In vitro and in vivo effects of kisspeptin antagonists p234, p271, p354, and p356 on GPR54 activation.

Kisspeptins (KPs) and their receptor (GPR54 or KiSS1R) play a key-role in regulation of the hypothalamic-pituitary-gonadal axis and are therefore interesting targets for therapeutic interventions in the field of reproductive endocrinology. As dogs show a rapid and robust LH response after the administration of KP10, they can serve as a good animal model for research concerning KP signaling. The aims of the present study were to test the antagonistic properties of KP analogs p234, p271, p354, and p356 in vitro, by determining the intracellular Ca2+ response of CHEM1 cells that stably express human GPR54, and to study the in vivo effects of these peptides on basal plasma LH concentration and the KP10-induced LH response in female dogs. Exposure of the CHEM1 cells to KP-10 resulted in a clear Ca2+ response. P234, p271, p354, and p356 did not prevent or lower the KP10-induced Ca2+ response. Moreover, the in vivo studies in the dogs showed that none of these supposed antagonists lowered the basal plasma LH concentration and none of the peptides lowered the KP10-induced LH response. In conclusion, p234, p271, p354, and p356 had no antagonistic effects in vitro nor any effect on basal and kisspeptin-stimulated plasma LH concentration in female dogs.

Validation of a noninvasive diagnostic tool to verify neuter status in dogs: The urinary FSH to creatinine ratio.

Determining the presence of functional gonadal tissue in dogs can be challenging, especially in bitches during anestrus or not known to have been ovariectomized, or in male dogs with nonscrotal testes. Furthermore, in male dogs treated with deslorelin, a slow-release GnRH agonist implant for reversible chemical castration, the verification of complete downregulation of the hypothalamic-pituitary-gonadal (HPG) axis can be difficult, especially if pretreatment parameters such as the size of the testes or prostate gland are not available. The aims of this study were to validate an immunoradiometric assay for measurement of FSH in canine urine, to determine if the urinary FSH to creatinine ratio can be used to verify the neuter status in bitches and male dogs, as an alternative to the plasma FSH concentration, and to determine if downregulation of the HPG axis is achieved in male dogs during deslorelin treatment. Recovery of added canine FSH and serial dilutions of urine reported that the immunoradiometric assay measures urinary FSH concentration accurately and with high precision. Plasma FSH concentrations (the mean of two samples, taken 40 minutes apart) and the urinary FSH to creatinine ratio were determined before gonadectomy and 140 days (median, range 121-225 days) and 206 days (median, range 158-294 days) after gonadectomy of 13 bitches and five male dogs, respectively, and in 13 male dogs before and 132 days (median, range 117-174 days) after administration of a deslorelin implant. In both bitches and male dogs, the plasma FSH concentration and the urinary FSH to creatinine ratio were significantly higher after gonadectomy, with no overlapping of their ranges. Receiver operating characteristic analysis of the urinary FSH to creatinine ratio revealed a cut-off value of 2.9 in bitches and 6.5 in males to verify the presence or absence of functional gonadal tissue. In male dogs treated with deslorelin, the plasma FSH concentrations and urinary FSH to creatinine ratios were significantly lower after administration of the implant, but their ranges overlapped. We conclude that the urinary FSH to creatinine ratio can be used to verify the neuter status of bitches and male dogs. However, it cannot be used for the assessment of complete downregulation of the HPG axis after administration of a deslorelin implant. The urinary FSH to creatinine ratio is preferable over the plasma FSH concentration because it involves only one sample that can be collected relatively easy and noninvasively.

The effects of kisspeptin agonist canine KP-10 and kisspeptin antagonist p271 on plasma LH concentrations during different stages of the estrous cycle and anestrus in the bitch.

