Estrous Period Research Articles

Various behavioral signs of estrous and their relationship with time of ovulation in dairy cattle

The objective of this study was to investigate the relationship between various behavioral signs of estrous and time of ovulation and, determine which behavioral estrous sign(s) best predicted time of ovulation. In total, 94 ovulations were observed in 67 Holstein-Friesian dairy cows. Different behavioral estrous signs were observed at 3-h intervals and their relation with time of ovulation (ultrasound examinations at 3-h intervals) was investigated. In all estrous periods, sniffing and chin resting was displayed, while mounting was displayed in 90% and standing heat in 58% of estrous periods. Estrus was more intense in primiparous cows compared to multiparous cows and when more animals were in estrus at the same time. Although, these factors influenced intensity of estrous behavioral signs, they did not influence time of ovulation. Ovulation occurred 30.0 ± 5.1 h after onset of estrus (ranging between 18.5 and 48.5 h) and 18.8 ± 4.4 h after end of estrus (ranging between 9.5 and 33.5 h). Although informative, these predictors are highly variable between individuals and the method used to determine the onset and end of estrus is time consuming this, therefore limits in their use as a practical predictor of ovulation. Sniffing and chin resting were displayed during the non-estrous period and are therefore, not useful predictors of ovulation time. For animals that displayed standing heat, onset of standing heat was a good predictor for ovulation time (occurring 26.4 ± 5.2 h before ovulation). However, standing heat was only displayed in a limited number of cows, especially when only one cow was in estrus at a time. Onset of mounting was the best predictor for time of ovulation (occurring 28.7 ± 5.3 h before ovulation), and it was displayed in 90% of the estrous periods. However, mounting cannot yet be assessed automatically, which limits its practical use as ovulation predictor.

Mating Tactics in Response to Costs Incurred by Mating with Multiple Males in Wild Female Japanese Macaques

We investigated the costs of mating with multiple males in terms of feeding time, traveling distances, sexual proceptivity, and male aggression, for wild female (Macaca fuscata yakui) on Yakushima Island, Japan. We analyzed all-day focal sampling data from 7 females during the mating season (Sept.-Nov. 1996). On days when estrous females copulated with multiple males, they decreased their feeding time to half that of anestrous days, traveled longer distances, showed more proceptive sexual behaviors and received more aggression from subordinate males than on days when they copulated with only the 1st-ranking male. On days when females copulated with only the 1st-ranking male, they showed no difference in feeding time with that of anestrous days, and expended less effort than the above mating pattern because of short traveling distances, diminished sexual proceptivity and a lower frequency of aggression received. The results suggest that the costs of estrous vary according to female sexual proceptivity and the number and social status of mating partners. Female Japanese macaques exhibit a mixed mating strategy over prolonged estrous periods, which may provide females with opportunities to maximize the benefits of copulating with multiple males and to minimize the costs of estrus by mating with only the 1st-ranking male. During an estrous cycle, females may be adjusting efforts for reproduction and survival; i.e., mating vs. feeding.

Effect of long-term treatment with low doses of the LHRH antagonist Cetrorelix on pituitary receptors for LHRH and gonadal axis in male and female rats.

Our previous studies showed that treatment of female rats with large doses of Cetrorelix, an antagonist of luteinizing hormone-releasing hormone (LHRH), reduces levels of serum LH, estradiol, progesterone, and the concentration of pituitary LHRH receptors (LHRH-Rs) and their mRNA expression. Serum LH and testosterone levels and pituitary LHRH-R in male rats are also decreased by high doses of Cetrorelix. This approach can be used for therapy of sex hormone-dependent cancers. However, in conditions where an incomplete hormone deprivation is indicated, lower doses of Cetrorelix may suffice. Thus, we investigated the effect of a 30-day treatment with a low-dose depot formulation of Cetrorelix (20-24 microg per kg per day) on the pituitary-gonadal axis of male and female rats. In both sexes, lower serum LH levels were observed on day 4 after administration. In males, LH returned to control levels by day 10, whereas in females, a rebound LH elevation occurred. Testosterone levels in male rats were decreased up to day 20, but on day 30, the values were similar to controls. In females, serum estradiol was reduced on day 4; however, by day 10 it returned to normal. Progesterone levels were diminished through the entire period. Female rats showed diestrous smears during the first week of treatment and prolonged estrous periods thereafter. The weights of testes and ovaries were significantly lower, but not the weights of prostate, seminal vesicles, and uterus. Pituitary LHRH-R mRNA and LHRH-R protein levels were not significantly different from the controls. Thus, the treatment with low doses of Cetrorelix did not seriously impair gonadal functions. The results suggest that Cetrorelix in low doses induces only a partial pituitary-gonadal inhibition and might be indicated for treatment of endometriosis, leiomyomas, and benign prostatic hyperplasia.

