Clinical use of Gonadotropin-Releasing Hormone (GnRH) Agonists in Companion Animals: An Overview

In dogs, cats, ferrets, and pet birds, reproductive physiology is under the control of the hypothalamic­pituitary­-gonadal (HPG) axis. Many hormones are responsible for estrus and reproduction, the most significant being luteinizing hormone (LH), follicle stimulating hormone (FSH), and gonadotropin-releasing hormone (GnRH). Short-lived GnRH is released in a pulsatile fashion from the hypothalamus and acts on the pars distalis of the pituitary gland to stimulate the synthesis and release of the gonadotropins, FSH and LH (Figure 1). Secretion of these gonadotropins into the circulation lead to changes gonadal hormone production and reproductive function.

Figure 1: Regulation of gonadal secretion via the hypothalamic-pituitary-gonadal axis.

Chemical modification of the native short-acting GnRH molecule has led to development of long-acting, potent GnRH agonists, which have been used as a medical means of management for a number of reproductive issues and diseases of companion animals (1-3). GnRH agonists may either stimulate estrus or effectively sterilize the patient, depending on the duration of action and the dosage applied. These agents work by initially stimulating gonadotrophin secretion, followed shortly thereafter with desensitization of the GnRH receptor to the GnRH agonist (Figure 2). This results in a temporary but long-term, fully-reversible down-regulation of gonadotrophin secretion, leading to suppression of reproduction function in both male and female animals (4).

Figure 2: GnRH agonists initially stimulate pituitary LH and FSH secretion, followed by desensitization and down-relation of gonadotrophin secretion.

In recent years, effective low-dose, slow-release implants containing potent GnRH agonists have been released for use in veterinary medicine, especially in Europe and Australia. In companion animals, the deslorelin implant (Suprelorin, Virbac) is the most commonly GnRH agonist used in small animals (5). Deslorelin implants work by lowering pituitary gonadotrophin section. This is not a permanent change but depending on the deslorelin dose, can last up to many months. The implant does not have to be removed, but subsequent doses are needed to sustain the effect.

Unfortunately, GnRH agonist availability is limited in the United States. Although there are GnRH agonists available that are approved for the treatment of human diseases, such as prostate cancer, they are costly and not financially feasible for a pet owner to consider. To date, deslorelin acetate (Suprelorin, Virbac Animal Health, Fort Worth, TX, USA) is the only GnRH agonist that is currently available in the United States but only for the treatment of adrenal disease in ferrets (6). However, it is not legal to use Suporelin in non-ferret species in the United States and extra-label use is explicitly prohibited.

The aim of this blog is to review the applications and treatments of the deslorelin (GnRH agonist) currently used in companion animal medicine.

Deslorelin Use in Intact Male Dogs
In male dogs treated with deslorelin, this GnRH agonist leads to decreased gonadotropins secretion and resultant lowered plasma testosterone concentrations, decreased testicular volume, and azoospermia (1-3,7-9). However, the response to this GnRH agonist can be very variable from one dog to another, and the duration of inhibition of testosterone secretion depends both on the concentration of the deslorelin implant and the size of the dog.

Many studies have confirmed that use of GnRH agonists for reversible chemical sterilization in male dogs is both safe and well-tolerated (7-9). Furthermore, repeated implantation can be used to maintain circulating testosterone at low concentrations. If the deslorelin implants are stopped, the treated dogs will regain normal serum testosterone levels within a few weeks, with full recovery of seminal quality once the GnRH implant has lost its efficacy (10,11).

In addition to contraception, GnRH agonists have also been used to reduce the size of the prostate gland, an effect that may be useful in dogs with benign prostatic hyperplasia (12-14).

Deslorelin in Intact Male Cats
As in dogs, GnRH agonists are gaining increased importance in feline reproductive medicine (2,3,15). In intact male cats, deslorelin implants induce chemical sterilization, as in dogs. In these cats, testosterone concentrations decline rapidly to undetectable values by 3 weeks after implantation and remain low for weeks in the majority of the tomcats treated. As the circulating testosterone falls, the testicular volume decreases and penile spines disappear.

However, high individual variability has been reported, with the duration of efficacy varying between 6 and 24 months (15-17). Similar to dogs, it is possible to use repeated implantation of deslorelin to sustain the drug’s effect.

Deslorelin in Intact Bitches
Although deslorelin implants are only approved for male dogs in Europe (and again, not at all in the USA), studies have been performed in the bitch to investigate its use either as a contraceptive or a method of estrus induction (1-3,18-20).

The first step in the mechanism of action of all GnRH agonists is the stimulation in FSH and LH secretion (so-called "flare-up effect") (4). This followed within a few days by a profound hypogonadal effect (i.e., decrease in FSH and LH levels), which is achieved through receptor down-regulation by internalization of receptors. Generally this induced and reversible hypogonadism is the therapeutic goal, as noted above for the male dogs and cats (1-3).

The initial stimulating effect on gonadotrophin secretion is more pronounced in females than in males (18,19). Thus, estrus induction will be observed in the majority of bitches implanted in anestrus. If pregnancy is achieved, most recommend removal of the deslorelin implant either at the beginning of proestrus, at the time of the LH surge, or at the time of ovulation (2,3,18,19). However, some have reported that some bitches carried their pregnancies to term without the implant being removed, suggesting that down-regulation of gonadotrophin secretion may not be strong enough to induce luteal failure in all bitches.

For use as a contraceptive method, the main problem with using deslorelin implants in female dogs is estrus induction, as discussed above (20). For this reason, deslorelin implants cannot be considered a viable alternative to other, current used contraception in bitches.

Deslorelin in Intact Queens
In contrast to female dogs, the main indication for the use of deslorelin in the female queen is estrus inhibition. Studies have confirmed that this GnRH agonist can be used to effectively suppress ovarian activity (15,20-22), but the duration of inhibition was highly variable among the individual queens depending on the dosage administered. However, deslorelin generally suppresses ovarian activity for many months.

