bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential Effects of Direct and Indirectly Acting Cholinergic Agonists on Growth Hormone Release in Man, and Lack of Effect on Cortisol Secretion
Authors: Casanueva F. F. , Villanueva L. , Cordido F. , Diaz Y. , Cabranes J. A. , Spiegel R. , Dieguez C.
Published: September 2006

Acetylcholine plays a key role in the modulation of growth hormone secretion. In fact, growth hormone release after provocative stimuli is blocked by muscarinic cholinergic antagonists, and conversely, indirect cholinergic agonists potentiate growth hormone secretion. To further understand the mechanism by which cholinergic pathways exert their effects, we have compared the growth hormone and cortisol secretion elicited on normal volunteers by pyridostigmine and by RS‐86 (2‐ethyl‐8‐methyl‐2,8‐diazaspiro‐((4,5))‐decan‐1,3‐dion hydrobromide). The former acts by inhibiting acetylcholinesterase, thus being an indirect muscarinic agonist, while the latter is a muscarinic receptor agonist which binds directly to and stimulates cholinergic receptors.

In six subjects, pyridostigmine (120 mg po) induced an increase of growth hormone of 11.0±2.4 μg/L at 90 min, significantly greater than following placebo administration (1.4 ± 0.3 μg/L). In another group of five volunteers, RS‐86 was administered in separate tests at a dose of 0.5, 1 and 2 mg po. Growth hormone levels were not altered by any RS‐86 dose compared with placebo values. Neither pyridostigmine nor RS‐86 altered cortisol values.

These results suggest that the mechanism of action of the cholinergic agonists is of great importance for their growth hormone‐releasing capabilities, and question the accepted view of a cholinergic regulation of cortisol secretion in man.
Hormonal Activation of Female Sexual Behavior is Accompanied by Hypothalamic Norepinephrine Release
Authors: Vathy Ilona , Etgen Anne M.
Published: September 2006

The present study employed the intracranial microdialysis technique to measure norepinephrine release in the ventrolateral dendritic fields of the ventromedial hypothalamus of freely‐moving animals before and during ovarian steroid (estradiol and progesterone) activation of female sexual behavior (lordosis). One day after implantation of a dialysis probe, animals were injected with 3 μg of estradiol benzoate followed 44 h later by 200 μg of progesterone. Introduction of a male rat 4 h after progesterone treatment was correlated with dramatic increases in extracellular norepinephrine levels measured in dialysates of the ventrolateral ventromedial hypothalamus of female rats which displayed high levels of lordosis behavior. In contrast, female rats given the same steroid treatment but which did not show lordosis responses did not have elevated norepinephrine levels in their dialysates. Moreover, animals that received an estrogen antagonist concurrently with the estrogen treatment had neither an increase in ventromedial hypothalamic levels of norepinephrine during behavior testing nor did they display lordosis. These results indicate a close relationship among ovarian steroids, noradrenergic transmission in the ventromedial hypothalamus, and the expression of female sexual behavior.
Role of the Hypothalamic Paraventricular Nucleus in Mediating Stress‐Related Litter Deficits in the Golden Hamster
Authors: Pratt N. C. , Lisk R. D.
Published: September 2006

Primiparous female hamsters (Mesocricetus auratus) were mated to proven breeders and stressed during early pregnancy. Females were housed singly throughout gestation except for Days 4, 5 and 6 when they were paired for ten‐min intervals three times each day with another female matched for age, weight and day of pregnancy. Within each of the pairs, one female was consistently dominant to the other. Controls were exposed to a novel area instead of a conspecific. At parturition, all pups were counted, sexed and weighed. There were no significant differences between control and dominant females' litter sizes or sex ratios (defined as percentage male). Subordinate females produced significantly smaller litters than control or dominant dams and significantly lower sex ratios than control dams. Subordinates produced fewer males than control or dominant dams, but there were no differences in the number of females produced. The paraventricular nucleus (PVN) of the hypothalamus has been implicated as a relay center for the physiological response to stress. Bilateral lesions of the PVN were performed on another group of females and the same protocol described above was followed after the females recovered from surgery. When this ‘stress relay center’ was lesioned, subordinates did not show the significant deficits in litter size and sex ratio. Sham‐operated females showed a similar response to social stress to that of intact females. These results suggest that subordinate dams produce smaller litters via selective resorption of males in utero and that the PVN may be a relay center for the mediation of this response.
An Electron Microscopic Immunocytochemical Study on the Tyrosine Hydroxylase‐Positive, Dopamine Beta‐Hydroxylase‐Negative Cells in the Pineal Gland of Golden Hamsters
Authors: Shiotani Yahe , Jin Kui Long , Kawai Yuriko , Kiyama Hiroshi
Published: September 2006

We have examined the ultrastructure of tyrosine hydroxylase‐positive, dopamine β‐hydroxylase‐negative neuron‐like cells in the pineal gland of golden hamsters by electron microscopic immunocytochemistry using antiserum to tyrosine hydroxylase. The tyrosine hydroxylase‐positive cells were scattered throughout the parenchyma of the pineal gland. Their cell bodies were oval or spindle‐shaped (maximum diameter 15 to 20 μm) with one or two short cell processes and a large round nucleus. They contained mitochondria, short rough‐surfaced endoplasmic reticulum, Golgi apparatus, and granulated vesicles (50 to 100 nm). The cell membrane was in close contact with the pinealocyte cell body, pinealocyte processes and tyrosine hydroxylase‐negative or ‐positive nerve terminals.
Effect of Tunicamycin, Swainsonine, Castanospermine, Beta‐Hydroxynorvaline and Monensin on the Post‐Translational Processing of Rat Prolactin Molecular Forms
Authors: Bollengier F. , Velkeniers B. , Mahler A. , Vanhaelst L. , Hooghe‐Peters E.
Published: September 2006

