Intrabursal transfer of spermatozoa (ITS): A new route for artificial insemination of mice

Artificial insemination (AI) by direct injection of epididymal spermatozoa into the reproductive tract of females is simpler and more convenient than in vitro fertilization (IVF) and subsequent transfer of fertilized eggs to recipient oviducts for simultaneous acquisition of a large number of pups. Introduction of epididymal spermatozoa into oviducts via the oviductal wall or via vaginal and intrauterine routes is currently the most commonly used method for AI in mice. In this study, we explored another route fur AI of the mouse and found that transfer of spermatozoa into a space near the infu ndibulum between the ovary and ovarian bursa enables in vivo fertilization of ovulated oocytes at the ampulla. When 1 muL of a sperm suspension contai ning 1 x 10(4) spermatozoa freshly isolated from B6C3F1 males was intraburs ally injected into superovulated B6C3F1 females on E (embryonic day) 0.4 (1 0:00 AM), 5 of 7 females yielded 2-cell embryos with rates of efficiency ra nging from 4 to 21% (11% on average), which were much lower than those (91% on average) for embryos obtained by natural mating. All the 2-cell embryos derived from injection of sperm developed in vitro to hatched blastocysts. Similar results were obtained from injection of 1 muL of sperm suspension containing 1 x 10(3) spermatozoa, although in vivo fertilizing ability was slightly improved (28% on average). When 1 muL of sperm suspension containi ng 1 x 104 spermatozoa was injected intrabursally into superovulated female s that had been mated with vasectomized males, 6 of 10 mice (60%) yielded 1 9 normal mid-gestational fetuses with an average litter size of 3.2, which was much lower than that (14.5) for embryos obtained by natural mating. Alt hough the present findings appear to be preliminary, this technique, based on the intrabursal transfer of spermatozoa, will be of practical use for AI in mice, particularly for transgenic and mutant mice that are often diffic ult to breed. (C) 2001 by Elsevier Science Inc.