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The aim of the study was to determine the effect of butyrate infusion into the rumen on butyrate flow to the duodenum, expression of short-chain fatty acid (SCFA) transporters (monocarboxylate transporter-1, -2, and -4) and receptors (G protein coupled receptor-41 and -43) in the duodenal epithelium and nutrient digestion in sheep. Eight wethers (39.0 ± 3.00 kg; mean ± SD) with ruminal and T-shape duodenal cannulas were allocated to 4 × 4 replicated Latin square design with each experimental period lasting for 21 d (12 d of adaptation and 9 d for data and sample collection). Experimental treatments were: 1) distilled water infusion into the rumen (CONT); 2) 15 g/d of butyric acid infusion into the rumen (BUT15); 3) 30 g/d of butyric acid infusion into the rumen (BUT30); and 4) 45 g/d of butyric acid infusion into the rumen (BUT45). The daily dose of butyrate was infused into the rumen via the rumen cannula, with 200 mL of solution of butyric acid and distilled water, at a constant rate (0.1389 mL/min) throughout the day using a peristaltic pump. Correspondingly, 200 mL/d of distilled water was infused into the rumen of CONT. The wethers were fed daily 900 g of chopped meadow hay and 200 g of concentrate in two equal meals at 0600 and 1800 h. Butyrate infusion into the rumen did not affect total SCFA concentration in the rumen fluid ( > 0.11). Molar proportion of butyrate in total SCFA linearly increased, and molar proportion of acetate and isovalerate linearly decreased ( ≤ 0.02) with an increasing amount of butyrate infused into the rumen. The molar proportion of butyrate in total SCFA in the duodenal digesta linearly increased ( < 0.01), and butyrate flow to duodenum tended to linearly increase ( = 0.06) with an increasing dose of exogenous butyrate delivered to the rumen. Butyrate infusion into the rumen did not affect ( ≥ 0.14) the mRNA expression of monocarboxylate transporter-2 and -4 and G protein coupled receptor-43 in the duodenal epithelium. The G protein coupled receptor-41 and monocarboxylate transporter-1 mRNA expression in the duodenal epithelium was very low in many of the analyzed samples. Digestibility of organic matter, neutral detergent fiber, and acid detergent fiber in the stomach (forestomach and abomasum) decreased for BUT15 and BUT30 and then increased for BUT45 (quadratic, ≤ 0.04); however, neither digestibility in the intestine nor total tract digestibility differed between treatments ( ≥ 0.10).

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Orexin A may play a special role in animals' sensitivity to the day length changes such as sheep. The localization of mRNA for prepro-orexin in the ovine hypothalamus was found to correspond to the pattern described in rodents. The results of that research also showed that the expression of the orexin gene depends on the length of a day and is higher during short days. Other study revealed that mRNA for orexin receptors (OxR)1 and OxR2 shows strong expression in the anterior, intermediate and posterior pituitary lobes of the rat. In addition, it was also found that in the anterior pituitary, OxR1 is more strongly expressed than OxR2. These observations indicate that the pituitary gland is capable of receiving the orexin signal. The aim of the study was to determine the interaction of season and orexin A on PRL and GH secretion by pituitary explants in short-term culture. Studies were carried out on pituitaries explants collected from lactating Polish Longwool sheep during the long (LD, May, n=5) and short day (SD, December, n=5). Glands were transected saggitally into halves, with each incubated in 2.5 ml of M-199 for 180-min in medium containing either 0 or 1000 ng/ml of orexin A. Treatment with orexin during LD increased significantly the secretion of PRL (P < 0.01) and GH (P < 0.05), compared to controls. In cultures from glands collected during SD, orexin significantly (P < 0.05) decreased the secretion of both hormones, compared to controls. We conclude that the secretion of PRL and GH from the ovine pituitary gland is negatively responsive to orexin A during SD; whereas orexin may stimulate PRL and GH secretion during LD.

