Protracted induction of parturition enhances placental maturation, but does not influence incidence of placental retention in cows.

As the etiopathology of retained placenta is still not resolved in cattle, we compared the effects of protracted induction of parturition (PIP) and conventional induction of parturition (SIP) on placental maturation and the occurrence of retained placenta. PIP was induced in 13 cows by administration of 1.3 mg dexamethasone im twice daily between Days 268 and 273 of gestation and 40 mg dexamethasone im on Day 274 of gestation. For SIP, 10 cows received a single injection of 40 mg dexamethasone on Day 274 of gestation. A third group (SPON, n = 11) served as a nontreated control group. Within 2 hours after birth, two placentomes were extracted from the uterus and used for assessment of placental maturation by histology and immunohistochemistry. Incidence of retained placenta was lower (P < 0.05) in group SPON (9%) compared with groups PIP (54%) and SIP (70%). Staining with Masson's trichrome and pan-cytokeratin indicated a higher degree of atrophy and flatness of the maternal crypt epithelium in cows with physiological release of fetal membranes (REL) compared with cows with retained placentae (RET). Staining with anti-caspase-3 ratified the observations as more apoptotic cells were detected in groups SPON and PIP compared with group SIP; however, data were not statistically significant. Additionally, the expressions of the potent vasodilators endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS) were evaluated. Both eNOS and iNOS were only expressed in chorionic tissue. Endothelin-1 (ET-1), a major vasoconstrictor, showed positive staining in maternal crypt epithelium and in chorionic epithelium. No differences were found for iNOS and eNOS and ET-1, neither among the experimental groups nor between RET and REL cows. These findings indicate that a PIP results in increased placental maturation, but does not influence the incidence of placental retention in cows. The expression of vasoactive substances does not seem to be related to the placental separation process.

Effects of a protracted induction of parturition on the incidence of retained placenta and assessment of uterine artery blood flow as a measure of placental maturation in cattle.

The objectives of the present study were to compare the effects of a protracted and a conventional induction of parturition on the incidence of retained placenta, and to evaluate the suitability of transrectal Doppler sonography of the uterine arteries as a noninvasive method for the assessment of placental maturation. Protracted induction of labor (PIP) was precipitated in 13 cows by the administration of 1.3 mg dexamethasone im twice daily between Days 268 and 273 of gestation, and 40 mg dexamethasone im on Day 274 of gestation. For conventional induction of labor (SIP), 10 cows received 40 mg dexamethasone on Day 274 of gestation. A third group was not treated and served as control (SPON; N = 11). Blood flow volume (BFV) and resistance index in the uterine arteries were measured with Doppler sonography once a day from Day 268 of gestation until labor. After each ultrasonographic examination, blood samples for determination of steroid hormones were taken. Incidence of retained placenta was lower (P < 0.05) in group SPON (9%) compared with groups PIP (54%) and SIP (70%). In the last 7 days before parturition uterine BFV and resistance index did not change (P > 0.05) and did not differ between groups SPON, PIP, and SIP (P > 0.05). Resistance index was higher (P < 0.001) in cows with retained placenta compared with cows with released placenta, and BFV did not differ (P > 0.05) between them. Total estrogen concentrations increased by 283% (P < 0.001) in group PIP and by 60% (P < 0.05) in group SPON between Days -7 and -1 before parturition. They stayed constant (P > 0.05) until Day -2 in group SIP, but increased (P < 0.05) after the high dosage of dexamethasone within 1 day by 140%. Total estrogen levels were higher (P < 0.05) in cows with released placenta than in cows with retained placenta. In conclusion, a protracted compared with a short induction of labor results in higher estrogen levels before term, but does not affect incidence of placental retention. Neither alterations in placental maturation nor changes in steroid hormones influenced uterine blood supply. Therefore, Doppler sonography of uterine arteries is unsuitable to investigate the process of placental maturation induced by glucocorticoids in cows. Nevertheless, disturbances in the placental maturation process in cows with retained fetal membranes after parturition can be detected before parturition by a higher uterine blood flow resistance in the uterine arteries.

Plasma progesterone concentrations in the mid-luteal phase are dependent on luteal size, but independent of luteal blood flow and gene expression in lactating dairy cows.