Kisspeptin (KP) plays a key role in the regulation of the hypothalamic-pituitary-gonadal axis via the release of GnRH. As normal KP signaling is essential for reproductive function, it could be an interesting new target for therapeutic interventions, e.g., nonsurgical contraception in dogs. The aims of the present study were to investigate the effect of KP-10 administration on plasma LH concentration in different stages of the reproductive cycle and to investigate the suitability of p271 as KP antagonist in the bitch. Two groups of six adult Beagle bitches were used. In one group, plasma LH concentration was determined before (40 and 0 minutes) and 10, 20, 40, and 60 minutes after the intravenous administration of 0.5-µg/kg body weight (BW) canine KP-10. In the other group, the bitches received a continuous intravenous infusion with p271 (50 µg/kg BW/h) for 3 hours, and 0.5-µg/kg BW canine KP-10 was administered intravenously 2 hours after the start of the p271 infusion. Their plasma LH concentration was determined before (-40 and 0 minutes) and 30, 60, 90, 120, 130, 140, 160, and 180 minutes after the start of the p271 infusion. In both groups, the experiments were performed during the follicular phase, the first and second half of the luteal phase, and during anestrus. Canine KP-10 induced an increase of plasma LH concentration during all estrous cycle stages and anestrus. There was no difference in LH response between the two groups. The lowest LH response was seen during the follicular phase and the highest response during anestrus. The area under the curve (AUC) for LH and LH increment in the follicular phase were lower than those in anestrus. The AUC LH and LH increment in the first half of the luteal phase were lower than those in the second half of the luteal phase and anestrus. The AUC LH and LH increment in the second half of the luteal phase were not different from those in anestrus. Continuous administration of the antagonist p271 did not alter basal plasma LH concentration and could not prevent or lower the LH response to KP-10 in any of the cycle stages and anestrus. It can be concluded that the LH response to KP-10 is dependent on estrous cycle stage and that peripheral administrated p271 cannot be used as KP antagonist in the dog. This provides new insight in reproductive endocrinology of the bitch, which is important when KP signaling is considered for therapeutic interventions, such as for estrus induction or nonsurgical contraception in the bitch.

Successful treatment for subinvolution of placental sites in the bitch with low oral doses of progestagen.

Subinvolution of placental sites (SIPS) is the major cause of persistent sanguineous vaginal discharge after parturition in the bitch. Spontaneous remission is common but may take several months, and hence, medical therapy to end the discharge is often requested. In this retrospective study, we evaluated the effect of treatment for SIPS with low oral doses of a progestagen. Nine bitches with SIPS, but otherwise clinically healthy, were found in the computer database of the Department of Clinical Sciences of Companion Animals. Seven of these bitches were treated with low oral doses of a progestagen (megestrol acetate, 0.1 mg/kg body weight (bw) once daily for the 1st week, then 0.05 mg/kg bw once daily for the 2nd week). The other two bitches were untreated. Treatment results were evaluated by a telephone questionnaire. Progestagen treatment was successful in all of the treated dogs; sanguineous vaginal discharge stopped within the treatment period. One of the two untreated dogs remained symptomatic until the next oestrus, approximately 120 days after parturition, and the other remained symptomatic until 6 weeks before the start of the next pro-oestrus, 270 days after parturition. No side effects of the progestagen treatment were observed. Subsequent gestations, parturitions and puerperal periods of 5 mated bitches were uneventful. One bitch did not become pregnant after mating. In conclusion, the results of this study indicate that oral administration of low doses of progestagen for 2 weeks is effective in stopping persistent sanguineous vaginal discharge in bitches with SIPS, with neither side effects nor reduced subsequent fertility.

Disorders of sexual development and associated changes in the pituitary-gonadal axis in dogs.

