Jiun-Yih Yeh

Effects of hyperprolactinemia on testosterone production in rat Leydig cells

The pathogenesis of hyperprolactinemia (hyperPRL) induced hypogonadism has been suggested to be related with a dysfunction of hypothalamus–pituitary–testis axis. While the direct inhibitory effects of prolactin (PRL) on testosterone (T) release have been demonstrated, the mechanism is still unclear. Our previous study demonstrated a diminished T release in the testicular interstitial cells (TICs) from the anterior pituitary (AP)‐grafted rats as compared with the control, and the pattern was in agreement with the in vivo model. However, TICs incubation cannot totally represent the response of the Leydig cells. Therefore, a Percoll gradient purified Leydig cell model was adopted to explore the response of T release under similar challenges in this study to investigate the effects of hyperPRL on the Leydig cells per se. HyperPRL in male rats was induced by grafting rat AP under the renal capsule. The control animals were grafted with rat brain cortex tissue (CX). Six weeks after grafting, the rats were sacrificed. Either TICs or Leydig cells were isolated, respectively, for in vitro incubation and challenge. Challenge drugs included human chorionic gonadotropin (hCG, 0.05 IU/ml), steroidogenic precursors (25‐OH‐cholesterol, 10−6 M; pregnenolone, 10−6 M), forskolin (an anenylyl cyclase activator, 10−4 M) and 8‐bromo‐3′:5′ cyclic adenosine monophosphate (cAMP) (8‐Br‐cAMP 10−4 M). T released by TICs or Leydig cells was determined by radioimmunoassay. The TICs from the AP‐grafted rats showed lower levels of T release than the control group while the purified Leydig cells demonstrated a reverse pattern in response to challenges of hCG, steroidogenic precursors, forskolin and 8‐Br‐cAMP. In hyperPRL rats, a paradoxical pattern of T release between TICs and purified Leydig cells is observed. The purified Leydig cells from AP‐grafted rats demonstrated a higher level amount of T release than the control after stimulation. The phenomenon can be attributed to the change of Leydig cell sensitivity to the stimulation after the effects of chronic hyperPRL. Moreover, another possibility is the role played by other interstitial cells to modulate steroidogenesis in Leydig cells. J. Cell. Biochem. 80:313–320, 2001.

The role of cyclic AMP production, calcium channel activation and enzyme activities in the inhibition of testosterone secretion by amphetamine

1 The aim of this study was to investigate the mechanism by which amphetamine exerts its inhibitory effect on testicular interstitial cells of male rats. 2 Administration of amphetamine (10−12–10−6 M) in vitro resulted in a dose‐dependent inhibition of both basal and human chorionic gonadotropin (hCG, 0.05 iu ml−1)‐stimulated release of testosterone. 3 Amphetamine (10−9 M) enhanced the basal and hCG‐increased levels of adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) accumulation in vitro (P<0.05) in rat testicular interstitial cells. 4 Administration of SQ22536, an adenylyl cyclase inhibitor, decreased the basal release (P<0.05) of testosterone in vitro and abolished the inhibitory effect of amphetamine. 5 Nifedipine (10−6 M) alone decreased the secretion of testosterone (P<0.01) but it failed to modify the inhibitory action of amphetamine (10−10–10−6 M). 6 Amphetamine (10−10–10−6 M) significantly (P<0.05 or P<0.01) decreased the activities of 3β‐hydroxysteroid dehydrogenase (3β‐HSD), P450c17, and 17‐ketosteroid reductase (17‐KSR) as indicated by thin‐layer chromatography (t.l.c.). 7 These results suggest that increased cyclic AMP production, decreased Ca2+ channel activity and decreased activities of 3β‐HSD, P450c17, and 17‐KSR are involved in the inhibition of testosterone production induced by the administration of amphetamine.

Direct effects of prolactin on corticosterone release by zona fasciculata‐reticularis cells from male rats

