O. Khorram

Role of arginine vasopressin in control of ACTH and LH release during stress

To determine the role of arginine vasopressin (AVP) in stress-induced release of anterior pituitary hormones, AVP antiserum or normal rabbit serum (NRS) was micro-injected into the 3rd ventricle of freely-moving, ovariectomized (OVX) female rats. A single 3 microliter injection was given, and 24 hours later, the injection was repeated 30 min prior to application of ether stress for 1 min. Although AVP antiserum had no effect on basal plasma ACTH concentrations, the elevation of plasma ACTH induced by ether stress was lowered significantly. Plasma LH tended to increase following ether stress but not significantly so; however, plasma LH following stress was significantly lower in the AVP antiserum-treated group than in the group pre-treated with NRS. Ether stress lowered plasma growth hormone (GH) levels and this lowering was slightly but significantly antagonized by AVP antiserum. Ether stress also elevated plasma prolactin (Prl) levels but these changes were not significantly modified by the antiserum. To evaluate any direct action of AVP on pituitary hormone secretion, the peptide was incubated with dispersed anterior pituitary cells for 2 hours. A dose-related release of ACTH occurred in doses ranging from 10 ng (10 p mole)-10 micrograms/tube, but there was no effect of AVP on release of LH. The release of other anterior pituitary hormones was also not affected except for a significant stimulation of TSH release at a high dose of AVP. The results indicate that AVP is involved in induction of ACTH and LH release during stress. The inhibitory action of the AVP antiserum on ACTH release may be mediated intrahypothalamically by blocking the stimulatory action of AVP on corticotropin-releasing factor (CRF) neurons and/or also in part by direct blockade of the stimulatory action of vasopressin on the pituitary. The effects of vasopressin on LH release are presumably brought about by blockade of a stimulatory action of AVP on the LHRH neuronal terminals.

Possible negative ultra-short loop feedback of luteinizing hormone releasing hormone (LHRH) in the ovariectomized rat

Abstract To determine if LHRH might act within the brain to modify its own release, repeated blood samples were removed from conscious ovariectomized rats and minute doses of LHRH were injected into the third ventricle (3V). The effect of these injections on plasma LH and FSH was measured by radioimmunoassay (RIA). The higher dose of intraventricular LHRH (10 ng in 2 yl) induced an increase in plasma LH within 10 min after its injection. Plasma LH decreased for the next 60 min. This was followed by restoration of LH pulses characteristic of the ovariectomized rat. This dose of LHRH slightly elevated plasma FSH concentrations. In stark contrast, a 10 fold lower dose of 1 ng of LHRH injected into the ventricle resulted in a highly significant decrease of plasma LH at 10 min following injection, followed by return of LH pulsations. There was no effect on the pulsatile release of FSH. The results are interpreted to mean that at the higher dose, sufficient LHRH reached the site of origin of the hypophyseal portal vessels in the median eminence so that it diffused into portal vessels and was delivered to the gonadotrophs to induce LH release. In contrast, the lower dose provided sufficient hypothalamic concentrations of the peptide to suppress the discharge of the LHRH neurons, thereby leading to a decline in plasma LH, indicative of an ultrashort-loop negative feedback of LHRH to suppress its own release.

Partial characterization of immunoreactive substance P in the rat pituitary gland

Two distinct carboxy-terminus-directed anti-substance P (SP) sera (R-1C and R-6G) were used to characterize immunoreactive SP (I-SP) in acetic acid extracts of anterior pituitary (AP) and posterior pituitary (PP) glands of adult male rats. The tissue concentrations of I-SP measured by R-1C and R-6G were comparable. The contents of I-SP were 600-1150 pg/AP and 25-52 pg/PP. I-luteinizing hormone releasing hormone and I-somatostatin (I-SOM) were undetectable in AP extracts, but PP extracts contained the equivalents of 325-785 pg I-SOM/gland. Serial dilutions of AP and PP extracts produced displacement curves with both SP antisera that were parallel to the respective synthetic SP standard and hypothalamic extract displacement curves. Gel filtrations of AP and PP extracts on a Sephadex G-25 column produced I-SP peaks eluting in the same fractions as synthetic SP and hypothalamic I-SP. However, the AP I-SP profile also revealed a side peak migrating between the void volume and the major I-SP peak. Neither immunoreactive species in the AP extract were eliminated when eluted with 6.0 M guanidine HCl, a strong denaturing agent. In vitro incubation of paired anterior hemipituitaries for 30 min in the presence of a 56 mM K+ concentration resulted in a significant (p less than .0001), 25-fold increase in the release of I-SP into the incubation medium above the mean control value. Radiofrequency lesions placed in the median eminence-arcuate region of male rats caused a significant (p less than .001) reduction of I-SP in both the AP and PP. These reductions were inversely related to the plasma prolactin values. The elevation in plasma prolactin was taken as an index of completeness of lesions. We conclude that: 1) the rat pituitary contains I-SP as assessed by its immunologic and chromatographic behavior, 2) K+ depolarization is a potent stimulator of the release of AP I-SP in vitro, 3) the ME-arcuate region is important for the maintenance of pituitary I-SP levels in the rat.