Uréia para vacas leiteiras no pós-parto: teor plasmático de uréia e pH uterino

O objetivo deste trabalho foi verificar os efeitos de níveis crescentes de uréia nas dietas de vacas leiteiras no pós-parto sobre os teores plasmáticos de N-uréia (NUP) e o pH uterino no estro e na fase luteal. Utilizaram-se dezesseis vacas mestiças (Holandês-zebu) com escore corporal ao parto entre 3,5 e 4,0 (escala 1 a 5), que foram alocadas em quatro tratamentos constituídos de dietas contendo volumoso (silagem de milho) e concentrado, na proporção 60:40: T1 = 0,0; T2 = 0,7; T3 = 1,4; e T4 = 2,1% de uréia na matéria seca total. No estro e na fase luteal, do segundo ou terceiro ciclo estral pós-parto, realizou-se a medição do pH uterino quatro horas após o fornecimento da dieta e; simultaneamente foram coletadas amostras de sangue para determinação dos teores plasmáticos de uréia. A ingestão de matéria seca dos 70 aos 110 dias de lactação reduziu a partir do nível 0,7% de uréia e a produção de leite não foi influenciada pelos níveis de uréia das dietas. Não houve influência dos níveis de uréia sobre os teores plasmáticos de N-uréia, em amostras de sangue coletadas no estro, que foram, em média, de 19,59 mg/dL. O pH uterino no estro (6,66) e na fase luteal (6,97) também não foi influenciado pelos diferentes níveis de uréia nas dietas.

EVOLUTION OF INDUCED OVULATION IN NORTH AMERICAN CARNIVORES

We tested the hypothesis that induced ovulation is adaptive in North American carnivores by providing a mechanism to overcome the risk of unsuccessful mating in solitary species living in seasonal environments and a method for females to evaluate male quality via sexual selection inside the reproductive tract. We obtained published data on North American carnivores and determined from their distribution a coefficient of seasonality and primary productivity. Using traditional statistics and comparative methods, we determined that induced ovulators lived in more seasonal environments apparently not influenced by energy. No statistical differences in behavioral traits existed between induced and noninduced ovulators, but trends in data were consistent with our predictions, with induced ovulators tending to have larger home ranges and longer estrous periods. Mating systems also differed between the groups: induced ovulators were characterized by mostly (93%) multimale mating systems, whereas noninduced ovulators were monogamous (42%), multimale (33%), or polygynous (25%). Also, induced ovulators exhibited larger sexual dimorphism and smaller neonates than noninduced ovulators or felids. We suggest that induced ovulation evolved through sexual selection as a reproductive strategy beneficial for males (assurance of egg fertilization during short pair bonds) and females (postcopulatory mate choice based on level of stimulation causing induced ovulation).

Manipulation of lactation and suckling on the resumption of postpartum reproductive activity in Damascus goats

The reproductive activity of Damascus goats was studied in does that were not allowed to mate (non-lactating does) (Group C) and lactating does with different suckling treatments (Groups A and B). Time resumption of sexual activity was not significantly different between lactating and non-lactating does or between does’ groups of different suckling treatments. All does started to demonstrate estrous behavior during autumn (September). In Groups A, B and C the mean length of the estrous cycle was 19.0±0.6, 19.5±0.3 and 22.0±2.0 days, duration of the estrous period 29.6±3.0, 31.0±5.3 and 25.3±5.8 h, mean corpus luteum (CL) counts 1.6±0.2, 1.8±0.2 and 1.8±0.3, while mean litter size was 1.1±0.1, 1.2±0.2 and 1.3±0.5, respectively. In the three groups an increase in the plasma progesterone concentrations started from summer (mid-June), indicating the occurrence of one or more ovulatory anestrus (silent ovulation). Plasma prolactin levels were higher ( P<0.01) in lactating does than in non-lactating does and at 4 weeks postpartum. Moreover, prolactin levels were significantly higher ( P<0.01) in does suckling their kids than in does being machine milked. Plasma progesterone levels during postpartum were negatively significantly correlated with prolactin levels. The study demonstrated that lactation and suckling lengths have no effect on the resumption of postpartum sexual activity in Damascus goats and that season is the major factor controlling the ovarian activity.

