Effect of neonatal androgenization on the testosterone feedback sensitivity in adult rats in two lighting conditions

Neonatally androgenized and intact adult male Wistar rats received daily, during 1 week, either testosterone propionate or sesame oil injections in periodic or constant light. Serum and pituitary gonadotropins and hypothalamic LHRH were measured. In periodic light, neonatal androgenization did not change the serum concentration or pituitary contents of gonadotropins, or the hypothalamic content of LHRH. Testosterone injections decreased serum concentration and pituitary content of gonadotropin of intact rats but failed to decrease the pituitary gonadotropin content of neonatally androgenized rats. In constant light, serum FSH was decreased in neonatally androgenized rats. Testosterone injections decreased both serum LH and FSH concentrations of intact rats but only serum LH of androgenized rats. Pituitary gonadotropin and hypothalamic LHRH contents remained unchanged. We conclude that neonatal androgenization renders the male rat hypothalamo-pituitary axis more resistant to changes of testosterone concentration in adulthood. Constant light did not sensitize the neonatally androgenized rats to testosterone, but on the contrary, testosterone injections were less effective in constant than in periodical light.     

Delay in the age of balano-preputial skinfold cleavage and alterations in serum profiles of testosterone, 5 alpha-androstane-3 alpha,17 beta-diol, and gonadotropins in adult rats treated during puberty with luteinizing hormone releasing hormone

Previous work has shown that chronic treatment of intact, immature male rats with luteinizing hormone releasing hormone (LHRH) decreases sex accessory gland weights and results in retardation of the normal developmental increase in the ratio of serum testosterone (T)/5 alpha-androstane-3 alpha,17 beta-diol (3 alpha-Diol) via an apparent enhancement of testicular 5 alpha-reductase or 3 alpha-hydroxysteroid oxidoreductase activities. In the present work, androgen dependent balano-preputial skinfold cleavage was significantly delayed by approximately one week in intact, immature male rats which were treated daily for two weeks with either 1.0 micrograms, 2.5 micrograms or 5.0 micrograms of LHRH during a discrete phase of pubertal development (28-41 days of age). In intact, adult (62 day old) animals which received LHRH treatments during pubertal development, serum T concentrations and sex accessory gland weights were reduced compared to control animal values. Serum 3 alpha-Diol content in the adult rats was either unaltered or increased significantly depending on the LHRH dosage employed during sexual development. Serum luteinizing hormone concentrations were not different between control and LHRH-pretreated adult rats whereas the highest dosage of LHRH employed (5.0 micrograms) during puberty resulted in a significant elevation of adult serum follicle stimulating hormone levels. It is suggested that chronic LHRH treatment of the male rat during puberty results in a perturbation in testicular androgen biosynthetic activities and an impairment of pituitary-testicular hormone feedback mechanisms which persist at least through early adulthood.     

Cysteamine reduces serum gonadotropin concentrations in adult male rats

We have examined the effects of cysteamine on the hypothalamic-pituitary-gonadal axis of the adult male rat. A single subcutaneous injection of cysteamine (300 mg/kg) reduces significantly (p less than or equal to 0.05 serum concentrations of LH, FSH and T. Cysteamine blocked LH secretion induced by castration and administration of naloxone and LHRH. Neither acute nor chronic treatment (7 days) altered the hypothalamic LHRH content. These results suggest that cysteamine acts to reduce pituitary responsiveness to LHRH, resulting in lower mean serum gonadotropin and testosterone concentrations. It is possible, however, that cysteamine acts also at the hypothalamus to reduce LHRH secretion and/or at the testes to reduce testosterone release.     