The role of prolactin (PRL) in the male is not fully defined. The aim of this study was to investigate the function and mechanism of PRL on the production of corticosterone by zona fasciculata‐reticularis (ZFR) cells in vitro. The ZFR cells were obtained from male rats under normal, hyperprolactinemic, or hypoprolactinemic situation. PRL stimulated the corticosterone release in a dose‐dependent pattern in the ZFR cells from normal male rats. The cellular adenosine 3′‐5′‐cyclic monophosphate (cAMP) concentration positively correlated with PRL concentration in the presence of forskolin or 3‐isobutyl‐1‐methylxanthine (IBMX). PRL enhanced the stimulatory effects of cAMP mimetic reagents, i.e., forskolin, 8‐bromo‐adenosine 3′,5′‐cyclic monophosphate (8‐Br‐cAMP), and IBMX on the release of corticosterone. The adenylate cyclase inhibitor (SQ22536) inhibited the corticosterone release in spite of presence of PRL. Nifedipine (L‐type calcium channel blocker) did not inhibit corticosterone release. The hyperprolactinemic condition was actualized by transplantation of donor rat anterior pituitary glands (APs) under kidney capsule. By comparison with the cerebral cortex (CX)‐grafted group, AP‐graft resulted in an increased release of corticosterone, 3β‐hydroxysteriod dehydrogenase (HSD) activity and cAMP production by ZFR cells. Acute hypoprolactinemic status was induced by bromocriptine for 2 days. The results showed the productions of corticosterone were lower in hypoprolactinemic group than in control group, which were persistent along with different ACTH concentrations. These results suggest that PRL increase the release of corticosterone by ZFR cells via cAMP cascades and 3β‐HSD activity. J. Cell. Biochem. 73:563–572, 1999.

Regulation of testosterone secretion by prolactin in male rats.

The goal of this study was to characterize the mechanism by which hyperprolactinemia alters testosterone production in rat testicular interstitial cells (TICs). Hyperprolactinemia was induced by grafting 2 anterior pituitary (AP) glands under the subcapsular space of the kidney in experimental rats. Control rats were grafted with brain cortex (CX). Six weeks post‐grafting, rats were challenged with human chorionic gonadotropin (hCG) then, the changes in either plasma testosterone or luteinizing hormone was measured. Additionally, TICs were isolated and challenged in vitro with hCG or prolactin, and the testosterone release measured by radioimmunoassay. Further investigation in signal transduction as intracellular 3′:5′ cyclic adenosine monophosphate (cAMP) production was observed under a regulation of forskolin or SQ22536. After the challenge of hCG or GnRH, the AP‐grafted rats showed a suppressed response in testosterone release as compared to those in the CX‐grafted group. The in vitro data from the AP‐grafted rats compared to the CX‐grafted animals showed a diminished response in testosterone release upon hCG stimulation. Administration of forskolin or SQ22536 disclosed dysfunction of adenylate cyclase in TICs from the AP‐grafted rats. When 8‐Br‐cAMP was incubated with TICs, the testosterone production was lower in the AP‐grafted compared to the CX‐grafted group. These results suggest that in addition to adenylate cyclase dysfunction, inefficiency of post‐cAMP pathways are also involved in the hypogonadism elicited by hyperprolactinemia in rats. J. Cell. Biochem. 74:111–118, 1999.

Role of testicular interstitial macrophages in regulating testosterone release in hyperprolactinemia

Hyperprolactinemia‐induced hypogonadism has been linked to a dysfunction of the hypothalamus‐pituitary‐testis axis. The direct inhibitory effects of prolactin on the testicular release of testosterone have also been demonstrated, though their mechanisms remain unclear. Incubation of rat testicular interstitial cells (TICs) with prolactin stimulated the release of testosterone. TICs from rats with anterior pituitary‐grafting‐induced hyperprolactinemia release lower amounts of testosterone than controls. However, Leydig cells isolated from anterior pituitary‐grafted rats release a greater amount of testosterone. These paradoxical observations have remained unexplained. This study examined the roles of testicular interstitial macrophages and of their product, tumor necrosis factor‐α (TNF‐α), in regulating Leydig cells under condition of hyperprolactinemia. Hyperprolactinemia was induced by grafting two anterior pituitary glands of rats under the renal capsule. Control animals were grafted with rat cortex tissue. The rats were sacrificed 6 weeks later. TICs and macrophages, and Leydig cells were isolated for in vitro incubation and drugs challenge. Testosterone released by testicular interstitial or Leydig cells was measured by radioimmunoassay. TNF‐α concentration in the medium of TICs or macrophages was measured by enzyme‐linked immunosorbent assay (ELISA). A dose‐dependent stimulation of TNF‐α secretion in the medium of TICs or macrophages by the prolactin challenge was observed. Higher amounts of TNF‐α were released by TICs in the anterior pituitary‐grafted rats than in the control group. In contrast, the release of TNF‐α by testicular interstitial macrophages isolated from the anterior pituitary‐ and cortex‐grafted groups was quantitatively similar. Challenge with human chorionic gonadotropin did not modify the TNF‐α release by testicular interstitial macrophages in either group. Challenge of Leydig cells with TNF‐α inhibited their release of testosterone stimulated by human chorionic gonadotropin, but not their basal testosterone release. These different patterns of testosterone release in TICs versus Leydig cells cultures in anterior pituitary‐grafted rats may be due to the influence of testicular interstitial macrophages. These observations correlate with in vivo conditions, where prolactin increases the release of TNF‐α by testicular interstitial macrophages, which, in turn, decreases the human chorionic gonadotropin‐stimulated release of testosterone by Leydig cells. In summary, hyperprolactinemia‐induced hypogonadism involves a mechanism of prolactin‐originated, macrophage‐mediated inhibitory regulation of testosterone release by Leydig cells. TNF‐α, one of the cytokines secreted by macrophages, may play a key role in this mechanism.

