Reproductive studies of Brahman cattle IV. Luteinizing hormone levels in ovariectomized Brahman, Brahman × Hereford and Hereford cows following a 20 mg dose of Estradiol-17β

Luteinizing Hormone (LH) levels were quantitated by radioimmunoassay (RIA) in six mature, long-term ovariectomized cows each of Brahman (B), Brahman × Hereford (B×H) and Hereford (H) breeding following an in-tramuscular injection of 20 mg of Estradiol-17_β (E) suspended in corn oil. After E administration all cows were bled via coccygeal venipuncture every two hours from 0–8 hours post-injection, every hour from 9–24 hours post-injection, concluding with bleedings every two hours from 26–36 hours post-injection. An LH surge was observed in 5/6 B cows, 6/6 B×H cows and 6/6 H cows. Basal LH levels (mean of first eight data points of each breed type) did not differ (P>.10) between B (3.5 ng/ml), B×H (2.4 ng/ml) and H (2.4 ng/ml). Elapsed time from E injection to peak LH value varied significantly (P<.05) between B, B×H and H, respectively (27.8 hrs, 23.8 hrs, 22.2 hrs). Peak LH values also varied between breed (B, 20.2 ng/ml; B×H, 36.0 ng/ml; H, 113.2 ng/ml: P<.005). The area under the LH curve differed significantly between B, B×H and H (P<.05), however, the duration of the LH surge was not different between breeds; B (13.2 hrs), B×H (16.2 hrs) and H (15.3 hrs). Overall significant period effects (P<.05), breed effects (P<.10) and period × breed interactions (P<.05) were found. In summary, B are less reactive to a 20 mg dose of E than are B×H or H using the following criteria: time to peak LH value, peak LH value and area under the LH curve. These data strongly indicate inherent differences between breeds regarding estrogen feedback mechanisms at the hypophysial-hypothalamic axis.     

Prolactin levels in ovariectomized Brahman, Brahman x Hereford and Hereford cows following a 20 mg dose of estradiol-17beta

Six Brahman (B), six Brahman x Hereford (BxH) and six Hereford (H) chronically ovariectomized cows were injected intramuscularly with 20 mg of estradiol-17beta (E2). The cows were bled via coccygeal vessel puncture immediately before E2 injection, every 2 hr from 0 to 8 hr post-injection, every hr from 9 to 24 hr post-injection and every 2 hr from 26 to 36 hr post-injection. Serum prolactin (PRL) concentrations were quantitated by a validated radioimmunoassay. All cows exhibited a PRL surge following the E2 injection. A PRL surge was defined as a sustained elevation in PRL of at least one standard deviation above the level of PRL before the rise. Nadir levels of PRL prior to the surge did not differ significantly between breeds. Time (hr) to the onset of the E2-induced PRL surge was 5.0, 5.0 and 6.2 in B, BxH and H, respectively (P<.10). Elapsed time (hr) from E2 injection to the PRL peak level varied (P<.01) between B (10.8) and H (17.8) and BxH (11.8) and H. Peak PRL levels (ng/ml) varied (P<.10) between breeds (B, 70.6; BxH, 123.9; H, 49.4). Area under the PRL curve (sq cm) varied (P<.05) between BxH (45.2) and H (24.7) but not between BxH and B (34.3; P>.10) or B and H (P>.10). Duration (hr) of the PRL surge did not differ significantly between breeds (B, 19.3; BxH, 20.5; H, 21.2). Overall, bleeding period effects (P<.01), breed effects (P<.10), and breed x period interactions (P<.01) were found.     