Alteration in the normal pattern of serum testosterone and 5α-androstane-3α,17β-diol in the immature male rat following chronic treatment with luteinizing hormone releasing hormone or luteinizing hormone

A major component of sexual maturation in the male rat is a progressive decline in serum concentrations of 5α-androstane-3α,17β-diol (3α-diol) and a concomitant increase in testicular testosterone biosynthesis and secretion. Chronic administration of synthetic luteinizing hormone releasing hormone (LHRH) or luteinizing hormone (LH)/human chorionic gonadotropin (hCG) to immature male rats has been shown to result in a delay in sexual maturation as evidenced by decreased sex accessory gland weights and altered testicular testosterone production. We have examined the postulate that such treatments may either reverse or retard the normal developmental pattern of serum testosterone and 3α-diol concentrations. Chronic $\text{in vivo}$ treatment of 28 day old immature male rats for 2 weeks with daily injections of either 0.5 μg of LHRH, 1.0 μg of LHRH, or 30 μg of LH was found to result in significant reductions in weights of the seminal vesicles and ventral prostate glands and diminutions in serum testosterone concentrations. Serum content of 3α-diol was either unchanged or slightly elevated in the LHRH treated animals and increased significantly in the LH treated animals. These data suggest that either a reversal of or retardation in the normal developmental pattern of serum testosterone and 3α-diol content has been achieved in the immature male rat by chronic LHRH or LH treatment.     

阉割和睾丸酮替代对雄性老年大鼠下丘脑和血浆促黄体生成...

Hypothalamic and plasma luteinizing hormone-releasing hormone (LHRH) levels following orchidectomy (ORDX) and testosterone (T)-replacement were compared between young (2-3 months old) and aged (24-26 months old) male rats by radioimmunoassay. Plasma T level and hypothalamic LHRH content are markedly decreased in the aged rat as compared to those of the young rat, whereas plasma LHRH levels are similar in the two groups. Following ORDX and ORDX plus T-replacement, plasma T levels in both groups are about the same, whereas the rates of variation of hypothalamic and plasma LHRH levels in the aged rat are significantly lower than those in the young rat. These results suggest that the negative feedback mechanism of the hypothalamic LHRHergic system is impaired in the aged rat, which may be one of the important reasons causing age-dependent deterioration of the functional control of hypothalamic-pituitary-testicular axis.     

Pituitary responsiveness to luteinizing hormone-releasing hormone in prepubertal and postpubertal male ferrets

The pituitary response to three different doses of exogenously administered LHRH was examined in prepubertal (9-wk-old) and postpubertal (32-wk-old) male ferrets. The doses of 5, 10, and 15 ng LHRH/kg body weight tested in this study produced dose-related increases in circulating LH concentrations in both pre- and postpubertal groups. In addition, a significant effect of age on LH response was observed, with the prepubertal animals demonstrating significantly greater serum LH values in response to the two higher doses than the postpubertal males. Prepubertal ferrets also exhibited a significant increase in endogenous LH pulse amplitude in sampling periods following exogenous administration of LHRH compared to baseline pulse amplitudes in periods prior to the LHRH infusions. These results suggest that the low frequency of endogenous LH pulses previously observed in prepubertal ferrets is not due to unresponsiveness of the pituitary gland to LHRH. Thus, suppression of the hypothalamo-hypophyseal axis observed in the prepubertal ferret is probably mediated at the level of the hypothalamus.     

Relationship between release of LH and incorporation of tritiated thymidine in the anterior pituitary gland of the castrated male rat: effect of LHRH and its highly active analogue buserelin

Castration of the adult male rat significantly (P less than 0.01) increased the concentration of LH in serum and the incorporation of (3H) thymidine into the pituitary DNA. The administration of a single dose of LHRH or its analogue buserelin stimulated the release of LH but it did not modify (3H) thymidine incorporation. When multiple doses of LHRH or buserelin were injected, there was a significant (P less than 0.01) inhibition of LH release and also the incorporation of (3H) thymidine into the DNA of the anterior pituitary gland was significantly (P less than 0.01) diminished. These observations are compatible with the idea of the close relationship between hormonal release and DNA synthesis in the anterior pituitary gland of the rat.     