Interaction of carbohydrate metabolism and rat liquid gastric emptying in sustained running

Background:  Moderate to severe running usually leads to gastrointestinal dysmotility and critical energy exhaustion. It is unknown whether the carbohydrate metabolism of runners can influence gastric emptying (GE). Using a running rat model, the present study explored the impact of exercise/carbohydrate metabolism on liquid GE.

Involvement of Cholecystokinin (CCK) Receptor in the Adaptation of Gastric Emptying Induced by Adrenocorticotropin (ACTH) in Male Rats

It has been shown that gastric emptying is delayed in stressed animals. This investigation was to study the effect of adrenocorticotropin (ACTH), a stress hormone, on the gastric emptying and intestinal transit in rats. Fasted male rats were injected intraperitoneally (i.p.) with saline, ACTH (10 or 20 μg/ml/kg), lorglumide (a selective potent cholecystokinin A receptor antagonist, 10 μg/ml/ kg) or ACTH (20 μg/ml/kg) and lorglumide (10 μg/ml/kg) at 30 min before decapitation. All rats were orally ingested with liquid meal containing radioactive Na_2 ^(51)CrO_4 (0.5 μCi/ml) and 10% charcoal at 15 min before decapitation, and blood samples were collected. Administration of ACTH significantly increased the concentrations of plasma corticosterone, but decreased the levels of plasma cholecystokinin (CCK) and gastric emptying as well as intestinal transit in rats. The decreased gastric emptying, but not intestinal transit, was reversed by the treatment of lorglumide. These results suggested that CCKA receptor was involved in the inhibitory effect of ACTH on the gastric emptying in rats.

Involvement of Cholecystokinin (CCK) Receptor in the Regulation of Dexamethasone on Gastric Emptying in Male Rats

Dexamethasone is a potent glucocorticoid, which has been used in the anti-inflammation therapy. It has been well known that the gastrointestinal (GI) motility is affected by stress. However, the role of glucocorticoids in regulating GI motility is unclear. The purpose of this study was to investigate the effect and action mechanism of dexamethasone on the GI motility. In the first study, the male rats were injected intraperitoneally (i.p.) with saline or dexamethasone (15 or 30 μg/ml/kg) 1 h before decapitation. In the second study, dexamethasone (30 μg/ml/kg) and lorglumide (a selective cholecystokinin A receptor, CCKA, antagonist) were injected intraperitoneally 60 min and 30 min prior to decapitation, respectively. All experimental animals were orally ingested with radioactive Na_2^(51)CrO_4 containing 10% charcoal by a PE-205 tubing into the stomach after a 24 h fast and then decapitated 15 min later. The small intestine was divided equally into ten segments. The radioactivity in stomach and each segment of intestine was counted by a gamma-counter. The ratios of gastric emptying and charcoal transit were calculated. Blood samples were collected. The concentrations of CCK and corticosterone in plasma samples were measured by radioimmunoassay, and those of serum adrenocorticotropin (ACTH) were measured by an enzyme immunoassay (EIA) kit. All data were expressed as mean ± s. e. mean, and analyzed by the analysis of variance (ANOVA). The differences between specific means were analyzed by Dunnetts test. The results demonstrate that dexamethasone dose dependently inhibited the level of serum ACTH and the gastric emptying, and the intestinal transit is also inhibited by dexamethasone at the dose of 15 μg/ml/kg. Plasma CCK and corticosterone concentrations were dose-dependently decreased by dexamethasone injection. Furthermore, after injection of lorglumide, the decreased gastric emptying caused by dexamethasone was restored to the control level. These results suggested that dexamethasone-induced inhibition of gastric emptying and intestinal transit is mediated by an action on CCKA receptor.