bluuebunny/crossref_metadata_2025 huggingface dataset:
Source of Stress‐Induced Increase in Plasma Met‐Enkephalin in Rats: Contribution of Adrenal Medulla and/or Sympathetic Nerves*
Authors: Barron B. A. , Pierzchala K. , Van Loon G. R.
Published: September 2006

The contribution of the adrenal medulla and/or the sympathetic nerves to the plasma levels of Met‐enkephalin was investigated. Rats were divided into four groups: sham‐operated/saline, sham‐operated/guanethidine, adrenal‐demedullated/saline, demedulla‐ted/guanethidine. After 4 weeks of injection with either saline or guanethidine (25 mg/kg/day), animals were cannulated in the left carotid artery for blood sampling. Three days later, blood samples were taken before and at 2 and 30 min of restraint stress. Adrenal demedullation lowered basal plasma epinephrine levels markedly and prevented entirely the increase induced by restraint stress. Chronic guanethidine treatment lowered basal plasma norepinephrine levels and decreased the response to stress. Guanethidine treatment increased the basal plasma epinephrine level without affecting the response to stress. The combination of guanethidine plus adrenal demedullation lowered basal plasma concentrations of all three catecholamines and further attenuated the norepinephrine response. Restraint stress increased plasma native and peptidase‐derivable Met‐enkephalin. Adrenal demedullation, resulting in greater than 95% depletion of adrenal catecholamines and significant depletion of adrenal Met‐enkephalin, did not inhibit the stress‐induced increase in plasma Met‐enkephalin, and in fact, was associated with a potentiated response to stress. Guanethidine treatment with or without demedullation increased baseline plasma native Met‐enkephalin and abolished the stress‐induced increase in plasma native and peptidase‐derivable Met‐enkephalin. Thus, the stress‐induced increase in plasma Metenkephalin results from release from sympathetic nerves, rather than adrenal medulla. However, the sympathetic nerves and adrenal medulla together do not appear to account entirely for basal concentrations of circulating Met‐enkephalin. Hepatic portal vein plasma concentration of native Met‐enkephalin was greater than that in the carotid artery, suggesting contribution from the gastrointestinal tract; however, evisceration did not decrease plasma native Met‐enkephalin. Some compensatory mechanism results in elevation of basal plasma native Met‐enkephalin in sympathectomized rats. Also, in the absence of the adrenal medulla there is a compensatory increase in the amount of Met‐enkephalin released into the circulation in response to stress.
Kappa‐Selective Opioid Receptor Agonists Leumorphin and Dynorphin Inhibit Supraoptic Neurons in Rat Hypothalamic Slice Preparations
Authors: Inenaga K. , Imura H. , Yanaihara N. , Yamashita H.
Published: September 2006

To clarify the effects of opioid peptides, and in particular the effects of kappa‐receptor agonists on the activity of supraoptic neurons, extracellular recordings were made from 71 spontaneously firing neurons in the rat hypothalamic slice preparation. Of 71 neurons, 28 showed a phasic firing pattern (phasic neurons: putative vasopressin neurons). The mean firing rate of phasic neurons was 2.6 spikes/s (intraburst firing rate 5.4 ± 2.2 spikes/s). The mean firing rate of neurons classified as non‐phasic neurons (putative oxytocin neurons) was 4.5 spikes/s. Following bath application of leumorphin (LM) at 10−7 M, which has potent opioid activity at kappa‐receptors, 17 (61%) of 28 phasic neurons were inhibited and 22 (51%) of 43 non‐phasic neurons were inhibited. Excitation was observed in only one non‐phasic neuron. The dose‐dependence of the response to LM was tested in five supraoptic neurons. There was an inverse relationship between LM concentration and percent change in firing rate. The threshold concentration of LM was approximately 10−8 M. The relatively selective kappa‐receptor antagonist, MR‐2266, completely blocked the LM‐induced responses. Its effects were long‐lasting and only partial recovery was observed 2 h after the application of MR‐2266. Dynorphin had similar inhibitory effects on supraoptic neurons to those obtained with LM when tested on the same neurons. In another series of experiments the mu‐receptor agonist morphine and the delta‐receptor agonist [D‐Ala, D‐leu]‐enkephalin (DADLE) were applied to 28 supraoptic neurons (12 phasic and 16 non‐phasic neurons) at 10−7 M and their actions compared directly with that of LM. Only two of 12 phasic neurons tested were inhibited by DADLE and none of five phasic neurons tested was inhibited by morphine, while eight of the 12 neurons tested were inhibited by LM. By contrast the non‐phasic neurons tested were inhibited by application of each of the peptides; seven of 16 neurons tested were not only inhibited by LM, but also five of 11 neurons by DADLE and seven of 15 by morphine. The magnitude of the responses varied from cell to cell. These results suggest that LM acts at the same receptors as dynorphin, and that opioids acting preferentially at kappa‐receptors inhibit both vasopressin and oxytocin neurons while delta‐ and mu‐receptor agonists inhibit primarily oxytocin neurons.
Pyridostigmine, an Acetylcholinesterase Inhibitor, Stimulates Growth Hormone Release, but has no Effect on Basal Thyrotrophin or Adrenocorticotrophin Levels, or the Thyrotrophin Response to Thyrotrophin‐Releasing Hormone
Authors: Freeman Emma , Touzel Rick , Grossman Ashley , Besser Michael , Ross Richard
Published: September 2006