The objective of the present study was to investigate if plasma progesterone (pP(4)) concentrations are dependent on luteal size, blood flow, or gene expression in luteal tissue. To induce cycles with high and low pP(4) concentrations, respectively, 20 lactating dairy cows received either a single treatment with 25 mg prostaglandin F(2α) (PGF(2α)) on Day 4 Hour 12 (PG1; n=8), or two treatments (25 mg PGF(2α) each) on Day 4 Hours 0 and 12 (PG2; n=12) of the estrous cycle (Day 1, Hour 0=ovulation). In four cows, ovulation occurred between 4 and 6d after the second PGF(2α) treatment; these cows and one lame cow were excluded from the study. In the 15 remaining cows with physiological interovulatory intervals, pP(4), area (LTA) and volume (LTV) of luteal tissue, as well as absolute (LBF) and relative (rLBF) luteal blood flow were determined on Day 9, and relative luteal P(4) (rLP(4)) as well as luteal mRNA expression of important receptors, angiogenic, vasoactive, and steroidogenic factors were quantified on Day 11 (±1) during two successive estrous cycles. Furthermore, rLP(4) was multiplied by LTV to produce a semiquantitative assessment of absolute luteal P(4) (LP(4)). There was no effect (P>0.05) of treatment (one or two PGF(2α) treatments), neither on pP(4) concentrations nor on any other parameter in the present study. Nevertheless, there was a lower LP(4) (P=0.01), LTA (P=0.03), and LTV (P=0.02), as well as tendencies of lower pP(4) (P=0.06) and LBF (P=0.09) at first compared with second diestrus. Plasma P(4) was related with LP(4) (r=0.43, P=0.04), LTA (r=0.65, P=0.0001), and LTV (r=0.43, P=0.02), but not with rLBF (r=-0.18, P=0.34). Furthermore, there was no significant correlation between gene expression of important steroidogenic factors and P(4) concentrations in luteal tissue. Results indicate that plasma P(4) concentrations in the mid-luteal phase were dependent on luteal size, but independent of blood flow and gene expression per luteal tissue unit.

Variability of mammary blood flow in lactating Holstein-Friesian cows during the first twelve weeks of lactation.

The goals of the present study were to measure mammary blood flow volume (BFV) during the first 12 wk of lactation in dairy cows by using color Doppler sonography and to determine what affects the mammary blood flow. Forty cows were examined via color Doppler sonography on d 1, 7, 14, 28, 56, and 84 after parturition (d 0). The total BFV (BFV(total)) to the 4 mammary glands was calculated by measuring time-averaged maximum velocities (TAMV) and cross-sectional areas (A) of the left and right pudendoepigastric trunks via transrectal color Doppler sonography. Because there were no significant differences in A, TAMV, and BFV between the right and left pudendoepigastric trunks, the means of A and TAMV, and the BFV(total) of both trunks were used for calculations. The intraindividual and interindividual variability of repeated BFV measures quantified by intraclass correlation coefficients were 96 and 98%, respectively. The BFV(total) ranged from 19.9 to 27.9 L/min, with a mean of 22.3+/-4.9 L/min. Interindividual differences in BFV values were attributable to variations in A and TAMV. The interindividual variability of the BFV(total), which was determined using the coefficients of variation of the BFV(total) on individual days, ranged from 16 to 28%. All the cows had similar changes in the BFV(total) during the study. Changes in BFV(total) were not correlated with changes in the mean of A, but there was a good correlation between changes in BFV(total) and in the mean of TAMV (r=0.94). The BFV(total) was highest on d 1 of lactation, decreased 28% by d 7, and remained at this level until d 28. By d 56, the BFV(total) had increased by 15% compared with d 14 and by 10% compared with d 28. The BFV(total) on d 84 was significantly different from all other days except d 56. There were moderate correlations between daily milk yield and BFV on individual days (0.24

Effects of a shortened preovulatory follicular phase on genital blood flow and endometrial hormone receptor concentrations in Holstein-Friesian cows.

