​Effect of medium milieu on sperm penetration and pronuclear formation of bovine oocytes matured in vitro. Byung-Ki Kima, Sang-Chan Leeb, Kwang-Sun Leea, Bok-Kyu Leea, Chang-Hee Hana, Jong-Heung Kimb, Chae-Sik Leeb​

Theriogenology 57:2093-2104, 2002

Effect of medium milieu on sperm penetration

and pronuclear formation of bovine

oocytes matured in vitro

Byung-Ki Kima, Sang-Chan Leeb, Kwang-Sun Leea,

Bok-Kyu Leea, Chang-Hee Hana, Jong-Heung Kimb,

Chae-Sik Leeb

aFaculty of Natural Science, Department of Biology, 

University of Dong-Eui,Kaya-Dong, Pusan 614-714,

South Korea, bSaewha Women's Clinic, Pusan, South Korea

Abstract

The purpose of the present study was to investigate the optimal 

concentration of osmolarity, calcium and bicarbonate for sperm penetration 

and formation of pronuclei(PN), and to investigate the time required for 

capacitation, penetration across the zona pellucida and formation of PN in 

bovine cumulus-free oocytes matured in vitro. Bovine follicular oocytes 

collected at slaughter were matured and fertilized in vitro. Bovine sperm 

penetrated the zona pellucida in medium containing 240 to 440 mOsm, 

whereas PN formation was observed in a narrow range of osmolarities, from 

280 to 360 mOsm. Maximal penetration by spermatozoa and PN formation was 

obtained in the medium with 2.5 mM calcium. High rates of spermatozoa 

penetration were observed in the medium with 37 to 49 mM NaHCO3. However, 

PN were formed regardless of the concentration of NaHCO3. The times 

required for sperm capacitation and penetration through the zona pellucida 

were 260 and 50 min, respectively. The first development of PN was 

recorded at 120 min after sperm penetration. Therefore, our study suggests 

that fertilization ability of spermatozoa in vitro appears to be more 

stable in high concentrations of NaCl. Oocytes are more sensitive to 

osmotic stress than spermatozoa. Calcium is required for both sperm 

penetration and PN formation in cumulus-free oocytes, but bicarbonate may 

be needed mainly for the penetration of spermatozoa.

Key words: Osmolarity; Calcium; Bicarbonate; Pronuclei