301 NEW APPROACHES TO CONTRACEPTION IN DOGS USING GnRH ANALOGUES, CALCIUM, AND NALOXONE

Abstract

The continuous administration of GnRH leads to the desensitization and down-regulation of GnRH receptors on pituitary gonadotropes (Parker and Schimmer 2001 in Goodman & Gilman's The Pharmacological Basis of Therapeutics, New York: McGraw-Hill, 1541–1562). In this study, we combined the use of a GnRH analogue with calcium/naloxone association (Sciorsci et al. 2000 Immunopharmacol. Immunotoxicol. 22, 575−626) to obtain a desensitizing effect on LH receptors in an attempt to regulate spermatogenesis in dogs. Eight post-pubertal mixed breed dogs were divided into two groups: (A) four subjects were pharmacologically treated with a 30 day administration of buserelin acetate (Suprefact Aventis Pharma, Italy) 0.3 mg/each s.c. TID, associated with 0.1-mL/kg/bw i.m. injection SID of calcium gluconate 20% (New ICC, Upjohn, Italy) in which 0.4 mg/mL of naloxone hydrochloride (Diosynth, The Netherlands) was dissolved; (B) four subjects received placebo (NaCl 0.9% s.c. TID for 30 days). At the end of the treatment, hemiorchidectomy was performed and 4 months later the second testis was removed. Clinical examination of the genital tract was done by ultrasound monitoring. Before and after pharmacological treatment and until total orchidectomy, semen was collected and evaluated for macroscopic and microscopic parameters. Plasma testosterone concentration was measured twice a week by RIA. Testicular specimens were fixed in formalin solution, embedded in paraffin wax and stained with hematoxylin–eosin. The presence of germ cells and the testicular tubule diameters were analyzed using a computer assisted image analyzer (MONO system, Italy). Data were analyzed by the ANOVA test. In the treated group, all dogs showed a reduction in testicular and prostatic diameters compared to the control group. Transient elevations in testosterone concentrations were observed in response to the early injections and reached baseline levels after 8–10 days. The testosterone decline resulted in lower ejaculate volume and, at the end of treatment, azoospermia was observed. At the end of treatment, testosterone values had progressively regained normal values over 2 months and, when measured immediately before total orchidectomy, sperm concentration and motility had returned to normal. At the histological observation, seminiferous tubules showed the block of spermatogenesis at the level of spermatocytes with a statistically significant cell reduction in spermatides and spermatozoa (P < 0.001) whereas the seminiferous tubular morphometry was normal. Total orchidectomy demonstrated that a normal histological testicular structure had been restored. In the control group, no modifications were observed. As observed in a previous study (Aiudi et al. 2005 Reprod. Fert. Dev. 17(2), 280–281), GnRH induced suppression of the reproductive function in male dogs. In this study, we demonstrated that GnRH analogue associated with calcium and naloxone induces the same effect in a shorter time and that this effect is reversible. These devices designed for the shutdown of gonadal steroidogenesis and spermatogenesis would be useful for treating sex hormone-dependent syndromes such as malignant and benign prostatic tumors.