Normal sexual differentiation depends on completion of chromosomal sex determination, gonadal differentiation, and development of the phenotypic sex. An irregularity in any of these three steps can lead to a disorder in sexual development (DSD). We examined nine dogs with DSD by abdominal ultrasonography, laparotomy, histologic examination of the gonads, and reproductive tract, cytogenetic analysis, and mRNA expression of the SRY gene. We also determined the plasma concentrations of luteinizing hormone (LH), estradiol-17ß, and testosterone before and after administration of gonadotropin-releasing hormone (GnRH) and compared these results with those obtained in anestrous bitches and male control dogs. The gonads of three dogs with DSD contained both testicular and ovarian tissue, while in the other six only testicular tissue was found. Each of the dogs had a uterus. Based on gynecologic examination, cytogenetic analysis, and the histology of the gonads, seven of the nine dogs appeared to be XX sex reversals. Three of these were XX true hermaphrodites and four were XX males; the other two dogs had incomplete XY gonadal dysgenesis. All seven XX sex-reversed dogs were found to be negative for the SRY gene by polymerase chain reaction. The basal plasma luteinizing hormone (LH) concentration was significantly higher in dogs with DSD than in anestrous bitches but not significantly different from that in male dogs. The basal plasma LH concentration increased significantly after GnRH administration in all dogs with DSD. The basal plasma estradiol concentration was significantly higher in dogs with DSD than in anestrous bitches but not significantly different from that in male dogs. The basal plasma testosterone concentration was lower in dogs with DSD than in male dogs. In all dogs with DSD both the basal and GnRH-induced plasma testosterone concentrations were above the upper limit of their respective ranges in the anestrous bitches. In conclusion, the secretion of LH and estradiol in these dogs with DSD, all of which had testicular tissue in their gonads, was similar to that in male control dogs. These results indicate that the basal and/or GnRH-stimulated plasma testosterone concentration might be used to detect the presence of testicular tissue in dogs with DSD.

Progesterone receptor isoforms in the mammary gland of cats and dogs.

Progesterone exerts its effect by binding to specific progesterone receptors (PR) within the cell. In dogs and cats, no data are available on PR isoforms as found in other species. We therefore investigated the sequence of the PR gene and encoded protein in dogs and cats, the expression of PR isoforms in mammary tissue using Western blots and the presence of PR in mammary tissue using immunohistochemistry. Comparison of the amino acid sequence of the canine and feline PR with human PR revealed major differences in the PR-B-specific upstream segment (BUS). However, the essential activation function 3 (AF3) domain was intact in the cat but mutated in the dog. The DNA and ligand-binding domains were highly similar among the species. In cats with fibroadenomatous hyperplasia (FAH), high expression of PR mRNA together with growth hormone (GH), GH receptor (GHR) and IGF-I mRNA was found in comparison with feline mammary carcinomas. Immunohistochemical analysis showed strong nuclear as well as cytoplasmic staining for PR in FAH. Western blot analysis revealed expression of the PR-A and PR-B isoforms in the feline mammary gland. In canine mammary tissue, the most abundant PR staining was found in proliferative zones of the mammary gland. Western blot analyses showed mainly staining for PR-A with lower PR-B staining. It is concluded that in dogs and cats both PR isoforms are expressed. The role of mutations found in the canine PR-B is discussed.

Effects of gonadotropin-releasing hormone administration on the pituitary-gonadal axis in male and female dogs before and after gonadectomy.

GnRH-stimulation tests were performed in 14 female and 14 male client-owned dogs of several breeds, before and 4 to 5 mo after gonadectomy. The aim of the study was to obtain more insight into the pituitary-gonadal axis in intact and neutered dogs and to establish reference values. Basal plasma luteinizing hormone (LH) and follicle-stimulating hormone (FSH) concentrations were increased significantly after gonadectomy in both bitches and male dogs. In both males and females ranges of the basal plasma FSH concentrations, before and after gonadectomy, did not overlap as opposed to the overlap in ranges of the basal plasma LH concentrations. Before gonadectomy basal plasma LH concentrations were lower and basal plasma FSH concentrations were higher in bitches than in male dogs. After gonadectomy these basal values did not differ significantly. GnRH administration before gonadectomy resulted in an increase in plasma LH and FSH concentrations in both genders. GnRH administration after gonadectomy produced an increase only in plasma LH concentrations in both genders, and a just significant increase in plasma FSH in castrated male dogs. GnRH administration before gonadectomy resulted in a significant increase in plasma testosterone concentration in both genders. In males ranges of basal and GnRH-stimulated plasma testosterone concentrations before and after gonadectomy did not overlap. Basal plasma estradiol concentrations were significantly higher in intact males than in castrated males and their ranges did not overlap. The basal estradiol concentrations in bitches before and after ovariectomy were not significantly different. At 120 min after GnRH administration, ranges of plasma estradiol concentration of intact and ovariectomized bitches no longer overlapped. In conclusion, basal plasma FSH concentration appears to be more reliable than basal plasma LH concentration for verification of neuter status in both male and female dogs. The basal plasma testosterone concentration appears to be reliable for verification of neuter status in male dogs. The plasma estradiol concentration at 120 min after GnRH administration can be used to discriminate between bitches with and without functional ovarian tissue.