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The incidence of polyspermy in lamb oocytes matured and fertilized in vitro is very high and this results in a reduced developmental potential of embryos arising from them. We have attempted to produce oocytes more resistant to this fertilization anomaly. The oocytes from prepubertal lambs 7-12 weeks old were matured in a medium supplemented with various blood sera and oviductal fluid and fertilized in vitro. Significantly higher monospermic penetration was found in a medium supplemented with BSA--3 mg/ml (63.9%) and OF--20% concentration (55.8%). Lower monospermy was recorded in the presence of 10% LS (44.6%) or 10% SS (40.8%), and particularly in a medium with 10% FCS (26.9%). In contrast, high monospermy (78.7%) was observed in oocytes from adult donors matured and fertilized in an identical system. In another set of experiments we estimated whether polyspermy can be reduced by improvement of the cytoplasmic maturation of prepubertal oocytes using a two-step maturation protocol. After artificial arrest of the maturation for 24 h with a specific cdk inhibitor--BL-I, 50 miocroM--more than 80% oocytes from prepubertal and adult donors did not resume meiosis. When incubated thereafter in a drug-free medium for another 24 h, the oocytes of both categories progressed to MII in the rate comparable with control (80% to 90% MII). However, after fertilization no significant differences in the level of monospermic penetration was recorded between the arrested group (59.8%) and control (58.8%), both matured in the presence BSA, and 46.6% and 52.3% after treatment with OF. Also, no significant difference was observed between the arrested and control oocytes from adult donors (72.6% and 84.8%, respectively). These results suggest that high polyspermy in prepubertal oocytes is caused by developmental imperfection and can't be fully eliminated either by modifying the composition of culture media or by prolongation of the culture interval.

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BACKGROUND: Helicobacter pylori infection might, in some instances, be considered as zoonosis. AIM: The aim of this study was to assess the H. pylori prevalence in Polish shepherds and in their families as compared to controls. Patients and methods. A total of 42 shepherds from Polish Tatra Mountains with regular contact with sheep, 28 members of their families with incidental contacts and 61 age- and gender-matched farmer controls without such contacts were involved in this study. H. pylori status was determined by 13C-urea breath test. Serology was used to measure anti-H. pylori and anti-CagA IgG. Plasma gastrin, interleukin-8 and tumor necrosis factor-alpha were also determined. RESULTS: The H. pylori prevalence reached 97.6% in shepherds, 86% in their family members, but significantly less, 65.1%, in controls without contact with sheep. Anti-H. pylori IgG, anti-CagA in contact groups were significantly higher than in controls. Also, plasma gastrin, interleukin-8 and tumor necrosis factor-alpha had significantly higher values as compared to controls. CONCLUSIONS: Shepherds showed almost 100% H. pylori prevalence and higher incidence of CagA seropositivity, plasma gastrin and pro-inflammatory cytokine levels. Considering 100% positive 13C-urea breath test in sheep, it may be reasonable to suggest that H. pylori infection in shepherds and their family members originates from sheep and H. pylori infection might, therefore, be considered as zoonosis.

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The severe loss of developmental competence affecting fertilized ova when removed from the oviductal environment suggests that this organ plays a functional role in early embryonic development. The purpose of this study was to determine the effect of sheep heat-inactivated OF on the mortality rate of ovine embryos produced in vitro and transferred into recipients. As control groups we used embryos fertilized and cultured in media supplemented with different kinds of proteins (FCS, BSA). Transfer of embryos in the two pronuclei stage to the oviducts of synchronized recipients resulted in 60% of successfully termed pregnancies after incubation of embryos in OF, 40% in BSA and only 10% after FCS. All ewes were further assessed for pregnancy by ultrasonography 33, 53 and 80 days after embryo transfer. The highest embryo mortality appeared between day 33 and 52. We concluded that incubation of ovine oocytes in OF during the final period of the maturation process may play a functional role at the time of fertilization and early embryonic development.

