Possible Involvement of inhibln in Altered Follicle-Stimulating Hormone (FSH) Secretion during Dissociated Luteinizing Hormone (LH) and FSH Release: Unilateral Castration and Experimental Cryptorchidism

BIOLOGY OF REPRODUC11ON 41, 967-981 (1989) Possible Involvement of inhibln in Altered FollicleStimulating Hormone (FSH) Secretion during Dissociated Luteinizing Hormone (LH) and FSH Release: Unilateral Castration and Experimental Cryptorchidism1 CATHERINE PJVIER,2’4 HELENE VERONICA Foundation Laboratories for ratory for Neuronal Structure La Jolla, California ROBERTS,5 and WYLIE VALE4 Peptide Biology’ and Functio& 92037 ABSTRACT Male rats were either unilaterally or bilaterally castrated, or were rendered cryptorchid when they were either 15 or 45 days old. Subsequently, blood was sampled over the next several weeks and plasma luteinizing hormone (LII), follicle-stimulating hormone (FSH), testosterone (T), and immunoreactive inhibin-a (irla) levels were measured by specflc radioimmunoassays (RIM). At the end of the experiment, gonadal expression of inhi bin-a, inhibin34, and inhibin-B subunits was measured by Si nuclease analysis and in situ hybridization. In both age groups, bilateral castration (BC) produced the expected marked (p0.0i) increases in plasma LII and FSH levels, and concomitant decreases in T and irks secretion within I 2 days after surgery. In 15-day-old animals, unilateral castration (UC) significantly increased FSH and decreased circulating levels of irks, but did not measurably alter LII or androgen production. At 7 days after surgery, the level of inhibin mRNA in the remaining testis was unchanged. In 45-day-old animals, UC caused a measurable increase in FSH, with little or no changes in the circulating levels of irks. Plasma T levels were lowered (p0.05) by UC; however, there were no statistical changes in LII levels in these UC rats. Finally, T administration markedly reversed UC- induced increase in FSH secretion in both age groups. Androgen therapy also interfered with in/ti bin release in 45-day-old, but not in 15-day-old rats. In rats 15 days old at the time of surgery, cryptorchidism produced a small but measurable increase (p.O.OS) in LII release at Week 6 only, which was accompanied by a significant (p0.01) decline in T secretion. Plasma FSH levels were elevated at all times in cryptorchid rats, and at 2, 4, and 6 wk, these levels were not statistically distinguishable (p>O.05)from those of castrated animals. In this group of rats, cryptorchidism caused a transient increase (pO.05) in irla values I wk after surgery, but no changes at later times. Finally, measurement of testicular inhibin-a subunit messenger RNA (mRWA) levels showed an approximately 2-fold increase compared to total RIIA levels in the testis. However, because of the signifi cant decrease in total RNA levels per testis caused by cryptorchidism, the absolute change in inhi bin-a subunit mR.NA levels per testis corresponded to an approximately 3-fold decrease. There were no measurable changes in inhibin-A and inhibin-B subunit mR.NA levels relative to total RWA levels in the testis. In 45-day-old rats, cryptorchidism induced a marked (p0.0i) increase in plasma FSH levels, as well as elevations (pO.OS) in LII secretion 4 and 6 wk after surgery and a transient, but significant (p0.05), decrease in plasma Tlevels at2 wk. As in the younger age group,plasma irla levels were only elevated (p0.05) at I wkpostsurgery, but an 80% increase in inhibin-a subunit expression by the testes, relative to total RNA levels, was observed at 6 wkpost-surgery. This apparent increase was accompanied by a significant decrease in total testicular Ri/A levels. The inhibin-13 subunit mRNA levels were not changed relative to total testicular Ri/A levels in the 45-day-old group. In this age group, gonadotropin-releasing hormone (GnRH) also caused a significantly (p0.05) larger increase in the plasma LII and FSH levels of cryptorchid rats versus intact animals. Histological examination of the testis showed marked degeneration of Sertoli cells 6 wk after cryptorchidism. However, in/ti bin subunit ,nRNAs continued to be expressed, and actually appeared to be enhanced for expression of the a-subunit Accepted July Received March l’p.; work was conducted in part Division (C. R. and supported by a Centennial Fellowship from the Medical Research Council of Canada and the I. Aaron Charitable Foundation. V. R. was supported by a fellowship from the National Institute of Health No. F32HW7172.0l. 2Rcprint requests. 3Prcsent address: Division of Endocrinology. Research Institute, The Hospital for Sick Children, Toronto, Canada M5G 1X8. 31, 1989. 21. 1989. supported by NIH Grants HD-13527 and DK-26741, and by The Clayton Foundation for Research, California W. V. are Clayton Foundation Investigators). It h& was 967 The Clayton and Labo MEUNIER,3’4 968 RIVIER ET AL. in the remaining Sertoli tissue near the basal lamina. The intensity of the mRNA signals for the inhibin-A and inhibin-B subunits did not change. Leydig cells and supportive tissue from control or operated rats did not appear to express any of the inhibin subunit mRNAs. Finally, we observed that the pituitary of cryptorchid rats showed a measurable increase in responsiveness to GnRH. We conclude from these results that measurement of irkt secretion and mR.NA levels have failed to denwns:rate a clear-cut inverse relationship between the circulating levels of this hormone (as followed by an antiserum directed against the a chain of inhibin) and FSH in either unilaterally castrated or ciyptorchid male rats. These results leave open the possibility that factors other than inhibin may mediate the changes in FSH secretion induced by unilateral castration and/or cryptorchidism. Even though the role of androgen feedback in modulating the increased FSH secretion of cryptorchid rats is not clear, we have shown that the pituitary of cryptorchid rats is more sensitive to GnRH than is the gland of intact animals. The initial proposal for the existence of inhibin relied on experiments showing that destruction of the germinal epitheium by irradiation caused pituitary hyperfunction in the absence of atrophy of the secondary sex glands (Motiram and Cramer, 1923), and that injection of aqueous testicular extracts to castrated rats prevented the appearance of the enlarged “castration cells” in the pituitary (Martins and Rocha, 1931; McCullagh, 1932). This suggested the existence of a specific feedback between the germinal epitheium and pituitary function, which was not mediated by androgens. Therefore, cxperiinental procedures such as unilateral castration and cryptoithidism, which are believed to specifically alter the function of the Sertoli cells (and consequently the secretion of inhibin as well as the release of fofficlestimulating hormone [FSH]), have been used to investigate possible functional relationships between these two hormones. In the sexually immature male rat (Ojeda and Ramirez, 1972; Gupta Ct al., 1975a Lindgren et al., 1976; Cunningham et al., 1978; Risbridger et a!., 1981a), but not in older animals (Howland and Skinner, 1975 (...truncated)