Reproductive studies of Brahman cattle II. Luteinizing hormone patterns in ovariectomized Brahman and Hereford cows before and after injection of gonadotropin releasing hormone

Six Brahman and six Hereford long-term ovariectomized cows were bled via tail vessel at 15 minute intervals for a period of 4 hours. Serum was collected and analyzed via radioimmunoassay (RIA) for luteinizing hormone (LH) to determine if ovariectomized Brahman and Hereford cows have pulsatile LH patterns and if breed of animal influenced LH patterns. Brahman and Hereford ovariectomized cattle did have pulsatile LH patterns. Although the trend was for higher LH levels in Hereford than Brahman cows there were no significant differences in mean serum LH levels, number or magnitude of LH peaks or serum LH pulse height.Six Brahman and five Hereford long-term ovariectomized cows were injected (IM) with a single dose of 500μg of gonadotropin releasing hormone (GnRH). Animals were bled via tail vessel at 15 minute intervals for a period of 6 hours. Serum was assayed for LH via RIA to determine if ovariectomized Brahman and Hereford cows differ in GnRH induced LH response. All animals showed increased serum LH in response to GnRH injection within the first 15 minute collection interval. There were no significant differences in duration of response between ovariectomized Brahman or Hereford cows. Ovariectomized Brahman cows had significantly lower (P<.005) Lh values per period than ovariectomized Hereford cows. It was therefore concluded that ovariectomized Brahman cows were significantly less responsive to GnRH induced LH release than were ovariectomized Hereford cows, although duration and shape of the response curves were not different.     

Luteolytic action of RU486: Modulation of luteal 3β-hydroxysteroid dehydrogenase and 20α-hydroxysteroid dehydrogenase activities in late pregnant rats

The effect of the synthetic antiprogestin RU486 on luteal function in late pregnant rats was studied by evaluating the activities of the enzymes 3β-hydroxysteroid dehydrogenase (3β-HSD) and 20α-hydroxysteroid dehydrogenase (20α-HSD). RU486 (2 mg/kg) administered to rats on day 18 of pregnancy at 10.00 h induced preterm delivery 26.4 ± 0.35 h (n = 8) after treatment. Luteal 3β-HSD activity increased 24 and 34 h after RU486 injection, but a significant and progressive decrease started at 48 h with the maximal reduction 72 h after RU486 treatment, when compared with controls. Serum progesterone concentration decreased at the time of 3β-HSD activity reduction. Interestingly, 20α-HSD activity started to increase 58 h after RU486 injection. The administration of the cyclooxygenase inhibitor, diclofenac (1.3 mg/kg), on days 17–19 of pregnancy to RU486-treated rats, delayed abortion and the duration of delivery, and prevented the decrease in 3β-HSD and the increase in 20α-HSD activities observed 58 h after antiprogesterone treatment. RU486 administered intrabursally (1 μg per ovary) on day 20 (14.00–15.00 h) increased 3β-HSD and decreased 20α-HSD luteal activities at 18.00 h on day 21 of pregnancy, without modifying serum progesterone concentration, when compared with normal pregnant rats. In conclusion, the luteolytic process after preterm delivery induced by RU486 administration in late pregnant rats is characterized by a decrease in luteal 3β-HSD activity and circulating progesterone, which may trigger the increase in luteal 20α-HSD activity. Prostaglandins seems to be involved in the increase of 20α-HSD activity and therefore, in the demise of corpora lutea.     

Pubertal traits and seasonal variation of the sexual activity in Brahman, Hereford and crossbred heifers

Age and weight at puberty and the pattern of ovulatory activity from puberty to 26 mo of age were studied in a total of 55 Brahman (B), Hereford (H), 5/8B-3/8H (B(5)H(3)) and 5/16B-11/16H (B(5)H(11)) heifers. The trial was conducted over two years. Heifers were kept during the whole experimental period on a paddock of native pasture at a stocking rate of 0.5 animals per hectare. Vasectomized bulls were used for estrus detection and blood was collected from the heifers twice weekly to monitor ovulatory activity through progesterone level as determinated by RIA. In addition to age and weight at puberty, the weights and dates at the time of cessation and resumption of ovulatory activity were also considered. There were significant effects (P<0.05 or P<0.01) of genetic group (purebreds vs crossbreds) for age and weight at puberty; the difference being 60 d and 18 kg, respectively, favoring the crossbred groups. After puberty attainment, an anestrous period was detected in all Brahman (118.6 d) and B(5)H(3) (113.4 d) heifers; anestrus was observed in only 14% of B(5)H(11) heifers and for a shorter (43.5 days) period. Hereford heifers did not exhibit interruption of ovulatory activity. In Brahman and B(5)H(3) heifers, the pattern of interruption and resumption of estrus and ovulatory activity correlated closely (0.89 and 0.95 for Brahman and B(5)H(3) heifers, respectively) with the photoperiod curve, suggesting the probable influence of this environmental factor on the regulation of estrous cycle activity in heifers with a higher proportion of Bos indicus genes. Thus, the genetic factor needs be taken into account when establishing programs in reproductive management.     

Resistance of Brahman and Hereford Cattle to African ticks with reference to serum gamma globulin levels and blood composition

Field collections of ticks from two breeds of cattle showed that the common species of ticks wereAmblyomma hebraeum, Rhipicephalus appendiculatus andRhipicephalus evertsi evertsi. The density of these species was higher on Hereford than on Brahman cattle. The results also indicated that the density of immature stages of the above-mentioned ticks is higher on Hereford than on Brahman cattle.A positive correlation was found between the number of ticks on the cattle and the serum gamma globulin levels, an indication of an increase in the production of antibodies. An inverse relationship was found between tick burden and red blood cell count and hemoglobin concentration. Other ablood components, such as basophils, eosinophils and lymphocytes were not affected by the changes in the density of the tick populations. It appears that resistance may have been acquired by the hosts and that Brahman cattle may acquire resistance to a higher degree than Herefords.     

Cortisol and prolactin concentrations during three different seasons in relocated Brahman and Hereford bulls

The objective of this study was to evaluate seasonal changes of cortisol and prolactin (PRL) concentrations in Brahman and Hereford bulls moved to locations that differ in geographical and environmental conditions. Postpubertal Hereford bulls from Montana (n = 15) and Nebraska (n = 15) and Brahman bulls from Texas (n = 18) were located in or relocated to Montana, Nebraska or Texas so that each location had 5 Montana Herefords, 5 Nebraska Herefords and 6 Texas Brahman bulls. Blood samples were collected at 20-minute intervals for 8 hours in November (Fall 1), April (Spring) and November (Fall 2) of the next year. These dates corresponded to 6, 12 and 18 months, respectively, after relocation in May of the first year. Cortisol concentrations were higher (P<0.05) in Fall 1 than in Fall 2 and were higher (P<0.05) for bulls in Montana than for bulls in Texas. The decrease in cortisol concentrations from Fall 1 to Fall 2 was negatively related (P<0.05) to age and weight. There was a three-way interaction (P<0.05) of breed-type origin, location and season for PRL concentrations. Seasonal patterns of PRL concentrations differed between relocated Texas Brahman and Hereford bulls, and patterns for relocated bulls differed from those of the nonrelocated bulls. Seasonal patterns of PRL were influenced to a greater extent by relocation in Texas Brahman bulls than in Hereford bulls.     

Expression of ghrelin and the ghrelin receptor in different stages of porcine corpus luteum development and the inhibitory effects of ghrelin on progesterone secretion, 3β-hydroxysteroid dehydrogenase (3β-honestly significant difference (HSD)) activity and protein expression

Recent studies have suggested that ghrelin plays a direct role in controlling female reproduction. The aim of the present study was to investigate the mRNA and protein expression of ghrelin and its receptor (via real time PCR, Western blot and immunohistochemistry analysis, respectively) in porcine corpora lutea (CL) collected during early (CL1: 1-2 days after ovulation), middle (CL2: 7-10 after ovulation), and late luteal phase (CL3: 13-15 after ovulation). Ghrelin expression and concentration of both acylated and unacylated forms of ghrelin significantly increased during CL development. Immunohistochemistry analysis shown localization of ghrelin protein in the cytoplasm of large luteal cells. No changes in the expression of the ghrelin receptor were observed. Direct in vitro effects of ghrelin on progesterone (P4) secretion and 3-beta-hydroxysteroid dehydrogenase (3β-honestly significant difference (HSD)) activity, which were measured by the conversion of pregnenolone (P5) to P4, and 3β-HSD protein expression were then analyzed. To assess 3β-HSD activities, mature luteal cells were first cultured for 24 h with ghrelin at 100, 250, 500 and 1000 pg/mL with P5, or with aminoglutethimide (AMG). AMG is an inhibitor of CYP11A1-mediated hydroxylation; an addition of AMG and P5 enabled P4 production to serve as an index of 3β-HSD activity. Inhibitory effects of ghrelin on P4 secretion, 3β-HSD activity and protein expression were observed. In conclusion, the presence of ghrelin and its receptor in porcine corpora lutea and the direct inhibitory effects of ghrelin on luteal P4 secretion and 3β-HSD suggest potential auto/paracrine regulation by ghrelin in the luteal phase of ovary function.     

Effect of alfaprostol, lasalocid, and once-daily suckling on postpartum interval in Brahman and Brahman crossbred cattle

Brahman cows (n = 49) and primiparous heifers (n = 11), Brahman x Hereford primiparous F1 heifers (n = 86) and Simmental x Brahman primiparous F1 heifers (n = 13) were randomly allotted by breed, age and date of calving to one of eight treatment groups: 1) control; 2) once-daily suckling; 3) lasalocid (200 mg/hd/d); 4) alfaprostol (5 mg intermuscular injections on Days 21 and 32 post partum); 5) lasalocid + once-daily suckling; 6) alfaprostol + once daily suckling; 7) alfaprostol + lasalocid; 8) alfaprostol + lasalocid + once daily suckling. All animals received 2.3 kg/hd/d of a concentrate (6 corn : 1 cottonseed meal) and lasalocid was mixed and fed in the concentrate. Body weights and condition scores were taken on Day 1 post partum and every 28 d thereafter. All animals were maintained with sterile marker bulls with Brahman and Simmental x Brahman cattle artificially inseminated at first estrus. Blood samples were collected at weekly intervals starting on Day 21 post partum until estrus and at nine to twelve days post estrus when the ovaries were palpated for corpora lutea. After the first postpartum estrus with a corpora lutea, cows were placed with fertile bulls. Mean serum progesterone concentrations were below 0.5 ng/ml prior to treatment. Calf weight gains to 90 d were not affected by age (P > 0.10) but were lower in the once-daily suckling group (P < 0.05). Treatment did not affect cow weight or condition score (P > 0.10). Cows had a shorter postpartum interval (P < 0.0001) than heifers. Once-daily suckling shortened postpartum interval (P < 0.0001) and positively influenced the cumulative frequency of return to estrus by 40 d post partum (P < 0.02). Alfaprostol did not affect postpartum interval (P > 0.10) but did increase the cumulative frequency of return to estrus by 90 d post partum (P < 0.03). Lasalocid did not affect postpartum interval or cumulative frequency of return to estrus (P > 0.10). Both once-daily suckling and alfaprostol were effective in increasing the numbers of animals inseminated by 90 d post partum. The once-daily suckling + alfaprostol treatment resulted in the shortest postpartum interval.     

Influence of Synchro-Mate-B and breed composition on estrous response and pregnancy rate in spring- and fall-bred Brahman crossbred beef cows

Estrous response, pregnancy rate, and the relationship between palpable corpora lutea (CL) and plasma progesterone concentrations were studied during the spring (n = 149) and fall (n = 146) breeding seasons using postpartum, crossbred beef cows consisting of 0-Brahman, 1 4 - Brahman , or 1 2 Brahman breeding. At the start of each breeding season, a jugular blood sample was collected for progesterone analysis; each cow was palpated for the presence of a CL and randomly alloted within breed to a non-treated control or Synchro-Mate-B (SMB) treatment. Fewer (P<0.01) fall than spring bred cows exhibited estrus within the first 21 d of breeding. Within 48 h of implant removal, 84 and 64% (spring and fall, respectively) of synchronized cows exhibited estrus. In both seasons, more SMB than control cows became pregnant by 7 d of breeding. Percentage of Brahman breeding did not influence estrous response or pregnancy rate. On the basis of 292 evaluations, palpation per rectum correctly assessed ovarian status 74% of the time. These data indicate that although season can influence response. SMB can be used effectively to synchronize Brahman crossbred cows. In addition, it was found that there is a close relationship between palpation per rectum and plasma concentrations of progesterone.     

An ultrasonographic study of luteal function in breeds of sheep with different ovulation rates

Development and demise of luteal structures were monitored using daily transrectal ultrasonography in 2 breeds of sheep differing in ovulation rates (nonprolific Western white-faced cross-bred, n = 12 and prolific pure-bred Finn sheep, n = 7), during 1 estrous cycle in the mid-breeding season. Jugular blood samples were collected once a day for radioimmunoassay (RIA) of progesterone. The mean diameter of ovulatory follicles was higher in Western white-faced than in Finn ewes (6.4 +/- 0.2 and 5.3 +/- 0.2 mm, respectively; P < 0.001). The mean volume of luteal structures was higher (P < 0.05) in Western white-faced compared with Finn sheep from Days 5 to 15 of the cycle (Day 0 = day of ovulation). This accounted for the higher (P < 0.05) total luteal volumes recorded in Western white-faced ewes on Day 7 and from Days 11 to 15, despite the higher ovulation rate in Finn ewes (2.7 +/- 0.3 and 1.7 +/- 0.2, respectively; P < 0.05). Mean serum progesterone concentrations were higher (P < 0.05) in Western white-faced than in Finn ewes from Days 4 to 14. Daily total luteal volumes were positively correlated with daily serum progesterone concentrations throughout the cycle in Finn sheep (r > or = 0.40, P < 0.02), and during luteal growth and regression (r > 0.60, P < or = 0.00001) but not during mid-cycle in white-faced ewes (r = 0.16; P = 0.22). During the growth of the corpora lutea (CL), luteal tissue volume increased faster (P < 0.05) than serum progesterone concentrations in both breeds of sheep. During luteolysis, the decrease in luteal volumes parallelled that in serum progesterone concentrations in Finn (P = 0.11) but not in Western white-faced ewes, where luteal volumes decreased more slowly (P = 0.02) in relation to progesterone secretion. Increased ovulation rate in prolific Finn ewes resulted in more but smaller CL, and lower serum progesterone levels compared with nonprolific Western white-faced ewes. We conclude that breed-specific mechanisms exist to control the formation of luteal tissue and progesterone secretion in cyclic ewes differing in prolificacy. The mechanisms may involve ovulation of Graafian follicles at different sizes and inhibitory paracrine effects of CL on co-existing CL.     

Plasma progesterone levels during the estrous cycle of Holstein and Brahman cows, Carora type and cross-bred heifers

Daily plasma progesterone (P(4)) was determined during one estrous cycle of 19 cows and 18 heifers of four different breeds: Holstein (H), Brahman (B), Carora-type (C) and crossbred (CB) females. Estrus detection was made by visual observation and using a teaser bull with a chin-ball marker. The P(4) profiles showed no differences among groups. In Group 1 (H), P(4) levels ranged from 0.5 ng/ml plasma on the day of estrus (Day 0) to 5.1 ng/ml at the luteal phase peak (Day 13). In Group 2 (B), P(4) levels ranged from 0.5 ng/ml on Day 0 to 9.2 ng/ml on Day 13. In Groups 3 (C) and 4 (CB), P(4) levels ranged from 0.5 ng/ml, on Day 0, to 13.7 ng/ml on Day 12 and 8.8 ng/ml on Day 13. These last two groups were moved to the same location and then compared. It was found that P(4) concentrations were significantly higher (P < 0.025) in Group 3 between Days 7 and 14 of the estrous cycle. In all groups, P(4) levels were lower than 1 ng/ml one day before the next estrus, and levels of 0.4, 0.5, 0.4 and 0.4 ng/ml were obtained the day of estrus in Groups 1 to 4, respectively. Results indicated that the pattern of P(4) for each one of the groups was similar to those reported by other investigators.     