Prolactin cells derived from the anterior pituitaries of female rats were cultured in the presence of tunicamycin, swainsonine, castanospermine, β‐hydroxynorvaline and monensin in order to study their effect on the post‐translational processing of the Mr 17,000, 23,000 and 26,000 prolactin molecular forms. Sodium‐dodecyl‐sulphate polyacrylamide electrophoresis and subsequent immunoblotting revealed that: 1) tunicamycin, swainsonine and castanospermine, compounds that are essentially known as inhibitors of the N‐glycosylation processus, had no effect on Mr 17,000, 23,000 and 26,000 rat prolactin; 2) βhydroxynorvaline, which has been assumed to inhibit processing of pre‐prolactin to mature 23,000 prolactin, did not increase the synthesis of 26,000 rat prolactin. In case of inhibition of the processing of a pre‐prolactin to mature prolactin, one would expect an increase of the pre‐prolactin; consequently, we could not establish the 26,000 rat prolactin, we revealed in immunoblotting, as a pre‐prolactin; 3) monensin affected the post‐translational processing of 17,000 and 26,000 rat prolactin, but left the 23,000 mature form intact. This is an important finding for the following reasons: monensin blocks the transport of secretory and membrane proteins, and this blockade prevents the cleavage of these molecules; indeed, production of 17,000 rat prolactin, a form of cleaved prolactin, was inhibited. Monensin also affects glycosylation and 26,000 rat prolactin has been identified as a presumably O‐iinked glycosylated variant. The fact that its synthesis is inhibited by monensin treatment, but not by inhibitors of the N‐linked process, particularly tunicamycin, and that 26,000 rat prolactin is susceptible to mild alkali and decomposition via β‐elimination are decisive arguments in favour of the O‐linked glycosidic linkage.
Development and Validation of a Two‐Site Immunochemiluminometric Assay for Rat Growth Hormone‐Releasing Hormone
Authors: Page M. D. , Lewis M. D. , Lewis B. M. , Weeks I. , Scanion M. F.
Published: September 2006

We describe the development and validation of a two‐site immunochemiluminometric assay for rat growth hormone‐releasing hormone (GHRH) based on the affinity purification of polyclonal rabbit antisera to rat GHRH using a human 1–29 GHRH affinity column. Assay sensitivity is 3.2 pg/ml using 100 μl of unextracted sample and the working range for the assay within 15% confidence limits is 64 to 5,000 pg/ml. Rat hypothalamic extract and secreted material demonstrated a single large peak of immunoreactive material coeluting with synthetic rat GHRH on high‐performance liquid chromatography with a smaller, earlier peak which probably represents methionine sulphoxide [Met(O)27] GHRH. Extracted material diluted in parallel to the standard curve. Incubated rat hypothalami readily released measurable amounts of rat GHRH which responded appropriately to depolarization with 60 mM K+ in a Ca2+‐dependent manner.
Glucocorticoid Receptor Immunoreactivity in the Rat Intermediate Lobe
Authors: Bertini Luca T. , Westphal Marines M. , de Kloet Ronald E. , Kiss Jozsef Z.
Published: September 2006

Cells containing Type II glucocorticoid receptor (GR) immunoreactivity were identified in the rat pituitary gland by immunocytochemistry using a specific monoclonal antibody. At light microscopic level, GR immunoreactive cells were located in the intermediate lobe in addition to the well known GR‐containing cell population in the anterior lobe. In both groups of cells GR appeared predominantly in the cell nuclei. Adrenalectomy resulted in a decrease in staining intensity of the anterior lobe and changed the pattern of fluorescence in a minority of cells where cytoplasmic staining became predominant. These changes appeared less marked in the intermediate lobe. Dexamethasone administration reversed the adrenalectomy‐induced alterations of GR staining in both lobes. At the electron microscopic level, GR immunoreactive sites were revealed by the protein A‐gold technique. In contrast to the distribution of fluorescence, GR was localized in cell nuclei as well as in the cytoplasm in both lobes. Quantitative estimates indicate that about 40% more immunoreactive sites are present in the anterior lobe than in the intermediate lobe. The presence of GR in the intermediate lobe suggests that this pituitary region, like the anterior lobe, is influenced by glucocorticoid hormones.
Embryonal Development of Arginine Vasotocin/Mesotocin Gene Expression in the Chicken Brain
Authors: Milewski N. , Lvell R. , Grossmann R. , Ellendorff F.
Published: September 2006

An ontogenic series of chick embryo brains (4, 6, 9,12,14,16 and 18 days of incubation, hatching day: 21) was coronally or saggitally sectioned and investigated for expression of the arginine vasotocin (AVT)/mesotocin (MT) gene. To this end a 39mer oligonucleotide recognizing the AVT/MT encoding sequence of their respective mRNAs was constructed employing optimized codon usage and was used for in situ hybridization. AVT/MT mRNA‐expressing neurons were first detected on embryonal day (E) 6 adjacent to the third ventricle. By E9 the periventricular nucleus had expanded in size but the hybridization signal was weaker. These results suggest a migration of cells in all directions away from the third ventricle into the diencephalon; some perikarya were even observed at the lateral pial surface above the optic chiasm. By E12, brain differentiation had advanced to distinct hypothalamo‐neurohypophyseal nuclei (supraoptic and paraventricular nuclei) as well as to accessory groups expressing AVT/MT mRNA. Two cell types were then distinguishable in the paraventricular nucleus; the specific mRNA was expressed either as a weak or a strong hybridization signal. Comparison with other studies suggested that differentiation had almost attained adult level though cell growth and differentiation of supraoptic and paraventricular nuclei still occurred. This was complete by E18 when individual cells were larger and the intensity of the hybridization signal became very strong.

