BackgroundUnlike other domestic mammals, in which metaphase-II oocytes are ovulated, canine ovulation is characterized by the release of primary oocytes, which may take 12 to up to 36 hours. Further 60 hours are needed for maturation to secondary oocytes which then remain fertile for about 48 hours. Oestrus takes 7 to 10 days on average and may start as early as a week before ovulation. This together with the prolonged process of post-ovulatory oocyte maturation requires an according longevity of spermatozoa in the female genital tract in order to provide a population of fertile sperm when oocytes have matured to fertilizability. Therefore the distribution and viability of spermatozoa in the bitch genital tract was examined during post-ovulatory oocyte maturation.MethodsThirteen beagle bitches were inseminated on the day of sonographically verified ovulation with pooled semen of two beagle dogs containing one billion progressively motile spermatozoa. Ovariohysterectomy was performed two days later (group 1, n = 6) and four days later (group 2, n = 7). The oviduct and uterine horn of one side were flushed separately and the flushing’s were checked for the presence of gametes. The oviducts including the utero-tubal junction and the uterine horns, both the flushed and unflushed, were histologically examined for sperm distribution.ResultsThe total number of spermatozoa recovered by flushing was low and evaluation of viability was limited. Prophase-I oocytes were collected from oviduct flushing in group 1, whereas unfertilized metaphase-II oocytes were detected in group 2. From day 2 to day 4 after ovulation a significant decrease in the percentage of glands containing sperm (P<0.05) and a marked reduction of the mean sperm number in uterine horn glands were observed. A concomitant diminution of spermatozoa was indicated in the utero-tubal junction accompanied by a slight increase in sperm numbers in the mid oviduct.ConclusionsOocyte maturation to metaphase-II stage is accompanied by a continuous sperm detachment and elimination in the uterine horns. Entrance of spermatozoa into the caudal oviduct seems to be steadily controlled by the utero-tubal junction thus providing a selected sperm population to be shifted towards the site of fertilization when oocyte maturation is completed.

Apoptotic cells were evaluated in the lamina epithelialis (LE) and lamina propria (LP) of the canine uterotubal junction and oviductal isthmus by means of the TUNEL assay and Caspase-3 detection regarding cyclic changes. Oviducts of bitches in anestrus, early and late follicular phase, 2 and 4 days after ovulation, mid-luteal phase, and endometrial reparation were available. The rate of apoptosis was generally low (min ¼ 0; max ¼ 33 apoptotic cells in 500 cells). Regardless of the stage of the estrous cycle, the number of apoptotic cells was significantly higher in the LE compared to the LP (P < 0.05). In anoestrus, early and late follicular phase as well as during endometrial reparation, the number of apoptotic cells was on a constantly low level. A significant increase was found 4 days after ovulation by caspase-3 detection coinciding with early luteal development and in mid-luteal phase. The results demonstrate that the number of apoptotic cells changes depending on the stage of the ovarian cycle. The coincidence of the increase in the number of apoptotic cells and the time of achievement of oocyte competence, which is known to occur in the bitch 3-4 days after ovulation, suggests a causal relationship between the postovulatory maturation stage of oocytes and the rate of apoptosis in the caudal oviduct. Furthermore, a specific relevance of apoptosis may exist in the process of fertilization regarding sperm storage and detachment of spermatozoa bound to the epithelium of the caudal oviduct, which has been identified as a functional sperm reservoir in other species. Anat Rec, 294:342-348, 2011. V V C 2010 Wiley-Liss, Inc.Key words: apoptosis; oviduct; uterotubal junction; isthmus; sperm reservoirThe oviduct plays an important role in reproduction by providing a suitable environment for oocyte pick up and maturation, sperm transport and storage, capacitation of spermatozoa, fertilization, and finally early embryonic development (Verhage et al., 1973a;Steinhauer et al., 2004;Rodriguez-Martinez, 2007). The canine estrous cycle differs considerably from that of other species, in that the follicular phase is prolonged and ovulation is followed by a prolonged luteal phase with an average duration of about 54-70 days in the nonpreg-*Correspondence to: