Effect of xylazine on interovulatory interval and serum progesterone concentrations in the horse mare

The objective of this study was to determine the effect of the alpha(2)-adrenergic agonist, xylazine, on interovulatory interval and progesterone concentrations in the horse mare. Mares were assigned to one of four treatments: Group 1 (controls) received an intramuscular injection (i.m.) of 5 cc saline (n=6), Group 2 received 10 mg prostaglandin F(2alpha) (PGF(2alpha)) i.m. (n=5), Group 3 received 500 mg xylazine i.m. (n=6) and group 4 received an intravenous injection (i.v) of 350 mg xylazine (n=6). Treatment was administered on Day 10 of the estrous cycle (Day 0 = Day of detected ovulation). There was no difference in length of interovulatory interval between PGF(2alpha)-treated mares and control mares (mean +/- SEM; 18.8 +/- 1.0 versus 21.7 +/- 1.6 d). When compared with either xylazine-treated group, PGF(2alpha)-treated mares had a shorter interovulatory interval (18.3 +/- 1.0 d versus 22.2 +/- 0.6 and 22.8 +/- 1.3 d, respectively; P < 0.05). There was no difference in the length of interovulatory interval between control mares and either xylazine-treated group. At the time of treatment all mares had progesterone concentrations > 10 ng/ml, therefore the onset of luteolysis was defined as the day of the estrous cycle when progesterone concentrations decreased below 10 ng/ml. In PGF(2alpha)-treated mares, this event occurred earlier than in any other group (Day 11.2 +/- 0.2 of the estrous cycle versus 16.0 +/- 1.3 for control, Day 15.7 +/- 0.2 for Group 3 and Day 15.2 +/- 0.6 for Group 4; P < 0.002). It was concluded that a single treatment with xylazine, either by an intramuscular or intravenous route, had no significant effect on interovulatory interval or progesterone concentrations in horse mares.     

Comparison of a solid-phase,no-extraction radioimmunoassay for progesterone with an extraction assay for monitoring luteal function in the mare,bitch, and cow

Equine, canine, and bovine plasma samples were assayed for progesterone (P₄) using extracted plasma in a liquid-phase assay and unextracted plasma in a solid-phase ¹²⁵I direct assay developed for human serum. The direct assay was also used to monitor P₄ levels in defatted milk. Results indicated that the direct-assay method was as reliable as the extraction assay for monitoring changes in plasma P₄ during the estrous cycle and early pregnancy. The regression equation specifying the relationship for the two methods was exponential. In this study, correlation coefficients from data subsets ranged from 0.93 to 0.99. The direct assay for P₄ did result in higher values than the extraction assay in plasma from diestrous animals. The use of diluted standards with stripped plasma from the species being assayed gave values that correspond more closely to the extraction assay. The comparisons in this study indicate that the direct radioimmunoassay (RIA) method offered a convenient, reliable method for monitoring luteal function in the species that were evaluated.     

Longitudinal studies of luteal function by salivary progesterone determinations

A 'normal range' for salivary progesterone concentrations has been established using data derived from women who were menstruating regularly and in whom dating of the cycle by accepted criteria was possible. Since these values are in agreement with those in the first 9 days of conception cycles and with those in cycles in which ovulation was confirmed by ultrasonography, they provide a reliable index of progesterone output compatible with fertility. Measurement of daily salivary progesterone values in subfertile women for time spans exceeding 3 months allowed accurate assessment of base-line ovarian activity and of the response to ovulation-induction therapy. Salivary sampling, by allowing collection of frequent samples with a minimum of time, stress and inconvenience, is ideally suited to longitudinal studies of ovarian activity. This sampling regimen is also applicable to the monitoring of progesterone output throughout pregnancy.     

Control of follicular development and luteal function in the mare: effects of a GnRH antagonist

Control of the equine estrous cycle was studied by suppressing gonadotropin secretion by administration of a GnRH antagonist to cyclic pony mares. Four mares received vehicle (control cycle) or a GnRH antagonist, Antarelix (100 microg/kg) on Day 8 of diestrus, and blood samples were collected at 15-min intervals from 0 to 16 h, 24 to 36 h, and daily until the next ovulation. Ovarian activity was monitored by transrectal ultrasonography, and measurement of plasma concentrations of progesterone and estradiol. Antagonist treatment eliminated large diestrous pulses of LH. Progesterone concentrations had fallen significantly in all mares by the day after treatment and, in three of the four mares, remained low until luteolysis. However timing of luteolysis (ie., progesterone concentrations <1 ng/mL) was not affected by antagonist treatment. The preovulatory surges of estradiol and LH were significantly delayed in the treatment cycle, as was the appearance of a preovulatory follicle >30 mm. Cycle length was significantly longer during the treatment than the control cycle. These results show that treatment of diestrous mares with a GnRH antagonist attenuated progesterone secretion, indicating a role for LH in control of CL function in the mare, and delayed ovulation presumably because of lack of gonadotropic support.     