RNase pretreatment depressed the signal by 100%. Northern blot hybridization of RNA extracted from newly hatched chicks verified the specificity of the probe. A single band was evident, indicating an mRNA of approximately 700 bases, which is only found in hypothalamus and not in muscle, liver or extra‐hypothalamic brain, and corresponds in size closely with mammalian vasopressin mRNA.

The data demonstrate a very early development (E6) of neurohypophyseal hormone gene expression in the chick embryo brain with important differentiation around E9. At E12 magnocellular neuronal arrangement resembled adult neurons but the gene expression signal increased in intensity until E18.
Cardiovascular Instability Induced by Intracerebroventricular Administration of Vasopressin or Corticotrophin‐Releasing Factor in Conscious Long Evans and Brattleboro Rats
Authors: Gardiner S. M. , Compton A. M. , Bennett T.
Published: September 2006

Conscious Long Evans and Brattleboro rats, chronically instrumented with pulsed Doppler probes around left renal and superior mesenteric arteries and the distal abdominal aorta, had vasopressin or corticotrophin‐releasing factor administered intracerebroventricularly. In Long Evans rats, vasopressin caused no cardiovascular instability, but in Brattleboro rats, vasopressin caused the development of marked oscillations in blood pressure and heart rate. Under the latter conditions, falls in blood pressure were accompanied by increases in hindquarter blood flow. Similar oscillations in blood pressure, heart rate and regional haemodynamics were seen in Long Evans and Brattleboro rats given corticotrophin‐releasing factor. These results indicate that centrally‐administered vasopressin and corticotrophin‐releasing factor can have an important influence on the control of regional haemodynamics. In some respects the effects of the peptides resemble the changes seen during the defence reaction.
Mechanism of the Haemotensive Action of Porcine Relaxin in Anaesthetized Rats
Authors: Parry L. J. , Poterski R. S. , Summerlee A. J. S. , Jones S. A.
Published: September 2006

Experiments were done to examine the pressor effect of iv porcine relaxin in anaesthetized rats. Acute injections of relaxin caused consistent and sustained rises in systemic blood pressure that were dose‐dependent within the physiological range. Pretreatment of rats with a specific vasopressin (V1) receptor antagonist, but not an α‐adrenoreceptor antagonist, substantially reduced the pressor effect of relaxin. After the vasopressin receptor antagonist, small rises in blood pressure occurred after a longer latent period, compared with the responses in intact animals. The data clearly indicate that acute injections of relaxin cause a pressor response that is predominantly affected via the release of vasopressin. The possible sources of the persistent hypertensive component are discussed and it is suggested that relaxin might act through the central angiotensinergic systems to release vasopressin and cause a pressor response.
Effect of Hypothalamic Deafferentation on the Pulsatile Secretion of Luteinizing Hormone in Ovariectomized Lactating Rats*
Authors: Tsukamura Hiroko , Maeda Kei‐ichiro , Ohkura Satoshi , Yokoyama Akira
Published: September 2006

The pulsatile luteinizing hormone (LH) secretion in ovariectomized lactating rats bearing complete (CD), anterior (AD), anterolateral (ALD), posterior (PD), or roof (RD) deafferentation of the hypothalamus was determined. All lactating rats were ovariectomized on Day 2 of lactation (Day 0, day of parturition). The deafferentation of nerve fibres to the mediobasal hypothalamus was performed on Day 6 or 7 of lactation. Twenty‐four h after the surgery, blood samples were taken through the indwelling atrial catheter every 6 min for 3 h. Plasma concentrations of LH and prolactin (PRL) were measured by radioimmunoassay. The loss of LH pulses associated with lactation was still apparent following AD, PD and sham‐deafferentation (SD); pulsatile LH secretion was, however, present in rats with CD, ALD and RD despite continued suckling. The only significant difference in plasma PRL concentrations among the various groups was a reduction in the PRL level in rats with RD in comparison to those with SD. We conclude that the neural signal responsible for the inhibition of pulsatile LH release by suckling is conveyed through the dorsal part of the hypothalamus and PRL does not mediate the suppression of LH pulses in mid‐lactation.
Inward Rectification (Ih) in Immunocytochemically‐ldentified Vasopressin and Oxytocin Neurons of Guinea‐Pig Supraoptic Nucleus
Authors: Erickson Kenneth R. , Ronnekleiv Oline K. , Kelly Martin J.
Published: September 2006

Intracellular recordings of magnocellular neurons from the supraoptic nucleus of guinea‐pigs were made with KCI/K citrate‐ and biocytin‐filled electrodes. Fifty of 99 cells exhibited a time‐dependent inward rectification (TDR). The TDR was activated during hyperpolarizing current pulses to membrane potentials more hyperpolarized than −75 mV. In voltage‐clamp recordings, an inward current appeared at voltage steps more hyperpolarized than −75 mV, with properties similar to the slow inward rectifier (Ih) described in other tissues. The Ih was blocked by 2 mM CsCI. BaCI2 (100 to 500 μM) did not block the Ih. Immunocytochemical identification of the recorded cells revealed that both vasopressin (AVP)‐ and oxytocin (OT)‐ containing neurons exhibited an Ih.
Voltage‐Clamp Recordings from Identified Dissociated Neuroendocrine Cells of the Adult Rat Supraoptic Nucleus
Authors: Cobbett Peter , Weiss Mark L.
Published: September 2006