Production and exocrine secretion of LHRH-like material by the male rat reproductive tract

Luteinizing hormone-releasing hormone (LHRH)-like immunoreactivity was localized in the male reproductive system of the rat. Epithelial cells of the epididymus, seminal vesicles and coagulation gland showed a strong reaction to anti-LHRH serum. Also the epithelia of the ductus deferens and the prostate gland appeared to be immunoreactive, albeit to a lesser extent. The LHRH-like substances are most likely secreted into the male tract, as can be concluded from the observation that the secretion product in the lumina of the seminal vesicles, coagulation gland and prostate gland was also immunopositive. The functional significance of these phenomena is discussed. No immunostaining was obtained with antisera to FSH, LH or beta-hCG.     

Prolactin receptor regulation by LH in the rat mammary gland

The aim of this study was to investigate hormonal factors responsible for the huge increase in PRL receptors on the day of estrus in the rat mammary gland. For this purpose, ovariectomized rats were primed with E2 so as to reach a physiological serum concentration of E2 (21.5 +/- 1.2 pg/ml) and high PRL serum values (72.8 +/- 21.9 ng/ml). In these conditions, PRL specific binding and capacity were respectively 22.8 +/- 8.3%/mg protein and 96 +/- 29 fm/mg protein. An injection of either LHRH (500 ng/rat) or LH (60 micrograms LH-RP1/rat) was capable of increasing significantly both PRL specific binding and capacity. Capacity reached the values of 498 +/- 103 and 507 +/- 240 fm/mg protein for LHRH and LH respectively. LHRH action appeared to be mainly mediated through LH secretion, since no difference was found between LHRH and LH. LHRH and LH injections alone were unable to modify PRL binding, suggesting that they only potentiate E2 and PRL action. These results show for the first time that LH is involved in the regulation of PRL receptors in the rat mammary gland.     

阉割和睾丸酮替代对雄性老年大鼠下丘脑和血浆促黄体生成素释放激素水平的影响

应用放射免疫分析(RIA)技术比较了年青(2—3月龄)和老年(24—26月龄)雄性大鼠下丘脑和血浆促黄体生成素释放激素(LHRH)水平及其在睾丸切除(ORDX)和睾丸酮(T)替代下LHRH水平的变化。老年大鼠血浆T水平明显降低,下丘脑LHRH含量亦呈明显下降趋势,但血浆LHRH水平与年青大鼠十分接近。在ORDX和T替代下,两组动物血浆T水平没有明显差别,但老年大鼠下丘脑和血浆LHRH的变化率却不同程度地低于年青大鼠。上述结果提示,老年雄性大鼠下丘脑LHRH神经元系统的负反馈能力明显削弱,这也许是下丘脑-垂体-睾丸轴呈增龄性衰变的重要原因之一。     

Correlation of hypothalamic LHRH, pituitary LH and plasma LH in developing rats.

Hypothalamic LHRH, pituitary LH and plasma LH levels were measured in rats of both sexes from day 5-60 after birth. The content of hypothalamic LHRH was very high in one-week-old male and female rats. It declined gradually till day 17 in the female rat and sharply on day 10 in the male rat. Subsequently the content of hypothalamic LHRH increased and showed peak values on day 25 in the female rat and on day 45 in the male rat. It decreased markedly at respective times of puberty in both sexes (day 37 in the female rat and day 52-60 in the male rat). Results of the study suggest that maturation of hypothalamo-hypophyseal-axis proceeds in three distinct stages. Observations on days 17, 25 and 37 in the female rat and on days 5, 7, 10 and 22 in the male rat clearly show an inverse relationship between hypothalamic LHRH and plasma LH and a parallel relationship between pituitary and plasma LH. Marked decline in the content of hypothalamic LHRH at respective times of puberty in both sexes indicates that the release of threshold levels of LHRH from the hypothalamus may apparently be the event initiating the pubertal changes in rat.     

