Tag Archives: Pig Oocytes

Parthenogenetic Activation of Pig Oocytes: DISCUSSION(10)

These results suggest that FCS also inhibits cortical and/or zona reactions in bovine oocytes. In the present study, we used BSA, not FCS, in our culture and fertilization media, and these conditions can produce embryos that are more viable by preventing polyspermic penetration. It is possible that these media induce more normal cortical and zona reactions than do other media that have been used in pig IVF Further, during normal fertilization (in the oviduct), sperm heads were observed only in the ZP, not in the PVS, suggesting that pig oocytes have a strong zona block to polyspermic penetration under physiological conditions. Although the reasons for high polyspermy with pig IVF are not completely understood, it is suggested that oviductal secretions may participate in the prevention of polyspermy. In our IVF studies, sperm begin to penetrate oocytes 3 h after insemination. Therefore, it is possible that by 3 h after A23187 treatment, a sufficient cortical reaction has occurred to induce a zona and/or PVS block, although the actual time was not determined. It has been reported that the zona reaction requires 17-35 min in the hamster, 60-120 min in the mouse, and 10-100 min in the rat. Further experiments are necessary to examine the zona reaction time of pig oocytes during fertilization. Recently, we also found that there were major differences between the extracellular matrices (ZP and PVS) of pig oocytes matured in vitro and those flushed from oviducts. The changes in the extracellular matrix may participate in the block to polyspermic penetration by an as yet unknown mechanism. buy asthma inhalers

Parthenogenetic Activation of Pig Oocytes: DISCUSSION(9)

These results indicate that FCS inhibits the zona reaction without preventing CG exocytosis. The same results have also been previously reported for mouse oocytes. If mouse oocytes were matured in a serum-free medium, zona hardening occurred and sperm penetration was prevented. However, when FCS or follicular fluid was included in the medium, zona hardening was inhibited, and the fertilizability of oocytes was maintained. Du-cibella et al. first demonstrated that premature CG exocytosis was the reason for zona hardening of mouse oocytes matured in a serum-free medium, because the limited CG exocytosis modified ZP proteins; thus sperm could not bind to ZP and penetrate oocytes. buy ortho tri-cyclen online

Parthenogenetic Activation of Pig Oocytes: DISCUSSION(8)

Recently, it has also been reported that in some mammals, the formation of the CG envelope, after CG exocytosis, in the perivitelline space (PVS) also plays an important role in the block to polyspermy. According to the study by Cran and Cheng in pig oocytes, the formation of the CG envelope may in fact represent the dispersion of CG exudates. Wells et al. reported that A23187 treatment of pig oocytes prevented sperm penetration if insemination was conducted soon after treatment, but this block disappeared if oocytes were cultured for > 2 h before insemination. Our present study (experiment 2) gave different results from those reported by Wells et al.. We found that even when the treated oocytes were cultured for 3.5 h before insemination, the oocytes still possessed the ability to resist sperm penetration. The differences between the two studies may be due to different media or to different supplementations. Although there was no information about the medium used in the study by Wells et al. buy asthma inhaler

Parthenogenetic Activation of Pig Oocytes: DISCUSSION(7)

One of the roles of [Ca2+]; transients in fertilized oocytes is to cause CG exocytosis, which in turn induces a zona reaction that participates in the block to polyspermy in most mammals. CG exocytosis induced by artificial stimulators is also the result of the rise in [Ca2+];. In the present study, we found that CG exocytosis was significantly affected by the amplitude of the Ca2+ transient. A lower concentration of A23187 induced little increase in [Ca2+]i5 which resulted in a small amount of uneven CG exocytosis and nuclear activation of only a few oocytes. By contrast, a higher concentration of A23187 induced a higher ratio of Ca2+ transient, which caused most of the CGs to be released in oocytes, and the cortical reaction prevented sperm penetration in ZP-intact oocytes. However, the plasma membrane block to sperm penetration of oocytes activated by A23187 was not observed in this study. flovent inhaler

Parthenogenetic Activation of Pig Oocytes: DISCUSSION(6)

