The objectives of this study were to (1) ascertain ovarian follicular status in heifers where the pituitary gonadotrope cells producing LH and FSH were downregulated by long-term treatment with a GnRH agonist and (2) characterize the acute superstimulation of follicular growth in response to FSH in heifers where the pituitary was downregulated with a GnRH agonist. At the start of the study, heifers (3 year old) were implanted with GnRH agonist (n = 20) or received no treatment (control, n = 5). After 6 months, follicular growth in heifers implanted with GnRH agonist was restricted to early antral follicles (2-4 mm). At this time, these heifers were allocated to four groups and follicular development was superstimulated with FSH as follows: group 1 (n = 5) and group 2 (n = 5), a conventional FSH protocol with injections over 4 days (Days 0-3) with Group 2 receiving two times the normal dose of FSH; group 3 (n = 5), a single injection of FSH in saline on Day 0; group 4 (n = 5), a single injection of FSH in slow-release polyvinylpyrrolidone on Day 0. Follicular growth in the control heifers (n = 5) was superstimulated using the conventional 4-day FSH protocol. On Day 5, heifers in groups 1 and 2 had similar (P > 0.05) numbers of follicles in the size categories 6 to 7 mm (12.8 +/- 3.0 follicles) and 8 to 9 mm (6.5 +/- 1.0) as control heifers (6-7 mm, 14.6 +/- 2.2; 8-9 mm, 6.2 +/- 2.2) and six of the former heifers had follicles 10 mm (3.2 +/- 1.4). Also on Day 5, follicular growth for heifers in group 3 was restricted to 4 to 5 mm (11.6 +/- 3.0) and 6 to 7 mm (7.6 +/- 3.7), whereas four out of five heifers in group 4 had follicles of 6 to 7 mm (8.2 +/- 2.3) and three heifers had follicles of 8 to 9 mm (4.6 +/- 2.2) and 10 mm (2.2 +/- 0.9). Injection of exogenous LH on Day 5 induced ovulation in 9 out of 10 heifers in groups 1 and 2, no heifers in group 3, and 2 of 5 heifers in group 4. Plasma concentrations of progesterone 6 days after ovulation were the same (P > 0.05) for heifers in groups 1 and 2 (8.9 +/- 0.9 ng/mL) and control heifers (10.0 +/- 0.9 ng/mL). This study has shown that heifers treated chronically with a GnRH agonist have suppressed ovarian follicular growth but are able to respond to acute superstimulation with FSH. Furthermore, follicles in heifers treated with a GnRH agonist undergo morphologically normal growth in response to FSH and can ovulate and develop into a CL The long-term GnRH agonist treated heifer provides a practical model for repeated ovarian follicular superstimulation, multiple ovulation, and embryo transfer, without the need to control stage of the estrous cycle. (C) 2013 Elsevier Inc. All rights reserved.

The present study reports on attempts to delay puberty in a model marsupial species using the gonadotrophin-releasing hormone (GnRH) agonist deslorelin. Female tammar wallaby pouch young received deslorelin (5 mg) or placebo implants (n=8/group) when they were 193±2 days old. Sexual maturity was significantly delayed in deslorelin-treated animals, with the first successful production of offspring in treated and control animals occurring at 813±62 and 430±42 days of age, respectively. This delay was associated with a period of retarded pouch and teat development. Progesterone concentrations remained at basal levels throughout the first breeding season, indicating the absence of luteal cycles in treated females. Recovery and maturation of the hypothalamic-pituitary axis was a gradual process. Treated animals failed to respond to GnRH challenge at 12 months of age and had a reduced LH response at 18 months of age, before attaining full responsiveness by 24 months of age. Despite this apparent pituitary recovery by 24 months of age, as evidenced by complete teat eversion and LH responsiveness to GnRH, the time to first parturition was significantly delayed beyond this time in three females. This suggests that there may be longer-lasting effects at the level of the ovary and/or on FSH secretion. The significant delay in the onset of sexual maturation in response to chronic GnRH agonist treatment in this model marsupial species may be of practical significance to the management of fertility in captive and semi-free range marsupial populations.

