Cortisol influence on testicular testosterone secretion in domestic cat: An in vitro study

Pesquisa Veterinária Brasileira

Genaro, Gelson¹  Franci, Celso R¹ 

[1] Universidade de São Paulo (FMRP-USP), Ribeirão Preto, Brazil

关键词: Testosterone;    domestic cat;    cortisol;    testis;    stress;    transport.;   

DOI  :  10.1590/S0100-736X2010001000013

来源: Colegio Brasileiro de Patologia Animal-CBPA

【 摘 要 】

The aim of the present experiment was to investigate the effect of corticosteroids (exogen) on in vitro testosterone secretion after stress by transportation (40 minutes). Feline testes (Felis silvestris catus) were incubated in the following media: TCM 199; TCM 199 + hCG 10_7M; TCM 199 + hydrocortisone 10_7M, or TCM 199 + hCG + hydrocortisone. The animals (n=21) were allocated into three groups: (S) that arrived at 3 h prior to surgery, (A) that remained in the laboratory for 36 h before being submitted to surgical procedure, and (C) that were also allowed to remain for 36 hours in the laboratory before the surgical procedure, but whose testes had been incubated with hydrocortisone prior to incubation in the referred media. The results showed that group S secreted higher levels of testosterone, regardless of the culture media. It is noteworthy that the suppressing action of hydrocortisone sodium succinate led to a reduction in the testosterone concentration, despite the presence of hCG.INDEX TERMS: Testosterone, domestic cat, cortisol, testis, stress, transport.RESUMOO objetivo deste trabalho foi investigar o efeito da hidrocortisona sobre a secreção de testosterona após cultivo in vitro dos testículos, em distintas situações de estresse (transporte) de gatos domésticos (Felis silvestris catus). Testículos foram incubados nos seguintes meios de cultura: TCM 199; TCM 199 + hCG 10_7M; TCM 199 + hydrocortisona 10_7M e TCM 199 + hCG + hidrocortisona. Os animais (n=21) foram alocados em 3 sub-grupos: (S) animal admitido 3 horas antes da orquiectomia, (A) animais orquiectomizados após 36 horas de permanência no biotério e (C) animais que permaneceram por 36 horas no biotério antes da cirurgia e que tiveram seus testículos pré-incubados em hidrocortisona. Os resultados demonstraram que o grupo S secretou maiores valores de testosterona em todas as condições estudadas. É válido mencionar que a supressão promovida pela hidrocortisona também promoveu redução na concentração de testosterona no meio TCM 199, a despeito da presença de hCG. TERMOS DE INDEXAÇÃO: Testosterona, gato doméstico, cortisol, testículo, estresse, transporte.     IntroductionDespite evidences that stress exerts influence on reproductive functions, the interactions between the hypo-thalamus-pituitary-adrenal (HPA) and gonadal (HPG) axes remain unclear (Moberg 1991, Baldwin et al. 1996, Fenske 1997, Ferasin 2001, Genaro et al. 2003, Lacerda Neto et al. 2004, Bridges et al. 2005). There are indications of both inhibiting (Bambino & Hsueh 1981, Welsh et al. 1982, Genaro et al. 2003) and stimulating (Faulborn et al. 1979, Welsh & Johnson 1981) effects of the HPA axis on HPG. The hypothalamus secretes the Gonadotrophin Releasing Hormone (GnRH), and the pituitary, which secretes the Luteinizing Hormone (LH) and the Follicle Stimulating Hormone (FSH), as well as testis (a gland that secretes testosterone (T) can be both the target of hormones belonging to the HPA axis (Willard et al. 1995).These hormonal interactions depend on various factors such as species, sex, age, social class, animal housing and others (Welsh & Johnson 1981, Brown et al. 1988, Tasker et al. 1999, Barber 2004, Genaro et al. 2004, 2007). Overall, hormone secretion by the HPA axis is inversely proportional to the activity of the HPG axis (Willemse et al. 1993). Therefore, the suppressing effect of stress on reproduction should be mainly due to the reduced secretion of sexual hormones (to a review: Moberg, 1991).The majority of studies on the domestic cat (Felis silvestris catus) focus on female endocrinology (Kirkpatrick 1985, Silva et al. 2009). Still, if species-specificities are considered, studies using domestic cats are a possible effective model for the investigation of various aspects of the reproductive function of the Felidae family, with many endangered species (Genaro et al. 2007).The present study aimed to evaluate the effect of cortisol on testicular testosterone secretion following acute stress by animal transportation and/or lack of environmental adaptation through an in vitro study in domestic cats.  