Questionnaire-based survey of parturition in the queen.

The lack of scientific data concerning whether parturition in the queen proceeds normally or not may prevent veterinarians and cat owners from recognizing parturition problems in time. A questionnaire-based study of parturition in 197 queens was performed to determine several parameters of parturition and their influence on its progress. The mean length of gestation was 65.3 days (range 57 to 72 days) and it decreased with increasing litter size (P = 0.02). The median litter size was 4.5 kittens (range 1 to 9), with more males (53%) than females (46%) (P = 0.05). Sixty-nine percent of the kittens were born in anterior presentation and 31% in posterior presentation, indicating that either can be considered normal in the cat. Males were born in posterior position (34%) more often than females (26%) (P = 0.03). The mean birth weight was 98 g (range of 35 to 167 g) and decreased with increasing litter size (P < 0.01). Mean birth weight was higher in males and kittens born in posterior presentation (P < 0.01). Forty-four (5%) of the 887 kittens were stillborn. This was not correlated with the presentation at expulsion but stillborn kittens were more often female (P = 0.02) and weighed less than those born alive (P = 0.04). The median interkitten time was 30 min (range 2 to 343 min) and 95% were born within 100 min after expulsion of the preceding kitten. The interkitten time as a measure of the progress of parturition was not influenced by the kitten's gender, presentation at expulsion, birth weight, or stillbirth, or by the parity of the queen. The results of this study can be used to develop reference values for parturition parameters in the queen, both to determine whether a given parturition is abnormal and as the basis for a parturition protocol.

Effects of the 3ß-hydroxysteroid dehydrogenase inhibitor trilostane on luteal progesterone production in the dog.

Interference with the pregnancy-maintaining influence of progesterone is the basis of most methods for termination of unwanted pregnancy in dogs. The currently available methods are based on induction of luteolysis or blocking of the progesterone receptor. Inhibition of progesterone synthesis using a competitive inhibitor of 3ß-hydroxysteroid dehydrogenase (3ß-HSD) could be another strategy to terminate unwanted pregnancies. In this study we investigated the effects of the 3ß-HSD inhibitor trilostane on corpus luteum function in non-pregnant bitches. Trilostane was administered orally for seven consecutive days in either the pituitary-independent part of the luteal phase (PIP, start of treatment on D11 after ovulation, n = 6) or the pituitary-dependent part (PDP, start of treatment on D31 after ovulation, n = 6), in an oral dose of about 4.5 mg/kg bw, twice daily. Results were compared with those obtained in control bitches (n = 6). ACTH stimulation tests were performed to assess adrenocortical reserve capacity. Trilostane caused no apparent side effects and ACTH stimulation tests revealed good suppression of cortisol secretion. Trilostane also caused a significant decrease in plasma progesterone concentration. When it was stopped during PIP, progesterone secretion was completely restored and there was no difference in the length of the luteal phase between those dogs and control dogs (99 days, range 70-138 d and 99 d, range 60-112 d, respectively). When trilostane was stopped during PDP there was no post-treatment recovery of progesterone secretion and although the luteal phase tended to be shorter (66 d, range 41-101 d) the difference was not significant (P = 0.09). Plasma prolactin concentration did not increase after the trilostane-induced decrease in plasma progesterone. The interoestrous interval in dogs treated during PIP (234 d, range 175-269 d) or PDP (198 d, range 120-287 d) was not significantly shorter than the control interval (247 d, range 176-313 d). In conclusion, trilostane treatment was effective in decreasing plasma progesterone concentration in bitches during the luteal phase, but the dose regimen used in this study produced less clear-cut inhibition of ovarian steroidogenesis than have other strategies to decrease plasma progesterone concentration. Further studies are warranted to determine whether trilostane can be used to terminate unwanted pregnancy in the bitch without inducing adrenocortical insufficiency.

