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In four experiments male and female calves, heifers and bulls were housed in a climate chamber and exposed to two weeks of hot environmental conditions (30–35°C and 60% relative humidity (RH) during the day; 25°C and 60–70% RH during the night). The animals were subjected to milder conditions (15–18°C and 70% during the day; 12–15°C and 70% during the night) during the two week period preceding and the two weeks following heat exposure. Serum prolactin increased significantly (P< 0.01) when ambient temperature was increased and decreased after reduction of temperature. No changes could be observed for GH, TSH, LH and FSH. There was a significant (P< 0.01) sex difference in GH levels (males < females) in pre-pubertal calves but this difference was not apparent in older animals.

Abstract. The presence of oxytocin in ovarian tissue was examined immunocytochemically. Bovine antral follicles and corpora lutea were fixed with glutaraldehyde, picric acid and acetic acid fixative and immuno-stained by the indirect peroxidase-antiperoxydase (PAP) technique. Immunoreactive oxytocin was demonstrated in the granulosa cells of small and large follicles, in the granulosalutein cells of the young corpus luteum and in the large luteal cells of the mature corpus luteum.The regressing corpus luteum was not stainable. It is discussed that these findings additionally support the view that oxytocin is actually synthesized in ovarian tissues.

Abstract Immunoreactive oxytocin was measured in ovaries (corpus luteum and follicular fluid) and adrenals of cows, and in testes, seminal vesicles, prostate gland and adrenals of bulls. Secretion of oxytocin was further measured after culture of whole follicles, granulosa cells and theca tissue. Concentrations of oxytocin increased in corpora lutea of cycling cattle until mid-luteal phase (447 ± 93 ng/g wet weight) and decreased afterwards. Low concentrations were found in corpora lutea of pregnant animals (6 ± 3 ng/g wet weight). Follicular fluid contains some oxytocin (on average 42–108 pg/ml) but concentrations were significantly higher in the fluid of ovarian cysts (190 pg/ml). After culture of follicles the amount of oxytocin released into the medium increased indicating de novo synthesis. The granulosa cells were the main source of follicular oxytocin. Production increased during luteinization indicating that luteinization is an important step for the production of oxytocin in ovaries. Tissues of testes (65 ± 10 pg/g wet weight) and adrenals from cows (122 ± 39 pg/g wet weight) and bulls (111 ±2 pg/g wet weight) contained oxytocin but at much lower concentrations compared to corpus luteum tissue. About 10 times higher concentrations of oxytocin were measured in the adrenal medulla (717 ± 96 pg/g wet weight) compared to the cortex (72 ± 11 pg/g wet weight). Seminal vesicles and prostate gland contained no measurable amounts of oxytocin (< 5 pg/g wet weight).

A radioimmunoassay for oxytocin in cow plasma is described. Antisera were raised in rabbits against synthetic oxytocin coupled to bovine thyroglobulin. Iodinated oxytocin free of unlabelled oxytocin and most likely also free of diiodo-oxytocin was used as radioactive tracer. The tracer showed a high degree of purity, and was stable on storage. It could be used in the assay for 2–3 months. The assay showed very little crossreactivity with vasopressin. Acetone was used for the extraction of oxytocin from plasma as well as from standards made of synthetic oxytocin in pooled cow plasma. Inhibition curves obtained with plasma collected from cows at parturition were parallel to those obtained with the oxytocin standard preparation. The mean recovery of oxytocin added to cow plasma was 106 % (sd= 14). The within-assay coefficient of variation (CV) varied from 5.2 to 10.9 %, and the between-assay CV was in the order of 13 %. The assay sensitivity was 1 pg (0.5 μU) per tube, corresponding to 3 pg/ml plasma.Around the time of milking the plasma oxytocin profile showed a strong response to the preparation for milking, and a further effect releated to the attachment of the teat cups of the milking machine. Peak concentrations were in the range of 15–50 pg/ml. During parturition there was a peak of oxytocin (65 pg/ml) coinciding with the expulsion phase. After this peak levels decreased but remained measurably elevated until the expulsion of the placenta. The plasma disappearance curve for immunoreactive oxytocin after the infusion of 100 IU oxytocin over a period of 1 h showed two components with apparent half-lives of 7–9 and 25 min, respectively.