In vitro intracellular recordings of membrane potential obtained from the oxytocin and vasopressin neurons of the mammalian hypothalamo‐neurohypophysial system in slices (1–3) and expiants (4, 5) have demonstrated many of the intrinsic properties of these magnocellular neuroendocrine cells (MNCs). Voltage‐clamp techniques, which are required to study directly the currents underlying intrinsic or transmitter‐evoked potential changes, have been applied to cultured embryonic (6) or neonatal supraoptic neurons (7–9) and have been successfully applied to adult supraoptic neurons in situ in only one laboratory (10, 11). We have modified a technique for dissociation of viable adult guineapig hippocampal neurons (12) to dissociate supraoptic MNCs from adult rats for voltage‐clamp studies. MNCs were selectively labelled with a fluorescent dye in vivo so that they could be identified after dissociation and prior to making recordings. These data have been published in abstract form elsewhere (13, 14).
Mu‐Selective Opioid Peptides Stimulate Prolactin Release in Lactating Rats
Authors: Baumann Michael H. , Rabii Jamshid
Published: September 2006

The fact that opiates elicit prolactin secretion is well known. However, we have recently discovered that morphine does not stimulate prolactin release in lactating rats. The physiological basis for this alteration in opiate sensitivity during lactation is not known. Since morphine‐induced prolactin secretion in male rats is mediated via the mu opioid receptor subtype, one possible explanation is that mu receptors are down‐regulated during lactation. To address this possibility, the effects of mu opioid peptides on prolactin secretion were examined in lactating rats. The presumed mu‐selective peptides DAGO ([D‐Ala2, Me‐Phe4, Gly‐ol5]‐enkephalin) and PLO‐17 ([NMe‐Phe3, D‐Pro4]‐morphiceptin) were administered to primiparous lactating rats and the resulting hormone responses measured. Both DAGO and PLO‐17 caused a rapid and significant rise in plasma prolactin during lactation. The prolactin‐releasing effects of both peptides were naloxone reversible, suggesting involvement of opioid receptors. Moreover, the DAGO‐induced secretion of prolactin could be completely abolished by pretreatment with the irreversible mu antagonist β‐funaltrexamine. In lactating rats, DAGO and PLO‐17 were poor growth hormone‐releasing agents, providing further evidence for the mu specificity of these peptides. These results imply that during lactation, as in other reproductive states, mu opioid receptor sites are positively coupled to the prolactin secretory mechanism. Thus, the previously observed inability of morphine to elicit prolactin release in lactating rats cannot be explained on the basis of down‐regulation of mu opioid receptors.
Identification and Localization of Insulin‐Like Growth Factor Receptors in Human Infant Pituitary Gland—Similar Distribution to Somatotrophs
Authors: Walker David A. , Hogg Annette , Haynes Kerry , Cox Alison , Jonas Helen , Mendelsohn Frederick A. O. , Werther George A.
Published: September 2006

Insulin‐like growth factors(IGF) are involved in feedback regulation of growth hormone(GH) secretion from the pituitary. Though receptors for IGF‐I and IGF‐II have been identified on particulate preparations of rat pituitary, their localization and relationship to GH‐secreting acidophils has not been determined. We used quantitative in vitro autoradiography and immunocytochemistry to simultaneously determine the distribution of IGF receptors and GH‐secreting cells in human infant pituitary gland. Frozen or fixed post‐mortem human infant pituitary glands were sectioned for binding studies, or immunocytochemistry. Binding for IGF‐I and IGF‐II showed characteristic specificity for respectively Type I and Type II receptors. Binding sites were visualized throughout the pituitary gland, with similar density and distribution for IGF‐I and IGF‐II receptor sites. Receptor density was two‐fold higher in anterior than posterior pituitary, with highest density in the lateral horns of the anterior pituitary. The distribution of GH‐containing cells (acidophils) was similar to IGF receptor distribution. Increased density of IGF receptors in regions of GH‐secreting cells may point to the mechanism whereby IGF uniquely inhibits synthesis of human GH in contrast to its promotion of synthetic processes in other cells.
The Origin and Regulation of Posterior Pituitary Vasopressin Ribonucleic Acid in Osmotically Stimulated Rats
Authors: Levy Andy , Lightman Stafford L. , Carter David A. , Murphy David
Published: September 2006

Vasopressin ribonucleic acid (VP RNA) accumulates in the posterior lobe of the rat pituitary in response to an osmotic stimulus. The accumulation of posterior pituitary VP RNA can be prevented by stalk transection or the intracerebroventricular injection of colchicine. The hypothalamic and pituitary VP RNAs however, have different structures and are independently regulated. Depletion of serotonin with parachlorophenylalanine blocks the osmotically induced accumulation of both VP and oxytocin RNA in the hypothalamus but does not affect the accumulation of VP RNA in the posterior pituitary gland. In Brattleboro rats, posterior lobe oxytocin RNA but not VP RNA, increases after osmotic stimulation. Although axonal transport of RNA may occur, our results are also consistent with VP RNA synthesis in pituicytes controlled by hypothalamic neuroendocrine factors released by neurohypophyseal terminals in response to osmotic stimuli.
Activation of Opioid Receptors in the Mediobasal Hypothalamus Stimulates Prolactin Secretion in the Conscious Rat
Authors: Kapoor Ranjna , Willoughby John O.
Published: September 2006

In an attempt to localize the opioid receptor(s) (mu, delta and kappa) involved in opioid‐stimulated prolactin release in the conscious male rat, opioid agonists were microinjected into the mediobasal hypothalamus and prolactin levels measured before and after injection.

The specific mu agonist, DAGO ((D‐Ala2, NMe‐Phe4, Gly‐ol5)‐enkephalin) was the most effective in eliciting prolactin release, the smallest effective dose being 0.01 nmoles. The specific delta agonist, DPDPE ((D‐Pen2, D‐Pen5)‐enkephalin) had no significant effect even at the highest dose of 10 nmoles. The specific kappa agonist, U50,488H ((trans‐3,4‐dichloro‐N‐methyl‐N‐(2‐(1‐pyrrolidinyl)‐cyclohexyl)‐benzene acetamide) was effective at the doses 1.0 and 10 nmoles.