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The objective of the first stage of these studies was to investigate whether temperature increases in the milk and body of cows during the early period of pregnancy. We studied 94 pregnant and 116 nonpregnant cows, and the temperatures were measured daily beginning 24 days after insemination. In addition, progesterone levels in milk were measured twice (on Days 21 and 24 after insemination), and examinations per rectum were conducted to determine pregnancy. Data analysis showed that in almost 90% of the pregnant cows milk temperature increased by 0.64 degrees C some time between Days 5 and 12 after insemination. This increase in milk temperature was highly significant and corresponded with a 0.46 degrees C increase in body temperature. There was no increase in the milk or body temperature in nonpregnant cows. After discovering in the first stage that a high percentage of pregnant cows experienced both milk and body temperature increases, we focused our attention in the second stage of the study on the question of whether this increase in temperatures might be due to an immune response of the mother to the entry of the embryo into the uterus. In this stage we conducted three different experiments on another group of animals consisting of 309 cows and heifers. The experiments included analysis of progesterone, estradiol, cortisol and PGE2 levels in the blood serum of the cows and heifers; an estimation of the pyrogenic activity of PGE1, PGE2 and progesterone; and measurements of body temperature in the heifers before and after embryo transfer. The results of these experiments suggested that an increase in milk and body temperature could be an indicator of an immune response of the mother to the entry of the embryo into the uterus.

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The study was designed to compare the direct effect of three prolactin-like hormones on steroidogenesis of ovine luteal cells collected at day 40-45 of pregnancy. 100 ng/ml of ovine placental lactogen or 100 ng/ml of ovine growth hormone or 100 ng/ml of ovine prolactin were added to the media of luteal cell cultures. After 48 h incubation, all cultures were terminated and the media were frozen until further steroid analysis. To determine to what extent growth hormone (GH), prolactin (PRL) and lactogen (PL) regulate the activity of 3 beta-HSD, an enzyme involved in progesterone synthesis, the classical steroidal competitive inhibitor of 3 beta-HSD trilostane, was investigated for its effects on basal and GH-, PRL-, and PL-stimulated progesterone biosynthesis since there is a possibility that the luteotropic effect of these hormones are mediated via 3 beta-HSD. oPL resulted in an increase of progesterone secretion in a statistically significant manner, while GH or PRL had no effect on progesterone secretion. A decrease in progesterone secretion as an effect of 100 mM trilostane was observed in all culture types. An explanation for the luteotropic effect of PL and the lack of this effect for GH is that the GH receptor associates with a different molecule within the ovarian tissue and forms a heterodimeric receptor for PL, and the possibility that physiological effects of native oPL may be mediated through its binding to specific PL receptors, which have low affinities for oGH and oPRL.

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The study was undertaken to investigate the steroid hormone production by sheep luteal cells. Corpora lutea were collected from 30 Olkuska sheep on Days 3, 6, 9, 12 and 15 of the estrous cycle during the reproductive season. In Experiment 1, steroid hormone concentration was estimated in extracts of CL. In Experiment 2, luteal cells were cultured in vitro for 24 h. Luteal cells isolated on Days 9 and 12 secreted high amounts of progesterone and androgens but smaller amounts of estradiol. Concentration of these steroids in CL extracts collected on the same days showed the same trend. In CL harvested on Day 15, a decrease in androgens and progesterone as well as a significant increase in estradiol were observed in culture media and in extracts. Judging from the high amounts of estradiol and low amounts of androgen observed at the end of the luteal phase, we speculate that the steroid hormones secreted by the regressing CL may play an active role in the regulation of the estrous cycle in the Olkuska sheep with autocrine influence on the luteal activity or a possible paracrine action on follicular growth. In the third Experiment, the possibility of heterogeneity in the multiple corpora lutea population of prolific Olkuska sheep was investigated. Differences were found in the level of progesterone and estradiol secretion by individual corpora lutea recovered from the same animal, which also varied in terms of weight. This is the first study which shows the existence of intra-ovarian and individual heterogeneity between corpora lutea recovered from ewes during the normal estrous cycle.