A sensitive and specific radioligand receptor assay (RRA) using rat testis homogenate as the receptor source is described for measurement of luteinizing hormone (LH) in bovine blood. Interfering and non-specific substances in blood were removed by means of ion-exchange chromatography on CM-Sephadex C-50. Criteria of validation such as recovery of added LH to plasma or serum, reproducibility, and specificity gave good results. Inhibition curves obtained with bovine plasma and serum were parallel to those obtained with the bovine standard preparation. The range of the dose-response curve was between 0.5–20 ng of bovine LH. The pattern of LH concentrations in purified serum samples under different physiological conditions such as during the oestrous cycle and after administration of GnRH showed a very close correlation whether measured by means of radioimmunoassay (RIA) or receptor assay. Values of RRA-LH were consistently higher than those of RIALH. Thus the lower the RIA-LH levels, the more pronounced were the discrepancies between results of both assay systems. The mean ratio of RRA-LH/RIA-LH for basal levels (less than 1 ng RIA-LH/ml plasma) was 17.8 as compared to a mean ratio for higher peak values (more than 20 ng RIA-LH/ml plasma) of only 1.2.

A synopsis is presented of hormonal profiles (peripheral plasma values of LH, FSH, progesterone; urinary oestrogens) during the oestrous cycle of cattle. After ovulation (day one) progesterone levels begin to increase from about day 4, reach a plateau around day 10, and decrease to basal concentrations starting on days 17 to 18. Corresponding with the progesterone decrease, urinary oestrogens show a main increase reaching its maximum one day before ovulation. LH levels are fairly constant from ovulation until day 5, small but significant peaks can be observed between days 6 and 10 in some cows (6 out of 21) and levels then decrease from day 11 to 13. A gradual additional increase begins on day 17, resulting in the preovulatory LH peak 25.7 ± 6.9 h (n = 75) before ovulation. This peak has a duration of 7.4 ± 2.6 h (n = 89) and occurs any time of the day but preferentially around 8 a. m. and 8 p. m. respectively. Likewise related are the two main periods for ovulation. During oestrus, (average duration of 16.9 ± 4.9 h (n = 28) the electrical resistance of the vaginal mucus decreases and the lowest values always correspond with the LHpeak. FSH levels show a wavelike pattern with peaks around days 4, 8, 12–13, 17, 18 and also coinciding with the pre-ovulatory LH peak. The sharp decrease in FSH between the two peaks on days 17 and 18 correlates with the decrease of progesterone during luteolysis. Obviously there is a parallelism between FSH elevations and follicular growth (determined by means of pelviscopic observations).

A specific heterologous radioimmunoassay for bovine follicle-stimulating hormone (bFSH) has been developed using guinea pig antiserum to ovine FSH with radio-iodinated ovine FSH. Slight cross-reactions with bovine TSH and LH could be eliminated by adding bovine TSH and LH to each tube during the assay. The labelling method with lactoperoxidase turned out to be more useful than the chloramine-T method. Inhibition curves obtained with male and female bovine plasma were parallel to those obtained with the bovine standard preparation. Exogenous bFSH could be quantitatively recovered from 200 μl of plasma or serum with a mean of 101.7 ± 7.7 %. The intra-assay coefficient of variation (CV) was ≦ 6 %. The inter-assay CV was 13.5 %.Sensitivity was 70 ng/ml NIH-FSH-B1. Mean plasma concentrations of FSH (NIH-FSH-B1) during the oestrous cycle were 303 ± 101 ng/ml. When plasma samples were collected in 6 h intervals from 17 oestrous cycles from 13 regularly cycling heifers and cows, a significant FSH peak occurred just before and during oestrus and coincided with the pre-ovulatory LH peak. Further FSH elevations could be measured on day 4, 8 and 12 after ovulation and 5 and 3 days before ovulation and showed a rhythmic pattern of four day intervals approximately.