Body temperature fluctuations in the periparturient horse mare

The objective of this study was to determine the pattern of mare deep body temperature fluctuations associated with parturition using biotelemetry. A radio transmitter was implanted in one flank in each of six mares. Telemetered data were received by a pair of antennae placed at right angles in a 3.3 x 6.6-m stall and stored on a computer hard disk. Hourly temperature data were recorded for the period of -168 through 168 hours post partum. A decrease of 0.76 degrees C in body temperature began at 4 hours prior to parturition (P < 0.1) then decreased rapidly between the 3 hours prior to and the time of parturition (Time 0). The lowest mean body temperature recorded was at the time of parturition (36.58 +/- 0.16 degrees C; P < 0.001). A supranormal increase in mean body temperature began one hour post partum, peaked at 38.02 +/- 0.08 degrees C and remained elevated for 48 hours post partum until gradually decreasing to the level of the prepartum mean by 106 hours.     

Salivary progesterone and luteal function in two low-fertility populations of Northeast Zaire

30 Ituri women (Zaire) -- 14 Efe and 16 Balese -- were targeted as subjects in this study designed to verify that, under field conditions, the salivary steroid method can reliably discern follicular and luteal levels of progesterone in normal menstrual cycles and to examine the hypothesis that infertility among these women is due to tubal factors. Findings of normal ovulatory function in fertile women would support the hypothesis; findings of abnormal gonadal function might either indicate a chronic endocrine imbalance or the short-term effects of nutritional and other stressors. All potential subjects ranged in age between 20-35 years, were involved in stable conjugal unions, had no nursing children, and reported either no births or none within the last 6 years. 25 women completed the study. The Boston field control subjects consisted of 18 volunteers ranging in age between 18-43 years. All reported a history of regular menstrual cycles and were nither using oral steroid contraceptives nor engaged in a regular exercise program. The African women had significantly lower luteal progesterone levels than did the Boston controls. Additionally, a significantly higher proportion of the African women failed to demonstrate clear luteal activity, suggesting that a higher rate of anovulation contributed to the low average luteal progesterone levels. The composite-cross-sectional profile for the Ituri Forest women suggests that the average luteal phase for this group was shorter than for the Boston controls. Further investigations need to determine whether gonadal dysfunction such as observed in this study is a regular feature of the reproductive physiology of women in the Ituri Forest, or whether it emerges only in periods of food shortage and significant weight loss. Gonorrhea may be the major cause of infertility in the Ituri region, but it is likely that other factors directly affecting gonadal function contributed to the observed pattern of low fertility. Clearly, the study illustrates the potential usefulness of salivary steroid assays.     

Correlation between homologous and heterologous enzyme immunoassays for progesterone determinations in milk from cows

Two enzymeimmunoassays, homologous and heterologous, have been used for measuring progesterone in unextracted bovine milk using HRP as the enzyme label. Antibody raised by immunization of the rabbit against 11 alpha-hemisuccinate-BSA was used for the homologous system (EIA-11 alpha) and 7 alpha-carboxyethylthioether-BSA (EIA-7 alpha) for the heterologous. The progesterone derivatives used for the enzyme-hormone conjugates were 11 alpha-hemisuccinate and 6 beta-OH-hemisuccinate respectively. Milk progesterone in 60 samples measured by EIA-11 alpha and EIA-7 alpha were highly correlated (r = 0.93). Both systems were further compared with a conventional direct progesterone radioimmunoassay (RIA) in regular use for the same samples showing a good correlation. The sensitivity estimated was much higher in the EIA-7 alpha (0.5 pg/well) than in the EIA-11 alpha (32 pg/well).     

Telemetric monitoring of body temperature in the horse mare

The objective of this study was to develop a radiotelemetric system capable of frequent monitoring of mare body temperature. A radio transmitter was implanted in the flank of each of four mares. Telemetered data were received by a pair of antennae placed at right angles in a 3.3 x 6.6-m stall and stored on a computer hard disk. The data were recorded every 5 minutes except when mares were out of the stall for a 1- to 2- hour exercise period. No effect of environmental temperature, ranging from 5 degrees C to 30 degrees C, on mare body temperature was apparent. The radiotelemetric system used in this study was effective for frequent measurement of mare body temperature.     