CONTENTS:Stray dogs are a significant problem in large cities. Contraception is an important and useful solution to control the growing population of these dogs. Early-age neutering is an effective technique for canine population control; however, surgical neutering may not be possible in various situations. GnRH-agonist implantation has been successful for long-term reversible contraception in dogs. The efficacy of GnRH-agonist implantation on long-term suppression of reproductive performance was observed in male dogs. Eleven 4-month-old dogs were implanted with 4.7, 9.4 mg deslorelin or placebo. Sexual behaviour and testicular size were monitored every 2 months. Ejaculates were collected and evaluated at 8, 12, 15, 18, 24, 30, 32, 34 and 36 months of age. Dogs implanted with placebo were found to be healthy and in normal reproductive status. Most dogs (3/4) implanted with 4.7 mg deslorelin showed male sexual behaviour at age of 34 months old. From this group, two dogs had normal semen quality, while semen could not be collected from the other dog, and after castration, no sperm were obtained following epididymal flushing. One dog implanted 4.7 mg deslorelin and four dogs implanted with 9.4 mg deslorelin remained in the non-pubertal reproductive status at 30-34 months. The delay to puberty was longer in dogs implanted with higher dose of GnRH agonist. Implantation of pre-pubertal dogs with high doses of GnRH agonist will delay the onset of puberty and may be an effective strategy to reduce the number of unwanted breedings.

Kangaroo populations frequently reach high densities under favourable conditions, particularly near urban areas, where sociopolitical constraints limit management options. This study investigates new approaches to kangaroo management that focus on controlling the fertility of females. In particular, it investigates long-acting contraceptive implants containing the gonadotrophin-releasing hormone agonist deslorelin (Suprelorin (R)) or the synthetic progestin, levonorgestrel. It shows that both contraceptive implants are effective in a model marsupial species, the tammar wallaby (Macropus eugenii), and gives a detailed understanding of the effects of these two agents on the physiology of this species. Captive and field investigations on the eastern grey kangaroo (Macropus giganteus) have shown that long-term fertility control can be achieved in that species using either of these contraceptive implants, with no detectible side effects. However, these two contraceptive implants have different modes of action. Levonorgestrel treatment has no effect on follicular development but the luteinising hormone (LH) surge, and hence ovulation, is inhibited. Suprelorin, on the other hand, inhibits the synthesis and release of LH and follicle-stimulating hormone from the pituitary, thereby inhibiting follicular development and ovulation. The methods of administration and contraceptive duration also differ. These differences can be exploited to tailor kangaroo management programs to specific populations to ensure long-term contraception and population management.

Contents We tested the effect of dose of GnRH superagonist on pituitary and testicular function in a study with four groups of four male dogs. The Controls received blank implants and the other three groups received implants containing 3, 6 or 12 mg deslorelin (d-Trp6-Pro9-des-Gly10-GnRH ethylamide). In all deslorelin-treated groups, there was initially an acute increase in plasma concentrations of LH and testosterone, followed by declines such that both hormones became undetectable after approximately 12 days. There was a dose-response in some of these early aspects of the hormone profiles. With respect to long-term effects of treatment, the 12-mg dose had significantly greater effects than the smaller doses for the duration of minimum testicular volume [366 +/- 77, mean +/- SEM (3 mg), 472 +/- 74 (6 mg), and 634 +/- 59 (12 mg) days], absence of ejaculate [416 +/- 88 (3 mg), 476 +/- 83 (6 mg), and 644 +/- 67 (12 mg) days], undetectable plasma concentrations of LH and testosterone [367 +/- 64 (3 mg), 419 +/- 72 (6 mg), and 607 +/- 69 (12 mg) days], the delay until complete recovery of LH and testosterone secretion [394 +/- 65 (3 mg), 484 +/- 72 (6 mg) and 668 +/- 47 (12 mg) days], and the delay until testes had regrown to normal volume [408 +/- 77 (3 mg), 514 +/- 74 (6 mg), 676 +/- 59 (12 mg) days]. The time taken to restore full ejaculates was also longest for the 12-mg dose: 716 +/- 67 (12 mg) days vs 440 +/- 66 (3 mg) and 538 +/- 83 (6 mg) days after implantation. There was no correlation between delay to recovery of normal ejaculate quality and body mass. We conclude that the dose-response relationship with deslorelin implants is not expressed with respect to the degree of suppression of reproduction, but on the maximum duration of suppression and thus to delay until recovery.