Animal and temporal effects on ovarian follicular dynamics in Brahman heifers

The aims of the current study were to determine if the pattern of ovarian follicular growth and development in Bos indicus heifers is different to that reported in Bos taurus breeds, and to examine the factors that determine which dominant follicle will ovulate. In addition, the extent to which variation in follicular dynamics is attributable to variation between animals and over time was evaluated. The ovaries of 17 Brahman heifers were examined daily by transrectal ultrasonography using a 7.5 MHz transducer for a total of 117 interovulatory intervals over a period of 10 months. Size and position of individual follicles ⪖5 mm in diameter, and size of corpora lutea (CL) were recorded. Circulating progesterone concentrations were determined from plasma samples obtained twice weekly. Although size of dominant follicles and CL within the ovaries of Bos indicus heifers were smaller than reported for Bos taurus breeds, the overall patterns of dominant follicle growth were similar. There were significant correlations between number of dominant follicles occurring prior to ovulation and time of appearance of the second dominant follicle, duration of detection of CL and size of the ovulatory follicle in the preceding oestrous cycle (P < 0.05). There were significant animal effects on a number of ovarian characteristics including number of dominant follicles per oestrous cycle (P < 0.001), with one heifer having four dominant follicles in more than a third of oestrous cycles observed. In addition, changes in daylength over the 10 month period were related to changes in duration of the interovulatory interval, persistence and maximum diameter of CL and size of ovulatory follicles. Liveweight change over the same period was related to changes in maximum diameter of the first dominant follicle.     

Type 2 11β-hydroxysteroid dehydrogenase in foetal and adult life

Two isoforms of 11β-hydroxysteroid dehydrogenase (11β-HSD) catalyse the interconversion of active cortisol to inactive cortisone; 11β-HSD1 is a low affinity, NADP(H)-dependent dehydrogenase/oxo-reductase, and 11β-HSD2 a high affinity, NAD-dependent dehydrogenase. Because of the importance of 11β-HSD in regulating corticosteroid hormone action, we have analysed the distribution of the 11β-HSD isoforms in human adult and foetal tissues (including placenta), and, in addition have performed a series of substrate specificity studies on the novel, kidney 11β-HSD2 isoform. Using an RT-PCR approach, we failed to detect 11β-HSD1 mRNA in any human mid-gestational foetal tissues. In contrast 11β-HSD2 mRNA was present in foetal lung, adrenal, colon and kidney. In adult tissues 11β-HSD2 gene expression was confined to the mineralocorticoid target tissues, kidney and colon, whilst 11β-HSD1 was expressed predominantly in glucocorticoid target tissues, liver, lung, pituitary and cerebellum. In human kidney homogenates, 11-hydroxylated progesterone derivatives, glycyrrhetinic acid, corticosterone and the “end products” cortisone and 11-dehydrocorticosterone were potent inhibitors of the NAD-dependent conversion of cortisol to cortisone. Finally high levels of 11β-HSD2 mRNA and activity were observed in term placentae, which correlated positively with foetal weight. The tissue-specific distribution of the 11β-HSD isoforms is in keeping with their differential roles, 11β-HSD1 regulating glucocorticoid hormone action and 11β-HSD2 mineralocorticoid hormone action. The correlation of 11β-HSD2 activity in the placenta with foetal weight suggests, in addition, a crucial role for this enzyme in foetal development, possibly in mediating ontogeny of the foetal hypothalamo-pituitary-adrenal axis.     

Relationship of oestrus synchronization method, circulating hormones, luteinizing hormone and prostaglandin F-2 alpha receptors and luteal progesterone concentration to premature luteal regression in superovulated sheep