Immunoreactive delta sleep-inducing peptide in the rat hypothalamus, pituitary and adrenal gland: effects of adrenalectomy

Immunoreactive (IR) delta sleep-inducing peptide (DSIP) was examined by immunocytochemistry in the rat pituitary and adrenal gland and found to be colocalized with IR thyroid-stimulating hormone in the pituitary and with noradrenaline in the adrenal medulla. IR-DSIP was also detectable in nerve fibers in the posterior pituitary. By radioimmunoassay, IR-DSIP was quantified in plasma and tissue extracts after uni- or bilateral adrenalectomy. Significantly elevated plasma levels of IR-DSIP were measured 5 days after bilateral adrenalectomy (p less than 0.001). IR-DSIP was increased (p less than 0.05) in pituitary extracts from bilaterally adrenalectomized rats after 5 days, but not after 14 or 28 days. Sham- and unoperated animals did not significantly differ in plasma or tissue concentration of IR-DSIP. High-performance liquid chromatography of C18 SEP-PAK purified hypothalamus extracts revealed a single peak of IR-DSIP material of lower hydrophobicity than synthetic DSIP. The elevated concentration of IR-DSIP in the rat pituitary and plasma after bilateral adrenalectomy is consistent with the previously suggested role of DSIP to influence the activity of the hypothalamic-pituitary-adrenal axis.     

A change in basal FSH release accompanies the onset of the second or selective phase of increased serum FSH in the cyclic rat

Experiments were undertaken to investigate if changes occur at the level of the anterior pituitary gland to result in selective follicle-stimulating hormone (FSH) release during late proestrus in the cyclic rat. At 1200 h proestrus, prior to the preovulatory luteinizing hormone (LH) surge in serum and the accompanying first phase of FSH release, serum LH and FSH concentrations were low. At 2400 h proestrus, after the LH surge and shortly after the onset of the second or selective phase of FSH release, serum LH was low, serum FSH was elevated about 4-fold, pituitary LH concentration was decreased about one-half and pituitary FSH concentration was not significantly decreased. During a two hour $\text{in}$$\text{vitro}$ incubation, pituitaries collected at 2400 h released nearly two-thirds less LH and 2.5 times more FSH than did pituitaries collected at 1200 h. Addition of luteinizing hormone releasing hormone (LHRH) to the incubations caused increased pituitary LH and FSH release. However, the LH and FSH increments due to LHRH in the 2400 h pituitaries were not different from those in the 1200 h pituitaries. The results indicate that a change occurs in the rat anterior pituitary gland during the period of the LH surge and first phase of FSH release which results in a selective increase in the basal FSH secretory rate. It is suggested that this change is primarily responsible for the selective increase in serum FSH which occurs during the second phase of FSH release.     

Rhythms in immunoreactive melatonin in the retina and harderian gland of rats: Persistence after pinealectomy

Immunoreactive melatonin levels were measured in the retina and Harderian gland of adult male rats throughout a 24 hour period. The animals were maintained under a light:dark cycle of 14:10 (lights on at 0600h). In intact animals, immunoreactive melatonin values in both organs exhibited a 24h rhythm with peak levels being measured at 0800h, 2 hours after lights on. Pinealectomy significantly increased peak levels at 0800h in both the retina and the Harderian gland. Gonadectomy abolished the peak retinal melatonin levels at 0800h. Likewise, continual light exposure for 1 week depressed the melatonin peak in the retina but not in the Harderian gland.     

The release of pituitary gonadotrophins by luteinizing hormone releasing hormone in the intact,castrated and aspermatogenic rat

The change in serum gonadotrophin concentration in response to synthetic Luteinizing Hormone Releasing Hormone (LHRH - 400 ng i.v.) was investigated under barbiturate anaesthesia in adult male rats either chronically castrated, rendered aspermatogenic by the administration of α-chlorohydrin 12–16 weeks previously (to remove inhibin), or treated with vehicle. A single injection of LHRH increased serum LH and FSH concentrations similarly in both intact and aspermatogenic rats. In castrated rats the amount of LH released was much greater and the FSH secretion sustained. A second injection produced a similar increase although a second peak of FSH could not be detected in castrated rats as the FSH level was still elevated. The increase in LH levels was two to three times larger in response to the second injection of LHRH than to the first in all groups. The results do not support the hypothesis that the enhanced gonadotropin response to castration in the aspermatogenic rat is due to increased pituitary sensitivity to LHRH.     

Effect of thyroidectomy on rhythmic gonadotropin release.