The goal of this study was to determine the effects of the length of the preovulatory phase on genital blood flow and mRNA expression of endometrial hormone receptors in cattle (Bos Taurus). Ovulation was synchronized in 50 Holstein-Friesian cows using a modified Ovsynch (ovulation synchronization) protocol, in which the second gonadotropin-releasing hormone (GnRH) administration was given 40h (G40, n=17) or 60h (G60, n=16) after the prostaglandin F(2alpha) (PGF(2alpha)) administration. The third group (S, n=17) did not receive a second GnRH administration. Transrectal color Doppler examinations were carried out 24h before (Day -1) and on Day 7 after ovulation (Day 0). Follicular size (FS) and luteal size (CLS) were quantified by measuring the areas of these structures on cross-sectional B-mode ultrasound images. Follicular blood flow (FB) and luteal blood flow (CLB) were quantified by determining the colored areas of these structures. Uterine blood flow was measured using the time-averaged maximum velocities (TAMVs) and the pulsatility indices (PIs) of both uterine arteries. Endometrial mRNA transcript abundance of estrogen receptors alpha and beta as well as oxytocin and progesterone receptor were determined on Days -1 and 7 in G40 and G60 cows. In all cows, plasma progesterone (P(4)) values were measured on Day 7. On Day -1, FS and FB values were lower (P < or = 0.05) in G40 cows compared with those in S cows but did not differ (P>0.05) between G60 cows on the one hand and G40 cows and S cows, respectively, on the other hand. On Day 7, CLS and P(4) did not differ (P>0.05) between cows of Groups G40, G60, and S; CLB was lower (P< or =0.05) in G40 cows than in G60 and S cows, but no difference (P>0.05) occurred between G60 and S cows. The uterine TAMV and PI values did not differ among the three groups (P>0.05). Gene expression of hormone receptors did not differ (P>0.05) between Groups G40 and G60 on Days -1 and 7. Results of this study indicate that a shortened preovulatory follicle phase primarily affects ovarian but not the measured uterine events in cows undergoing synchronization of ovulation.

Effects of human chorionic gonadotropin on luteal blood flow and progesterone secretion in cows and in vitro-microdialyzed corpora lutea.

To check human chorionic gonadotropin (hCG) effects on luteal blood flow (LBF) and progesterone (P(4)) synthesis, six cows received either 3000 IU hCG or saline (NaCl) on Day 7 (Day 1=ovulation) during two estrous cycles. Plasma P(4) and LBF were measured before (0h) and up to 48h after treatment. Luteal blood flow increased by 51% (P<0.05) at 1h after hCG administration and returned to baseline levels thereafter. Plasma P(4) levels were increased from pretreatment levels by 30% at 1h (P=0.05) and 81% at 48h (P=0.02) after hCG treatment. In contrast, NaCl did not cause changes in LBF and P(4) (P>0.05). Additionally, central and peripheral parts of 14 abattoir-derived corpora lutea of the mid-luteal phase (Day 8 to 12) were perfused with Ringer solution in an in vitro microdialysis system, supplemented with 50 or 150 IU/mL hCG for 1h. Application of 50 IU/mL hCG showed no influence on P(4) response (P>0.05) in both central and peripheral parts, whereas 150 IU/mL hCG resulted in an increase of P(4) synthesis (P=0.002) in the central parts only. In vivo, hCG provoked an immediate and long-term rise in P(4) but only a temporary elevation of LBF. Luteal blood flow itself does not seem to be the exclusive cause for an increase in P(4), because the in vitro data clearly showed direct effects of hCG on P(4) secretion. Interestingly, different P(4) secretion patterns could be found between central and peripheral parts of the corpus luteum in both control and hCG perfused corpora lutea.

Relationships between ovarian blood flow and ovarian response to eCG-treatment of dairy cows.

The goal of the present study was to investigate ovarian blood flow and ovarian response in cows undergoing a gonadotropin treatment to induce a superovulatory response, using transrectal colour Doppler sonography. Forty-two cows including 19 cross-bred, 14 German Holstein and 9 German Black Pied cows were examined sonographically before hormonal stimulation on Day 10 of the oestrous cycle, three days after administration of eCG (Day 13) and seven days after artificial insemination (Day 7(p.i.)). After each Doppler examination, blood was collected for determination of total oestrogens (E) and progesterone (P4) in peripheral plasma. The blood flow volume (BFV) and pulsatility index (PI), which is a measure for blood flow resistance, were determined in the ovarian artery, and B-mode sonography was used to count dominant follicles and corpora lutea. Important criteria to assess the ovarian response following the hormonal treatment were the number of follicles >5mm in diameter on Day 13 and the number of corpora lutea on Day 7(p.i.) per cow. The number of follicles ranged from 2 to 61 (mean+/-S.E.M.: 17.5+/-1.7) and corpora lutea from 0 to 50 (mean+/-S.E.M.: 17.0+/-1.6). The BFV increased from 28.4 to 45.0 ml/min between Days 10 and 13 and reached a maximum of 108.5 ml/min on Day 7(p.i.) The PI decreased from 6.25 on Day 10 to 4.70 on Day 13 and to 2.10 on Day 7(p.i.) The BFV and PI on Day 13 did not correlate with the number of follicles (P>0.05). However, on Day 7(p.i.) the number of corpora lutea correlated positively with the BFV (r=0.64; P<0.0001), and an inverse relationship was found for the PI (r=-0.51; P=0.0005). There were no correlations (P>0.05) between the BFV and PI on Day 10 and the number of follicles on Day 13 or the number of corpora lutea on Day 7(p.i.) Results of the present study show that in cows, a hormonal treatment to induce a superovulatory response yielded a marked increase in BFV and a marked decrease in PI in the ovarian artery. However, there was no correlation between BFV and PI in the ovarian arteries before hormonal stimulation and the number of follicles and corpora lutea that developed after stimulation. Thus BFV and PI measured in the ovarian arteries have limited diagnostic value to predict the outcome of a gonadotropin treatment.