Another difference is that most of the electrically activated pig oocytes did not form a male pronucleus after sperm penetration; our unpublished data), but most of the A23187-treated pig oocytes did (Tables 2-4). In the present study, sperm may have penetrated only the oocytes that had actually not been activated, because A23187 induced nuclear activation in only 19.8-62.3% of the oocytes, and the sperm penetration rates were also very low (341%). It is also possible that sperm penetration occurred before the oocytes were fully activated by A23187. buy ortho tri-cyclen

Parthenogenetic Activation of Pig Oocytes: DISCUSSION(5)

A single electrical pulse induces only one Ca2+ transient, and this transient is sufficient to induce CG exocytosis and resumption of oocyte meiosis in pig oocytes. When A23187 was used in the present study, we also observed one Ca2+ transient in the pig oocytes. From this, it seems that there is similarity between the two stimulators. However, some differences were observed between the two artificial stimulators. One is that electrically activated pig oocytes showed the same penetrability by spermatozoa after in vitro insemination as that in control oocytes, but A23187-treated oocytes showed significantly lower penetrability in a concentration-dependent manner (Tables 2 and 3). Buy Asthma Inhalers Online

Parthenogenetic Activation of Pig Oocytes: DISCUSSION(4)

However, cortical reaction, not nuclear activation, was induced with A23187 in Ca2+-free medium in mouse oocytes, indicating that extracellular calcium is necessary for full activation of the oocytes. Calcium transients were not observed in pig oocytes either by electrical pulse or A23187 (data not shown) in Ca2+-free medium. It is possible that the mechanisms of oocyte activation by A23187 or other artificial stimulations are different among species. Our present results indicate that A23187-induced Ca2+ transients started with a gradual increase phase and this increase reached the peak value within 60-160 sec, depending on the concentration of A23187 used, and then showed a very slow recovery phase that lasted 1-3 min. levitra super active plus

Parthenogenetic Activation of Pig Oocytes: DISCUSSION(3)

This medium contains 7.5 mM extracellular Ca2+, which is a concentration much higher than that used in previous studies. We found that most parameters (peak ratio value of Ca2+ transient, CG exocytosis, and nuclear activation) measured in the present study were concentration-dependent; i.e., higher concentrations of A23187 induced a higher ratio of Ca2+ transient, which in turn induced higher rates of CG exocytosis and nuclear activation. The differences observed in the present study from previous reports may be due to the composition (such as concentration of calcium) of the media used by different researchers or the quality of oocytes matured under different conditions. The oocytes matured in our present study can develop to blastocysts (-30% of blastocyst formation of inseminated oocytes) after IVF/in vitro development and also to term after IVF/embryo transfer. It is well-known that cytoplasmic maturation is significantly affected by the culture conditions in pig oocytes. Various cytoplasmic factor(s) may affect the response of oocytes to external stimulators. buy diabetes drugs

Parthenogenetic Activation of Pig Oocytes: DISCUSSION(2)

Although A23187 is widely used for the activation of oocytes in many animals, the effectiveness is different among species. In the present study, nuclear activation rates were still low even when the concentration was increased to 100 ц,М. Studies in cattle with either ethanol or A23187 indicate that the activation rates are dependent upon the age of oocytes. Also in cattle, the activation rates of oocytes were increased by culture of the oocytes in media containing the protein synthesis inhibitor cycloheximide. However, according to the study by Hagen et al., oocyte age-dependent activation of pig oocytes was not observed by electrical pulse. The possibilities of oocyte age-dependent activation by A23187 and/or by the combination of A23187 and cycloheximide for the activation of pig oocytes remain to be determined. ventolin inhaler

Parthenogenetic Activation of Pig Oocytes: DISCUSSION(1)

It has been reported that the effect of A23187 is the direct induction of extracellular calcium influx and intracellular H+ efflux, thus resulting in increases in [Ca2+]j and intracellular pH ([pH],). Although in the present study we did not examine the [рЩ, [Ca2′ ], measurement indicated that activation of pig oocytes by A23187 was related to increased [Ca2′]; and that this increase was dependent upon the concentration of A23187 used. Also we found that the peak ratio values of the transients were positively correlated to the ability of oocytes to release CGs and form a pronucleus(ei). In addition, the A23187-induced cortical reaction modifies ZP, which in turn participate in inhibition of penetration of ZP-intact oocytes by spermatozoa. birth control pills

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