Changes in sexual behavior and fecal steroid hormone concentrations during the breeding season in female Hokkaido brown bears (Ursus arctos yesoensis) under captive condition.

The longitudinal changes in fecal steroid hormone concentrations and sexual behavior in 2 mated/pregnant and 3 non-mated female Hokkaido brown bears were investigated during the breeding season. Behavioral estrus (standing) lasted for 14 and 32 days in the mated females and for 25 to 36 days in the non-mated females. In non-mated females, sexual behavior, such as female-female mounting and masturbation, was observed for several days before and after the estrous period. In mated females, mean fecal estradiol-17beta concentrations were higher in the estrous period than in the post-estrous period, while fecal progesterone concentrations were higher in the post-estrous period than in the estrous period. The similar trends of steroid hormone changes were observed in the non-mated females.

Endocrine monitoring and its application to the management of the giant panda

AbstractUrinary estrogens (n=10) and progestins (n=4) were monitored in a total of 10 giant panda females. Estrogens were evaluated with two different immunoassays during the estrous period. The two different methodologies gave similar hormonal profiles, but different quantitative results due to differences in the crossreactivity of the different assay systems. High‐pressure liquid chromatography (HPLC) analysis of peak estrogen urine revealed that estrone sulfate was the major component, and estrone glucuronide was a minor component (97:3). A comparison of days of insemination in females that conceived and delivered offspring showed a correlation to insemination on days 1 and 2 post‐estrogen peak. An analysis of urinary progestins revealed that they were elevated over baseline concentrations following the estrogen surge during estrus (P&lt;0.0004). Zoo Biol 22:389–400, 2003. © 2003 Wiley‐Liss, Inc.

Repeat breeding in dairy heifers: follicular dynamics and estrous cycle characteristics in relation to sexual hormone patterns

Repeat breeding occurs at an incidence of 10% in the Swedish dairy cow population. Evidence is available for a hormonal asynchrony around estrus in repeat-breeder heifers (RBH). This asynchrony seems to be the underlying cause for a series of dysfunctions such as prolonged standing estrus and delayed ovulation, leading to fertilization failure. For further determinations of repeat-breeder estrous cycle characteristics, seven strictly selected RBH and six virgin heifers (VH) were studied during 3–7 consecutive cycles, with particular attention paid to the estrous period. Follicular dynamics were studied by ultrasonography and related to estrous behavior and pattern of sexual hormones (progesterone, estradiol-17β, and LH) in peripheral circulation. Mean group data were compared and a classification model was designed. The most prominent findings for RBH were prolonged duration of estrus, delayed LH peak, prolonged lifespan of the preovulatory follicle, and a late postovulatory rise in plasma progesterone. There was also a strong tendency for peri-ovulatory suprabasal progesterone levels in RBH. It is suspected that these deviations cause changes in the microenvironment of the preovulatory follicle, negatively affecting the final maturation of the oocyte. The heterogeneity of the RBH group underlines the multifactorial cause of the repeat-breeder syndrome. The VH formed a homogenous group with data varying within physiological limits. A classification model based on three characteristic variables managed to identify 81% of the VH and 79% of the RBH correctly. Results from this study propose that some heifers have general, consistent problems in synchronizing estrous events, displayed as varying symptoms in the course of consecutive estrous cycles. These subfertile animals could be classified as repeat-breeders.