We conclude that mu and kappa opioid receptors in the mediobasal hypothalamus are involved in opioid‐stimulated prolactin release and that delta receptors are not.
Neuronal Uptake of Monoclonal Anti‐Vasopressin Antibodies in vivo and Relationship with the Physiological Status of Vasopressin Neurosecretory Cells
Authors: Burlet A. , Arahmani A. , Tankosic P. , Burlet C.
Published: September 2006

Mouse monoclonal anti‐vasopressin antibody was injected just above one hypothalamic paraventricular nucleus of the rat brain. Immunocytochemistry and morphometric analysis showed that the antibody was taken up by neurons with the size and stereotaxic distribution of vasopressin‐producing neurons. Labelled neurons were counted with an electronic image analyser. When the rats were deprived of drinking water for 48 h before injection, specific uptake was significantly increased in the caudal part of the nucleus. Conversely, rehydration following dehydration or chronic treatment with 1‐deamino‐8‐D‐arginine vasopressin significantly decreased neuronal uptake, mainly in the rostral subdivision of the nucleus. By contrast, bilateral adrenalectomy performed 2 weeks before injection did not modify the number of labelled magnocellular neurons, though the accumulation of injected antibody in the external layer of the median eminence indirectly demonstrated the stimulation of antibody uptake by parvocellular vasopressin neurons.

The number of labelled neurons was therefore directly related to the physiological state of the vasopressin‐producing neurons. Further investigations will have to be performed to prove that the immunological targeting of peptidergic neurons offers a new tool to act in vivo on central neurons.
Anatomical Localization of Corticotropin‐Releasing Factor and Arginine Vasopressin in the Human Hypothalamus; the Effect of Corticosteroids on their Concentrations in Human and Rat Hypothalami
Authors: Pralong François P. , Linton Elizabeth A. , Favrod‐Coune Charles A. , Lowry Phillip J. , Muller Alex F. , Gaillard Rolf C.
Published: September 2006

In this study, we have determined the distribution of corticotropin‐releasing factor and vasopressin in the human hypothalamus, and investigated the effect of glucocorticoid administration on the concentrations of both peptides. Corticotropin‐releasing factor and vasopressin were measured by a two‐site immunoradiometric assay and/or radioimmunoassay. The presence of both peptides was studied in extracts of eleven areas of the human hypothalamus as well as in the pituitary stalk from autopsied patients who had been free of chronic steroid administration (n = 14) or had received Corticosteroids (n = 5). Unlike vasopressin, corticotropin‐releasing factor was detected in all extracts: the highest concentration was found in the pituitary stalk, whilst the lowest detectable amounts occurred in the supraoptic and lateral areas and in the mammillary bodies. This pattern of distribution is similar to that reported for the rat hypothalamus. The excellent correlation (R = 0.994) between corticotropin‐releasing factor data obtained by immunoradiometric assay and by radioimmunoassay renders the presence of a corticotropin‐releasing factor precursor molecule in the extracts highly unlikely. In the human brain extracts, glucocorticoid treatment affected neither the content, nor the distribution of corticotropin‐releasing factor and vasopressin. In the rats, dexamethasone administration produced a 50% decrease in the vasopressin content (P < 0.05) of the basomedial and dorsal parts of the hypothalamus and had no effect on the corticotropin‐releasing factor content of these areas. These results show that the distribution of corticotropin‐releasing factor is similar in both human and rat hypothalami. The rat data suggest that negative feedback effects of glucocorticoids involve changes in hypothalamic vasopressin content.
Central Oxytocin Stimulates Luteinizing Hormone Release in the Marmoset, a Primate which Fails to Show Lactationally‐lnduced Infertility
Authors: O'Byrne K.T. , Lunn S. F. , Coen C. W.
Published: September 2006

In several species, including humans, suckling is known to suppress pulsatile gonadotrophin secretion and substantially reduce fertility (1). During suckling high titres of oxytocin are present in both plasma (2, 3) and cerebrospinal fluid (CSF) (4) and there is some evidence that oxytocin is inhibitory to gonadotrophin secretion by an action on t h e central nervous system, (5, 6). The common marmoset, however, has a different physiological response and fails to show lactational infertility; ovarian cyclicity is resumed immediately post‐partum (7). We have therefore examined the effect of centrally administered oxytocin on luteinizing hormone (LH) secretion in this species.

We report that LH release was stimulated with a rapid:esponse and in a dose‐dependent manner by oxytocin injected into t h e third cerebral ventricle. In contrast, oxytocin administered intravenously was without effect on LH secretion. These results clearly indicate, for the first time, a stimulatory role for oxytocin in LH secretion in the marmoset, a species which fails to demonstrate lactational infertility.
Arachidonate Metabolism in the Anterior Pituitary: Effect of Arachidonate Inhibitors on Basal and Stimulated Secretion of Prolactin, Growth Hormone and Luteinizing Hormone. I. Hormone Release from Incubated or Perifused Pituitary Fragments
Authors: Wandscheer Dolores E. , Bihoreau Claire , Bertrand Philippe , Clauser Hubert , Kordon Claude
Published: September 2006

The potential involvement of arachidonic acid metabolites in the regulation of adenohypophyseal secretion was analysed on pituitary glands from male rats incubated in the presence of various inhibitors with different mechanisms of action: two inhibitors of phospholipase A2 (parabromophenacylbromide, PB and compound CB 874), an inhibitor of cyclooxygenase‐ and lipoxygenase‐catalysed pathways (5, 8, 11, 14‐eicosatetraynoic acid, ETYA) and an inhibitor of cyclooxygenase (ε‐lysyl acetylsalicylate, ASP). Under conditions which minimize side effects of the drugs, all inhibitors reduced prostaglandin synthesis and release, without affecting the metabolic integrity of the tissues (assessed by their intracellular adenosine triphosphate levels).