Ewes were treated with exogenous follicle-stimulating hormone (FSH) and oestrus was synchronized using either a dual prostaglandin F-2 alpha (PGF-2 alpha) injection regimen or pessaries impregnated with medroxy progesterone acetate (MAP). Natural cycling ewes served as controls. After oestrus or AI (Day 0), corpora lutea (CL) were enucleated surgically from the left and right ovaries on Days 3 and 6, respectively. The incidence of premature luteolysis was related (P less than 0.05) to PGF-2 alpha treatment and occurred in 7 of 8 ewes compared with 0 of 4 controls and 1 of 8 MAP-exposed females. Sheep with regressing CL had lower circulating and intraluteal progesterone concentrations and fewer total and small dissociated luteal cells on Day 3 than gonadotrophin-treated counterparts with normal CL. Progesterone concentration in the serum and luteal tissue was higher (P less than 0.05) in gonadotrophin-treated ewes with normal CL than in the controls; but luteinizing hormone (LH) receptors/cell were not different on Days 3 and 6. There were no apparent differences in the temporal patterns of circulating oestradiol-17 beta, FSH and LH. High progesterone in gonadotrophin-treated ewes with normal CL coincided with an increase in total luteal mass and numbers of cells, which were primarily reflected in more small luteal cells than in control ewes. Gonadotrophin-treated ewes with regressing CL on Day 3 tended (P less than 0.10) to have fewer small luteal cells and fewer (P less than 0.05) low-affinity PGF-2 alpha binding sites than sheep with normal CL. By Day 6, luteal integrity and cell viability was absent in ewes with prematurely regressed CL. These data demonstrate that (i) the incidence of premature luteal regression is highly correlated with the use of PGF-2 alpha; (ii) this abnormal luteal tissue is functionally competent for 2-3 days after ovulation, but deteriorates rapidly thereafter and (iii) luteal-dysfunctioning ewes experience a reduction in numbers of small luteal cells without a significant change in luteal mass by Day 3 and, overall, have fewer low-affinity PGF-2 alpha binding sites.     

Demonstration of luteotrophic responses of human recombinant gamma interferon in porcine corpora lutea using an in-vivo microdialysis system

Conceptuses from several mammalian species prior to implantation secrete proteins belonging to the family of interferons. The main species of interferons known to be secreted by the pig blastocyst is interferon gamma (IFNγ), the precise role of which is unclear. We decided to explore its effects on corpus luteum (CL) function using the novel microdialysis technique in vivo. Six cycling miniature pigs were monitored for estrus by daily plasma progesterone analysis and visual symptoms. On day nine of the cycle (day zero being the day of ovulation) the animals underwent surgery, and microdialysis tubing (vitafiber, Amicon U.S.A, cut off mol. wt. 1 million) were implanted in 17 corpora lutea. The inlets and outlets of all tubings were exteriorized and the entry and exit points of tubings in the CLs sealed with tissue glue. The afferent extension tubings were connected to a fraction collector and the system was continuously flushed with Ringer at a flow rate of 2.4 ml/h. After an initial flushout phase of 8 h, fractions were collected every half hour over 3 days. On days 10, 11 and 12 post estrus 12 CLs were stimulated for 4 h with 10⁻⁷ M, 2 × 10⁻⁷ M and 4 × 10⁻⁷ M human recombinant IFNγ (Pharma Biotechnologie) respectively. Simultaneously, fractions were also collected from the remaining five unstimulated corpora lutea which served as controls. Progesterone concentrations in the dialysates were estimated by a sensitive enzymeimmunoassay (EIA). A significant increase (P < 0.01) in progesterone release was observed in all 3 days following stimulation. The progesterone increase was more marked on the first day of stimulation (1 × 10⁻⁷ M) with the hormone levels rising further even after the end of stimulation. The overall increase in progesterone concentration was 2-fold on day 10 in comparison to 15–30% on subsequent days even though IFN concentrations for stimulation were 2- and 4-fold higher. In the unstimulated CLs, a gradual decline (P < 0.01) in progesterone levels were observed over days. In conclusion, these data provide evidence that the early conceptus signals its presence by way of IFNγ to maintain the CL in pigs.     

Effect of prostaglandin F2α on the hCG-stimulated progesterone production by human corpora lutea

The effect of prostaglandin PGF_2α on the hCG stimulated and basal progesterone production by human corpora lutea was examined . hCG (40 i.u./ml) stimulated progesterone formation in corpora lutea of early (days 16–19 of a normal 28 day cycle), mid (days 20–22) and late (days 23–27) luteal phases. This stimulation was inhibited by PGF_2α (10 μg/ml) in corpora lutea of mid and late luteal phases. PGF_2α alone did not show a consistent effect on basal progesterone production. The inhibition of hCG stimulated progesterone production by PGF_2α at times corresponding to luteolysis indicates a role for that prostaglandin in the process of luteolysis in the human corpus luteum.     