Abstract. The levels of oxytocin receptor (OTR), cytosolic progestin receptor (cPR), cytosolic and nuclear estrogen receptor (cER, nER) were measured in the endometrium of 28 heifers that had been slaughtered on a defined day of the estrous cycle, In an additional, trial endometrial tissue obtained from 78 heifers or cows at the abattoir was analyed for OTR. OTR was absent during the luteal phase (after day 6), but a minor elevation was observed after day 15. OTR increased rapidly after luteolysis on days 17–18 reaching a maximum during estrous on day 21, and decreased again during days 1–6. cER and cPR were different to OTR but followed a similar pattern with maximal levels during days 1–8 of the estrous cycle. At day 12 both receptors were minimal and increased again towards day 21. nER was maximal at day 19–21 coinciding with maximal estradiol levels and estrous. Our data indicate that owing to an increasing sensitivity of the endometrium to progesterone and estradiol after day 12, these steroids may be mainly responsible for the initiation of first PGF2αsurges and luteolysis. Oxytocin seems to be of minor importance at this stage owing to low sensitivity of the endometrium for oxytocin.

Six Holstein bulls were housed in a climate-chamber under constant light regimen and after two weeks of preconditioning at 15°C, 60% relative humidity RH (day) and 12°C, 60% RH (night) were subjected to two weeks of heat exposure. This involved one week at 30°C and 60% RH (day) and 25°C and 60% RH (night) and a further week at 35°C, 60% RH (day) and 30°C, 60% RH (night). Three bulls were untreated (group I) and 3 bulls were treated (group II) just before and during heat exposure with a prolactin inhibitor to study the possible physiological role of prolactin on the regulation of water, potassium and sodium.Serum prolactin levels increased significantly (P< 0.01) in group I from the control value of 6 ng/ml to 33 and 44 ng/ml when the ambient temperature was increased (weeks 3 and 4) and then decreased to 21 and 12 ng/ml after reduction in temperature during weeks 5 and 6, respectively.For group II prolactin values decreased under the treatment with the prolactin inhibitor to 0.5 ng/ml and remained at this level throughout the experiment. GH levels were unaffected by heat treatment or by treatment with prolactin inhibitor. There were no differences between groups I and II in respiratory rate, pulse rate and rectal temperature. Water intake increased in both groups under heat exposure but decreased significantly afterwards only in group II. Differences in urinary excretion volume and blood serum osmolality were not significant. Urinary potassium and sodium excretion were unchanged in group II but increased with heat exposure in group I. During heat exposure 2 bulls of group II lost weight despite maintaining food intake.

Abstract. An improved RIA for measurement of oxytocin in blood is described by using an extraction method with SEP-PAK C18cartridges, which allows also concentration of the sample, a new antiserum with a higher sensitivity to standard oxytocin and preparation of the standard curve in buffer. The lower limit of assay sensitivity was 0.25 pg/tube, corresponding to 0.25–1.0 pg/ml plasma depending on the amount of plasma extracted. Hence, it was no problem to measure oxytocin basal concentrations in peripheral blood in the range of 0.6–4 pg/ml plasma depending on the stage of the oestrous cycle. The highest oxytocin concentrations occurred during the early and mid-luteal phase. The method has been applied also for samples from women, sheep, pigs and horses. Mean (± sd) recovery of oxytocin added to plasma or only buffer after extraction was 71.3 ± 8,1%, and the coefficient of variation (CV) = 11.4% (n = 27 assays). The intra-assay CV of two control samples was 7.9 ± 2.8 and 7.8 ± 2.4% (n = 17 assays). The inter-assay CV of 5 control samples with low and high oxytocin concentrations varied between 10.8 ± 17.3% (n = 25 assays). The 50% intercept was 2.5 ± 0.3 pg, CV = 11.3% (n = 29 assays).

Abstract. Active and passive immunization of ewes against oxytocin prolonged the luteal phase of the oestrous cycle, as evaluated by oestrus behaviour and determination of progesterone. Between the animals there was a high variability in response. In general animals with prolonged cycles had lower concentrations of free oxytocin. But in some animals oxytocin antibodies had no effect on cyclicity. Therefore, an individual minimal threshold concentration for oxytocin was presumed. In control or treated animals circulating oxytocin concentrations increased parallel with progesterone concentrations but decreased earlier during the mid luteal phase of the cycle. A new increase in oxytocin concentrations was only observed if a new corpus luteum was formed.