Deslorelin in Spayed Bitches with Urinary Incontinence
Ovariectomy results in elevated circulating concentrations of pituitary LH because of the lack of gonadal negative-feedback on the pituitary gland. LH receptors are present throughout the canine urinary tract (23-25), and it has been postulated that elevated gonadotropins may contribute to the development of urethral sphincter mechanism incompetence (26,27).

Treatment of bitches with long-acting GnRH agonists, such as delorelin, downregulates LH secretion for prolonged time periods and temporarily restores continence to incontinent bitches for varying durations, ranging from 50-738 days (26,27). Similar to alpha-adrenergic agonists (e.g., phenylpropanolamine; PPA), GnRH agonists are not completely effective for the treatment of this urinary incontinence. However, unlike PPA, no adverse effects to GnRH agonists have been reported.

Deslorelin in Ferrets with Adrenal Disease
As in dogs and cats, deslorelin is also a promising and suitable method for contraception in ferrets (28-31). However, GnRH agonists are useful in medical management of ferrets suffering from adrenal disease (hyperadrenocorticism) a common disease in castrated males and females (32-34). In one study of ferrets with adrenal disease, the clinical signs (e.g., vulvar swelling, pruritus, sexual behavior, and aggression) were reduced or markedly suppressed within 14 days of implantation of the deslorelin (34). The time for signs to recur in these ferrets ranged from 8.5–20.5 months (34).

References:

1. Trigg TE, Doyle AG, Walsh JD, et al. A review of advances in the use of the GnRH agonist deslorelin in control of reproduction. Theriogenology 2006;66:1507-1512. 
2. Fontaine E, Fontbonne A. Clinical use of GnRH agonists in canine and feline species. Reprod Domest Anim 2011;46:344-353. 
3. Lucas X. Clinical use of deslorelin (GnRH agonist) in companion animals: a review. Reprod Domest Anim 2014;49 Suppl 4:64-71. 
4. Ortmann O, Weiss JM, Diedrich K. Gonadotrophin-releasing hormone (GnRH) and GnRH agonists: mechanisms of action. Reprod Biomed Online 2002;5 Suppl 1:1-7. 
5. Suprelorin (deslorelin acetate). Summary report from the European Medicines Agency. 
6. Suprelorin F. Package insert. Fort Worth, Texas: Virbac Animal Health 
7. Junaidi A, Williamson PE, Martin GB, et al. Pituitary and testicular endocrine responses to exogenous gonadotrophin-releasing hormone (GnRH) and luteinising hormone in male dogs treated with GnRH agonist implants. Reprod Fertil Dev 2007;19:891-898. 
8. Junaidi A, Williamson PE, Martin GB, et al. Dose-response studies for pituitary and testicular function in male dogs treated with the GnRH superagonist, deslorelin. Reprod Domest Anim 2009;44:725-734. 
9. Romagnoli S, Siminica A, Sontas BH, et al. Semen quality and onset of sterility following administration of a 4.7-mg deslorelin implant in adult male dogs. Reprod Domest Anim 2012;47 Suppl 6:389-392. 
10. Trigg TE, Wright PJ, Armour AF, et al. Use of a GnRH analogue implant to produce reversible long-term suppression of reproductive function in male and female domestic dogs. J Reprod Fertil Suppl 2001;57:255-261. 
11. Gentil M, Hoffmann B, Spang A, et al. Restart of steroidogenesis in dogs during recrudescence of testicular function following downregulation with a GnRH-agonist implant. Cell Tissue Res 2012;350:513-523. 
12. Vickery BH, McRae GI, Bonasch H. Effect of chronic administration of a highly potent LHRH agonist on prostate size and secretory function in geriatric dogs. Prostate 1982;3:123-130. 
13. Nizanski W, Levy X, Ochota M, et al. Pharmacological treatment for common prostatic conditions in dogs - benign prostatic hyperplasia and prostatitis: an update. Reprod Domest Anim 2014;49 Suppl 2:8-15. 
14. Polisca A, Orlandi R, Troisi A, et al. Clinical efficacy of the GnRH agonist (deslorelin) in dogs affected by benign prostatic hyperplasia and evaluation of prostatic blood flow by Doppler ultrasound. Reprod Domest Anim 2013;48:673-680. 
15. Goericke-Pesch S, Wehrend A, Georgiev P. Suppression of fertility in adult cats. Reprod Domest Anim 2014;49 Suppl 2:33-40. 
16. Goericke-Pesch S, Georgiev P, Antonov A, et al. Clinical efficacy of a GnRH-agonist implant containing 4.7 mg deslorelin, Suprelorin, regarding suppression of reproductive function in tomcats. Theriogenology 2011;75:803-810. 
17. Goericke-Pesch S, Georgiev P, Fasulkov I, et al. Basal testosterone concentrations after the application of a slow-release GnRH agonist implant are associated with a loss of response to buserelin, a short-term GnRH agonist, in the tom cat. Theriogenology 2013;80:65-69. 
18. Volkmann DH, Kutzler MA, Wheeler R, et al. The use of deslorelin implants for the synchronization of estrous in diestrous bitches. Theriogenology 2006;66:1497-1501. 
19. Fontaine E, Mir F, Vannier F, et al. Induction of fertile oestrus in the bitch using Deslorelin, a GnRH agonist. Theriogenology 2011;76:1561-1566. 
20. Maenhoudt C, Santos NR, Fontaine E, et al. Results of GnRH agonist implants in oestrous induction and oestrous suppression in bitches and queens. Reprod Domest Anim 2012;47 Suppl 6:393-397. 
21. Goericke-Pesch S. Reproduction control in cats: new developments in non-surgical methods. J Feline Med Surg 2010;12:539-546. 
22. Goericke-Pesch S, Georgiev P, Atanasov A, et al. Treatment of queens in estrus and after estrus with a GnRH-agonist implant containing 4.7 mg deslorelin; hormonal response, duration of efficacy, and reversibility. Theriogenology 2013;79:640-646. 
23. Coit VA, Dowell FJ, Evans NP. Neutering affects mRNA expression levels for the LH- and GnRH-receptors in the canine urinary bladder. Theriogenology 2009;71:239–47.
24. Ponglowhapan S, Church DB, Khalid M. Differences in the expression of luteinizing hormone and follicle-stimulating hormone receptors in the lower urinary tract between intact and gonadectomised male and female dogs. Domest Anim Endocrinol 2008;34:339-351. 
25. Reichler IM, Welle M, Sattler U, et al. Comparative quantitative assessment of GnRH- and LH-receptor mRNA expression in the urinary tract of sexually intact and spayed female dogs. Theriogenology 2007;67:1134–42.
26. Reichler IM, Hubler M, Jöchle W, et al. The effect of GnRH analogs on urinary incontinence after ablation of the ovaries in dogs. Theriogenology 2003;60:1207–16.
27. Reichler IM, Jöchle W, Piché CA, , et al. Effect of a long-acting GnRH analogue or placebo on plasma LH/FSH, urethral pressure profiles and clinical signs of urinary incontinence due to sphincter mechanism incompetence in bitches. Theriogenology 2006;66:1227–36.
28. Schoemaker NJ, van Deijk R, Muijlaert B, et al. Use of a gonadotropin releasing hormone agonist implant as an alternative for surgical castration in male ferrets (Mustela putorius furo). Theriogenology 2008;70:161-167. 
29. Prohaczik A, Kulcsar M, Trigg T, et al. Comparison of four treatments to suppress ovarian activity in ferrets (Mustela putorius furo). Vet Rec 2010;166:74-78. 
30. Goericke-Pesch S, Wehrend A. The use of a slow release GnRH-agonist implant in female ferrets in season for oestrus suppression. Schweiz Arch Tierheilkd 2012;154:487-491. 
31. van Zeeland YR, Pabon M, Roest J, et al. Use of a GnRH agonist implant as alternative for surgical neutering in pet ferrets. Vet Rec 2014;175:66. 
32. Rosenthal KL, Peterson ME, Quesenberry KE, et al. Hyperadrenocorticism associated with adrenocortical tumor or nodular hyperplasia of the adrenal gland in ferrets: 50 cases (1987-1991). J Am Vet Med Assoc 1993;203:271-275. 
33. Schoemaker NJ, Teerds KJ, Mol JA, et al. The role of luteinizing hormone in the pathogenesis of hyperadrenocorticism in neutered ferrets. Mol Cell Endocrinol 2002;197:117-125. 
34. Wagner RA, Piche CA, Jochle W, et al. Clinical and endocrine responses to treatment with deslorelin acetate implants in ferrets with adrenocortical disease. Am J Vet Res 2005;66:910-914. 

Top Endocrine Publications of 2014: Canine and Feline Reproductive Endocrinology

As I've done for the last 5 years, I’ve now finished compiling a fairly extensive list of references concerning canine and feline endocrinology that were written last year (in 2014). I’ll be sharing these with you over the next few months, as well as reviewing a few of the best papers from my lists of clinical endocrine publications.

In my last post, I provided my last list for the 2013 papers on canine and feline endocrine reproduction, so I've decided to start this year off with papers that deal with the same theme of endocrine disorders of the canine and feline gonads, prostate, and mammary gland.

Listed below are 23 papers published in 2014 that deal with a variety of topics of importance for reproductive endocrinology in dogs and cats. These range from the identification and study of kisspeptin (a protein ligand that activate GnRH neurons) in dogs (1) to the use of relaxin measurements to diagnose pregnancy status (2); as well as from a study of the effects of GnRH agonist and antagonists during the postnatal period in cats (3) to the effects of GnRH immunization for treatment of urinary incontinence in spayed bitches (4).

Other publication included studies dealing with suppression of fertility in dogs and cats (4-6,12,13,14,21) to endocrinologic investigations of pyometra (9), ovarian cysts (10), mammary neoplasia (16,19), and benign prostatic hyperplasia and prostatitis (17); and finally, from a study of oxytocin and social bonding in dogs (20) to a review of the influence of sex hormones on seizures in dogs and man (22).

References:

1. Albers-Wolthers KH, de Gier J, Kooistra HS, et al. Identification of a novel kisspeptin with high gonadotrophin stimulatory activity in the dog. Neuroendocrinology 2014;99:178-189.
2. Bergfelt DR, Peter AT, Beg MA. Relaxin: a hormonal aid to diagnose pregnancy status in wild mammalian species. Theriogenology 2014;82:1187-1198.
3. Carranza A, Faya M, Merlo ML, et al. Effect of GnRH analogs in postnatal domestic cats. Theriogenology 2014;82:138-143.
4. Donovan CE, Gordon JM, Kutzler MA. Gonadotropin-releasing hormone immunization for the treatment of urethral sphincter mechanism incompetence in ovariectomized bitches. Theriogenology 2014;81:196-202.
5. Fagundes AK, Oliveira EC, Tenorio BM, et al. Injection of a chemical castration agent, zinc gluconate, into the testes of cats results in the impairment of spermatogenesis: a potentially irreversible contraceptive approach for this species? Theriogenology 2014;81:230-236.
6. Favre RN, Bonaura MC, Praderio R, et al. Effect of melatonin implants on spermatogenesis in the domestic cat (Felis silvestris catus). Theriogenology 2014;82:851-856.
7. Goericke-Pesch S, Wehrend A, Georgiev P. Suppression of fertility in adult cats. Reprod Domest Anim 2014;49 Suppl 2:33-40.
8. Hoglund OV, Hagman R, Olsson K, et al. Intraoperative changes in blood pressure, heart rate, plasma vasopressin, and urinary noradrenalin during elective ovariohysterectomy in dogs: repeatability at removal of the 1st and 2nd ovary. Veterinary Surgery 2014;43:852-859.
9. Jitpean S, Holst BS, Hoglund OV, et al. Serum insulin-like growth factor-I, iron, C-reactive protein, and serum amyloid A for prediction of outcome in dogs with pyometra. Theriogenology 2014;82:43-48.
10. Knauf Y, Bostedt H, Failing K, et al. Gross pathology and endocrinology of ovarian cysts in bitches. Reprod Domest Anim 2014;49:463-468.
11. Kobayashi M, Hori T, Kawakami E. Efficacy of low-dose human chorionic gonadotropin therapy in dogs with spermatogenic dysfunction: a preliminary study. Reprod Domest Anim 2014;49:E44-47.
12. Lucas X. Clinical use of deslorelin (GnRH agonist) in companion animals: a review. Reprod Domest Anim 2014;49 Suppl 4:64-71.
13. Maenhoudt C, Santos NR, Fontbonne A. Suppression of fertility in adult dogs. Reprod Domest Anim 2014;49 Suppl 2:58-63.
14. Marino G, Rizzo S, Quartuccio M, et al. Deslorelin implants in pre-pubertal female dogs: short- and long-term effects on the genital tract. Reprod Domest Anim 2014;49:297-301.
15. Meloni T, Comin A, Rota A, et al. IGF-I and NEFA concentrations in fetal fluids of term pregnancy dogs. Theriogenology 2014;81:1307-1311.
16. Michel E, Rohrer Bley C, Kowalewski MP, et al. Prolactin--to be reconsidered in canine mammary tumourigenesis? Vet Comp Oncol 2014;12:93-105.
17. Nizanski W, Levy X, Ochota M, et al. Pharmacological treatment for common prostatic conditions in dogs - benign prostatic hyperplasia and prostatitis: an update. Reprod Domest Anim 2014;49 Suppl 2:8-15.
18. Parker K, Snead E. Atypical presentation of ovarian remnant syndrome in a dog. J Am Anim Hosp Assoc 2014;50:e1-5.
19. Queiroga FL, Perez-Alenza MD, Gonzalez Gil A, et al. Clinical and prognostic implications of serum and tissue prolactin levels in canine mammary tumours. Vet Rec 2014;175:403.
20. Romero T, Nagasawa M, Mogi K, et al. Oxytocin promotes social bonding in dogs. Proc Natl Acad Sci U S A 2014;111:9085-9090.
21. Schafer-Somi S, Kaya D, Gultiken N, et al. Suppression of fertility in pre-pubertal dogs and cats. Reprod Domest Anim 2014;49 Suppl 2:21-27.
22. Van Meervenne SA, Volk HA, Matiasek K, et al. The influence of sex hormones on seizures in dogs and humans. Vet J 2014;201:15-20.
23. Volta A, Manfredi S, Vignoli M, et al. Use of contrast-enhanced ultrasonography in chronic pathologic canine testes. Reprod Domest Anim 2014;49:202-209.

Top Endocrine Publications of 2013: Canine and Feline Reproductive Endocrinology

In my tenth compilation of the canine and feline endocrine publications, I’m moving on to endocrine disorders of the canine and feline gonads and mammary gland. Listed below are 20 papers published in 2013 that deal with a variety of topics of importance for reproductive endocrinology in dogs and cats.

Of all of these publications, one of the most common and clinically useful topics involves the use of gonadotropin-releasing hormone (GnRH) agonists for management of a variety of disorders. These include the use of GnRH agonist for treatment of queens in estrus and after estrus (3), for reproductive control in queens (4) and tom cats (5), and in dogs affected by benign prostatic hyperplasia (12).

On my next blog post, I will review the mechanism of action for the GnRH agonists, along with their many potential uses and dosage forms.

References:

1. Adams GP, Ratto MH. Ovulation-inducing factor in seminal plasma: a review. Anim Reprod Sci 2013;136:148-156. 
2. Faya M, Carranza A, Miotti R, et al. Fecal estradiol-17beta and testosterone in prepubertal domestic cats. Theriogenology 2013;80:584-586. 
3. Goericke-Pesch S, Georgiev P, Atanasov A, et al. Treatment of queens in estrus and after estrus with a GnRH-agonist implant containing 4.7 mg deslorelin; hormonal response, duration of efficacy, and reversibility. Theriogenology 2013;79:640-646. 
4. Goericke-Pesch S, Georgiev P, Atanasov A, et al. Treatment with Suprelorin in a pregnant cat. J Feline Med Surg 2013;15:357-360. 
5. Goericke-Pesch S, Georgiev P, Fasulkov I, et al. Basal testosterone concentrations after the application of a slow-release GnRH agonist implant are associated with a loss of response to buserelin, a short-term GnRH agonist, in the tom cat. Theriogenology 2013;80:65-69. 
6. Greenberg M, Lawler D, Zawistowski S, et al. Low-dose megestrol acetate revisited: a viable adjunct to surgical sterilization in free roaming cats? Vet J 2013;196:304-308. 
7. Leroy C, Conchou F, Layssol-Lamour C, et al. Normal canine prostate gland: repeatability, reproducibility, observer-dependent variability of ultrasonographic measurements of the prostate in healthy intact beagles. Anat Histol Embryol 2013;42:355-361. 
8. Luu VV, Hanatate K, Tanihara F, et al. The effect of relaxin supplementation of in vitro maturation medium on the development of cat oocytes obtained from ovaries stored at 4 degrees C. Reprod Biol 2013;13:122-126. 
9. Marino G, Zanghi A. Activins and inhibins: expression and role in normal and pathological canine reproductive organs: a review. Anat Histol Embryol 2013;42:1-8. 
10. Mattoso CR, Takahira RK, Beier SL, et al. Evaluation of von Willebrand factor during pregnancy, lactation and oestrous cycle in bitches affected and unaffected by von Willebrand disease. Reprod Domest Anim 2013;48:416-422. 
11. Nishida CR, Everett S, Ortiz de Montellano PR. Specificity determinants of CYP1B1 estradiol hydroxylation. Mol Pharmacol 2013;84:451-458. 
12. Polisca A, Orlandi R, Troisi A, et al. Clinical efficacy of the GnRH agonist (deslorelin) in dogs affected by benign prostatic hyperplasia and evaluation of prostatic blood flow by Doppler ultrasound. Reprod Domest Anim 2013;48:673-680. 
13. Poppl AG, Mottin TS, Gonzalez FH. Diabetes mellitus remission after resolution of inflammatory and progesterone-related conditions in bitches. Res Vet Sci 2013;94:471-473. 
14. Rota A, Tursi M, Zabarino S, et al. Monophasic teratoma of the ovarian remnant in a bitch. Reprod Domest Anim 2013;48:e26-e28. 
15. Serafim MK, Silva GM, Duarte AB, et al. High insulin concentrations promote the in vitro growth and viability of canine preantral follicles. Reprod Fertil Dev 2013;25:927-934. 
16. Sozmen M, Kabak YB, Gulbahar MY, et al. Immunohistochemical characterization of peroxisome proliferator-activated receptors in canine normal testis and testicular tumours. J Comp Pathol 2013;149:10-18. 
17. Spankowsky S, Heuwieser W, Arlt SP. Does oral administration of the amino acid tyrosine affect oestradiol-17beta concentration and sexual behaviour in the bitch? Vet Rec 2013;172:212. 
18. Trisolini C, Albrizio M, Roscino MT, et al. Leptin and queen ovary: new insights about ovulation. Res Vet Sci 2013;94:707-710. 
19. Tvarijonaviciute A, Carrillo-Sanchez JD, Ceron JJ. Effect of estradiol and progesterone on metabolic biomarkers in healthy bitches. Reprod Domest Anim 2013;48:520-524. 
20. Wongbandue G, Jewgenow K, Chatdarong K. Effects of thyroxin (T4) and activin A on in vitro growth of preantral follicles in domestic cats. Theriogenology 2013;79:824-832. 

What's the Best Way to Confirm Ovarian Remnant Syndrome in Cats?

In the queen, the most common cause of behavioral and physical signs of estrus after ovariohysterectomy is remnant ovarian tissue that has regained folliculogenesis and production of estrogens (1). 

In most cats, the "ovarian remnant syndrome" certainly is caused by failure to completely remove both ovaries at ovariohysterectomy. It is possible to have a small piece of ovarian tissue inadvertently drop back into the abdominal cavity during the spaying procedure and subsequently revascularize, resulting in signs of estrus (2). However, this syndrome can be also associated with the presence of accessory ovarian tissue that can be extremely difficult to detect at the time of routine ovariohysterectomy.

Whether entrapped in a ligature or accidentally dropped into the peritoneal cavity, revascularization of remnants by the omental blood supply can occur (2). The ovarian tissue can remain hormonally functional and even ovulate.  Although this condition is generally called the ovarian remnant syndrome, there is much contention among veterinary surgeons that it is not a syndrome but a surgical error

Diagnosis of ovarian remnant syndrome can sometimes be challenging but is based on a combination of history and clinical signs, vaginal cytology, and hormonal testing (1,3-6).

Vaginal cytology

The first step in the workup of a cat in which ovarian remnant or ectopic ovarian tissue is suspected is to perform vaginal cytology during the time your cat is showing signs of estrus. The only thing that causes a cat's vaginal smear to contain a high number of cornified vaginal epithelial cells is the presence of estrogen. This finding on vaginal cytology is a great bioassay for the presence of high levels of estrogen in the circulation (6).

In general, the use of vaginal cytology as a bioassay for feline estrogen is more accurate than a single serum sample. Vaginal cytological changes in queens in estrus tends to be more subtle than those of dogs but usually include an increase in cornified vaginal epithelial cells and clearing of normal background mucous (6).

Serum estradiol concentrations

Why not simple measure serum estradiol, the main estrogen in the cat? In queens during estrus, estradiol levels rise sharply to more than 20 pg/ml before returning rapidly to basal concentrations in as few as 48 hours. Thus blood sampling for estradiol is generally not helpful because the timing of the sample may not coincide directly with the period of peak estrus activity.

Therefore, the finding of high serum estradiol concentrations (>20 pg/ml) in a cat showing sign of estrus is consistent with ovarian remnant syndrome, the diagnosis can never be excluded if lower levels of estradiol are measured (1).

Serum luteinizing hormone (LH) concentrations

The hormonal serum test that is most useful in this situation is a basal luteinizing hormone (LH) determination (7). For this LH testing, we generally use a commercially available LH assay from Synbiotics. Although originally designed as a canine ovulation timing aid, this LH test may be used to distinguish between ovariohysterectomized and sexually-intact queens (or bitches). One advantage of using the serum LH test over vaginal cytology is it can be run at any time, not just when the cat is showing signs of estrus. If you do LH test, the manufacturer recommends two tests at least 24 hours apart to rule out the LH surge.

In the intact queen, LH is maintained at basal concentrations through the negative feedback influence of ovarian estradiol secretion on the pituitary gonadotropins. Following ovariectomy, this control is lost and LH concentrations increase resulting in a positive test.

Therefore, if a high serum LH concentration is found, then it is unlikely that the cat has any ovarian tissue. On the other hand, if the serum LH is low (less than 1 ng/ml), that indicates there is estrogen present in the circulation suppressing pituitary LH secretion (7). The source of this circulating estrogen could be from either endogenous or exogenous sources. If it's of endogenous origin, that means that the cat does indeed have an ovarian remnant or ectopic ovarian tissue.

The only problem with the use of serum LH determination is that some cats ultimately found to have an ovarian remnant will have falsely high LH values. The reason for this discrepancy in the expected pituitary-ovarian feedback loop is unknown, but it is clear that the finding of a high LH value alone can never rule out an ovarian remnant with 100% certainty. So it's still best to use vaginal cytology in conjunction with the serum LH measurements.