Seasonal variations in characteristics of estrous cycles in pubertal Brahman heifers

A group of pubertal Brahman heifers (n = 16) was monitored from October through March to investigate the seasonal changes in estrous cyclicity. The heifers had a mean age of 16.7 +/- 0.3 mo at the initiation of the experiment. They were kept on pasture with vasectomized marker bulls. Supplemental feed to meet NRC requirements was provided. Estrus occurrence was checked once a day and blood samples were taken weekly by tail venipuncture from heifers that had been in estrus 7 to 14 d earlier. Samples were processed to yield serum and were assayed for serum progesterone by radioimmunoassay. A high proportion of heifers (88%) had abnormalities such as estrus without the formation of a functional corpus luteum (CL) or anestrus, with a distribution of the two abnormalities as follows: October 0 and 0, November 50 and 25, December 0 and 50, January 0 and 50, February 18 and 31 and March 0 and 7%, respectively. Mean serum progesterone concentrations during the luteal phase differed by month: 4.26, 1.50, 3.25, 2.27, 2.65 and 3.70 ng/ml for October, November, December, January, February and March, respectively (P<0.001). Heifers that went into anestrus had lower mean serum progesterone concentrations than heifers that had regular estrous cycles throughout the study period (1.35 vs 2.22 ng/ml; P<0.0005). The months with the shortest daylengths (December and January) had the highest incidence of anestrus. Transitional periods (November and February) seemed to occur before and after the months with the highest occurrence of anestrus. During this transitional period a high incidence of estrus without the formation of a functional CL was detected. Serum progesterone concentrations were lower in all heifers during the months with high occurrence of abnormal estrous cycles.     

In vivo intra-luteal implants of prostaglandin (PG) E(1) or E(2) (PGE(1), PGE(2)) prevent luteolysis in cows. I. Luteal weight, circulating progesterone, mRNA for luteal luteinizing hormone (LH) receptor, and occupied and unoccupied luteal receptors for LH

Previously, it was reported that chronic intra-uterine infusion of PGE(1) or PGE(2) every four hours inhibited luteolysis in ewes. However, estradiol-17β or PGE(2) given intra-uterine every 8h did not inhibit luteolysis in heifers, but infusion of estradiol+PGE(2) inhibited luteolysis in heifers. The objective of this experiment was to determine whether and how intra-luteal implants containing PGE(1) or PGE(2) prevent luteolysis in Angus or Brahman cows. On day-13 post-estrus, Angus cows received no intra-luteal implant and corpora lutea were retrieved or Angus and Brahman cows received intra-luteal silastic implants containing Vehicle, PGE(1), or PGE(2) and corpora lutea were retrieved on day-19. Coccygeal blood was collected daily for analysis for progesterone. Breed did not influence the effect of PGE(1) or PGE(2) on luteal mRNA for LH receptors or unoccupied or occupied luteal LH receptors did not differ (P>0.05) so the data were pooled. Luteal weights of Vehicle-treated Angus or Brahman cows from days-13-19 were lower (P<0.05) than those treated with intra-luteal implants containing PGE(1) or PGE(2). Day-13 Angus luteal weights were heavier (P<0.05) than Vehicle-treated Angus cows on day-19 and luteal weights of day-13 corpora lutea were similar (P>0.05) to Angus cows on day-19 treated with intra-luteal implants containing PGE(1) or PGE(2). Profiles of circulating progesterone in Angus or Brahman cows treated with intra-luteal implants containing PGE(1) or PGE(2) differed (P<0.05) from controls, but profiles of progesterone did not differ (P>0.05) between breeds or between cows treated with intra-luteal implants containing PGE(1) or PGE(2). Intra-luteal implants containing PGE(1) or PGE(2) prevented (P<0.05) loss of luteal mRNA for LH receptors and unoccupied or occupied receptors for LH compared to controls. It is concluded that PGE(1) or PGE(2) alone delays luteolysis regardless of breed. We also conclude that either PGE(1) or PGE(2) prevented luteolysis in cows by up-regulating expression of mRNA for LH receptors and by preventing loss of unoccupied and occupied LH receptors in luteal tissue.