The Role of Brain Peptides in the Control of Anterior Pituitary Hormone Secretion

A host of peptides act within the hypothalamus and also directly on the pituitary to control the secretion of anterior pituitary hormones. The classical releasing and inhibiting hormones act directly on the pituitary to inhibit or stimulate the release of anterior pituitary hormones. They appear also to act within the brain to modulate their own release and that of other releasing factors. With the discovery of many additional brain peptides, most of which were found initially in the gastrointestinal tract, many of these have also been examined, and it is apparent that there are important hypothalamic actions of a number of these peptides to alter pituitary hormone secretion (McCann et al., 1984b). In this chapter, we review some of our recent work in this area beginning with an examination of possible additional peptidic releasing or inhibiting factors that may directly alter pituitary hormone secretion and continuing with an examination of the intrahypothalamic action of the various peptides to alter their own release and that of pituitary hormones.

Recent studies on the role of brain peptides in control of anterior pituitary hormone secretion.

Recent work in our laboratory on the role of peptides to influence release of pituitary hormones by direct action on the gland and also some of the interactions of these peptides at the hypothalamic level to alter release of pituitary hormones will be reviewed. Considerable evidence from hypothalamic stimulation and lesion studies suggests the existence of a separate FSH-releasing factor (FSHRF). We have been able to purify a bioactive FSHRF which appears to be distinct from LHRH. Consequently, we believe that a distinct FSHRF will ultimately be isolated. With regard to prolactin, it is now clear that it is under dual control by both prolactin-inhibiting (PI) and prolactin-releasing factors (PRF). Although dopamine acts as a PIF, our recent fractionation studies indicate the existence of a peptidic PIF in hypothalamic extracts which can be separated from dopamine and GABA. The peptidic PIF is eluted from Sephadex in the same position originally described by us a number of years ago. Thus, inhibitory control is probably mediated by a combination of factors which would include dopamine, possibly GABA and a peptidic PIF. A number of peptides have been shown to have PRF activity which include TRF and also VIP. In recent studies, we have shown a prolactin-releasing action of oxytocin on male hemipituitaries or dispersed pituitary cells. Furthermore, high doses of oxytocin given intravenously released prolactin in male rats. There is a correlation between estrogen-induced prolactin release and an increase in plasma oxytocin and a correlation between suckling-induced oxytocin and prolactin release. These results suggest that oxytocin may be an important PRF.(ABSTRACT TRUNCATED AT 250 WORDS)

Simultaneous Induction of Neuropeptide Y and Gonadotropin-Releasing Hormone Release in the Rabbit Hypothalamus

Neuropeptide Y (NPY) can induce the release of endogenous mediobasal hypothalamic gonadotropin-releasing hormone (MBH-GnRH) and pituitary gonadotropins, especially LH. In these studies, we monitored changes in endogenous NPY concentrations at 20-min intervals for 6-8 h during push-pull perfusion (PPP) in both the mediobasal hypothalamus (MBH) and the third cerebroventricle (3VT) of ovarian intact, conscious rabbits. Because previous studies had shown that copper ion can induce hypothalamic GnRH release, cupric acetate (CuAc) was administered either intravenously or intraventricularly during the PPP to manipulate changes in NPY concentrations. Our results show that NPY concentrations in both MBH and 3VT PPP samples were detectable by radioimmunoassay. Administration of CuAc sharply increased hypothalamic NPY release within the same time interval as that for induction of hypothalamic GnRH release. The results are consistent with the hypotheses that NPY may act as a neuromodulator for hypothalamic GnRH secretion, or that common mechanisms drive secretion of these two neuropeptides.