Uterine blood flow during the first 3 weeks of pregnancy in dairy cows.

Transrectal color Doppler sonography was used to compare changes in uterine blood flow between cyclic and early pregnant lactating dairy cows. Examinations were carried out in 53 multiparous lactating cows on days 3, 6, 9, 11, 13, 15, 18 and 21 (day 0=estrus). Fourteen cows were examined during the estrous cycle and thirty-nine cows after insemination with frozen/thawed sperm. Uterine blood flow was reflected by the time-averaged maximum velocity (TAMV) and the pulsatility index (PI) in the uterine artery ipsilateral to the corpus luteum. Twenty-one cows that were not pregnant on day 25 were excluded from the study. There was high inter-individual variability in PI- (CV: 22-62%) and TAMV-values (CV: 22-42%) at all examinations. In cyclic cows, TAMV values increased between days 13 and 18 and PI values decreased between days 15 and 21 (P<0.05). In pregnant cows (n=18), TAMV values increased from days 9 to 11 and then decreased to minimal values by day 18 (P<0.05). The PI values decreased between days 3 and 11 and then increased to maximum levels on day 18 (P<0.05). On day 21, both variables reached (P<0.05) values that did not differ (P>0.05) from those on day 11. The changes in TAMV were correlated with estrogen and progesterone concentrations (r=0.69 and -0.70, respectively; P<0.05) in cycling cows, but not in pregnant cows (P>0.05). The PI values did not correlate with steroid hormone levels (P>0.05). Differences in uterine blood flow between cycling and early pregnant cows were observed only on day 18 (P<0.05). The results show that in pregnant cows changes in uterine blood supply can be detected already in the second week after insemination; these changes do not occur in the second week of the cycle.

Doppler sonography of the uterine arteries during a superovulatory regime in cattle. Uterine blood flow in superovulated cattle.

Transrectal color Doppler sonography was used to investigate the effects of a gonadotropin treatment to induce superovulation on uterine blood flow and its relationship with steroid hormone levels, ovarian response and embryo yield in dairy cows. The estrous cycle of 42 cows was synchronized by using PGF(2alpha) during diestrus and GnRH 48 h later (Day 0). Cows were examined on the day of eCG (2750 IU)-administration (Day 10), 3 days after eCG (Day 13) and 7 days after artificial insemination (Day 22), including the determination of total estrogens (E) and progesterone (P(4)) in peripheral plasma. Eight days after insemination (Day 23) the uterus was flushed and the number of total ova and embryos as well as transferable embryos was determined. The ovarian response was defined by the number of follicles>5.0mm in diameter on Day 13 and the number of corpora lutea on Day 22. Uterine blood flow was reflected by the blood flow volume (BFV) and the pulsatility index (PI) in the uterine arteries. Both variables showed distinct changes throughout the superovulatory cycle: BFV increased by 94% and PI decreased by 30% between Days 10 and 22 (P<0.0001). On Day 13, BFV but not PI correlated with follicle numbers (r=0.35; P<0.05); no correlation was found with E and P(4) (P>0.05). On Day 22, BFV correlated positively and PI correlated negatively with the number of corpora lutea (r=0.45 and r=-0.37; P<0.05) and P(4) (r=0.39 and r=-0.30; P<0.05). The number of transferable embryos was solely related to BFV measured on Day 13 (r=0.32; P<0.05). Our results show for the first time that in cows a superovulatory treatment is associated with a marked increase in BFV and a marked decrease in PI in the uterine arteries, concurrent with the development of multiple follicles and corpora lutea. However, transrectal color Doppler sonography of the uterine arteries does not facilitate the prediction of embryo yields following superovulatory treatment.