The pituitary-ovarian axis in dogs with remnant ovarian tissue.

It can be difficult to confirm the presence of remnant ovarian tissue (ROT) in bitches that are presumed to be ovariohysterectomised. A GnRH stimulation test can be used to distinguish ovariectomised bitches from those in anoestrus, but it is uncertain whether the GnRH-induced changes in plasma LH and oestradiol concentrations that occur in intact bitches also occur in ROT-bitches. We report here eighteen ROT-bitches and compare the results of GnRH stimulation tests with those of six ovariectomised and six bitches in anoestrus. The basal (n = 17) and/or GnRH-stimulated (n = 18) plasma oestradiol concentration was above the detection limit of the assay, i.e., < 7 pmol/l, in all ROT-bitches but below the detection limit in all ovariectomised bitches. Basal plasma LH concentration was significantly higher in ROT-bitches (4.1 ± 0.7 µg/L) than those in anoestrus (0.64 ± 0.04 µg/L), and significantly lower than in ovariectomised bitches (20.2 ± 3.6 µg/L). Basal plasma LH concentration was relatively high in bitches in which there was a long interval between ovariectomy and appearance of oestrus. GnRH administration resulted in a significant increase in plasma LH and oestradiol concentrations in ROT-bitches. The GnRH-induced increase and subsequent decline in plasma LH concentration were significantly less in ROT-bitches than in either ovariectomised bitches or those in anoestrus. The GnRH-induced increase in plasma oestradiol concentration was significantly smaller in ROT-bitches than in those in anoestrus. In conclusion, the results of this study demonstrate that in dogs ROT is associated with noticeable changes in the pituitary-ovarian axis and suggest that a GnRH stimulation test may be used to distinguish between completely ovariectomised bitches and those with ROT.

Alterations of the pituitary-ovarian axis in dogs with a functional granulosa cell tumor.

Information on the pituitary-ovarian axis in dogs with a granulosa cell tumor (GCT) is lacking. Therefore, we investigated the plasma concentrations of luteinizing hormone (LH) and estradiol before and after gonadotropin-releasing hormone (GnRH) administration in seven bitches with a functional GCT (GCT-total), of which three were intact (GCT-intact) and four had remnant ovarian tissue (GCT-ROT). The results of the GnRH stimulation test were compared with those in six anestrous and six ovariectomized bitches. The most noteworthy results were as follows. The basal plasma LH concentrations of the GCT-ROT bitches were higher (P<0.05) than those of the anestrous bitches. The increment in the plasma LH concentration after GnRH administration in the GCT-total bitches was lower (P<0.001) than the increments in both the anestrous and ovariectomized bitches. The basal plasma estradiol concentrations in the GCT-total bitches were higher (P<0.001) than those in the anestrous and ovariectomized bitches. In conclusion, the pituitary-ovarian axis is affected in bitches with a functional GCT and is characterized by relatively high plasma LH concentrations in GCT-ROT bitches and a subnormal LH response to GnRH stimulation in all GCT bitches compared with those in anestrous and ovariectomized bitches. The relatively high proportion of dogs with remnant ovarian tissue among the GCT bitches suggests a pathogenetic role for elevated gonadotropin secretion in the pathogenesis of GCT.

Persistent Mullerian duct syndrome in a Miniature Schnauzer dog with signs of feminization and a Sertoli cell tumour.