Effect of α-Melanocyte-Stimulating Hormone on Basal and Stimulated Release of Prolactin:Evidence for Dopaminergic Mediation

The effects of alpha-melanocyte-stimulating hormone (alpha-MSH) on basal and stimulated of prolactin were studied. Intraventricular (i.v.t.) injection of 2 microgram of alpha-MSH in ovariectomized (OVX) rats produced a 50% depression of plasma prolactin levels within 10 min, whereas 100 ng of alpha-MSH or saline had no effect. 2 microgram of alpha-MSH (i.v.t.) produced a more dramatic and sustained depression of prolactin levels if initial prolactin concentrations were elevated. This dose of alpha-MSH was also effective in lowering plasma prolactin levels in OVX rats, primed 72 h earlier with 50 microgram of estradiol benzoate (s.c.), although this depression was of lower relative magnitude than in the unprimed OVX animals. The intravenous injection of a dopamine receptor blocker, spiroperidol (0.1 mg/kg), prior to (i.v.t.) alpha-MSH blocked the inhibitory effect of alpha-MSH on prolactin release. In vitro, alpha-MSH in doses ranging from 0.5 to 5 microgram did not alter prolactin release from hemipituitaries of male or OVX female rates and from dispersed pituitary cells from female rats. These results suggest that alpha-MSH can modulate prolactin release possibly via the hypothalamic dopaminergic system, and that this modulation may be of greater significance during stress.

Changes in hypothalamic and pituitary content of immunoreactive alpha-melanocyte-stimulating hormone during the gestational and postpartum periods in the rat.

Abstract α-Melanocyte-stimulating hormone (α-MSH) was measured in the mediobasal hypothalamus (MH), median eminence (ME), preoptic-suprachiasmatic area (POA–SCN), anterior (AL), and posterior lobes (PL) of the pituitary gland during the gestational and postpartum periods in the rat. The content of α-MSH in the MH and POA–SCN compared to estrous levels was lower during the later days of gestation and decreased further in the MH during lactation in association with the elevated plasma prolactin (Prl). Distinct increases in the ME content of α-MSH compared to estrous levels occurred on Days 8 and 12 of the gestational period and Day 14 of the postpartum period. A significant increase in PL content of α-MSH compared to Days 5–11 and 17–20 occurred on Day 4 of gestation, and no significant changes were detected in the AP concentration of α-MSH throughout the period studied. In vitro, PLs and ALs from females on Day 4 of gestation secreted more α-MSH into the incubation medium than tissues from animals on Day 20. These results suggest that α-MSH of both brain and pituitary origin may play a role in mediating some of the physiological changes which occur during pregnancy and lactation.

Blockade of stress induced prolactin release in monosodium glutamate-treated rats

To elucidate the role of the medial basal hypothalamus (MBH) in stress-induced prolactin (Prl) release, adult male rats which had received monosodium glutamate (MSG 4 mg/g) on alternate days for the first 10 days of life were subjected to ether stress for 2 min. Blood samples were drawn at 0, 15, 30 and 60 min after etherization through an indwelling jugular catheter implanted 1 day before the experiment. Sixty minutes after etherization the dopamine receptor blocker spiroperidol (0.1 mg/kg) was injected intravenously and another blood sample was withdrawn 60 min later. Although there were no differences in basal levels of plasma Prl between control and MSG-treated animals, a dramatic elevation of plasma Prl was observed in control rats 15 min after etherization, whereas no significant change was seen in MSG-treated animals. Spiroperidol significantly increased plasma Prl in both groups of animals, but the magnitude of the increase in Prl levels of MSG-treated rats was roughly one-third (p less than 0.005) that of control animals. When morphine (3 mg/kg) was injected to test its capability to stimulate Prl release in MSG-treated rats, significant increases of similar magnitude of plasma Prl were observed in both groups. These results show that (1) the tuberoinfundibular dopaminergic system is functioning in MSG-treated rats to maintain Prl at low levels, (2) the stimulatory effect of morphine on Prl release is intact in MSG-treated rats, and (3) the tuberoinfundibular dopaminergic system is not responsible for stress-induced Prl release, but rather another mechanism located within the arcuate nucleus (ARC) and highly sensitive to destruction by MSG mediates stress-induced Prl release.

On the Presence of a Nondopaminergic, Peptidergic Prolactin Release-Inhibiting Factor in Hypothalamic Extracts of Infantile Rats

Crude hypothalamic extracts prepared from brains of 1-day-old rats produced a dose-dependent inhibition of prolactin (Prl) release by adult male hemipituitaries, and to a lesser extent by hemipituitaries of adult ovariectomized (OVX), estrogen-primed rats. These extracts contained 6-fold lower levels of dopamine than adult hypothalami. The inhibitory effect of the adult hypothalamic extracts, contrary to infantile hypothalamic extracts could be blocked by spiroperidol. Digestion of the infantile hypothalamic extracts with pronase totally abolished their Prl release-inhibiting activity, indicating the peptidic nature of this inhibitory substance. In contrast to their effect on Prl release by hemipituitaries, infantile hypothalamic extracts stimulated Prl release from dispersed anterior pituitary cells of OVX estrogen-primed rats, pointing to the importance of estrogen in modulating prolactin release-inhibiting factor (PIF) activity and the possibility that the PIF receptor is trypsin-sensitive.