ESTROUS CYCLES, COPULATION, AND PREGNANCY IN THE FENNEC FOX (VULPES ZERDA)

Characteristics of estrus, mating, and pregnancy were studied in captive fennec foxes. The monestrous cycles had mean intervals of 9.9 (±1.2) months, whether or not pups survived to weaning. Proestrus, judged by vulval swelling, began 6.5 (±0.7) days before estrus and was not accompanied by sanguineous discharge. Percentage cornified epithelial cells in vaginal smears increased at the time of estrus. In 8 of 10 estrous periods, a single mating followed by an extremely long copulatory lock (mean 1 h 58 min) was observed during continuous video monitoring. In the other two cycles, copulation occurred 2 and 3 times. Patterns of both fecal and serum estradiol and progesterone concentrations during estrous cycles were similar to those reported for other canids.

Longitudinal study of delayed reproductive success in a pair of white‐cheeked gibbons (Hylobates leucogenys)

AbstractConcern over a lack of breeding success in a pair of white‐cheeked gibbons (Hylobates leucogenys) prompted a four‐part study of gibbon behavior and physiology at the Lincoln Park Zoo. Data were collected to determine the female intermenstrual interval (IMI) and identify periods of peak receptivity. Subsequent behavioral data were gathered during both female menses and estrous periods to formulate a behavioral profile for each gibbon. The female experienced a species‐typical IMI of 21.6 days and exhibited heightened levels of behavioral receptivity toward the male during estrus. The male exhibited high levels of human‐directed behavior throughout the day. Closing the exhibit to visitors did not result in higher levels of social proximity or prosocial behavior between the pair, although it did eliminate the male's human‐directed behavior during the experimental condition only. The female exhibited higher levels of social grooming, solicitation of the male, and proximity to the male when the pair was locked indoors. Based on those findings, the pair was moved to an exclusively indoor exhibit. The female's visibility to the public decreased substantially in the alternate exhibit, but there were no other significant behavioral changes. Nearly one year after the exhibit change, the female gibbon was confirmed to be pregnant. Although the results of this study cannot be used to infer causal conditions that resulted in the pregnancy, they do support the notion that systematic documentation of animal behavior and physiology may be used as a tool for monitoring reproductive behavior and informing management decisions. Zoo Biol 21:413–434, 2002. © 2002 Wiley‐Liss, Inc.

Seasonal effects on estrous behavior and time of ovulation in nonlactating beef cows.

Estrous behavior and time of ovulation relative to the onset of estrus were determined in mature Angus x Hereford cows (n = 17 to 21 each season) during summer, winter, and spring for 2 yr. Estrous behavior was evaluated during the first of two consecutive estrous periods, and time of ovulation was determined during the second estrus. Concentrations of progesterone were quantified in twice weekly blood samples to ensure all cows had normal estrous cycles. The HeatWatch system was used to measure the duration of estrus, number of mounts received per estrus, and duration of the longest interval between mounts received. Commencing 16 h after the onset of the second estrus, transrectal ultrasonography was performed every 4 h until the dominant follicle was no longer present on the ovary, and time of ovulation was defined as 2 h preceding the absence of the dominant follicle. There was a seasonal effect on the duration of estrus; cows were estrus longer in summer (17.6 +/- 0.8 h) than in winter (15.5 +/- 0.8 h; P = 0.07) or spring (13.9 +/- 0.9 h; P < 0.05). Cows were mounted more times per estrus (P < 0.05) in winter (59.0 +/- 5.3) than in summer (43.6 +/- 5.3) or spring (38.2 +/- 5.8). Intervals between mounts of estrous cows were longer (P < 0.05) in summer (4.1 +/- 0.4 h) than in spring or winter (2.7 +/- 0.4 h). During all seasons, cows were mounted more times (P < 0.01) between 0600 to 1200 (3.2 +/- 0.2 mounts received/h of estrus) than during other times of the day (2.1 +/- 0.2 mounts received/h of estrus). Cows ovulated 31.1 +/- 0.6 h after the onset of estrus, and time of ovulation was not influenced by season. We conclude that season influences estrous behavior of beef cows; cows are mounted more times per estrus in winter than in summer or spring. Time of ovulation relative to the onset of estrus is constant during all seasons and averages 31.1 h.

Reproductive behavioral changes during the ovarian cycle of lesser bushbabies (Galago moholi) in captivity.