All agents tested (PB, ETYA, ASP) suppressed prolactin secretion induced either by thyrotropin‐releasing hormone or vasoactive intestinal peptide. Basal prolactin secretion was sensitive to phospholipase A2 inhibitors. Similar inhibitions were obtained with ETYA and CB 874 on growth hormone secretion under basal conditions as well as after stimulation by growth hormone‐releasing factor, thyrotropin‐releasing hormone, or vasoactive intestinal peptide.

In contrast, luteinizing hormone secretion, stimulated or not by gonadotropin‐releasing hormone, was not sensitive to any of the agents used.

It is concluded that, in intact male hemipituitaries, arachidonic acid metabolism is involved in the stimulation of prolactin and growth hormone secretion by neuropeptides. In contrast, luteinizing hormone release does not seem to depend on that mechanism. It has been verified that the inhibitors of arachidonic acid metabolism do not directly interfere with adenylate cyclase, or with the activation of protein kinase C, two enzymes which are involved in the regulation of secretory mechanisms.
Progestin Receptor Induction and Sexual Behavior by Estradiol Treatment in Male and Female Rats
Authors: Coirini Héector , McEwen Bruce S.
Published: September 2006

Sex differences in the induction of cytosolic progestin receptors (CPR) by estrogen priming were correlated with the sex differences in behavioral responses. We evaluated the temporal relationship between CPR in several brain regions and pituitary and the time‐course of 17β‐estradiol (E2) activation of female sexual behavior in gonadectomized male and female rats implanted with subcutaneous E2 Silastic capsules for 6 h, 24 h and 48 h. Both CPR levels and mating behavior increase monotonically with the time of E2 exposure. Induction of CPR was observed in the periventricular region of the preoptic area (PVPOA), arcuate nucleus (ARC), ventromedial nuclei (VMN) and pituitary in both sexes. A small induction of CPR was found in parietal cortex. The VMN in female rats showed a significant E2‐induced CPR increase at all times of exposure, while in male rats this induction was only significant after 24 h. Significant sex differences in absolute CPR levels and E2‐induced receptors were found in the following structures: VMN, 18 h after 6 h of E2 treatment and after 24 h and 48 h of continuous E2 exposure; PVPOA, only after 48 h of continuous E2 exposure; ARC at 24 h and 48 h; and pituitary after all E2 treatment. Mating behavior was tested under two conditions: E2 alone (2 h after removal of E2 capsules) and E2+progesterone (2 h after a progesterone injection given 10 min after concluding the first test). Receptivity was first observed after 24 h E2 exposure in female rats, whereas in male rats a small response appeared only after 48 h of E2 exposure. After progesterone priming, the time of E2 exposure necessary for expression of female sexual behavior was reduced to 6 h in females and 24 h in males. The appearance of mating behavior appears to follow that of inducible CPR in the VMN in both sexes. In addition, the CPR levels associated with the first receptivity either by male rats (16.6 fmol/mg protein) or female rats (15.3 fmol/mg protein) are very similar suggesting the presence of a threshold level controlling the expression of feminine sexual behavior. It is likely that inhibitory neural input plays a role in determining the threshold level of E2‐induced CPR, which is sufficient to trigger lordosis behavior.
Effects of Cysteamine Pretreatment and Hypothalamic Periventricular Nucleus Lesion on the Cold‐Stimulated Thyrotropin Responses to Intracerebroventricular 5‐Hydroxytryptamine in Male Rats
Authors: Toivonent M. , Tuomainen P. , Rauhala P. , Männistö P. T.
Published: September 2006

The hypothalamic somatostatinergic system was devitalized in male rats by intracerebroventricular (icv) cysteamine (CSH) pretreatment (250 μg/rat/day into the third ventricle) on 4 consecutive days or by a limited lesion of the hypothalamic periventricular nucleus (PeVNx). The acute effect of icv serotonin (5‐HT) on the cold‐stimulated thyrotropin (TSH) and prolactin responses were studied in these animals. The experiments were performed 24 h after the last saline or CSH infusions and 7 days after the sham‐ or PeVN‐lesions. CSH and PeVNx decreased the hypothalamic somatostatin content by 44% to 57% and 19% to 28%, respectively. PeVNx did not affect hypothalamic thyrotropin‐releasing hormone content. 5‐HT infusion (9 μg/rat icv) into the anterior third ventricle elevated, although not significantly, TSH levels in both saline‐ or CSH‐pretreated rats. 5‐HT infusion into the anterior third ventricle did not affect TSH in sham‐operated rats. However, 5‐HT augmented the cold‐stimulated TSH levels after PeVNx compared to sham‐lesion. Inversely, 5‐HT infusion (9 μg/rat) into the posterior third ventricle inhibited TSH secretion irrespective of the pretreatment or lesion. The inhibitory action of 5‐HT on TSH was significantly suppressed by CSH. 5‐HT infusions elevated serum prolactin levels irrespective of the infusion site, pretreatment or lesion. 5‐HT infusion into both the anterior and the posterior third ventricle decreased rectal temperature in saline‐pretreated, sham‐ and PeVN‐lesioned rats. The hypothermie effect of 5‐HT was weakened by CSH. The hypothalamic levels of noradrenaline, dopamine and their metabolites were not significantly affected by CSH and PeVNx. 5‐HT infusion into the anterior third ventricle decreased hypothalamic dopamine content in both saline‐ and CSH‐pretreated rats. However, such an effect was not seen in sham‐ or PeVN‐lesioned animals. Although CSH is an inhibitor of dopamine‐β‐hydroxylase, this activity was not reflected in serum TSH or prolactin levels. The results support our hypothesis of the site‐dependent action of icv 5‐HT or TSH secretion. The elevation of TSH levels may arise from the inhibition of somatostatin release from rostral anterior hypothalamus. The inhibition of TSH secretion may result from the inhibition of thyrotropin‐releasing hormone release from more caudal periventricular structures of the hypothalamus.
Effects of Stimulation of the Superior Cervical Ganglia and Local Application of Noradrenaline on Electrical Activity of the Syrian Hamster Pineal Gland
Authors: Patel S. , Demaine C.
Published: September 2006