Direct radioimmunoassay of progesterone in mare plasma

A rapid and low cost radioimmunologic procedure for progesterone assay in mare plasma is proposed. Radioimmunoassay is performed directly on 10 microliter of unextracted plasma. Free progesterone is adsorbed on dextran-charcoal, then the aqueous phase is decanted and extracted by 1 ml of scintillation fluid. Counting is performed directly on this two-phase system. Results are comparable to those obtained with radioimmunoassays using extracted plasma.     

Effect of periovulatory prostaglandin F2alpha on pregnancy rates and luteal function in the mare.

The objective of this study was to determine whether periovulatory treatments with PGF2alpha affects the development of the CL, and whether the treatment was detrimental to the establishment of pregnancy. Reproductively sound mares were assigned randomly to one of the following treatment groups during consecutive estrus cycles: 1. 3,000 IU hCG within 24 hours before artificial insemination and 500 microg cloprostenol (PGF2alpha analogue) on Days 0, 1, and 2 after ovulation (n=8), 2. 2 mL sterile water injection within 24 hours before artificial insemination and 500 microg cloprostenol on Days 0, 1, and 2 after ovulation (n=8); 3. 3,000 IU hCG within 24 hours before artificial insemination and 500 microg cloprostenol on Day 2 after ovulation (n=8); or 4. 3,000 IU hCG within 24 hours before artificial insemination and 2 mL of sterile water on Days 0, 1, and 2 after ovulation (controls; n=8). Blood samples were collected from the jugular vein on Days 0, 1, 2, 5, 8, 11, and 14 after ovulation. Plasma progesterone concentrations were determined by the use of a solid phase 125I radioimmunoassay. All mares were examined for pregnancy by the use of transrectal ultrasonography at 14 days after ovulation. Mares in Group 1 and 2 had lower plasma progesterone concentrations at Day 2 and 5, compared to mares in the control group (P < 0.001). No difference was detected between group 1 and 2. Plasma progesterone concentrations in group 3 were similar to the control group until the day of treatment, but decreased after treatment and were significantly lower than the control group at Day 5 (P < 0.001). Plasma progesterone concentrations increased in all treatment groups after Day 5, and were comparable among all groups at Day 14 after ovulation. Cloprostenol treatment had a significant effect on pregnancy rates (P < 0.01). The pregnancy rate was 12.5% in Group 1, 25% in Group 2, 38% in Group 3, and 62.5% in Group 4. It was concluded that periovulatory treatment with PGF2alpha has a detrimental effect on early luteal function and pregnancy.     

Monitoring luteal function in the lion-tailed macaque (Macaca silenus) through urinary progesterone metabolite measurements

A direct radioimmunoassay for measuring urinary 20-hydroxyprogesterone cross-reactivity to monitor and assess luteal function and detect pregnancy in the lion-tailed macaque (Macaca silenus) is described. Urine samples were collected daily during ten nonconceptive and five conceptive ovarian cycles of five dult female lion-tailed macaques. Urine was analyzed for concentrations of 20α-hydroxypro-gesterone cross-reactivity, estrone conjugates, and creatinine. The strength of the luteal phase in normal nonconceptive cycles (n = 8) is characterized by a maximum sevenfold increase (day 9) in mean 20α-hydroxyprogesterone cross-reactivity over follicular phase levels; the duration, by a 13-day sustained elevation of mean 20α-hydroxyprogesterone cross-reactivity levels. Pregnancy is detectable from 20α-hydroxyprogesterone cross-reactivity values approximately 20 days after the periovulatory estrone conjugate peak (n = 4). Apparent anovulation (n = 1), extended follicular phase (n = 1), and early abortion (n = 1) also are detectable using 20α-hydroxyprogesterone cross-reactivity measurements.     

Evidence that progesterone may influence subsequent luteal function in the ewe by modulating preovulatory follicle development

Ovulation was induced in seasonally anoestrous ewes by repeated 2-h injections of 250 ng Gn-RH, after 12 days (Group 1, N = 7; Group 2, N = 8), 2 days (Group 3, N = 8) or no (Group 4, N = 7) progesterone pretreatment. A preovulatory LH peak occurred spontaneously at a mean (+/- s.e.m.) time of 43.1 +/- 2.0 h, 38.5 +/- 3.1 h and 26.8 +/- 1.7 h after the start of Gn-RH treatment in Groups 1, 3 and 4 respectively, and was artificially induced in ewes in Group 2, after 24 h of treatment, by a single i.v. injection of 150 micrograms Gn-RH. Normal luteal function occurred in all progesterone-pretreated ewes, but in only 1/7 animals not treated with progesterone. These results demonstrate that, although normal luteal function in progesterone-primed ewes induced to ovulate with repeated injections of low doses of Gn-RH is associated with a delayed preovulatory LH peak, it is not this extended period of follicle development which is responsible for functional competence of the resultant corpus luteum. Since as little as 2 days of exposure to elevated plasma progesterone concentrations is effective, it is suggested that progesterone may act directly on the preovulatory follice.     