The technique of radioimmunoassay was used to determine prolactin concentration in plasma taken from bulls during and after infusions of oestradiol-17β and progesterone. During the infusion of oestradiol-17β (0.5, 1 and 2 mg) for 1 h the concentration of prolactin in the plasma fell, but showed an immediate increase of up to 6-fold after the infusion was completed. A dose-dependent increase (up to 20-fold) in prolactin concentration was observed after the infusion for 1 h of 5, 10, 40 and 80 mg progesterone. In those experiments in which 40 and 80 mg were administered the depression normally seen during the infusion was sustained for 2–3 h and the subsequent increase during the next few hours was comparatively small.

Abstract. Oxytocin and progesterone were measured radioimmunologically during a control cycle, early pregnancy and during several time courses until term in 4 ewes of a local Maroccan breed of sheep. Oxytocin concentrations increased in a similar matter after day 3–4 of the control cycle or after mating and decreased on days 13–15 of early pregnancy about 2 days earlier than during the oestrous cycle. After days 18–19 of pregnancy oxytocin concentrations remained low until term at concentrations as observed during the non-luteal phase of the oestrous cycle. Contrary progesterone concentrations remained high throughout gestation. Maternal recognition of pregnancy seems to have only a slight effect on the secretion pattern of oxytocin for few days after day 12. The results indicate a similar unknown mechanism during the cycle and early pregnancy for the vanishing oxytocin secretion and perhaps synthesis.

Oxytocin concentration in jugular vein blood was measured radioimmunologically with a detection limit of 3 pg/ml plasma in male and female cattle.Five bulls were tested; during mating a cow in oestrus with intromission and ejaculation, during mounting a dummy or another bull with ejaculation into an artificial vagina or during false mounts. No increase in oxytocin concentrations could be observed, but stimulation with an electro-ejaculator caused an increase ranging from 5–84 pg/ml after a latent period of 3–5 min. A similar response was observed in two cows following the same procedure.The contact with a bull, false mount or mating with intromission and ejaculation was not followed by a measurable oxytocin release in 5 test cows.The following stimulation techniques, massage of vulva and clitoris, massage of cervix and uterus per rectum, artificial insemination, introduction of a speculum into the vagina or insufflation of air into the vagina were performed with 5 cows and 5 heifers.Insufflation of air into the vagina was the most effective stimulus, eleciting an oxytocin release up to 588 pg/ml. All 5 heifers responded positively, as well as 4 cows in oestrus. The other manipulations caused an oxytocin response mainly in heifers (whether in oestrus or dioestrus), whereas only one cow in oestrus responded with an oxytocin release. In general, oxytocin concentrations increase about 30–90 s after the start of the stimulus.

Using the biotin-streptavidin amplification technique, highly sensitive specific second-antibody enzyme immunoassays for determining LH in bovine plasma with long (48 h) and short (4 h) incubation periods were developed. Biotin was linked to bLH by the N-hydroxysuccimidine method and the product (biotinyl-bLH) used to bridge between streptavidin-peroxidase and the immobilised bLH antibody in competitive tests. The assays were validated and their performance compaired with a radioimmunoassay currently in use. The sensitivities of the long and short incubation enzyme immunoassays (8 pg and 15 pg/well, respectively) were superior to that of 5-day incubation radioimmunoassay (100 pg/tube). Plasma interference in both assays were acceptable and volumes of 5 to 40 μl gave parallel standard curves and comparable LH levels, 10–20 μl plasma was sufficient to measure LH baseline levels by the long incubation enzyme immunoassay. The mean recovery of added standard bLH to plasma samples containing different endogenous LH was >90% (range 91.7–112) in both assays. The intra- and inter-assay variations of both assays were less than 10 and 17%, respectively. When both enzyme immunoassay and radioimmunoassay were used to measure LH in cyclic cows, the basal levels measured by enzyme immunoassay were lower than that measured by radioimmunoassay. Enzyme immunoassay offers an attractive alternative to the lengthy radioimmunoassay in current usage, with an added advantage of employing non-isotopic label.