Behavioral patterns were quantified in seven heterosexual lesser bushbaby (Galago moholi) pairs during the estrous cycle to determine the relative significance of behavioral and nonbehavioral components of female sexuality in mate attraction. Increases in the occurrence of several male behaviors indicating female attractiveness were initiated during vaginal swelling when the female was sexually nonreceptive. Female behavioral estrus, as indicated by intromission, was confined to a portion of vaginal opening coinciding with proestrous and vaginal estrous smears. Female attractiveness was maintained for much of the period of vaginal opening, whereas female receptivity ended a day or two earlier than attractiveness. Female receptive and proceptive behaviors were not well defined or extensive, and few female behaviors exhibited significant changes during the cycle. Scent-marking behaviors, such as urine washes, and male grooms, were generally elevated outside the behavioral estrous period. In G. moholi, male sexual arousal appears to be elicited primarily by female attractiveness, while behavioral components of female sexuality appear to be less important in influencing mate attraction.

Detection and Characterization of Estrus in Dairy Cattle with an Electronic Heatmount Detector and an Electronic Activity Tag

The length and onset of estrus was studied in 71 lactating dairy cows using an electronic heatmount sensor (HeatWatch; DDx Inc., Boulder, CO, DeForest, WI) and an electronic activity tag (Heat Seeker, Boumatic, Madison, WI). Three methods were used to determine estrus: 1) the electronic heatmount system, 2) an increased activity ratio algorithm determined by the Heat Seeker, and 3) an increased activity count algorithm calculated for each estrous period. Mounting and physical activity variables were characterized, and the effects of synchrony, parity, and weather on these variables were determined with data from two different trials. Cows in trial 1 were not synchronized, while cows in trial 2 were synchronized. The results of the study were consistent as follows: mean numbers of mounts were 6.70±0.7 and 5.42±0.80 for trials 1 and 2, respectively; each mount lasted 3.20±0.19s (trial 1) and 3.36±0.42s (trial 2). Total mounting activity averaged 5.83±0.78h per estrous period in trial 1 and 5.57±1.02h in trial 2. Estrus identified by the increased activity count algorithm corresponded more closely to standing mount activity (determined by the HeatWatch System) than did the increased activity ratio algorithm. Synchrony, parity, and weather did not have a direct effect on physical activity. Hot weather decreased the duration of standing mount activity significantly, but did not affect the number or duration of individual mounts. All three methods of estrus detection employed improved the efficiency of detection over visual observation.

Physical properties of bovine cervical mucus during normal and induced (progesterone and/or PGF2α) estrus

Ninety two Friesian cows were used to determine physical properties of cervical mucus collected during normal estrus and estrus induced. Estrus was induced using either progesterone (P 4) releasing intravaginal devices (PRID) and/or prostaglandin F2α (PGF2α). The animals were assigned to 4 groups (no treatment, a PRID for 12 days plus an injection of 1000 IU PMSG at the removal of the PRID, a double injection of 3 mL PGF2α 11 days apart, and a PRID for 7 days plus an injection of PGF2α 24 h before the removal of PRID). A number of cows with normal estrus exhibited three consecutive estrus cycles after calving. Cows that had not shown estrus for three months after calving had their reproductive system palpated twice at 10-day intervals, to determine their ovarian activity. Then PRID and/or PGF2α was administered to cows that were found to have a palpable corpus luteum in one of two palpations (cycling cows). The cows of the three induced estrous groups were artificially inseminated (AI) twice, while those with normal estrus received only a single AI. Cervical mucus samples were collected from all cows 5 to 30 min before the first AI. Additionally, samples of cervical mucus were collected from 20 cows at their first estrus after the induced estrus. The results are summarized as follows: 1) The physical properties of cervical mucus were similar in the first three normal consecutive estrus cycles after calving. 2) The physical properties of cervical mucus in normal estrus after calving were similar to those in the first estrus after an induced estrus. 3) The pH values for normal estrus were similar to those for induced estrus. 4) Viscosity of cervical mucus in the normal estrous group was significantly lower than that in the induced estrus. Furthermore, significant differences were noticed among the three induced estrous groups. 5) Spinnbarkeit, crystallization and receptivity of cervical mucus (penetration test) were significantly higher in the normal estrous group than in the induced estrous groups, while no difference was detected among induced estrus groups. 6) Pregnancy rates in physical properties of cervical mucus at the time of the second AI were similar to those for normal estrus but this suggestion needs further investigation. On the other hand, at the time of the first AI very few cows in the induced estrus groups produced cervical mucus similar to that of cows with normal estrus. The physical properties that differed the most were viscosity, spinnbarkeit, and the penetration test. Other authors found that cows do not exhibit estrus at the same determined time (11,26). On the basis of this finding physical properties of cervical mucus could be used as an indicator for the time of estrus manifestation, in order to increase pregnancy rates by applying AI at an optimal time. In summary, the results from the present study showed that the physical properties of cervical mucus in the first three consecutive normal estrous periods after calving are similar; that the first estrus after an induced estrus is a normal estrus; that viscosity, spinnbarkeit, crystallization and receptivity of cervical mucus (penetration test) are significantly different for normal than for induced-estrus groups at the time of first AI; and a small percentage of cows with induced estrus produce cervical mucus with parameters of viscosity, spinnbarkeit and receptivity similar to those found in normal estrus. These properties could be used as indicators for the quality of cervical mucus produced by the cows with induced estrus.