The objective of this study was to evaluate the expression of estrogen receptor alpha (ERα) and progesterone receptor (PR) in normal mammary tissues of bitches before and after the bitch was implanted with the GnRH agonist, deslorelin.Fifteen mature bitches with no pathological condition of the mammary gland, and during anestrus, were selected and divided into two groups: five bitches were implanted with placebo (placebo group); and the ten bitches were implanted with 9.4 mg deslorelin (deslorelin group).Mammary tissues were collected from all bitches before implantation and at 2 and 12 weeks after implantation.The expressions of ERα and PR were investigated by using immunohistochemistry.The highest scores of both ERα and PR were found at 2 weeks after deslorelin implantation which differed significantly from those of other stages.At 12 weeks after implantation, ERα score decreased to the same level as before implantation while PR score tended to decrease, though not significantly differed from at 2 weeks after implantation.In placebo group, neither difference of ERα nor PR scores was observed at any time of the implantation.This finding indicated that deslorelin may effect the expression of ERα and PR in the bitch mammary tissue, in which it may stimulate the expression of ERα and PR at 2 weeks after deslorelin implantation.On the other hand, at 12 weeks after deslorelin implantation, ERα returned to the same level as before deslorelin implantation while PR decreased but not significantly different.This may indicate that ERα was more sensitive to deslorelin implantation than PR.In addition, the similar level of PR expression between the deslorelin and placebo group at 2 weeks after the implantion also suggests that some other factors besides deslorelin, or together with deslorelin, may have the effects on PR expression in the bitch mammary tissue.

The present study is part of a programme of research designed to evaluate the efficacy of the GnRH superagonist,deslorelin (D-Trp6-Pro9-des-Gly10-LHRH ethylamide), as a contraceptive for male dogs. Adult dogs were assigned to a completely randomized design comprising six groups of four animals. Each dog in the control group received a blank implant (placebo) and each dog in the other five groups received a 6 mg deslorelin implant. One group of deslorelin treated dogs was sacrificed on each of days 16, 26, 41, 101 and 620, and testicular and prostate tissues were collected for study by light and electron microscopy. On days 16 and 26 after implantation, we observed partial disruption of the seminiferous tubules, with early spermatids shed into the lumen. On days 41 and 101 after implantation, 90–100% of the seminiferous tubules were atrophic and aspermatogenic.On day 101 after implantation, 99% of all sections showed atrophy of the epithelium and shrinkage of epithelial height in the ductus epididymides. On days 41 and 101 after implantation, prostate tissue showed complete atrophy of the glandular epithelium (100% of sections) and an apparent increase in the relative proportion of connective tissue. At the electron microscopic level, in dogs treated with deslorelin for 41 and 101 days, the Sertoli cells were smaller and their nucleoli appeared smaller than in the control dogs. The nucleoli of the Leydig cells were atrophied and prostate glandular epithelium showed reduced epithelial height, a trophy of the nucleolus and an absence of secretory granules.Tissues collected during the recovery phase revealed a complete recovery of spermatogenesis. In conclusion, slow release implants containing deslorelin induce a striking a trophy of the testes and prostate gland by 26 days after implantation, explaining the previously reported loss of ejaculate and arrest of sperm output. At histological level,the entire process appears to be completely reversible, in accordance with data on endocrine variables and semen production.

Surgical castration in ferrets has been implicated as an etiological factor in the development of hyperadrenocorticism in this species due to a castration-related increase in plasma gonadotropins. In search for a suitable alternative, the effect of treatment with the depot GnRH-agonist implant, deslorelin, on plasma testosterone concentrations and concurrent testes size, spermatogenesis, and the typical musky odor of intact male ferrets was investigated. Twenty-one male ferrets, equally divided into three groups, were either surgically castrated, received a slow release deslorelin implant or received a placebo implant. Plasma FSH and testosterone concentrations, testis size and spermatogenesis were all suppressed after the use of the deslorelin implant. The musky odor in the ferrets which had received a deslorelin implant was less compared to the ferrets which were either surgically castrated or had received a placebo implant. These results indicate that the deslorelin implant effectively prevents reproduction and the musky odor of intact male ferrets and is therefore considered a suitable alternative for surgical castration in these animals.