The effects of electrical stimulation of either or both superior cervical ganglia and the effects of local application of L‐noradrenaline on the spontaneous electrical activity of hamster pineal cells were evaluated. Extracellular recordings from pineals of anaesthetized hamsters revealed that the spontaneous electrical activity was mainly regular with interspike intervals attributed between 12 to 20 ms during the daytime and mainly irregular with interspike intervals of 1 to 250 ms during the night. Following stimulation of the superior cervical ganglia, either unilaterally or bilaterally, or local application of noradrenaline, the responses of these pineal cells fell into three major categories: A) non‐responsive, B) excited, and C) inhibited. There was no relationship between the magnitude or form of response and the source of stimulus i.e. the right superior cervical ganglia or the left superior cervical ganglia. Almost all inhibited responses from electrical stimulation of the superior cervical ganglia could be correlated with inhibited responses from the local application of noradrenaline whereas excited responses could not. In general, these results suggest that the spontaneous electrical activity of some pineal cells is influenced by inputs from the superior cervical ganglia and that the inhibitory input is likely to be mediated through the release of noradrenaline. The excitatory input from the superior cervical ganglia is probably mediated by another neurotransmitter. The heterogeneity of responses suggests that different receptors or different cell types may be involved.
Temperature and Protein Kinase C Modulation of Gonadotropin‐Releasing Hormone Receptors in Pituitary Cells from Intact or Castrated Male Rats
Authors: Leblanc Pierre , Haour France , I'Heritier Andrée , Mounier Françoise , Kordon Claude
Published: September 2006

Binding constants of [125 I]Des‐Gly10‐(D‐Ala6)‐gonadotropin‐releasing hormone‐ethylamide (GnRHa) to dispersed pituitary cells were evaluated in a 4‐day culture. Cells were sampled either from intact or from castrated male rats and binding was measured at various temperatures before or after treatment with phorbol‐12‐myristate‐13‐acetate (PMA) or 1‐5‐(isoquinolinyl‐sulfoxyl) 2‐methylpiperazine (H7), which activate or inhibit, respectively, protein kinase C (PKC). In cells from intact rats incubated with increasing concentrations of ligand at 21 °C for 25 min, the Scatchard plot was not linear and calculation of the Hill coefficient (NH) was indicative of positive cooperativity (NH= 1.26 ± 0.02). Such non‐linearity was not observed when cells were incubated at 0.5 °C for 3 h. In that condition the maximal number of binding sites measured at equilibrium (Bmax) increased (15.1 ± 0.05 versus 9.3 ± 0.5 fmoles × mg−1 proteins at 21 °C). Two control experiments permitted us to rule out the possibility that lower Bmax at 21 °C might reflect internalization: 1) Cells were first incubated with the ligand at 21 °C for 25 min and subsequently for 3 additional hours at 0.5 °C. Preincubation did not affect the Bmax obtained at 0.5 °C; 2) when the radioligand bound to the cell surface was washed out with an acidic buffer, only 13% of the specific radioactivity was retained irrespective of the ligand concentration applied, a much lower value than the 40% binding difference observed between 0.5°C and 21°C. When the cells were incubated with PMA, the Scatchard plot was linearized and the Bmax recorded at 21 °C increased by 50% over control cells (13 ± 0.7 fmoles × mg−1 proteins). Conversely, inhibition of PKC by H7, a preferential PKC inhibitor, was ineffective. In contrast, cells sampled from castrates exhibited linear and comparable Scatchard plots at either 0.5 ° or 21 °C, with Bmax values of 14.4 ± 0.3 and 15 ± 0.34 fmoles × mg−1 proteins, respectively. PKC activation did not affect binding in that model, but H7 decreased the number of sites (Bmax= 10.7 ± 0.9) and induced appearance of positive cooperativity (NH= 1.36 ± 0.07). Taken together, these experiments reveal a pool of GnRH receptors in the pituitary of intact rats that recognizes the ligand in a phosphorylation‐dependent manner. These binding sites also became evident when biological properties of the membrane were modified by temperature or cell homogenization. After castration, PKC activation was no longer a prerequisite for recruitment of the total population of receptors whereas protein kinase inhibition resulted in a reduction of maximal binding. Finally, our observations demonstrate that GnRH binding can exhibit positive cooperativity, either on normal cells or on cells from castrates after protein kinase inhibition, suggesting that such a cooperativity is not related to a GnRH‐dependent phosphorylation.
Effects of Chronic Treatment of Rats with Dopamine Receptor Drugs on the Post‐Translational Processing of Beta‐Endorphin in the Neurointermediate Lobe of the Pituitary Gland
Authors: Sweep C. G. J. (Fred) , Wiegant Victor M.
Published: September 2006