Sexual behavior of Zebu cattle ( Bos indicus) following estrous induction by Syncro-Mate B, with or without estrogen injection

The effect of estrogen administered with norgestomet implants in the expression of behavioral estrous of Zebu cattle was studied in a herd of 18 cycling Brahman cows, in two trials. In the first and second trials, six different cows were treated with progestagen on successive days. In addition, in the first trial, estrogen injection was applied with the treatment. With the purpose of enhancing mounting behavior in treated and/or non-treated animals, on the third day of each trial, three other cows with an active CL were injected with 25 mg of PGF 2α. All animals were examined three times a week by rectal ultrasound, and a blood sample was also taken for progesterone analysis. Sexual activity was monitored by a closed-circuit TV from 1500 to 0600 h. Direct observation was carried out from 0900 to 1200 h. A total of 15 cows displayed 36 mounting periods (11 with ovulation). Twenty-five annovulatory estrous periods were observed, 77% supported by large follicles. More cows (66% vs. 16%) showed mounting activity when estradiol valerate was used as part of the progestagen treatment ( P=.03). Ovulation rate in response to the effect of norgestomet implant was 16%, independent of treatment. It is concluded that estrogen administered with norgestomet implants increases the expression of behavioral estrous in Bos indicus cattle but does not improve ovulation rate.

Shrinkage in the horizontal dimensions of the vulva (vulvar shrinkage) as an indicator of standing heat in the beagle.

During the proestrous and estrous periods in 12 beagles, the following parameters were measured daily: the horizontal dimensions of the vulva, vaginal cytology, and serum luteinizing hormone (LH) and progesterone concentrations. Measurements of serum LH concentrations allowed for the identification of the LH surge and the optimal time for artificial insemination (AI). Nine out of the 12 beagles became pregnant through AI and completed a gestation. Shrinkage in the horizontal dimensions of the vulva (i.e., vulvar shrinkage) was primarily observed prior and subsequent to the LH surge. In six of the nine (66.7%) beagles that became pregnant, vulvar shrinkage was observed on the days in which the LH surge was confirmed, and the rate of vulvar shrinkage tended to be greater at higher serum LH concentrations. Further vulvar shrinkage was identified in all nine beagles within two days of the LH surge. An increase in the serum progesterone concentration was observed after the LH surge in each of the beagles that became pregnant, together with clinical signs of estrous behavior (i.e., standing heat) as well as a change in vulva condition from swollen to soft. This demonstrates that vulvar shrinkage is induced in response to the onset of the LH surge and that the LH surge can be predicted through the measurement of the horizontal dimensions of the vulva, vaginal cytology, and the assessment of serum progesterone concentrations in beagles.