Contraception is an essential tool for controlling reproduction in captive and free-ranging lions. This paper describes the treatment and contraception of 23 captive and 40 free-ranging lionesses (Panthera leo) and four captive tigers (Panthera tigris) in South Africa using 3 × 4.7 mg, 2 × 4.7 mg, 9.4 mg or 4.7 + 9.4 mg deslorelin implants. Thirty-one lionesses were treated more than once at 11- to 60-month intervals. In Brazil, two lionesses were treated with 9.4-mg implants and faecal progesterone and oestradiol concentrations were monitored for 920 days. All combinations of deslorelin showed the length of contraception to be around 30 months with one 3 × 4.7 mg treatment lasting 40 months in one captive lioness. The mean time taken to reconception was 30.1 months for the 3 × 4.7 mg combination. The faecal analyses of the lionesses in Brazil reflected quiescent ovarian activity for periods of 17 and 30 months, respectively, when small oestradiol peaks but no progesterone peaks started to appear. This confirmed the field observations in South Africa. No side effects occurred although several of the lionesses were treated repeatedly for up to 8 years. Deslorelin (Suprelorin) is a safe and effective means of controlling reproduction in captive or free-ranging populations of lions. Where contraception is to be maintained, the implementation of implants at 24-month intervals is recommended.

The present study evaluated the effects of chronic GnRH agonist (deslorelin) treatment on sexual maturation in the male tammar wallaby. Slow-release deslorelin or placebo implants were administered to male pouch young (n = 10/group) when they were between 180 and 200 days old, to determine if disruption of the pituitary-testicular axis during development altered the timing of sexual maturation or had long-term effects on adult reproductive function. Deslorelin treatment caused retardation of testicular growth and reduced the serum FSH and testosterone concentrations between 12 and 24 mo of age. Maturation of the hypothalamic-pituitary-testicular axis was also delayed in treated animals at 13 and 19 mo of age. Despite these alterations in the pattern and timing of neuroendocrine development, sexual maturation was not permanently blocked in these animals and deslorelin-treated animals reached sexual maturity at the same age as treated animals, as evidenced by a fully functional pituitary-testicular axis and proven fertility at 25 mo of age. The ability of the treated animals to reach puberty at the same time as control animals, despite delayed maturation of the hypothalamic-pituitary-testicular axis, suggests that puberty in the male tammar wallaby is additionally regulated by other, gonadotropin-independent factors.

The present study tested whether exogenous gonadotrophin-releasing hormone (GnRH) and luteinising hormone (LH) can stimulate LH and testosterone secretion in dogs chronically treated with a GnRH superagonist. Twenty male adult dogs were assigned to a completely randomised design comprising five groups of four animals. Each dog in the control group received a blank implant (placebo) and each dog in the other four groups received a 6-mg implant containing a slow-release formulation of deslorelin (d-Trp6-Pro9-des-Gly10-LH-releasing hormone ethylamide). The same four control dogs were used for all hormonal challenges, whereas a different deslorelin-implanted group was used for each challenge. Native GnRH (5 microg kg(-1) bodyweight, i.v.) was injected on Days 15, 25, 40 and 100 after implantation, whereas bovine LH (0.5 microg kg(-1) bodyweight, i.v.) was injected on Days 16, 26, 41 and 101. On all occasions after Day 25-26 postimplantation, exogenous GnRH and LH elicited higher plasma concentrations of LH and testosterone in control than deslorelin-treated animals (P < 0.05). It was concluded that, in male dogs, implantation of a GnRH superagonist desensitised the pituitary gonadotrophs to GnRH and also led to a desensitisation of the Leydig cells to LH. This explains, at least in part, the profound reduction in the production of androgen and spermatozoa in deslorelin-treated male dogs.

