Abstract. Production of hyaluronan (hyaluronic acid: HA) was demonstrated in denuded mouse oocytes (DOs) by the enzyme-linked immunosorbent assay, and the role of HA in enlargement of the perivitelline space in the oocytes was examined. The incidence of polyspermy following insemination was also observed in DOs in which HA synthesis was inhibited. HA was not detected in culture medium containing DOs immediately after collection. After culture for 7 h, 4.75 pg of HA per DO was detected in the medium, and the mean amount of HA significantly increased to 20.78 pg 14 h after culture. When DOs were cultured in medium containing 0.25 mM 4-methylumbelliferone (MU), an inhibitor of HA synthase, the mean amount of HA in the culture medium with DOs was 8.61 pg, which was significantly smaller than the amount in the control medium with non-treated DOs (21.59 pg). The mean size of the perivitelline space in oocytes cultured with cumulus cells (5.40 μm) did not differ from that (5.08 μm) of DOs. The mean size of the perivitelline space was significantly smaller in the MU-treated DOs (3.58 μm) than in the control DOs (4.65 μm). The fertilization rate did not differ between the MU-treated DOs (84.9%) and control DOs (81.0%), whereas the incidence of polyspermy was significantly higher in the MU-treated DOs (13.3%) compared with the control DOs (2.1%). These findings clarified that the HA involved in enlargement of the perivitelline space in oocytes is synthesized and secreted by the oocytes themselves. They also suggest that there is a close relationship between the size of perivitelline space and the incidence of polyspermy in mouse oocytes. Key words: Enzyme-linked immunosorbent assay, Hyaluronan production, Incidence of polyspermy, Mouse oocyte, Size of perivitelline space (J. Reprod. Dev. 55: [496][497][498][499][500][501] 2009) he perivitelline space is the gap between the plasma membrane and zona pellucida in oocytes. Recently, it has been reported that the incidence of polyspermy in porcine and mouse oocytes with larger perivitelline space is significantly lower than that in oocytes with a smaller perivitelline space [1][2][3][4]. Therefore, it has been suggested that there could be a relationship between the size of the perivitelline space and the incidence of polyspermy in porcine and mouse oocytes.It is known that glycosaminoglycans including hyaluronan (hyaluronic acid: HA) synthesized in cumulus cells by the stimulation of FSH and LH are secreted [5,6] and that the extracellular matrix in cumulus cells is occupied by the secreted HA [7][8][9][10]. HA has been found to also exist in the perivitelline space [11][12][13][14], and one of its properties is retention of large volumes of water [14]. Therefore, when HA accumulates in the perivitelline space, its role is to absorb water, which causes the perivitelline space to enlarge.On the other hand, with regard to the secretory source of HA in the perivitelline space, Salustri et al. [15] used [ 3 H] glucosamine as a metabolic precursor to examine the production of H...

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Background: Several studies reported that abnormal behavior was noted in pediatric patients receiving several drugs including neuraminidase inhibitors (NIs). However, information on drugs associated with abnormal behavior in a real-world setting remains limited. The purpose of this study was to clarify drugs associated with abnormal behavior using a spontaneous reporting system database. Methods: We performed a retrospective pharmacovigilance disproportionality analysis using the Japanese Adverse Drug Event Report database. Adverse event reports submitted to the Pharmaceuticals and Medical Devices Agency were analyzed. Drug associated with abnormal behavior were estimated using disproportionality analysis with calculation of the reporting odds ratio and 95% confidence interval. Results: A total of 1,144 reports of abnormal behavior were identified. Signals were detected showing the association of 4 including neuraminidase inhibitors (oseltamivir, zanamivir, laninamivir, and peramivir) with abnormal behavior, and these signals were stronger for oseltamivir than other neuraminidase inhibitors. Signals were also detected for acetaminophen and montelukast. Conclusion: Our results should raise physicians’ awareness of drugs associated with abnormal behavior, but further investigation of these medications is warranted.

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Cryopreservation of mammalian metaphase-II (M-II) oocytes is still impractical compared to that of early stage embryos. In this study we examined the effects of delipation and mitotic spindle stabilization in order to improve the post-vitrification survival rate of in vitro-matured (IVM) porcine oocytes at the M-II stage. Cumulus–oocyte complexes that had been collected from slaughterhouse ovaries were matured in vitro in NCSU23 supplemented with 0.6 mm cysteine, 10 ng mL–1 epidermal growth factor (EGF), 10% porcine follcular fluid (PFF), and 10 IU mL–1 eCG and hCG. The denuded M-II oocytes were vitrified in the presence of 30% ethylene glycol and 0.5 m sucrose using the minimum volume cooling (MVC) method with a MVC plate (Cryotop�; Kitazato Supply, Tokyo, Japan). Vitrified embryos were rewarmed by immersing the MVC plate directly into rewarming solution containing 1 m sucrose and 20% calf serum at 39�C for 1 min, followed by stepwise dilution of the cryoprotectants. We compared the effects of previtrification treatments, namely, (1) delipation, (2) mitotic spindle stabilization, (3) delipation + mitotic spindle stabilization, and (4) no treatment. For delipation, we used a noninvasive method (Esaki et al. 2004 Biol. Reprod. 71, 432–437) that we had published previously with slight modification. The embryos were treated with 4% trypsin at 38�C for approximately two min to expand the zona pellucida, and then centrifuged (12 000g, 38�C 23 min) with 7.5 µg mL–1 cytochalasin B to polarize cytoplasmic lipid droplets within the perivitelline space. For mitotic spindle stabilization, M-II oocytes were vitrified in the presence of 1 µm paclitaxel. After the oocytes were rewarmed, electrical activation of the oocytes (150 V mm–1, 100 µs, one time) was carried out to induce parthenogenesis. These parthenogenetic embryos were cultured in PZM-5 for 7 days, and the number of vitrified embryos that developed into blastocysts with respect to each treatment was determined. The blastcyst formation rate and mean cell numbers of the blastcysts were compared among the treatment groups (chi-square test, Tukey's test). Of the 50 M-II oocytes that had been vitrified without pretreatment, only one oocyte (2.0%) developed into a blastocyst with 20 cells. By contrast, the number of vitrified embryos that developed into blastocysts was significantly high when they were delipated prior to vitrification (37.8%, 14/37, 64.0 � 9.6; P < 0.01). Mitotic spindle stabilization also improved the survival rate of vitrified oocytes (18.6%, 21/113, 56.7 � 9.6; P < 0.01). The combination of delipation and mitotic spindle stabilization produced the highest number of vitrified oocytes that developed into blastocysts (43.8%, 35/80, 69.4 � 6.4), although the difference between the combination group and the delipation group was not significant. These results indicate that blastocysts can be produced very efficiently from IVM porcine oocytes that have been vitrified at the M-II stage using both noninvasive delipation and mitotic spindle stabilization procedures.