Provocative testing with hCG or GnRH analogues

The most efficient diagnostic aid for ovarian remnant syndrome is a hormone challenge testing (1,3-5). The goal of these stimulation tests is to cause ovulation of a follicle by exogenous LH-like compounds, thus confirming the presence of functional ovarian tissue. 

For these protocols, it is important that the queen be in true estrus. The procedure for testing is as follows:

1. Collect blood needed to measure the basal concentration of serum progesterone (and estradiol if desired).
2. Inject the cat with human chorionic gonadotropin (hCG), 44 IU/kg BW or 250-500 IU total dose, IM.
3. Alternatively, inject the cat with gonadotropin-releasing hormone (GnRH), 2 μg/kg BW or 25 μg total dose, IM.
4. Collect a post-stimulation blood sample for serum progesterone 1-3 weeks later.

Documenting a high serum progesterone level (>2 ng/ml) after hGC or GnRH stimulations confirms a diagnosis of ovarian remnant syndrome (1).

GnRH stimulation testing for cats not currently in estrus

Finally, a recent study demonstrated that measurement of plasma estradiol 2 hours after stimulation with a gonadotropin-releasing hormone (GnRH)-analogue appeared to be a reliable method to diagnose the presence of ovarian tissue in the female cat not currently showing signs of estrus (8).  With this protocol, a serum estradiol concentration greater than 3 pg/ml is consistent with the presence of ovarian tissue.

References:

1. Little SE. Female reproduction. In: Little SE (ed). The Cat: Clinical Medicine and Management. Saunders Elseiver 2012; 1195-1227.
2. DeNardo GA, Becker K, Brown NO, et al. Ovarian remnant syndrome: revascularization of free-floating ovarian tissue in the feline abdominal cavity. Journal of the American Animal Hospital Association 2001;37:290-296.
3. Wallace MS. The ovarian remnant syndrome in the bitch and queen The Veterinary Clinics of North: America Small Animal Practice 1991;21:501-507. 
4. Miller DM. Ovarian remnant syndrome in dogs and cats: 46 cases (1988-1992). Journal of Veterinary Diagnostic Investigation 1995;7:572-574.
5. Ball RL, Birchard SJ, May LR, et al. Ovarian remnant syndrome in dogs and cats: 21 cases (2000-2007). Journal of the American Veterinary Medical Association 2010;236:548-553
6. Mills JN, Valli VE, Lumsden HH. Cyclical changes of vaginal cytology in the cat. Canadian Veterinary Journal 1979;20:95–101.  
7. Scebra LR, Griffin B. Evaluation of a commercially available luteinizing hormone test to distinguish between ovariectomized and sexually intact queens. Proc Am Coll Vet Intern Med Forum 2003. 
8. Axner E, Gustavsson T, Strom Holst B. Estradiol measurement after GnRH-stimulation as a method to diagnose the presence of ovaries in the female domestic cat. Theriogenology 2008;70:186-191.

Q & A: High Serum Estradiol in a Neutered Male Dog with Alopeica

My patient is a 2-year old male castrated Alaskan Malamute that presented with the main clinical sign of patches of nonpruritic alopecia on the dorsal lumbar area. There is no other hair loss or clinical signs. 

The serum chemistry panel, total and free T4, and TSH concentrations are all normal.

I did an ACTH stimulation test and send serum to the Clinical Endocrinology laboratory at the University of Tennessee for the adrenal sex hormone panel. These are the results, before and after ACTH stimulation:

• Cortisol — Basal value 19 ng/ml (reference range, 2-56 ng/ml) Post-ACTH 96 ng/ml (reference range, 70-151 ng/ml)
• Androstenedione — 0.15 ng/ml (reference range, 0.05-0.3 ng/ml) Post-ACTH 0.76 ng/ml (reference range, 0.24-2.0 ng/ml)
• Progesterone — Basal 0.06 ng/ml (reference range, 03-.17 ng/ml) Post-ACTH 64 ng/ml (reference range, 22-1.45 ng/ml)
• Aldosterone — Basal value 19 ng/ml (reference range, 2-56 ng/ml) Post-ACTH 96 ng/ml (reference range, 70-151 ng/ml)
• Estradiol — Basal value 85 pg/ml (reference range, 23-65 pg/ml) Post-ACTH 75 pg/ml (reference range, 23-69 pg/ml)

My question: what would be the source of the high serum estradiol concentrations? Does it mean anything?

Should I start the dog on melatonin or mitotane?

My Response:

Estradiol (17β-estradiol) is a sex hormone produced in the ovary in the female. In the intact male, estradiol is also present, being produced as an active metabolic product of testosterone (1). In neutered dogs, The adrenal glands do not secrete estradiol directly but do normally secrete adrenal androgens (i.e., androstenedione and testosterone), even in the neutered dogs. A fraction of these adrenal androgens undergoes conversion to estradiol by an enzyme called aromatase in peripheral tissues (especially fat cells).

Conversion of circulating testosterone (steroid on left) to estadiol (on right).
The enzyme aromatase transforms the left-hand ring (the A-ring) of steroids to an aromatic state (hence the name) through oxidation and subsequent elimination of a methyl group.

So the source of the circulating estradiol that you are measuring could indirectly be the adrenal glands, but whether or not the high value found in this dog is clinically significant is difficult to know. These adrenal panels are difficult to interpret, and they are often abnormal in dogs even without clear evidence of typical or atypical Cushing's syndrome (2).

In a recent study of normal dogs published out of the University of Tennessee, they showed that the there was a wide range of variability in estradiol concentration both within and between the dogs (3). In addition, they reported that these estradiol concentrations often exceed the normal ranges established by the laboratory! This may account for why most of the adrenal panels I see have an abnormal sserum estradiol concentration.

Based on that study (3), I would ignore the results of these adrenal panels when the only abnormality is a high serum estradiol concentration.