A 5-year-old male Miniature Schnauzer was presented with unilateral cryptorchidism and signs of feminization. Abdominal ultrasonography revealed an enlarged right testis and a large, fluid-filled cavity that appeared to arise from the prostate. Computed tomography revealed the cavity to be consistent with an enlarged uterine body, arising from the prostate, and showed two structures resembling uterine horns that terminated close to the adjacent testes. The dog had a normal male karyotype, 78 XY. Gonadohysterectomy was performed and both the surgical and the histological findings confirmed the presence of a uterus in this male animal, resulting in a diagnosis of persistent Mullerian duct syndrome (PMDS). The enlarged intra-abdominal testis contained a Sertoli cell tumour. Computed tomography proved to be an excellent diagnostic tool for PMDS.

Minimal external masculinization in a SRY-negative XX male Podenco dog.

Normal mammalian sex differentiation takes place in three genetically controlled steps: chromosomal sex determination (XX or XY), gonadal differentiation and development of the phenotypic sex. Animals are considered to be sex reversed if chromosomal sex determination and gonadal development are not in agreement. In this report, sex reversal is described in a 1.5-year-old Podenco dog that was referred because of suspected recurrent growth of a previously removed os clitoridis in the vulva. With that exception the dog was phenotypically female, but had never been in oestrus and exhibited male behaviour. Abdominal ultrasonography showed a small tubular structure dorsal to the bladder, consistent with a uterus. An ovoid structure resembling a gonad was visible between the right kidney and inguinal canal. Plasma testosterone concentrations before and after GnRH administration indicated the presence of functional testicular tissue. Two testes, each with its epididymis and ductus deferens, and a complete bicornuate uterus were removed surgically. Cytogenetic analysis of peripheral blood lymphocytes showed a normal female karyotype (78, XX). These findings are consistent with the diagnosis of an XX male. PCR analysis of genomic DNA revealed that the SRY gene was absent. In summary, this report describes the first SRY-negative XX male Podenco dog with an almost complete female phenotype despite high basal and stimulated plasma testosterone concentrations. It is hypothesized that the clinical observations in this dog may have been caused by reduced and delayed Müllerian-inhibiting substance secretion and the absence of conversion of testosterone to dihydrotestosterone due to 5alpha-reductase deficiency.

Physiology of the canine anoestrus and methods for manipulation of its length.

Progression from early to late anoestrus is characterized by the appearance of a larger number of gonadotrophin-releasing hormone (GnRH) pulses with a higher amplitude, an increase in the sensitivity of the pituitary to GnRH, an increase in ovarian responsiveness to gonadotrophins, and an increase in basal plasma follicle-stimulating hormone (FSH) concentration. A period of increased luteinizing hormone (LH) pulsatility has been observed shortly before the onset of pro-oestrus. Apart from these changes in the hypothalamus-pituitary-ovary axis, the initiation of a new follicular phase in the bitch is also stimulated by dopaminergic influences other than the accompanying plasma prolactin decrease. Metergoline, a drug which in a low dosage lowers the plasma prolactin concentration via a serotonin-antagonistic pathway, does not shorten the anoestrus; while bromocriptine, in a dosage insufficient to cause a decrease in the plasma prolactin concentration, does prematurely induce a follicular phase. These observations indicate that it is not the decrease in the plasma prolactin concentration, but another dopamine-agonistic influence that plays a crucial role in the transition to a new follicular phase. The dopamine-agonist induced oestrus is associated with a rapid rise in the basal plasma FSH concentration, similar to what is observed during the physiological late anoestrus. Administration of GnRH, eCG and oestrogens may also be used to induce oestrus but with variable results. Oestrus can be prevented surgically or medically, for which purpose progestagens are the most important drugs. The mechanism is still unclear, although it has been demonstrated that with continuing medroxyprogesterone acetate (MPA) treatment the FSH response to GnRH stimulation decreases and changes occur in the pulsatile release of the gonadotrophins. In general, LH pulses coincide with a FSH pulse, but during MPA treatment, LH pulses were observed while there was such a small increase in FSH that it was not recognized as significant FSH pulse.

