In vivo Activity of the LHRH Pulse Generator as Determined with Push-Pull Perfusion of the Anterior Pituitary Gland of Unrestrained Intact and Castrate Male Rats

Estradiol induces surges of prolactin (PRL) and luteinizing hormone (LH) secretion as well as lactotrope proliferation in female rats. We examined whether these hypothalamus-dependent events require the direct action of estradiol on the anterior pituitary gland by selective blockade of its peripheral actions, using ICI182,780 (ICI), an antiestrogen that cannot cross the blood-brain barrier. Injection of ICI into ovariectomized rats, at a dose of 250 µg/day for 4 days, almost completely inhibited estradiol-induced growth of the uterus, proliferation of lactotropes as determined by bromodeoxyuridine incorporation, and afternoon surges of LH secretion. However, ICI only partially inhibited estradiol-induced surges of PRL secretion and had no effect on estradiol-induced tonic inhibition of LH secretion even at the highest dose of 1,000 µg/day. The inhibitory effects of ICI found at 250 µg/day were attributable to its selective peripheral, but not central actions since ICI did not alter hypothalamic expression of progesterone receptors, an estradiol-dependent brain process. Estradiol-induced increases in the number of progesterone receptor-immunoreactive cells in the hypothalamic ventromedial nucleus and the medial preoptic area were not inhibited by this dose of ICI but were inhibited by 500 µg/day tamoxifen, an antiestrogen that can cross the blood-brain barrier. Treatment of cycling female rats with 250 µg/day ICI beginning from diestrus day 2 was also effective in blocking estrous lactotrope proliferation and preovulatory surges of LH secretion but not PRL secretion. Finally, in ovariectomized estradiol-treated pup-deprived lactating rats, ICI did not affect suckling-induced PRL secretion but completely blocked lactotrope proliferation. These results suggest that a direct estradiol action on the anterior pituitary gland is required for lactotrope proliferation and the positive feedback action on LH secretion but not for the secretory surges of PRL or for negative feedback.

Section: Discussionmentioning

confidence: 99%

Direct Actions of Estradiol on the Anterior Pituitary Gland Are Required for Hypothalamus-Dependent Lactotrope Proliferation and Secretory Surges of Luteinizing Hormone but Not of Prolactin in Female Rats

Yin

Kawashima

2002

“…Effects of steroids on the synthesis of pro-GnRH and its processing into native GnRH have also been reported, but remain controversial [15, 19, 20, 21, 22, 23]. Similarly, increased [24, 25]or decreased [19, 26, 27]GnRH secretion has been observed after gonadectomy. Finally, it has been shown that testosterone influences the PKC-dependent metabolic cascade, thereby affecting the secretion of pro-GnRH-derived peptides triggered by activation of this signalling pathway [15].…”

Section: Introductionmentioning

confidence: 99%

In vitro Secretion of Gonadotropin-Releasing Hormone (GnRH) and [Hydroxyproline<sup>9</sup>]GnRH from the Rat Hypothalamus Exhibits a Differential Sensitivity to Castration and Second Messengers

Rochdi¹,

Theraulaz²,

Enjalbert³

et al. 1998

The decapeptide [hydroxyproline⁹]GnRH (HypGnRH) has been characterized as an endogenous posttranslational product of the gonadotropin-releasing hormone (GnRH) precursor in a wide range of mammalian brains. Despite consistent biological effects, its secretion by the hypothalamus remains hypothetical. We report here in vitro secretion of HypGnRH and GnRH by the hypothalamus from intact and castrated male rats and provide evidence that they are differentially regulated. Both peptides were identified by two anti-GnRH antibodies of different specificities after separation under two high-performance liquid chromatography conditions. Calcium dependency of HypGnRH release was demonstrated under stimulation with KCl in the absence or presence of Ca²⁺, as well as with Bay K 8644, veratridine, methoxyverapamil, or tetrodotoxin. Activation of signaling pathways involving adenylate cyclase and protein kinases A and C (PKC) induced HypGnRH release. Expression of data as percentage of release over tissue stores revealed a two- to threefold higher release of HypGnRH than of GnRH under the different modes of stimulation used, except under PKC activation which triggered a comparable recruitment of both peptides. Castration selectively affected PKC-coupled GnRH secretion which showed a twofold lesser release than in intact rats, while the HypGnRH release was unaffected. We conclude that HypGnRH and GnRH are not secreted from the hypothalamus according to the same mechanisms.

“…One to two weeks prior to use in the experiments male SpragueDawley rats (250-300 g) were fitted with a PPC as described by Dluzen and Ramirez [5,6], The animals were caged singly and maintained in a temperature-controlled room with a photoperiod of 14 h light: 10 h darkness (lights on at 05:00 h). Food and water were available ad libitum.…”

Section: Animalsmentioning

confidence: 99%

“…A miniaturized PPC [5] was stereotaxically implanted into each rat and directed toward the rostral portion of the anterior pituitary gland [6,7]. Details of cannula construction, implantation and technical vali dation have been previously reported .…”

Section: Implantation O F Ppcmentioning

confidence: 99%

Roles of Somatostatin and Growth Hormone-Releasing Factor in Ether Stress Inhibition of Growth Hormone Release

Aguila

Pickle²,

Yu³

et al. 1991

In order to evaluate the release of somatostatin (SRIF) and growth hormone-releasing factor (GRF) into the pituitary gland in response to ether stress, a push-pull perifusion (PPP) technique has been used in freely moving rats. Push-pull cannulae (PPC) were implanted into the anterior pituitary (AP) glands of male rats. After a 7-day recovery period the rats were fitted with indwelling jugular catheters. The next day the animals were subjected to PPP of the AP during 1 h followed by ether stress (2 min) and another hour of perifusion. The perifusion flow was 20 µl/min and 10-min fractions were collected and assayed for SRIF and GRF by RIA. Plasma growth hormone (GH) levels were assayed every 10 min. At the end of the experiments, the accuracy of PPC tip placements was ascertained with a dissecting microscope. Under basal conditions there were 2.9 pulses/h of SRIF with an amplitude of 12.76 ± 0.46 pg. The output of SRIF and GRF in the 10-min period beginning with application of ether was increased 2-fold (p < 0.005 and p < 0.01, respectively). Interestingly, the increased release of SRIF continued for an additional 10 min, whereas GRF output decreased and was almost undetectable. The release of both GRF and SRIF had returned to basal values 20–30 min after stress. Mean plasma GH levels were significantly lowered 10 min after stress. Each of the 9 animals showed a restoration of pulsatile GH release to basal levels within 20–30 min after stress. Our findings provide compelling evidence that SRIF plays a prominent role in stress-induced inhibition of GH release in the rat by blocking the response to the transient elevation of GRF and continuing to suppress GH release for 20 min.