Pyridostigmine, an acetylcholinesterase inhibitor, stimulates growth hormone (GH) release and is thought to act by inhibiting hypothalamic somatostatin release. There are few data concerning the effect of pyridostigmine on other pituitary hormones apart from GH. We have studied the effect of pyridostigmine on basal GH, thyrotrophin (TSH), prolactin, adrenocorticotrophin and cortisol release, and thyrotrophin‐releasing hormone (TRH)‐stimulated TSH and prolactin release, in two studies involving nine healthy male subjects. Pyridostigmine stimulated GH release in all subjects but had no effect on adrenocortocotrophin or cortisol levels, or basal or TRH‐stimulated TSH and prolactin levels.

There are some data to suggest that somatostatin inhibits TRH‐stimulated TSH release. Our findings, however, suggest that either endogenous somatostatin tone has little effect on the TSH response to TRH compared to its effects on GH or pyridostigmine acts through a mechanism other than altering somatostatin tone. Pyridostigmine did not alter adrenocorticotrophin or cortisol levels in the presence of a clear action on GH release, providing further evidence that the previously reported effects of cholinergic drugs on cortisol release are stress‐related.
Ovarian Steroid Regulation of Glutamic Acid Decarboxylase Gene Expression in Individual Hypothalamic Nuclei
Authors: Leigh A J. , Carter N.D. , Horton R. , Silverligh J.J. , Wilson C.A.
Published: September 2006

The possibility that steroids exert their effects on luteinizing hormone (LH) and prolactin release and female sexual behaviour via altering γ‐aminobutyric acid (GABA) synthesis within the hypothalamus was investigated. Ovariectomized rats were treated with either oil, 5 μg oestradiol benzoate (OB) or 5 μg OB plus 0.5 mg progesterone (P) 48 h later; autopsy was carried out 54 h after the OB injection. At this time, both steroid treatments stimulated prolactin release and lordotic activity. OB alone exerted a negative feedback effect on LH release while OB plus P stimulated an LH surge.

The brains of the rats in these three treatment groups were microdissected and glutamic acid decarboxylase messenger nbonucleic acid (GAD mRNA) was estimated in specific hypothalamic areas obtained from individual brains; these areas included the preoptic area (POA), ventromedial nucleus (VMN), anterior medial portion of the zona incerta (Zl) and the arcuatemedian eminence area (ARC/ME). GAD mRNA concentrations were assessed by the slot‐blotting technique and expressed as a ratio of GAD mRNA: β‐actin mRNA.

Both steroid treatments significantly reduced GAD mRNA in the ARC/ME and Zl and OB plus P also reduced the concentration in the POA, while OB alone had no effect in this area. Neither treatment affected GAD mRNA in the VMN. These results indicate that when steroids stimulate LH and prolactin release and female sexual behaviour, they reduce GABA activity probably by reducing GABA synthesis in the POA, Zl and ARC/ME. This suggests that GABA normally has an inhibitory influence on these parameters. GABA synthesis does not however, appear to be involved in the negative feedback effects of steroids on LH release as measured 54 h after OB treatment.
Release of Vasopressin from the Septum and Hippocampus of Freely Behaving Male Rats: Effect of Bacitracin and Synthetic Vasopressin
Authors: Ramirez V. D. , Ramirez A. D. , Rodriguez F. , Poncett C. , Vincent J. D.
Published: September 2006