Material and methods Twenty-one male cats (2-3 years of age, body weight 4-5 kg), kindly provided by non-governmental organizations were allocated equally into three experimental groups: A (n=7), C (n=7), and S (=7), as described in Table 1 (July-December).After 40 minutes of transportation, the animals were housed indoors in individual 1-m3 stainless-steel cages with commercial dry food and water provided ad libitum.The three experimental groups underwent two distinct pre-surgical treatments that differed in the number of hours allowed for the adaptation to the laboratory housing. Animals from Groups A and C arrived 36 hours prior to the surgical procedure and were kept at the laboratory housing of the Physiology Department (FMRP-USP) in the conditions described in the paragraph above Group S arrived at the laboratory 3 hours before the surgery. All animals were maintained under controlled temperature (23 ± 2oC) and fasted for 12h before surgery irrespective of the adaptation period length.Animals were anesthetized with a combination of ketamine HCl (20mg/kg, im, Francotar, Virbac-Brasil) and xylazine (1mg/kg, im, Coopazine, Coopers-Brasil). An attempt was made to minimize animal disturbance before the injections. Immediately before the surgical procedure and after anesthesia, 1.0ml of venous blood was collected from the right forelimb (Cephalic vein) for determination of plasma testosterone.Following orchiectomy, right and left testis were denuded and divided into four fragments. These fragments (one from each testis) were pre-incubated pair wisely for 60 minutes in TCM 199 culture medium (Gibco-USA, containing 0.1mg/ml of bovine serum albumin, fraction V, Hepes 15mM in pH 7.4). The testes from animals belonging to Group C (only) received hydrocortisone (10-7M) during the pre-incubation period. After pre-incubation, each fragment (To Groups: A, C and S) was submitted to two periods of sixty-minute (first and second hour) of incubation in one of the following conditions:1) TCM 199 culture medium;2) TCM 199 culture medium containing human Chorionic Gonadotrophin 10-7M (hCG-Pregnyl 1500 UI, Akzo Nobel Ltda, Organon, Brazil);3) TCM 199 culture medium containing hydrocortisone 10-7M (Hydrocortisone Sodium Succinate, União Química Farmacêutica Nacional S/A, Brazil);4) TCM 199 culture medium containing hydrocortisone 10-7M and hCG 10-7M.Both the pre-incubation of C group and the incubation of A, C and S were carried out under continuous stirring (80 cycles/ minute) at a temperature of 38.8oC (without CO2 control). At the end of each sixty-minute period, the total medium volume (1 ml) was collected and substituted for an equal volume of the same medium containing the same initial compounds. The medium obtained after the pre-incubation period was discarded. The medium collected after the incubations was frozen (-20oC) for further evaluation of testosterone by radioimmunoassay.Testosterone was determined by radioimmunoassay using 3H-testosterone from NEN Life Science Products (Boston, USA). The specific antibody was kindly provided by Dr. José Antunes-Rodrigues (USP-Ribeirão Preto, Brazil). All samples were analyzed in a single assay. The lower detection limit was 0.16ng/ml. The intra-assay variation coefficient was 5.4%.The data (mean ±SEM) for each group (according to experimental groups or medium) were analyzed by one-way ANOVA followed by the Tu

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