The marmoset monkey (Callithrix jacchus) is an important model species for the development of reproductive technologies in humans and endangered primates. Obtaining sufficient sperm of high quality is a limiting factor in implementing marmoset IVF. A method for the collection of marmoset semen, using penile vibratory stimulation (PVS), has recently been described (1). Due to the high rate of copulation in marmosets, the pairing of males with females may affect semen collection. The aim of the present study was to determine whether the quantity and motility of sperm collected by PVS is affected by animal pairing. A total of 10 adult male marmosets were used, of which 3 were paired with another adult or juvenile male, and 7 were paired with an adult female. Semen was collected from each male on up to 5 separate occassions in sterile glass tubes. PVS involved the application of successive sequences of increasing vibration to the penis using a FertiCare personal vibrator. Immediately following collection, pre-warmed Hepes-buffered TALP medium (200 μl) was added to the ejaculate. Sperm suspensions were evaluated for total sperm count and sperm motility. Ejaculates were obtained from male-paired males on every attempt (12 of 12), whereas 5 of 23 attempts failed to yield an ejaculate from female-paired males. The number of sequences of stimulation needed to obtain an ejaculate differed between males but was unaffected by animal pairing. The ejaculates collected from female-paired males had lower total numbers of sperm (3.9���1.4�×�106 v. 10.1���2.2�×�106; P�<�0.05) with a lower percentage motile (35���9% v. 85 � 13%; P<0.01), compared with those from male-paired males. We conclude that housing males separately from females increases the quantity and motility of sperm collected by PVS.
(1) Schneiders, A., Sonksen, J., Hodges, J. K. (2004) J. Med. Primatol. 33, 98–104.
Oocyte paracrine signalling is necessary for mouse cumulus cell expansion, an important preovulatory process. The oocyte-secreted factor growth differentiation factor-9 (GDF-9) signals through the bone morphogenetic protein receptor-II (BMPR-II) and is currently the primary candidate molecule for the cumulus expansion enabling factor (CEEF). The present study was conducted to determine whether in the mouse GDF-9 is the CEEF. Cumulus oocyte complexes (COC) were collected from eCG-primed mice and the oocyte was microsurgically removed to generate an oocytectomised complex (OOX). An established scoring system was used to measure FSH-induced cumulus expansion; 0 (no expansion) to +4 (maximum expansion). OOX complexes treated with FSH alone failed to expand (score: 0), whereas expansion was significantly (P�<�0.05) induced by either recombinant mouse GDF-9 (score; mean +/– SEM: 2.7 +/– 0.1), recombinant TGF-μ1 (score: 2.6 +/– 0.2) or co-culture with oocytes (score: 2.3 +/– 0.2). A GDF-9 neutralising antibody mAb-53, raised against hGDF-9, was effective in neutralising the response of OOX complexes to GDF-9 (score: 0.1 +/– 0.1), but had no significant effect on the expansion of OOX complexes co-cultured with oocytes (score: 2.3 +/– 0.2). Likewise, a TGF-μ antagonist neutralised (P�<�0.05) TGF-μ-induced, but not oocyte-induced, expansion of OOX complexes. A soluble portion of the BMPR-II ectodomain, a known GDF-9 antagonist, failed to neutralise oocyte-induced cumulus expansion (P�>�0.05) at the highest dose implying that BMPR-II is not a critical receptor involved in regulating cumulus expansion. Using real-time RT-PCR, hyaluronan synthase-2 (HAS2) mRNA expression by OOXs was upregulated 6- to 7-fold by oocytes and GDF-9. The GDF-9 neutralising antibody mAb-53, partially neutralised GDF-9-induced OOX HAS2 expression, but not oocyte-induced HAS2 expression. This study provides evidence that like TGF-μ1, GDF-9 can enable FSH-induced cumulus expansion, however more importantly demonstrates that neither GDF-9 nor TGF-μ1 alone account for the crucial oocyte-secreted factor regulating cumulus expansion in the mouse.
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