Intracytoplasmic Sperm Injection in the Mouse1

Abstract

Intracytoplasmic sperm injection (ICSI) was successful in the mouse when a piezo-driven micropipette was used instead of a mechanically driven conventional pipette. Eighty percent of sperm-injected oocytes survived, and approximately 70% of them developed into blastocysts in vitro. When 106 embryos at the 2- to 4-cell stage were transferred to eight naturally mated foster mothers, 30% of the embryos (25-43%, depending on the host) reached the full term. Except for two that were cannibalized soon after birth, all of the young (30 pups) grew into normal adults. Studies of this type on the mouse may increase understanding of the fertilization process and of how ICSI works.

Chromosome analysis of mouse zygotes produced by intracytoplasmic injection of spermatozoa exposed to acrosome reaction inducing agents methyl-β-cyclodextrin and calcium ionophore A23187

“…This modified TYH with two-fold of Ca 2+ was designated as mTYH. Hepes-buffered H-mCZB [18] was used for preparing oocytes where appropriate. Temporary storage of oocytes and cultivation of fertilized ova were performed using bicarbonate-buffered CZB [19] with modification by addition of 5.56 mM glucose.…”

“…A small amount (approximately 5 μl) of diluted sperm suspension was then transferred into a droplet (20-30 μl) of H-TYH containing 10% PVP under paraffin oil, which was prepared in a plastic chamber on an inverted microscope with a piezo micromanipulator. A motile spermatozoon with morphologically normal feature was selected under ×200 magnification, and the head was separated from the tail by applying piezo pulses, then immediately injected into an oocyte using a piezo micromanipulator as described by Kimura and Yanagimachi [18]. Injected oocytes were transferred into mCZB for cultivation within 30 min of the manipulation.…”

Cryo-survival, fertilization and early embryonic development of vitrified oocytes derived from mice of different reproductive age

“…With the sperm/PVP suspension in the same dish, only highly motile sperm with a morphologically normal head were selected, and the head was separated from the tail by applying a few Piezo pulses. The sperm head was injected into the oocyte using a Piezo drive unit [21]. After injection, the oocytes were washed thoroughly with mHTF medium and then transferred into a droplet containing 50 μL of embryo maintenance medium (SAGE Media, USA) and cultured in the medium for 120-122 h under paraffin oil at 37°C in 5% CO 2 in humidified air.…”

Pronuclear formation of freeze-dried canine spermatozoa microinjected into mouse oocytes

“…A spermatozoon was aspirated into the injection pipette tail first in H-TYH containing 10-12% polyvinyl pyrrolidone (molecular weight: 360000; Nacalai Tesque, Kyoto, Japan), and the tail was cut at the mid-piece by applying a few piezopulses. The tail-cut spermatozoon was individually injected into a mouse oocyte according to the method of Kimura and Yanagimachi [32]. The ICSI series of experiments was finished within 1 h of sperm preparation.…”