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Chlortetracycline fluorescence patterns and in vitro fertilisation of frozen-thawed boar spermatozoa incubated under various bicarbonate concentrations

Published online by Cambridge University Press:  26 September 2008

Lalantha R. Abeydeera ,
Hiroaki Funahashi ,
Nam-Hyung Kim  and
Billy N. Day
Lalantha R. Abeydeera
Affiliation:
Department of Animal Sciences, University of Missouri-Columbia, Columbia, Missouri, USA
Hiroaki Funahashi
Affiliation:
Department of Animal Sciences, University of Missouri-Columbia, Columbia, Missouri, USA
Nam-Hyung Kim
Affiliation:
Department of Animal Sciences, University of Missouri-Columbia, Columbia, Missouri, USA
Billy N. Day*
Affiliation:
Department of Animal Sciences, University of Missouri-Columbia, Columbia, Missouri, USA
*
Billy N. Day, University of Missouri-Columbia, Department of Animal Sciences, 159 Animal Sciences Research Center, Columbia, MO 65211, USA. Tel: +1(573)-882-7555. Fax: +1(573)-884-7827.
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Summary

Porcine oocyte-cumulus complexes were cultured in bovine serum albumin(BSA)-free North Carolina State University (NCSU)23 medium containing porcine follicular fluid (10%), cysteine (0.1 mg/ml)and hormonal supplements (eCG and hCG: 10IU/ml each) for 22h. They were then cultured in the same medium but without hormonal supplements for an additional 22h. After culture, cumulus cells were removed and oocytes were co-incubated with frozen-thawed ejaculated boar spermatozoa in tissue culture medium (TCM)199 containing caffeine (5mM), fetal calf serum (FCS; 10%) and varying concentrations (26–56 mM)of NaHCO3 for 9h(experiment 1). In experiment 2, chlortetracycline (CTC) was used to assess the functional state of spermatozoa incubated under different NaHCO3 concentrations. Experiment 3 examined the effect of FCS (1% and 10%)and NaHCO3 (26 and 46 mM) on fertilisation parameters. Compared with 26 mM, penetration rate was significantly higher (p<0.05)at 36–56mM NaHCO3. Polyspermy showed a similar pattern although no difference was observed between 26 and 36 mM. At 46mM NaHCO3, the mean number of spermatozoa (MNS) penetrated per oocyte increased significantly (p< 0.05). A significantly higher proportion of spermatozoa were capacitated and acrosome reacted at 46 and 56mM NaHCO3, respectively. The fertilisation medium containing 46mM NaHCO3 and 1% FCS showed a higher penetration rate (84%)with a relatively low incidence of polyspermy (39%). The results indicate that NaHCO3 stimulates capacitation and/or the acrosome reaction of boar spermatozoa in a dose-dependent manner and thus affects fertilisation parameters.

Keywords

Acrosome reactionBicarbonateCapacitationIn vitro fertilisationPig oocytes
Type
Article
Information
Zygote , Volume 5 , Issue 2 , May 1997 , pp. 117 - 125
Copyright
Copyright © Cambridge University Press 1997