Post-transcription Regulation of mRNA and Hormone Synthesis and Release at the Anterior Pituitary Gland: Information derived from the recovery of pituitary gonadotrope cell function after therapy with a GnRH agonist

Eastern grey kangaroos are widespread on the east coast of Australia and frequently reach high densities in reserves and parkland near urban areas. Management of these populations is highly contentious and non-lethal fertility-control technologies are sought as an alternative option to manage population size. This study evaluated the potential of slow-release gonadotrophin- releasing hormone agonist (deslorelin) implants to inhibit reproduction in female kangaroos. Deslorelin treatment effectively inhibited reproduction in adult females for periods of 559 +/- 111 days (n = 6) and 651 +/- 21 days ( n = 5) after administration of one or two 10-mg implants respectively. Animals treated with the lower dosage tended to resume breeding earlier than those that received a total of 20 mg of deslorelin ( minimum duration of 18 months). Deslorelin treatment had no effect on blastocyst reactivation in a single treated female and repeat treatment had no negative side-effects. This study has demonstrated that slow-release deslorelin implants can successfully inhibit reproduction for extended periods in the female eastern grey kangaroos. This approach may have potential application in reproductive management of problem kangaroo populations.

The prevention of breeding in animals using GnRH analogues has been the object of research over many years. Recently, a new drug delivery formulation was developed which enabled the development of products that could be commercialised for veterinary use. The formulation has now been approved in certain countries for use in male dogs, and applications are being expanded to cover repeat usage, extended duration, use in females, other indications and other animal species. With respect to repeat usage, dogs have been re-implanted for four consecutive doses and monitored until they returned to normal steroidogenesis. All dogs returned to normal steroidogenesis following cessation of treatment. In females, it was previously shown that implanted bitches with progesterone <5 ng/mL at the time of implantation had an induced estrus. In a new study at Chulalongkorn University, implanting female pups at around 4 mo prevented this occurrence, whereas implantation at 7 mo did not.

The contraceptive and endocrine effects of long-term treatment with implants containing the GnRH agonist deslorelin were investigated in female tammar wallabies (Macropus eugenii). Fertility was successfully inhibited for 515 +/- 87 days after treatment with a 5 mg deslorelin implant (n = 7), while control animals gave birth to their first young 159 +/- 47 days after placebo implant administration (n = 8). The duration of contraception was highly variable, ranging from 344 to 761 days. The strict reproductive seasonality in the tammar wallaby was maintained once the implant had expired. This inhibition of reproduction was associated with a significant reduction in basal LH concentrations and a cessation of oestrous cycles, as evidenced by low progesterone concentrations. There was evidence to suggest that some aspect of either blastocyst survival, luteal reactivation, pregnancy or birth may be affected by deslorelin treatment in some animals. These results show that long-term inhibition of fertility in the female tammar wallaby is possible using slow-release deslorelin implants. The effects of deslorelin treatment were fully reversible and there was no evidence of negative side effects. Slow-release GnRH agonist implants may represent a practicable method for reproductive management of captive and semi-wild populations of marsupials.

Gonadotropin releasing hormone (GnRH) has long been recognized as a potential target for the control and management of fertility in female animals. Attempts to apply GnRH-based technology to manage fertility have focussed on the development of GnRH agonists, antagonists and vaccines. All of these methods have potential, but the widespread application of these technologies has been limited to date. The greatest advance in the use of GnRH-based technology for long-term fertility control in recent years has been the development and commercialization of depot formulations that release GnRH agonists for periods of up to 1 year. These products have a broad range of potential applications in production and domestic animal management. The further development and commercialization of GnRH vaccines has been hampered by the variability of response between individual animals. The need to use adjuvant and multiple boosters also make this a less attractive option than the current GnRH agonist technology. However, GnRH vaccines have the advantage that they do not induce the initial stimulatory response that follows GnRH agonist administration. GnRH antagonists and GnRH-toxin conjugates show promise but are in an earlier phase of development. To date, no depot or long-acting formulations of antagonists have been developed. GnRH-toxin conjugates have yet to achieve permanent sterilization, but further dose-response trials may advance this approach.