Alopecia X is a term that has been used to describe dog that show dermatological signs that look similar to that seen with Cushing's disease (e.g., bilaterally symmetric alopecia and hyperpigmentation). This syndrome is common seen in the Nordic breeds, Pomeranians, and Chow chows. The cause of alopecia X is not understood (thus the term alopecia X!), but it is associated with hair cycle arrest.  Adrenal sex hormone abnormalities are no longer believed to play a role (4-6).

Since other labs are normal and since the dog is an Arctic breed, Alopecia X is certainly possible, and a relatively benign treatment such as melatonin can be tried. Click here to see a nice website with more info for vets and owners about alopecia X in Malamutes.

References:

1. Rijnberk A. Kooistra HS. Testes, In: Clinical Endocrinology of Dogs and Cats: An Illustrated Text. Second Edition. Schluetersche 2009.
2. Behrend EN, Kennis R. Atypical Cushing's syndrome in dogs: arguments for and against. The Veterinary Clinics of North America: Small Animal Practice 2010;40:285-296.
3. Frank LA, Mullins R, Rohrbach BW. Variability of estradiol concentration in normal dogs. Veterinary Dermatology 2010;21:490-3.
4. Frank LA, Hnilica KA, Oliver JW. Adrenal steroid hormone concentrations in dogs with hair cycle arrest (Alopecia X) before and during treatment with melatonin and mitotane. Veterinary Dermatology 2004;15:278-284.
5. Frank LA, Donnell RL, Kania SA. Oestrogen receptor evaluation in Pomeranian dogs with hair cycle arrest (alopecia X) on melatonin supplementation. Veterinary Dermatology 2006;17:252-258.
6. Frank LA. Oestrogen receptor antagonist and hair regrowth in dogs with hair cycle arrest (alopecia X). Veterinary Dermatology 2007;18:63-66.

Q & A: Diagnosis of Ovarian Remnant Syndrome in Cats

My patient is a 2-year old, female spayed DSH cat who presented with a 10-day history of vocalizing, rolling, and allowing a male altered cat in the household to mount her, especially at night. The cat has gone through two other similar episodes about 1 and 3 months ago, each lasting about 1 to 2 weeks in duration. The cat is otherwise completely healthy, and her behavior is totally normal between each of the episodes.

This certainly sound like the cat is coming into heat. I did the ovariohysterectomy on the cat myself when she was 10-months old, and the surgery was uneventful. I suspect that the cat either has an ovarian remnant or ectopic ovarian tissue.

My question is this — what's the best way to diagnosis this condition? I don't want to explore the cat without some evidence that the cat indeed does have ovarian tissue.

My Response:

Diagnosis of ovarian remnant syndrome is based on a combination of history and clinical signs, vaginal cytology, and hormonal testing (1-4).

Vaginal cytology: The first step in the workup of a cat in which ovarian remnant or ectopic ovarian tissue is suspected is to perform vaginal cytology during the time your cat is showing signs of estrus. The only thing that causes a cat's vaginal smear to contain a high number of cornified vaginal epithelial cells is the presence of estrogen. This finding on vaginal cytology is a great bioassay for the presence of high levels of estrogen in the circulation (5).

In general, the use of vaginal cytology as a bioassay for feline estrogen is more accurate than a single serum sample. Vaginal cytological changes in queens in estrus tends to be more subtle than those of dogs but usually include an increase in cornified vaginal epithelial cells and clearing of normal background mucous[3].

Serum estradiol concentrations: Why not simple measure serum estradiol, the main estrogen in the cat? In queens during estrus, estradiol levels rise sharply to more than 20 pg/ml before returning rapidly to basal concentrations in as few as 48 hrs. Thus blood sampling for estradiol is generally not helpful because the timing of the sample may not coincide directly with the period of peak estrus activity.

Serum luteinizing hormone (LH) concentrations: The hormonal serum test that is most useful in this situation is a basal luteinizing hormone (LH) determination (6). For this LH testing, we generally use a commercially available LH assay from Synbiotics. Although originally designed as a canine ovulation timing aid, this LH test may be used to distinguish between ovariohysterectomized and sexually-intact queens (or bitches). One advantage of using the serum LH test over vaginal cytology is it can be run at any time, not just when the cat is showing signs of estrus. If you do LH test, the manufacturer recommends two tests at least 24 hours apart to rule out the LH surge.

In the intact queen, LH is maintained at basal concentrations through the negative feedback influence of ovarian estradiol secretion on the pituitary gonadotropins. Following ovariectomy, this control is lost and LH concentrations increase resulting in a positive test.

Therefore, if a high serum LH concentration is found, then it is unlikely that the cat has any ovarian tissue. On the other hand, if the serum LH is low (less than 1 ng/ml), that indicates there is estrogen present in the circulation suppressing pituitary LH secretion (6). The source of this circulating estrogen could be from either endogenous or exogenous sources. If it's of endogenous origin, that means that the cat does indeed have an ovarian remnant or ectopic ovarian tissue.

The only problem with the use of serum LH determination is that some cats ultimately found to have an ovarian remnant will have falsely high LH values. The reason for this discrepancy in the expected pituitary-ovarian feedback loop is unknown, but it is clear that the finding of a high LH value alone can never rule out an ovarian remnant with 100% certainty. So it's still best to use vaginal cytology in conjunction with the serum LH measurements.

GnRH stimulation testing: Finally, a recent study demonstrated that measurement of plasma estradiol 2 hours after stimulation with a gonadotropin-releasing hormone (GnRH)-analogue appeared to be a reliable method to diagnose the presence of ovarian tissue in the female cat (7). Again, I would still perform vaginal cytology together with the test, since it is such a nice bioassay for the presence of estrogen.

I know that you are probably feeling guilty about this situation, but the presence of ovarian remnant(s) in a spayed cat is not always the result of "surgeon's error."  In most cats, it certainly is caused by failure to completely remove both ovaries at ovariohysterectomy. It is possible to have a small piece of ovarian tissue inadvertently drop back into the abdominal cavity during the spaying procedure and subsequently revascularize, resulting in signs of estrus (8). However, this syndrome can be also associated with the presence of accessory ovarian tissue that can be extremely difficult to detect at the time of routine ovariohysterectomy.