Abstract. Seasonal levels of androstenedione and testosterone were investigated in plasma of mature intact and castrated male white-tailed deer. In four intact bucks, androstenedione concentrations were low in February and March (around 1 nmol/l) and then increased significantly (P< 0.05) from April to November (peak 2.34 nmol/l). Testosterone remained low (below 3.5 nmol/l) from February to August and then rose significantly (P< 0.01) till November (peak 36.78 nmol/l). Both hormones declined from November to February. In three castrates, androstenedione levels remained virtually unchanged (averaging around 0.5 nmol/l) between January and September. After a rapid significant increase (P< 0.05) till November (peak value 2.45 nmol/l), androstenedione concentrations declined quickly to a baseline level. Testosterone in castrates remained around 0.3 to 0.7 nmol/l for most of the year with a non-significant peak (1.45 nmol/l) in October.These data indicate that the spring and summer increase in androstenedione in the intact deer is of testicular origin; the fall peak, however, may be a result of increased production in the adrenal glands.It can be speculated that the increase in androstenedione in the blood of the male deer during the spring may be responsible for the seasonal initiation and support of antler growth as well as being supportive to the re-activation of the reproductive system.

Prolactin in blood plasma samples obtained from cows was determined by radioimmunoassay. During infusion of 2 mg or 4 mg 17β-oestradiol/hour there was a depression of the prolactin level, which seemed to be pronounced during infusions of 6 or 12 mg 17β-oestradiol/3 hours. In the experiments in which 2-6 mg 17β-oestradiol were administered a 2-5 fold increase in the plasma prolactin concentrations occurred after the infusions were completed. In the experiment with 12 mg 17β-oestradiol the depression was sustained for 3 hours after infusion; the subsequent increase the next few hours was comparatively small.

Plasma samples from jugular vein blood of cows before, during and after normal or glucocorticoid induced parturition and also in cases treated with a prolactin inhibitor were collected. Total oestrogens, prolactin and LH were determined by radioimmunological techniques, corticoids by competitive protein binding and for progesterone analysis both techniques were used. Progesterone began to decrease 1–2 day before parturition while the corticoid values increased concomitantly. The oestrogen levels remained high, almost until parturition had occurred; they began to decrease very rapidly approximately 5 hours before delivery. Prolactin showed the expected peak just before delivery and LH was constantly low. In the animals treated with a synthetic glucocorticoid or the prolactin inhibitor a temporary suppression of endogenous corticoids or of prolactin resp. was observed; no significant differences could be seen in the values of the other hormones and the physiological performance as compared to a normal delivery. It is concluded that a possible synergistic action of oestrogens and glucocorticoids is involved in the termination of pregnancy in the bovine species.

The peripheral blood level of prolactin has been examined in the bovine by means of radioimmunological method. Prolactin values of lactating animals (kept in an open stable) show a seasonal influence with higher basic values during the summer and lower basic values during the winter. Normal milking gives a potent short stimulus for the release of prolactin. Repeated milking also leads to a rise in prolactin, although the absolute value of the peaks seems to decrease with the number of stimuli. Massage of the udder, washing of the teats and a short premilking before actual milking lead to an increase of prolactin in most animals. A very short release of prolactin can also be detected after rectal massage of the cervix and of the vagina. Exogenous administration of oxytocin in a dose of 5 to 80 IU does not produce any increase of prolactin. After injections or infusions of 120 IU and 160 IU, only the latter dosage could induce a noticeable increase.

For radioimmunoassay of LH in bovine blood the preparation LER-LH-791-l as antigen for radioiodination and the double antibody technique using filtration for separation was applied. LH-determinations were carried out in about 700 serum samples from 5 cows and 1 heifer during a total of 13 oestrous cycles. Basic levels normally ranged between 0.4 and 2.7 ng LH/ml serum. Distinct peaks, which were represented by values between 3 and 62 ng/ml, appeared no longer than 6 hours.Commonly, one peak could be demonstrated around day 8 or 9 of the cycle, another 15 – 22 hours prior to ovulation; the latter is sometimes accompanied by movements of the LH level, performing also peaks a day before and/or after this more distinct preovulatoric peak. In some cases an extra peak occurred 3 – 6 days before ovulation. The finding of LH release twice (or more times) during the bovine cycle is in agreement with preliminary results obtained by earlier studies with the OAAD-test