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The luteotropic activity of ovine placental lactogen (oPL) on different days of gestation in ewes was assessed using in vitro methods. Corpora lutea (CL) harvested on Days 45, 70, 95, 120 and 135 of gestation and during parturition were enzymatically dispersed and plated on multiwell plates. After 48 h of incubation, all cultures were terminated and media were frozen for further steroid analysis. Cells were cultured in control medium, with addition of oPL alone, or in combination with PGE2 or PGF2alpha. Supplementation of culture media with oPL increased basal progesterone secretion by cells isolated on Days 45 and 70 of gestation. There was no effect on progesterone secretion by cells isolated on other days of gestation; PGE2 added to the culture media increased progesterone production only by cells isolated on Day 70 of pregnancy. Simultaneous oPL treatment with PGE2 had a statistically significant and stimulatory effect on progesterone production by luteal cells collected on Days 70 and 95 of pregnancy. In contrast, PGF2alpha alone in culture media decreased progesterone secretion by cells isolated on Days 45, 70 and 95 of gestation, while oPL plus PGF2alpha on Days 70 and 95 of gestation protected against luteolytic action of PGF2alpha. The results showed 1) a direct effect of the oPL on luteal cells isolated on Days 45 and 70 of gestation; 2) synergism between PL and PGE2 in progesterone production; by cells isolated on Day 70; 3) and a luteoprotective effect of oPL against the luteolytic action of prostaglandin F (PGF2alpha) observed on Days 70 and 95 of gestation.

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Opioid peptides derived from several genes are present in the reproductive system and are known to be involved in the regulation of reproduction. This work shows that the level of alpha-neoendorphin which derives from prodynorphin changes in folliculogenesis. The lowest level was observed in the small follicles, increased in the medium, and reached the highest level in the preovulatory follicles. The presence of the kappa-opioid receptor in granulosa cells, obtained from follicles of varying sizes was demonstrated. The monoclonal anti-kappa-opioid receptor antibody was used to reveal the expression of kappa-opioid receptor. The results obtained suggest changes in alpha-neoendorphin levels and kappa-opioid receptor density during maturation of the follicles.

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The objective of this study was to identify whether high ovulation rate in high fecundity Polish Olkuska Sheep results from within ovary differences by comparing the number, size and estradiol concentration in ovarian follicles obtained from high fecundity ewes of Olkuska breed, non prolific Polish Mountain Sheep and the crossbreed between them. The ovaries were obtained after laparotomy from the anesthetized animals on the 17th day after the onset of estrus. The diameter of individual follicles was estimated with the use of operating microscope with a calibrated scale in the ocular. Dissected follicles were stored at -20 °C for the estradiol estimations in the homogenates. The mean number of highly estrogenic and medium follicles in crossbreed was significantly lower than that in Olkuska and significantly higher than that in Polish Mountain. However, mean estradiol level in high estrogenic and medium follicles in three investigated groups was not significantly different. Mean ovulation rates were 3.1 for Olkuska, 1.5 for Polish Mountain and 2.2 for crossbreed. It is concluded that, like in Booroola sheep, the increased number of follicles that mature and ovulate results from the differences within the ovary. The recruitment of ovulatory follicles in high fecundity Olkuska sheep, lasts longer than in non prolific Polish Mountain sheep and similar tendency has been observed in crossbreed.