To investigate whether chronic changes in the activity of proopiomelanocortin cells in the neurointermediate lobe (NIL) of the pituitary gland are associated with changes in the enzymatic processing of β‐endorphin (βE), the effects of treatment of rats with the dopamine receptor antagonist haloperidol or the dopamine receptor agonist bromocriptine (2.5 mg.kg−1 sc, once daily for 21 days) were studied on the content of βE‐related peptides in the NIL and on the release of these peptides from NILs in an in vitro superfusion system. Treatment with haloperidol increased, and with bromocriptine decreased the tissue content and the release of Nα‐acetyl‐, β‐, γ‐ and α‐endorphin‐immunoreactivity (AcE‐, βE‐, γE, and αE‐IR). The endorphin‐IR was further characterized using reversed‐phase high‐performance liquid chromatography and specific radioimmunoassay systems, and the following peptides were identified: des‐tyrosine α‐endorphin (DTαE), αE, AcαE, γE, AcγE, βE‐(1–31), AcβE‐(1–31), AcβE‐(1–27), AcβE‐(1–26) and βE‐(1–26)/βE‐(1–27) (the latter peptides were not separated with the high‐performance liquid chromatography system used). Analysis of NIL superfusates indicated that all peptides found in the tissue were released in vitro. In addition, an as yet unidentified acetylated IR‐endorphin component was found which was not observed in extracts of NIL tissue, and therefore was probably formed during release. Following haloperidol treatment, the levels of all βE‐related peptides detected were increased in the tissues as well as superfusates, the increase in AcβE‐(1–27) being most and that in βE‐(1–26)/βE‐(1–27) least pronounced. Following bromocriptine treatment, the concentrations of all peptides in tissues and superfusates were decreased as compared to vehicle controls. The acetylated endorphins, in particular AcβE‐(1–27), were most affected and βE‐(1–26)/βE‐(1–27) least affected. The results indicate that chronic modulation of the synthesizing and secretory activity of proopiomelanocortin cells in the NIL is parallelled by changes in the enzymatic processing of βE.
The Characteristics of Medial Septal Neurons Antidromically Identified as Projecting to the Median Eminence and their Response to Gonadal Steroids
Authors: Donevan Sean D. , Ferguson Alastair V.
Published: September 2006

We have obtained extracellular single unit recordings from 129 neurons in the medial septum of the male rat that projected to the median eminence. These cells were identified on the basis of their antidromic responses to electrical stimulation in the median eminence. The antidromically‐evoked spike was always followed by a period of inhibition (−100 ms) which often (80%) persisted at stimulus intensities below threshold for antidromic activation. The identified neurons displayed one of three patterns of activity, the relative proportions of which were dependent upon the anesthetic. In rats anesthetized with urethane, 15% of the antidromically identified septal neurons displayed an oscillating, 25% a continuous, and 61% a low irregular pattern of spontaneous activity or no activity. In contrast, in barbiturate‐anesthetized animals only 6% exhibited an oscillating and 3% a continuous pattern of activity. The remaining neurons (91%) displayed either a low firing rate or were silent. We then assessed the responses of these identified neurons to systemic administration of gonadal steroids. Systemic administration of 17β‐estradiol elicited a rapid decrease in the firing rate of 9 of 29 neurons tested. The inhibitory response was specific to those cells displaying an oscillating (6/9) or continuous (3/6) firing pattern of which 60% were inhibited. In contrast, systemic administration of testosterone failed to elicit such changes in the activity of the identified septal neurons (n = 12) including one neuron which was subsequently shown to be inhibited by estradiol. As such identified septal neurons may contain gonadotropin‐releasing hormone, these results suggest a role for estradiol but not testosterone in the short‐term regulation of the secretion of this releasing hormone in the male rat.
Gonadotropin‐Releasing Hormone and Thyrotropin‐Releasing Hormone Regulation of G Protein Function in the Rat Anterior Pituitary Lobe
Authors: Ravindra R. , Aronstam R. S.
Published: September 2006

In order to evaluate the role of guanine nucleotide‐dependent transducer proteins (G proteins) in hormone‐mediated signal transduction in the anterior pituitary lobe, we examined the effect of gonadotropin‐releasing hormone (GnRH) and thyrotropin‐releasing hormone (TRH) on two parameters of G protein function, namely [35 S]GTPγS binding and low KmGTPase activity. Plasma membranes were prepared from anterior pituitary lobes of adult male rats using conventional procedures. GTP binding was determined by incubating 2 to 5 μg membrane protein with approximately 100,000 cpm [35 S]GTPγS in a buffer containing 20 mM Tris‐ HCl, 1 mM EDTA, 1 mM dithiothreitol, and 100 mM NaCI at a pH of 7.4 for 10 or 15 min at 37 °C GnRH agonist and TRH stimulated high affinity [35 S]GTPγS binding in a concentration‐dependent manner. GTP binding was maximally stimulated by GnRH agonist (1 μM) and TRH (0.1 μM) by up to 27% and 34%, respectively. A time‐course study revealed that 1 μM GnRH agonist stimulated GTP binding by 30% at 15 min; 0.1 μM TRH stimulated GTP binding by 23% at 1 min, 18% at 5 min and 25% at 10 min. A stable GTP analog, 5′‐guanylylimidodiphosphate, inhibited GnRH‐ as well as TRH‐stimulated GTP binding. GnRH antagonist did not affect GTP binding. However, in the presence of the antagonist, stimulation of GTP binding by the GnRH agonist was completely blocked.

The low KmGTPase activity (EC 3.6.1.‐), another parameter of G protein function, was assayed in 2 to 5 μg membrane protein using [γ‐32 P]GTP at 37 °C in an ATP‐regenerating buffer containing 1 μM unlabeled GTP. GnRH agonist (0.1 μM) and TRH (1 μM) maximally stimulated this GTPase activity by up to 50% and 40%, respectively. GnRH agonist (1 μM) stimulated the GTPase activity by 30% at 10 min and 48% at 30 min. TRH (1 μM) stimulated the GTPase activity at all time points monitored; stimulation was 46% at 5 min, 49% at 20 min, and 41% at 30 min. Interestingly, the GnRH antagonist stimulated GTPase activity by about 20%, but inhibited GnRH agonist‐stimulated GTPase activity in a concentration‐dependent manner. These results indicate that the binding of GnRH and TRH to their receptors results in interaction of the receptor with a G protein and activation of the G protein cycle.