Monitoring of estrous cycle of ewes by ram-seeking behavior

For 6 years adult, non-synchronized Hampshire ewes ( n=2, 5, 8, 8, 9 and 12) and rams ( n=6, 10, 12, 11, 11 and 9) were maintained separately and released to contiguous pastures having in common a woven-wire fence about 80 m long and 1.1 m high. Natural estrous cycles were monitored daily by observations of ram-seeking behavior as defined as a ewe being persistent in staying at the common fence with attention focused on the ram to the virtual exclusion of grazing. Day 0 of the estrous cycle is the first day that ram-seeking is observed. One hundred and thirty four fence stays and 115 estrous cycles have been observed. For cycles falling within the normal range, the mean cycle duration (±SE) was 16.53±0.03 days. During the first four study years, a copulation test with an intact ram, carried out on a spot-check basis, indicated that all tested ewes showing ram-seeking, except one, were in estrus. Ram-seeking never occurred outside of the estrous period of the cycle. During the fifth year, daily copulation tests with an intact ram and daily observations for ram-seeking has indicated that estrus generally lasts about two days while ram-seeking lasts for one of these days, so that the two behaviors while overlapping are not coextensive. During the sixth year, daily copulation tests with a vasectomized ram showed again a partial coincidence of the two behaviors, but also that occasionally they appeared to have the same duration. It is concluded that the utilization of ram-seeking provides an alternative, practical means to monitor the estrous cycle.

Immunohistologic Detection of Estrogen Receptor Alpha in Canine Mammary Tumors: Clinical and Pathologic Associations and Prognostic Significance

Eighty-nine canine mammary tumors and dysplasias of 66 bitches were investigated to determine the immunohistochemical expression of classical estrogen receptor (ER-alpha) and its clinical and pathologic associations and prognostic value. A complete clinical examination was performed and reproductive history was evaluated. After surgery, all animals were followed-up for 18 months, with clinical examinations every 3-4 months. ER-alpha expression was higher in tumors of genitally intact and young bitches (P < 0.01, P < 0.01) and in animals with regular estrous periods (P = 0.03). Malignant tumors of the bitches with a previous clinical history of pseudopregnancy expressed significantly more ER-alpha (P = 0.04). Immunoexpression of ER-alpha decreased significantly with tumor size (P = 0.05) and skin ulceration (P = 0.01). Low levels of ER-alpha were significantly associated with lymph node involvement (P < 0.01). Malignant tumors had lower ER-alpha expression than did benign tumors (P < 0.01). Proliferation index measured by proliferating cell nuclear antigen immunostaining was inversely correlated with ER-alpha scores (P = 0.05) in all tumors. Low ER-alpha levels in primary malignant tumors were significantly associated with the occurrence of metastases in the follow-up (P = 0.03). Multivariate analyses were performed to determine the prognostic significance of some follow-up variables. ER-alpha value, Ki-67 index, and age were independent factors that could predict disease-free survival. Lymph node status, age, and ER-alpha index were independent prognostic factors for the overall survival. The immunohistochemical detection of ER-alpha in canine mammary tumors is a simple technique with prognostic value that could be useful in selecting appropriate hormonal therapy.

Effect of α-tocopherol on plasma testosterone and plasminogen activator activity or inhibition in ram spermatozoa

The objective of this study was to evaluate the effect of α-tocopherol on blood testosterone and specific proteolytic enzymes in spermatozoa and seminal plasma, with final aim the increase of sperm fertilizing ability with a nutritional supplement. The effect of α-tocopherol supplementation on testosterone parameters (mean value, basal level, peak number, mean concentration value during peaks, peak amplitude, peak duration) and plasminogen activator activity (PAA), plasminogen activator inhibition (PAI) and plasmin inhibition (PI) of spermatozoa and seminal plasma was studied in the ram during autumn (estrous period for the sheep in Greece) and spring (anestrous period). Treated animals showed a marked increase in serum α-tocopherol. Testosterone parameters were not affected by the α-tocopherol in either autumn or spring, however, the spermatozoal PAA and PAI (anti-tPA) were increased in the spring but not in autumn. These enzymes and their inhibitors are normally increased in autumn (the breeding season) and low in spring. If PAA can improve the fertilizing ability of spermatozoa in the spring, our finding may mean that a nutritional supplement, such as α-tocopherol, could provide rams for an accelerated onset of the breeding season in the ewe.