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References

Austin, C.R (1951). Observations on the penetration of sperm into the mammalian egg. Aust. J. Sci. Res. B4, 581–96.Google Scholar
Beckmann, L.S. & Day, B.N. (1993). Effect of media NaC1 concentration and osmolarity on culture of the early stage porcine embryo and viability of embryos cultured in a selected superior medium. Theriogenology 39, 611–22.CrossRefGoogle Scholar
Bhattacharyya, A. & Yanagimachi, R. (1988). Synthetic organic pH buffers can support fertilization of guinea pig eggs, but not as effectively as bicarbonate buffer. Gamete Res. 19, 123–9.CrossRefGoogle Scholar
Boatman, D.E. & Robbins, R.S. (1991). Bicarbonate: carbondioxide regulation of sperm capacitation, hyperactivated motility, and acrosome reactions. Biol. Reprod. 44,806–13.CrossRefGoogle ScholarPubMed
Chang, M.C. (1951). Fertilizing capacity of spermatozoa deposited into Fallopian tubes. Nature 168, 697–8.CrossRefGoogle ScholarPubMed
Chang, M.C. (1959). Fertilization of rabbit ova in vitro. Nature 184, 466–7.CrossRefGoogle Scholar
Cran, D.G. & Cheng, W.T.K. (1986). The cortical reaction in pig oocytes during in vivo and in vitro fertilization. Gamete Res.. 13, 241–51.CrossRefGoogle Scholar
Fraser, L.R. (1984). Mechanisms controlling mammalian fertilization. In Oxford Reviews of Reproductive Biology, ed. J.R., Clarke, pp. 174–225. Oxford: Oxford University Press.Google Scholar
Fraser, L.R. (1987). Minimum and maximum extracellular Ca2+ requirements during mouse sperm capacitation and fertilization in vitro. J. Reprod. Fertil.. 81, 77–89.CrossRefGoogle ScholarPubMed
Fraser, L.R. & Monks, N.J. (1990). Cyclic nucleotides and mammalian sperm capacitation. J. Reprod. Fertil., Suppl. 42, 9–12.Google ScholarPubMed
Fraser, L.R., Umar, G. & Sayed, S. (1993). Na2+-requiring mechanisms modulate capacitation and acrosomal exocytosis in mouse spermatozoa. J. Reprod. Fertil. 97, 539–49.CrossRefGoogle ScholarPubMed
Fraser, L.R.Abeydeera, L.R. & Niwa, K. (1995). Ca2+-regulating mechanisms that modulate bull sperm capacitation and acrosomal exocytosis as determined by chlortetracycline analysis. Mol. Reprod. Dev. 40, 233–41.CrossRefGoogle Scholar
Funahashi, H. & Day, B.N. (1993). Effects of follicular fluid at fertilization in vitro on sperm penetration in pig oocytes. J. Reprod. Fertil. 99, 97–103.CrossRefGoogle ScholarPubMed
Funahashi, H., Cantley, T.C., Stumpf, T.T., Terlouw, S.L. & Day, B.N. (1994). Use of low-salt culture medium for in vitro maturation of porcine oocytes is associated with elevated oocyte glutathione levels and enhanced male pronuclear formation after in vitro fertilization. Biol. Reprod. 51, 633–9.CrossRefGoogle ScholarPubMed
Garty, B.N. & Salomon, Y. (1987). Stimulation of partially purified adenylate cyclase from bull sperm by bicarbonate. FEBS Lett. 218, 148–52.CrossRefGoogle ScholarPubMed
Hamner, C.E. & Fox, S.B. (1969). Biochemistry of oviductal secretions. In The Mammalian Oviduct, ed. Hafez, E.S.E. & Blandau, R.J., pp. 333–55. Chicago: University of Chicago Press.Google Scholar
Harrison, R.A.P., Mairet, B. & Miller, N.G.A. (1993). Flow cytometric studies of bicarbonate-mediated Ca2+influx in boar sperm populations. Mol. Reprod. Dev. 35, 197–208.CrossRefGoogle ScholarPubMed
Hunter, R.H.F. & Nichol, R. (1988). Capacitation potential of the fallopian tube: a study involving surgical insemination and the subsequent incidence of polyspermy. Gamete Res.. 21, 255–66.CrossRefGoogle ScholarPubMed
Hyne, R.V. (1984). Bicarbonate- and calcium-dependent induction of rapid guinea pig sperm acrosome reactions by monovalent ionophores. Biol. Reprod. 31, 312–23.CrossRefGoogle ScholarPubMed
Kovachev, K.D., Zagorski, D.I., Ivanova, M.G. & Bobadov, N.D. (1994). Cryogenic damage to boar spermatozoa frozen in pellets and in tubes. Theriogenology 42, 1369–79.CrossRefGoogle Scholar
Lee, M.A. & Storey, B.D. (1986). Bicarbonate is essential for fertilization of mouse eggs: mouse sperm requires to undergo the acrosome reaction. Biol. Reprod. 34, 349–56.CrossRefGoogle ScholarPubMed
Mahi, C.A. & Yanagimachi, R. (1973). The effects of temperature, osmolality and hydrogen ion concentration on the activation and acrosome reaction of golden hamster spermatozoa. J. Reprod. Fertil. 35, 55–66.CrossRefGoogle ScholarPubMed
