Hayasaka A , N. Yoshimoto A , Y. Mori A , K. Suzuki A , R. Honda B , H. Okamura B , Y. Ide C , W. Sakamoto C , T. Nakashima C , N. Nakagata C , R. Torii D and Y. Yoshikawa EA Kumamoto Primates Research Park, Sanwa Kagaku Kenkyusho Co., Ltd., Kumamoto, Japan. email: hayasaka@skk-net.com;
B Department of Obstetrics and Gynecology, Kumamoto University, Kumamoto, Japan;;
C Center for Animal Resources & Development, Kumamoto University, Kumamoto, Japan;;
D Research Center for Animal Life Science, Shiga University of Medical Science, Shiga, Japan;;
E Graduate School of Agricultural and Life Sciences/Faculty of Agriculture, University of Tokyo, Tokyo, Japan.
Reproduction, Fertility and Development 16(2) 256-256 https://doi.org/10.1071/RDv16n1Ab272
Submitted: 1 August 2003 Accepted: 1 October 2003 Published: 2 January 2004
Abstract
In the present study, we report on oocyte collection, intracytoplasmic sperm injection and early embryogenesis in chimpanzees. Eight adult female chimpanzees, 11–27 years of age, received a single s.c. injection of 3.75 mg GnRH (Leuplin, Takeda Co. Ltd., Osaka, Japan) 1 to 3 days after the beginning of menstruation. Daily i.m. injections of hMG (Humegon, Nippon Organon K.K., Tokyo, Japan) were initiated the following day. The dose of hMG was altered from 75 to 300 IU according to serum estradiol levels. When at least one follicle of 17 mm or more in diameter was observed, 10 000 IU of hCG (Pregnyl, Nippon Organon K.K.) were administered by i.m injection. Oocytes were recovered by ultrasound-guided transvaginal follicular aspiration 30.5 to 35.5 h after hCG injection. Mature oocytes were denuded of cumulus cells by treatment with 0.1% hyaluronidase, and injected with a frozen-thawed or fresh spermatozoan using a Piezo-driven micromanipulator. Zygotes were cultured in Quinn’s Advantage Fertilization Medium (Cooper Surgical, Inc., Trumbull, CT, USA) with 10 serum protein substitute (SPS) at 37°C in a 5% CO2 atmosphere until the pronucleus stage. The medium was replaced by Quinn’s Advantage Cleavage Medium with 10 SPS from the pronuclear to 8-cell stage, and Quinn’s Advantage Blastcyst Medium with 10 SPS, thereafter. Mild ovarian hyperstimulation syndrome (OHSS) occurred in one female chimpanzee with estradiol levels of 7520 pg mL−1. No oocytes were collected from 2 chimpanzees in which large follicles were observed. Thirty-five mature oocytes, one immature oocyte and 6 degenerate/fragmented oocytes were retrieved from 6 chimpanzees, including the one with OHSS. Among 35 mature oocytes injected with spermatozoa, 26 oocytes (74%) produced two pronuclei;; 23 zygotes (66%) cleaved to the 2-cell stage, 22 (63%) to the 4-cell stage, 14 (40%) to the 8-cell stage, and 9 (26%) to the morula stage. Seven zygotes (20%) developed to the blastocyst stage by 120 h. There were no differences in fertilization rate or early embryogenesis between frozen and fresh spermatozoa. Results indicate that techniques used for human-assisted reproduction may be applicable to the chimpanzee to help preserve this endangered species.