Nonstress blood samples were obtained from intact and thyroidectomized (TE) male rats at 3-hr intervals over a 24-hr period via rapid decapitation. The animals were thyroidectomized when 40 days old and used 6 weeks later. Intact animals showed periodicity in serum LH (P less than 0.01) and prolactin (P less than 0.01). Both gonadotropins began increasing after 8 PM and peak levels occurred at 11 PM. In contrast, 24-hr periodicity was not observed in serum FSH. Corticosterone levels in these same serum samples showed the expected circadian periodicity. After TE, the 24-hr pattern in all gonadotropins was altered significantly. Serum LH increased (P less than 0.01) and circadian periodicity appeared to be absent. FSH and prolactin levels were increased and decreased, respectively (P less than 0.01), with serum prolactin showing a 9-hr phase shift. Prolactin began increasing at 2 AM and reached a peak at 8 AM. Corticosterone in TE animals showed a 24-hr rhythm similar to that of intact rats. These findings confirm our previous observations that nonstress serum LH and prolactin levels fluctuate with a 24-hr periodicity and suggest that the level of, and the phase angle betweeen, these rhythms is markedly influenced by pituitary-thyroid activity.     

Adrenalectomy does not influence basal secretion of testosterone in rat in vivo

We have studied the effect of adrenalectomy on the testicular secretion of testosterone in the rat. In the acute period following adrenalectomy plasma testosterone levels were reduced but this was no different from those levels in appropriate sham-operated controls. This reduction in plasma testosterone levels is probably a result of direct effects of anaesthesia and surgical stress. Whilst studies on the late effect of adrenalectomy avoided this problem, plasma testosterone levels were normal in both adrenalectomised and sham-operated animals. Resetting of anterior pituitary-gonadal relationships may mask the absence of any contribution made by the adrenal gland to testicular steroidogenesis. In contrast to previous data we were unable to demonstrate that adrenalectomy influenced the secretion of testosterone in the male rat.     

The effect of ketamine/xylazine and carbon dioxide on plasma luteinizing hormone releasing hormone and testosterone concentrations in the male Norway rat

The effect of a commonly used anaesthetic, ketamine/xylazine and/or carbon dioxide (CO(2)) on plasma luteinizing hormone releasing hormone (LHRH) and testosterone concentrations was determined in male Sprague-Dawley rats. These values were compared with values obtained from pre-anaesthetic control samples. Ketamine/xylazine treatment did not significantly affect testosterone concentrations. In contrast, LHRH started to decrease one hour after ketamine/xylazine administration and continued to significantly decrease after 24 h. In addition, in the CO(2) euthanasia-only group, LHRH concentrations were also significantly decreased. These results suggest that ketamine/xylazine anaesthesia followed by CO(2) euthanasia 24 h later is exerting a significant effect on LHRH concentrations 24 h after anaesthetizing, while only having a slight effect on testosterone, and that CO(2) is exerting an immediate significant effect on LHRH. In conclusion, LHRH analysis should be avoided after ketamine/xylazine anaesthesia and CO(2) euthanasia.     

Estrogen and the subcellular distribution of luteinizing hormone releasing hormone: Rate sedimentation studies

Rate sedimentation of the 900×G supernatants (S₁) of hypothalamic homogenates from untreated male rats or ovariectomized rats with or without 5 μg estradiol benzoate (EB) revealed two populations of LHRH particles: a minor, slowly sedimenting one (peak 1) and a major, more rapidly sedimenting one (peak 2). Some LHRH-containing material also sedimented to the bottom of the gradient. The ovariectomized rats displayed more heterogeneity of particulate LHRH than did the male rats. Furthermore, the administration of EB to ovariectomized rats altered the relative sedimentation pattern of LHRH. In ovariectomized rats, hypotonic shock of S₁ prior to rate sedimentation eliminated peak 2 and post-peak 2 LHRH and increased free LHRH at the top of the gradient. Peak 1 LHRH was still present and was elevated after EB treatment. Also, EB treatment lowered the free LHRH at the top of the gradient. These data demonstrate that the administration of EB to an ovariectomized rat alters the subcellular distribution of LHRH.