Pulsatile plasma profiles of FSH and LH before and during medroxyprogesterone acetate treatment in the bitch.

The aim of this study was to investigate the effect of medroxyprogesterone acetate (MPA) on pulsatile secretion of gonadotropins in the bitch. Five intact Beagle bitches were treated with MPA in a dose of 10mg/kg body weight subcutaneously at intervals of 4 weeks for a total of 13 injections, starting during anestrus. The 6-h plasma profiles of luteinizing hormone (LH) and follicle stimulating hormone (FSH) were determined before, and 3, 6, 9, and 12 months after the start of MPA treatment. After 6 months of MPA treatment basal plasma LH concentration was transiently increased significantly. Basal plasma FSH concentration and the area under the curve above the zero level (AUC0) for FSH were significantly higher after 3 months of MPA treatment than before or after 9 and 12 months of treatment. MPA treatment did not significantly affect pulse frequency, pulse amplitude, or AUC above the baseline for either LH or FSH. During treatment 58 significant LH pulses were identified, and although each LH pulse coincided with an increase in plasma FSH concentration, in 17 cases the amplitude of the increase was too small to be recognized as a significant FSH pulse. In conclusion, MPA treatment did not suppress basal plasma gonadotropin levels in the bitches. On the contrary, it caused a temporary rise in the basal concentration of both FSH and LH, which may have been due to a direct effect of MPA on the ovary. In addition, several LH pulses were not accompanied by a significant FSH pulse, suggesting that MPA treatment attenuated the pulsatile pituitary release of FSH.

Hormonal changes in spontaneous and aglépristone-induced parturition in dogs.

To increase our understanding of the endocrine changes associated with parturition in dogs, plasma concentrations of progesterone (P4), 15-ketodihydroprostaglandin F(2alpha) (PGFM), estradiol-17-beta (E2beta), cortisol, ACTH, prolactin (PRL), LH, and FSH were measured in six spontaneously whelping bitches and in six bitches in which parturition was induced with the progesterone-receptor blocker aglépristone on day 58 of pregnancy. Expulsion of pups in the induced group took place in the presence of P4 concentrations that were still elevated. PGFM concentrations increased before parturition in both groups, but levels were lower in the induced bitches. PGFM levels reached a maximum in both groups during parturition and quickly decreased in the spontaneously whelping group after parturition, but remained elevated in the induced group. In both groups, cortisol concentrations reached similar maximum levels during the last 30 h before the onset of expulsion. During the 3 days postpartum, cortisol concentrations were higher in the induced group. The highly variable ACTH concentrations did not differ significantly throughout the study within or between groups. In both groups, E2beta concentrations decreased and PRL concentrations increased between the late gestational period and the 30-h period before parturition. Concentrations of both LH (spontaneously whelping group) and FSH (both groups) decreased between late gestation and the postpartum period. The results of this study illustrate the hormonal changes around parturition in the bitch, and reveal that aglépristone-induced parturition is associated with still incomplete luteolysis, an altered PGFM profile, and elevated postpartum cortisol concentrations as compared with spontaneously whelping dogs.

Estriolum treatment in the bitch: a risk for uterine infection?

Purulent vaginal discharge in a bitch in which ovariohysterectomy has been performed is often caused by inflammation of the uterine stump. The inflammation is due to either cystic endometrial hyperplasia (CEH) induced primarily by progesterone from remnant ovarian tissue or exogenous progestagens, or it is due to the presence of unabsorbed suture material. This report describes a 9-year-old Irish setter with hemopurulent vaginal discharge and non-pruritic symmetrical alopecia, which had undergone ovariohysterectomy 3.5 years ago and which had been treated with estriolum daily for the past 2.5 years because of urinary incontinence. Vaginoscopy revealed hemopurulent discharge throughout the vagina and vestibule. Cytological examination of ultrasound-guided fine-needle aspiration biopsies of a large mass in the hypogastricum, which appeared to be the uterine cervical stump, revealed septic purulent inflammation. The concentration of plasma progesterone was low and the concentration of plasma 17-ss oestradiol did not increase after gonadotrophin-releasing hormone administration. No remnant ovarian tissue was found by abdominal ultrasonography, laparotomy, or histological examination of mesovarian pedicles. Laparotomy revealed uterine stump empyema. Histological examination of the surgically removed mass excluded both CEH and unabsorbed suture material as the cause of the stump empyema. Instead, it is hypothesized that the long-term treatment with estriolum was a causative factor. This suggests that bitches treated with estriolum should be examined regularly.