This paper describes the release characteristics of arginine vasopressin (AVP) from the dorsal septum and dorsal hippocampus of freely behaving male rats using miniaturized push‐pull cannulae and a slow rate of perfusion to minimize tissue damage. The major findings were that) The spontaneous release of immunoreactive AVP (l‐AVP) from the dorsal hippocampus was episodic in nature and did not require the presence of bacitracin in the perfusion medium to reliably demonstrate its pattern of release; 2) despite the fact that the septum has close to seven times more I‐AVP per/mg of tissue, the dorsal septum perfusates had undetectable levels of the neuropeptide; 3) under these conditions hypertonic saline intraperitoneally did not alter the spontaneous release of this peptide; and 4) only when the perfusion medium contained effective concentrations of bacitracin (10−4 M) were reliable and detectable levels of I‐AVP measured in perfusates from the dorsal septum and exogenous synthetic AVP had a robust positive feedback action on its own release. The physiological relevance and importance of this robust and novel effect of AVP remains to be elucidated.
Effects of Alpha‐ and Beta‐Adrenergic Agonists and Antagonists on Growth Hormone Secretion in Man
Authors: Ghigo E. , Bellone J. , Arvat E. , Mazza E. , Cellat S. G. , Brambilla F. , Müller E. E. , Camanni F.
Published: September 2006

It is well known that the adrenergic system has both stimulatory and inhibitory influences on growth hormone (GH) secretion probably by modulating GH‐releasing hormone (GHRH) and/or somatostatin release. To better understand the mechanisms by which these influences take place, we investigated the effects of α‐ and β‐adrenergic agonists and antagonists on both basal and GHRH‐induced GH release in 23 male adult volunteers. The GH‐releasing effect of clonidine (0.15 mg infused iv over 10 min), an α2‐adrenergic agonist, was significantly blunted by yohimbine (30 mg orally at −50 min), a relatively specific α2‐adrenergic antagonist area under the response curve, mean±SEM: 672.6 ± 143.0 versus 219.6 ± 16.7 μg/l/h; P<0.05). On the other hand, the GHRH (1 μg/kg iv as a bolus)‐induced GH increase was unaffected by yohimbine (339.3 ± 19.1 versus 518.1±172.8 μg/I/h). Concomitant blockade of α1‐/α2‐adrenoreceptors by phentolamine (0.5 mg/ml/min infused iv from −60 to +30 min) abolished the GHRH‐induced GH rise (645.5± 106.0 versus 189.0±58.8 μg/l/h; P<0.01). Finally, the GHRH‐stimulated release was blunted by β2‐adrenergic stimulation with salbutamol (10 μg/min infused iv from −5 to +15 min) (324.3 ± 99.7 versus 112.7 ± 48.8 μg/l/h; P<0.02).

In conclusion: 1) The evidence that yohimbine is able to blunt the clonidine‐induced GH release but fails to inhibit the GHRH‐induced GH rise indicates that, as in animals, in man too the GH‐releasing effect of clonidine is specifically mediated by α2‐receptor activation, and may occur via endogenous GHRH release; 2) the inhibitory effect on GH release of β, namely β2, receptor activation is probably mediated by the somatostatinergic system; 3) an unopposed β‐adrenergic activation would account for the inhibitory effect on GHRH‐induced GH release of concomitant α1–/α2‐adrenoreceptor blockade by phentolamine.
The Possible Role of Androgenic Hormones in Maturation of Alpha2‐Adrenergic Mechanisms in the Rat
Authors: Kiem D. T. , Bartha L. , Makara G. B.
Published: September 2006

In order to get better characterization of androgenic hormones on the functionality of the α2‐adrenergic system and widen our previous studies, we investigated the effect of clonidine on the growth hormone (GH) secretion in male Wistar rats of various ages: 7‐, 14‐ and 30‐day old and adult, adult male castrated rats with and without testosterone treatment.

Two different patterns of the GH response to clonidine have been observed in the control and testosterone‐treated young animals: clonidine at the dose 15 μg/kg intraperitoneally had no effect on the GH secretion in 7‐ to 14‐day old rat pups. In contrast, its effect appeared following the increase in the plasma testosterone concentration induced by pretreatment with testosterone (5 mg/kg subcutaneously for 4 days) in these animals. In 30‐day old rats clonidine affected GH secretion and this influence was more pronounced in the testosterone‐treated animals than in the controls.

The decrease in the circulating testosterone levels caused by castration in adult male rats caused a decreased GH response to clonidine. Moreover, there was a tendency for the GH response to return in 4‐week old animals. The effect of clonidine has been restored by testosterone replacement of castrates.