Mattioli, M., Bacci, M.L., Galeati, G. & Seren, E. (1989). Developmental competence of pig oocytes matured and fertilized in vitro. Theriogenology 31, 1201–7.CrossRefGoogle ScholarPubMed
Miyamoto, H. & Chang, M.C. (1973). Effect of osmolality on fertilization of mouse and golden hamster eggs in vitro. J. Reprod. Fertil. 33, 481–7.CrossRefGoogle ScholarPubMed
Nagai, T. (1994). Current status and perspectives in IVM-IFV of porcine oocytes. Theriogenology 41,73–8.CrossRefGoogle Scholar
Nagai, T., Takahashi, T., Masuda, H., Shioya, Y., Kuwayama, M., Fukushima, M., Iwasaki, S. & Hanada, A. (1988). In vitro fertilization of pig oocytes by frozen boar spermatozoa. J. Reprod. Fertil. 84, 585–91.CrossRefGoogle ScholarPubMed
Nagai, T., Ding, J. & Moor, R.M. (1993). Effect of follicle cells and steroidogenesis on maturation and fertilization in vitro of pig oocytes. J. Exp. Zool. 266, 146–51.CrossRefGoogle ScholarPubMed
Neill, J.M. & Olds-Clarke, P. (1987). A computer-assisted assay for mouse sperm hyperactivation demonstrates that bicarbonate but not bovine serum albumin is required. Gamete Res. 18, 121–40.CrossRefGoogle Scholar
Niwa, K. (1993). Effectiveness of in vitro maturation and in vitro fertilization techniques in pigs. J. Reprod. Fertil., Suppl. 48, 49–59.Google ScholarPubMed
O'Donnell, J.M. (1972). Behaviour of bovine spermatozoa in media of varying osmolality and tonicity. Exp. Cell Res. 71, 297–306.CrossRefGoogle ScholarPubMed
Okamura, N., Tajima, Y., Soejima, A., Masuda, H. & Sugita, Y. (1985). Sodium bicarbonate in seminal plasma stimulates the motility of mammalian spermatozoa through direct activation of adenylate cyclase. J. Biol. Chem. 260, 9699–705.CrossRefGoogle ScholarPubMed
Olds, D. & VanDemark, N.L. (1957). Composition of luminal fluids in bovine female genitalia. Fertil. Steril. 8,345–54.CrossRefGoogle ScholarPubMed
Petters, R.M. & Wells, K.D. (1993). Culture of pig embryos. J. Reprod. Fertil. Suppl. 48, 61–73.Google ScholarPubMed
Singh, J.P., Babcock, D.F. & Lardy, H.A. (1978). Increased calcium-ion influx is a component of capacitation of spermatozoa. Biochem. J. 172, 549–56.CrossRefGoogle ScholarPubMed
Suzuki, K.Ebihara, M., Nagai, T., Clarke, N.G.E. & Harrison, R.A.P. (1994). Importance of bicarbonate/CO2 for fertilization of pig oocytes in vitro, and synergism with caffeine. Reprod. Fertil. Dev.. 6, 221–7.CrossRefGoogle ScholarPubMed
Tajima, Y., Okamura, N. & Sugita, Y. (1987). The activating effect of bicarbonate on sperm motility and respiration at ejaculation. Biochim. Biophys. Acta 924, 519–29.CrossRefGoogle ScholarPubMed
Vijayaraghavan, S., Critchlow, L.M. & Hoskins, D.D. (1985). Evidence for a role for cellular alkalinization in the cyclic adenosine 3',5'-monophosphate-mediated initiation of motility in bovine caput spermatozoa. Biol. Reprod. 32, 489–500.CrossRefGoogle ScholarPubMed
Wang, W.H., Niwa, K. & Okuda, K. (1991). In-vitro penetration of pig oocytes matured in culture by frozen-thawed ejaculated spermatozoa. J. Reprod. Fertil. 93, 491–6.CrossRefGoogle ScholarPubMed
Wang, W.H., Abeydeera, L.R., Fraser, L.R. & Niwa, K. (1995). Functional analysis using chlortetracycline fluorescence and in vitro fertilization of frozen-thawed ejaculated boar spermatozoa incubated in a protein-free chemically defined medium. J. Reprod. Fertil. 104, 305–13.CrossRefGoogle Scholar
Ward, C.R. & Storey, B.T. (1984). Determination of the time course of capacitation in mouse spermatozoa using a chlortetracycline fluorescence assay. Dev Biol. 104, 287–96.CrossRefGoogle ScholarPubMed
Yanagimachi, R. (1982). Requirements of extracellular calcium ions for various stages of fertilization and fertilization-related phenomena in the hamster. Gamete Res. 5, 323–44.CrossRefGoogle Scholar
Yanagimachi, R. (1994). Mammalian fertilization. In The Physiology of Reproduction, ed. E., Knobil & J.D., Neill. pp. 189–317. New York: Raven Press.Google Scholar
Yoshida, M., Mizoguchi, Y., Ishigaki, K., Kojima, T. & Nagai, T. (1993). Birth of piglets derived from in vitro fertilization of pig oocytes matured in vitro. Theriogenology 39, 1303–11.CrossRefGoogle Scholar