Plasma oxytocin concentrations during late pregnancy and parturition in the dog.

While oxytocin is widely used in the treatment of dystocia in dogs, there is little information about its secretion before and during normal unassisted whelping. We therefore measured plasma oxytocin concentrations during late pregnancy and the expulsive stage of parturition. Blood samples were collected from eight dogs at 3-min intervals during a 42-min period between the 2nd and 14th day before whelping and during parturition after the birth of 1-3 pups. The litters consisted of 5-15 pups and the progression of the expulsive stage was linear and nearly parallel in the eight bitches. The overall mean (+/-S.D.) plasma oxytocin concentration during late pregnancy was 3.6+/-2.1pg/ml. Mean values in individual dogs ranged from 1.2 to 7.4 pg/ml, but the intra-animal variation was rather small. During the expulsive stage the overall mean (+/-S.D.) plasma oxytocin concentration was 12.9+/-13.9 pg/ml, with mean values in individual dogs ranging from 3.5 to 46 pg/ml. The mean area under the oxytocin curve for parturient dogs was significantly higher (P<0.05) than for pregnant dogs. During the expulsive stage, the peak plasma oxytocin level in individual dogs ranged between 10 and 117 pg/ml. In six of the eight dogs a pup was born during blood collection and in five of these animals the plasma oxytocin concentration increased temporarily during periods of abdominal straining and expulsion. However, straining efforts and expulsion were not consistently associated with a rise in the circulating oxytocin level. It is concluded that in the dog plasma oxytocin levels are higher and more variable during the expulsive stage of parturition than during late pregnancy. Interrelationships between the secretion pattern of oxytocin, the level of uterine contractility, and the progress of fetal expulsion in dogs need further exploration.

Basal and GnRH-induced secretion of FSH and LH in anestrous versus ovariectomized bitches.

The basal and gonadotropin releasing hormone (GnRH)-induced plasma concentrations of follicle stimulating hormone (FSH) and luteinizing hormone (LH) were studied in four anestrous and four ovariectomized (OVX) bitches. Blood samples were obtained via jugular venipuncture 40min before and 0, 10, 20, 30, 60, 90, and 120min after the i.v. administration of synthetic GnRH in a dose of 10microg/kg body weight. The basal plasma FSH and LH concentrations were significantly higher in the OVX bitches than in the anestrous bitches. In the anestrous bitches, the plasma FSH concentration was significantly higher than the pretreatment level at 10, 20, and 30min, whereas the plasma LH concentration was significantly elevated at 10 and 20min. The maximal GnRH-induced plasma FSH concentration in the anestrous bitches did not surpass the lowest plasma FSH concentration in the OVX bitches, whereas the GnRH-induced plasma LH concentrations in the anestrous bitches overlapped with the basal plasma LH concentrations in the OVX bitches. In the OVX bitches, GnRH administration did not induce a significant change in the plasma FSH concentration, whereas the plasma LH concentration increased significantly at 10 and 20min. In conclusion, the results of the present study indicate that in anestrous bitches GnRH challenge results in increased plasma levels of both FSH and LH, whereas in the OVX bitches, in which the basal plasma FSH and LH concentrations are higher, only a rise in the plasma LH concentration is present after GnRH stimulation. The results also suggest that a test to measure plasma concentration of FSH in single samples appears to have potential in verification of neuter status in bitches.