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The sympatho-adrenal and pituitary-adrenal cortex axes are the most sensitive, and specific indicators of stress in animals. Increased plasma levels of catecholamines and glucocorticoids are generally considered as the classical response to stress. Most experiments on immobilization have been performed on rats and only a few of them concerned domestic animals. In this experiment we want to learn whether short-term restraint-a stressor most commonly used in animal husbandry-is a stressor for sheep (ewes) like in rats. For this reason we measured adrenaline (A), noradrenaline (NA) (radioenzymatic method), cortisol (RIA method), glucose and free fatty acids (FFA). Unlike in rats, in stressed sheep the peak of A appeared earlier than the NA peak, i.e. at 2 and 5 min. of stress, respectively. In contrast to rats, the basal and stress levels of NA exceeded the corresponding level of A. Cortisol concentration rose 7 fold above baseline and maximal concentration appeared at a time (15-30 min.) observed in other animal species. A similar time-related increase was observed in the plasma FFA concentration. It increased maximally 3.2 fold at 15 min. of stress. A significant correlation coefficient was found between plasma cortisol and FFA (r = 0.91) what may suggest the lipolytic effect of ACTH and/or a positive feedback of FFA on the hypophysis-adrenal axis. The plasma glucose of stressed animals rose only 1.47 fold above the basal level. A significant correlation was found between cortisol and glucose (r = 0.53) whereas no correlations have been obtained between A, NA and glucose or FFA.(ABSTRACT TRUNCATED AT 250 WORDS)

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The aim of this study was to determine the efficiency of a porcine pituitary gonadotrophin extract with a defined pLH content in the superovulation of sheep. Estrus was synchronized in 61 Polish Mountain ewes with intravaginal fluorogestone acetate sponges. Twenty-four hours before the sponges were removed, the ewes underwent different superovulatory treatments: Group I 250 IU of pFSH with 250 IU of pLH (n=19); Group II 500 IU of pFSH with 500 IU of pLH (n=19); and Group III 750 IU of pFSH and 750 IU of pLH (n=18). Gonadotrophine was administered intramuscularly twice a day over a 3-day period in decreasing dosages. A control group of ewes (n=5) was treated with saline. In most of the ewes estrus began about 20 hours after sponges were removed. All the ewes were bred naturally every 12 hours. Superovulation was confirmed in 75% of the treated animals. The ewes receiving 250 IU each of pFSH and pLH produced an average of 7.6 +/- 3.1 corpora lutea (CL), 6.3 +/- 2.4 ova and 4.3 +/- 4.1 transferable embryos. Group II (500 IU of pFSH and pLH) produced 8.5 +/- 4.0 CL, 7.6 +/- 4.1 ova, and 4.1 +/- 2.9 transferable embryos. Group III (750 IU each of pFSH and pLH) produced 8.3 +/- 5.2 CL, 7.5 +/- 5.5 ova and 5.2 +/- 5.1 transferable embryos. The mean embryo recovery rate was 87% for all three groups. Differences in superovulatory response and embryo recovery rate among the groups were not statistically significant (P>0.05).

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The effect of hydrocortisone and dexamethasone on superovulation was examined in 12 cows. On the day PMSG was given, each animal received either the first of five daily doses of 250 mg succinate hydrocortisone or one injection of 30 mg dexamethasone. In the 48-hr interval between the injection of PMSG and PGF(2)alpha, the concentration of progesterone rose from 6.97 to 10.22 ng/ml in the experimental groups and only to about 2.8 ng/ml in the control group. In the following days progesterone increased even more, from 15.7 to 26.0 ng/ml seven days after estrus in the experimental group and to 19.25 ng/ml in the control group. The group which received dexamethasone had an average of 4.7 corpora lutea and one embryo flushed per animal. The hydrocortisone group had an average of 2.5 corpora lutea and one cow had two embryos. The control group had 6.2 corpora lutea and 5.2 embryos per animal.

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Measurements of hematocrit, total plasma protein and red cell volume were made during sexual excitement and copulation in boars. Red cell volume (RCV) was determined by isotope dilution technique using endogenous 51Cr-tagged red cells. Basing on these data changes in the total blood volume (TBV) and plasma volume (PV) were calculated by 2 indirect methods. RCV increased by 12% to 16% in the intial phase of ejaculation and remained increased during ejaculation and 40 minutes after copulation. TBV and PV decreased during copulation, greatest drop being found in the final phase of ejaculation. After ejaculation the TBV was increasing, first to the resting value before copulation (about 20 minutes after copulation), thereafter it became markedly higher than during the resting period. Depending on the method used for calculation significant differences were found in the quantity of TBV and PV drop during ejaculation.

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