A comparison between in vitro fertilization and microinjection of immobilized spermatozoa from bulls producing spermatozoa with defects

Abstract

The objectives of this study were to compare the fertilization rate of bovine in vitro matured oocytes by in vitro fertilization (IVF) and by microinjection of a single spermatozoon (MI) and to relate these rates with fertility reported for these bulls in artificial breeding. Bull A (Holstein) had a nonreturn rate of 75%. Semen from this bull is routinely used in our standard IVF procedure. Bull B (Ayrshire), used regularly in artificial breeding and related to bull D, had a nonreturn rate of 69.2%. Bull C (Brown Swiss), with a chromosomal translocation and trisomy, achieved a nonreturn rate of 42%. Bull D (Ayrshire) produced nonmotile spermatozoa (SPZ) and had an abnormality described as "tail stump defect." No pregnancies sired by bull D have been reported. Oocytes were either fertilized in vitro by capacitated SPZ or by microinjection of a single immobilized SPZ into the ooplasm. SPZ were treated with 0.1 microM A23187 and used for IVF. For microinjection SPZ were cocultured for 5 h with bovine oviduct epithelial cells (BOEC) and then immobilized by freezing and thawing twice without cryoprotectant. A single batch of killed SPZ (stored at -25 degrees C) was used for all microinjections. All oocytes were cultured in Medium 199 for 22 h at 39 degrees C and subsequently fixed, stained, and examined for evidence of fertilization (i.e., female and male pronucleus formation, SPZ decondensation). Fertilization rates following IVF with semen from bulls A, B, C, and D were 80%, 54%, 1%, and 2%, and following microinjection were 39%, 22%, 21%, and 34%, respectively.

Effects of bull, sperm type and sperm pretreatment on male pronuclear formation after intracytoplasmic sperm injection in cattle

“…The reason for the variability, however, is still unknown. Heuwieser et al (1992a) used sperm from infertile bulls for ICSI with in vitro matured bovine oocytes and achieved fertilization rates comparable to those with sperm from bulls proven to be fertile by IVF. This indicates that the fertilizability of sperm in ICSI is not coincident with their performance in IVF.…”

Induction of the Acrosome Reaction and Zona‐Free Hamster Oocyte Penetration by a Bull With Complete Teratospermia Versus a Half Brother With Normal Sperm

“…The sterile bull with 100% ter-atospermia in the present study appears to exhibit the effects of an autosomal recessive gene transmitted through heterozygous males and females . Heuwieser et al (1992) reported that bull sperm exhibiting similar teratospermia and sperm with normal morphology produced by a subfertile bull with a transmissable chromosomal anomaly (Kovács et al, 1992) could fertilize few oocytes in an IVF system, but this was increased by microinjection. Lonergan et al (1994) reported that sperm from a subfertile bull carrying a 1/29 chromosomal translocation fertilized fewer bovine oocytes than sperm from a control bull.…”

Microfluidic jet injection for delivering macromolecules into cells

“…[10] Its advantages make it the method of choice for very delicate operations such as nuclear transfer in the context of generation of transgenic animals. [11] Microinjection would likely be the method of choice for intracellular delivery of macromolecules if its main disadvantages of being slow and expensive were eliminated. Efforts to reduce the cost and increase throughput include automated microinjection devices based on robotic systems[12] and microfluidic systems that move cells onto fixed microneedles.…”