Follow-Up:

I did a vaginal smear with cytology, which showed a large population of increase in cornified vaginal epithelial cells consistent with estrus. I also performed the two serum LH tests within 24 hours of each other. The serum LH concentrations on both samples were undetectable, again consistent with presence of estrogen.

Time to go to surgery? If so, is it best to do the exploratory while the queen is exhibiting signs of heat? I assume the ovarian tissue would be easier to identify at that time, correct?

My Response:

Now that we know the cat has circulating estrogen, the next step is to exclude any exogenous exposure of the cat to human skin to which estrogen hormone replacement therapy (HRT) cream, spray, or gel has been applied (9). If your history rules out that possibility, then you should explore the cat  to look for an ovarian remnant.

And yes, having the cat in heat at the time of surgery should make it easier to find the ovarian remnant, so a few days delay after onset of the next estrus would be best. Alternatively, you can administer human chorionic gonadotropin (hCG), 44 IU/kg or 250 IU total dose, IM while the queen is in estrus to cause a LH surge and induce ovulation, and then look for the luteal tissue at exploratory (2).

References:

1. Muram D, Drouin P. Ovarian remnant syndrome. Canadian Medical Association Journal 1982;127:399-400.
2. Wallace MS. The ovarian remnant syndrome in the bitch and queen The Veterinary clinics of North America Small animal practice 1991;21:501-507. 
3. Miller DM. Ovarian remnant syndrome in dogs and cats: 46 cases (1988-1992). Journal of Veterinary Diagnostic Investigation 1995;7:572-574.
4. Ball RL, Birchard SJ, May LR, et al. Ovarian remnant syndrome in dogs and cats: 21 cases (2000-2007). Journal of the American Veterinary Medical Association 2010;236:548-553
5. Mills JN, Valli VE, Lumsden HH. Cyclical changes of vaginal cytology in the cat. Canadian Veterinary Journal 1979;20:95–101.  
6. Scebra LR, Griffin B. Evaluation of a commercially available luteinizing hormone test to distinguish between ovariectomized and sexually intact queens. Proc Am Coll Vet Intern Med Forum 2003. 
7. Axner E, Gustavsson T, Strom Holst B. Estradiol measurement after GnRH-stimulation as a method to diagnose the presence of ovaries in the female domestic cat. Theriogenology 2008;70:186-191.
8. DeNardo GA, Becker K, Brown NO, et al. Ovarian remnant syndrome: revascularization of free-floating ovarian tissue in the feline abdominal cavity. Journal of the American Animal Hospital Association 2001;37:290-296.
9. Parker-Pope T. When hormone creams expose others to risk. The New York Time. October 25, 2010.

Top Endocrine Publications of 2010: The Canine and Feline Reproductive Endocrinology

 Fertility axis. GnRH, LH & FSH
stimulate secretion of testosterone,
progesterone, estrogen, & inhibin.
Controlled via negative feedback
on hypothalamus & pituitary gland.

In my 9th compilation of the canine and feline endocrine publications of 2010, I’m moving on to endocrine disorders of the canine gonads and mammary gland.

Listed below are 9 research papers written in 2010 that deal with a variety of issues of clinical importance in reproductive endocrinology in dogs.

These range from the investigations of the pituitary-ovarian axis in dogs with a functional granulosa cell tumor (1) to studies of the expression of estrogen and progesterone receptors in sebaceous gland tumors (2); and from studies of gonadotropin-releasing hormone (GnRH)-agonist implants in the treatment of reproductive disorders in the male dog (3) to a review of non-surgical methods for reproductive control in cats (4).

Finally, a number of studies investigate markers of malignancy for mammary gland tumors (6,7,8). Finally, the last paper reviews alternative ways to treat benign prostatic hyperplasia, including hormonal suppression of testicular function with GnRH-depot-analogues, inhibition of the 5-alpha-reductase, or peripheral androgen action with anti-androgens (9).

References:

1. Buijtels JJ, de Gier J, Kooistra HS, et al. Alterations of the pituitary-ovarian axis in dogs with a functional granulosa cell tumor. Theriogenology 2010;73:11-19.
2. Ginel PJ, Lucena R, Millan Y, et al. Expression of oestrogen and progesterone receptors in canine sebaceous gland tumours. Veterinary Dermatology 2010;21:297-302.
3. Goericke-Pesch S, Wilhelm E, Ludwig C, et al. Evaluation of the clinical efficacy of Gonazon implants in the treatment of reproductive pathologies, behavioral problems, and suppression of reproductive function in the male dog. Theriogenology 2010;73:920-926.
4. Goericke-Pesch S. Reproduction control in cats: new developments in non-surgical methods. J Feline Med Surg. 2010 12:539-46
5. Groppetti D, Pecile A, Arrighi S, et al. Endometrial cytology and computerized morphometric analysis of epithelial nuclei: a useful tool for reproductive diagnosis in the bitch. Theriogenology 2010;73:927-941.
6. Klopfleisch R, Klose P, Gruber AD. The combined expression pattern of BMP2, LTBP4, and DERL1 discriminates malignant from benign canine mammary tumors. Veterinary Pathology 2010;47:446-454.
7. Millanta F, Caneschi V, Ressel L, et al. Expression of vascular endothelial growth factor in canine inflammatory and non-inflammatory mammary carcinoma. Journal of Comparative Pathology 2010;142:36-42.
8. Queiroga FL, Perez-Alenza D, Silvan G, et al. Serum and intratumoural GH and IGF-I concentrations: prognostic factors in the outcome of canine mammary cancer. Research in Veterinary Science 2010;89:396-403.
9. Renggli M, Padrutt I, Michel E, et al. Benign prostatic hyperplasia: treatment options in the dog. Schweizer Archiv fur Tierheilkunde 2010;152:279-284.