Luciferin derivatives in bioluminescence-enhanced enzyme immunoassays

Ultrasensitive bioluminescence immunoassays for the determination of peptides and proteins (illustrated with human urinary kallikrein, bradykinin and the determination of human urinary kallikrein antibody titres) have been developed. The usable ranges of the standard curves are from 5 pg to 5000 pg per litre. The relative intra-assay coefficients of variation of the tests were between 2% and 6%, and the inter-assay coefficients of variation between 4% and 12%.     

Influence of exogenous progesterone on early embryonic development in the mare

The influence of exogenous progesterone on the development of equine oviductal embryos was determined based upon the recovery of Day-7 uterine blastocysts from treated mares (n=13) that were given 450 mg progesterone daily between Days 0 and 6 and from untreated control mares (n=13). Daily administration of 450 mg progesterone in oil significantly (P<0.02) increased serum progesterone concentrations in the treated mares. There was no significant difference in the recovery rate of Day-7 embryos between treated and control mares (8/13 versus 6/13, respectively). Embryonic development, assessed by morphologic evaluation, embryo diameter, and number of cell nuclei was not significantly different for embryos from treated and from control mares. The results of this study indicate that administration of progesterone beginning on the day of ovulation does not affect the embryo recovery rate or embryonic development, based on evaluation of uterine blastocysts recovered at Day 7 after ovulation.     

Gossypolone suppresses progesterone synthesis in bovine luteal cells

Gossypolone, a proposed major metabolite of gossypol, was synthesized and investigated for its effect on progesterone synthesis in cultured bovine luteal cells. Gossypolone inhibited human chorionic gonadotropin(hCG)-stimulated progesterone secretion, reduced substrate-enhanced conversions of 25-hydroxycholesterol to pregnenolone and of pregnenolone to progesterone in a dose-dependent fashion. These findings indicate that gossypolone inhibits not only 3β-hydroxysteroid dehydrogenase (3β-HSD) activity, as gossypol does, but also side-chain cleavage enzyme complex (cytochrome P450scc activity. However, the two compounds appear to have a similar potency in inhibiting progesterone secretion. Both gossypolone and gossypol (8.5 μM) induced morphological changes in cellular organelles.     

Assessment of progesterone profiles and postpartum onset of luteal activity in spring calving Hereford beef suckler cattle

Background  

Reproduction is the single greatest factor limiting beef cattle production. Previous research on beef suckler luteal activity has largely focused on the mechanisms, and duration, of postpartum anoestrus. However, the temporal pattern of luteal activity after resumption of post-partum ovarian activity, and the impact of pattern type on days open (DO) in purebred beef suckler cows, are unknown.     

Cloprostenol induced luteal regression in the beef cow. II. Histological evaluation of corpora lutea and correlation with luteal progesterone

Forty-two cycling, multiparous beef cows (percentage-Brahman) were injected twice at 11-d intervals with 500mug Cloprostenol (a prostaglandin F(2alpha) analog) to induce luteolysis. Cows were randomly assigned for ovariectomy at 12 hr intervals from 0 to 72 hr post-injection. Corpora lutea were excised and one-half of the corpus luteum was stored in phosphate-buffered formalin until mounting and staining with hematoxylin and eosin. The other half of the CL was snap-frozen for determination of progesterone content and concentration. Luteal cell density increased following Cloprostenol injection and was significantly correlated with a shift from predominantly healthy Type I cells to predominantly degenerating Types III and V cells. Cell mitosis tended to decrease by 12 hr and was lower by 24 hr post-injection. Cell pyknosis increased by 24 hr post-injection and was correlated with the decrease in percentage of healthy luteal cells. No pattern was detectable in cell karyorrhexis. Histological regression of the CL was inversely correlated with CL progesterone content. Therefore, we conclude that a reduction in cell mitosis is the earliest morphological sign of degeneration of the CL and that the CL follows a well-defined sequence of regression which is accompanied by a decrease in progesterone content.