Testosterone administration decreased basal plasma GH levels in the pups. However, it triggered the ultradian surges of GH secretion which were absent in the young animals.

Clonidine had no effect on the corticosterone secretion in 7‐day old animals. Testosterone treatment induced a response in the 7‐day old rat and markedly potentiated its effect on the secretion of this hormone in 14‐ and 30‐day old animals, respectively.

Neither progesterone nor hydrocortisone influenced the GH‐releasing effect of clonidine. Hydrocortisone markedly inhibited the basal‐ and clonidine‐induced corticosterone secretion.

The results of the present study indicate an important role of androgenic hormones in inducing and/or maintaining the effectiveness/sensitivity of the α2‐adrenergic receptor system and suggest a possible role for the androgenic hormones in the maturation of α‐adrenergic mechanisms in the rat.
Regional Changes in Brain Oxytocin Receptors Post‐Partum: Time‐Course and Relationship to Maternal Behaviour
Authors: Insel Thomas R.
Published: September 2006

In a previous report, receptors for oxytocin in rat brain were reported to increase during the early post‐partum period. The current study set out to replicate this finding using the novel, highly selective oxytocin receptor ligand, [125 l]d(CH2)5[Tyr(Me)2, Thr4, Tyr‐NH29]OVT ([125 I]‐OTA). Binding was measured using in vitro receptor autoradiography in rat brain on Day 15 of pregnancy, on Days 1 and 6 post‐partum, and at least 6 days following the end of lactation. Relative to pregnancy, oxytocin receptor binding was increased on Day 1 (but not Day 6) post‐partum in the bed nucleus of the stria terminalis and the ventromedial nucleus of the hypothalamus. In several other regions, including the anterior olfactory nucleus, the central nucleus of the amygdala, and the ventral subiculum binding was equivalent across the groups. Most surprising, binding in the bed nucleus of the stria terminalis was lowest in the post‐lactating group (post‐partum Day 1 group was 87% higher than post‐lactating group). Saturation studies suggested that binding differences reflected changes in number and not affinity of sites. These findings are consistent with earlier studies of changes in brain oxytocin receptors following exogenous gonadal steroid administration. To determine if maternal behaviour elicited by nonhormonal means (i.e. concaveation) was also associated with increased oxytocin receptors, virgin females with extensive pup exposure were studied. No change in oxytocin receptor binding in the bed nucleus of the stria terminalis was noted in virgin, maternal females—demonstrating that the increase observed post‐partum was not essential for the onset of maternal behaviour.
Effects of Somatostatin on the Growth Hormone‐Releasing Factor‐Induced Growth Hormone Secretion in Rats with Electrical and Chemical Inhibitions of Endogenous Growth Hormone‐Releasing Factor and Somatostatin
Authors: Sato Makoto , Takahara Jiro , Niimi Michio , Irino Shozo
Published: September 2006

The episodic pattern of growth hormone (GH) secretion of the male rat was simulated in rats exhibiting impaired GH‐releasing factor (GRF) and Somatostatin (SRIF) secretion, by administering various combinations of human GRF‐(1–44)NH2 (hGRF) and SRIF. Electrical lesions of the arcuate nucleus resulted in a marked decrease in the amplitude of GH secretory bursts, while the administration of cysteamine (200 mg/kg) did not change the GH secretory profile in rats with arcuate nucleus lesions. Immunohistochemical examinations revealed a marked decrease of GRF and SRIF immunoreactivity in the median eminence of the cysteamine‐treated rats with arcuate nucleus lesions. The intravenous injection of 5 μg of hGRF every 3 h caused equivalent surges of GH in the cysteamine‐treated rats with arcuate nucleus lesions. The additional infusion of 4 μg/h of SRIF during the trough periods of GH secretion did not affect the amplitude of the GH surges. Hourly injection of 5 μg of hGRF caused transient desensitization to hGRF. Interestingly, the additional infusion of 4 μg/h of SRIF every 3 h enhanced the amplitude of the GH bursts induced by the fourth and the seventh hGRF injections. However, the more frequent injection of 5 μg of hGRF every 30 min caused constant desensitization to hGRF with time, and the additional infusion of 4 μg/h of SRIF every 3 h did not change the attenuated responses to hGRF. These results indicated that the simultaneous administration of hourly GRF and continuous SRIF with brief pauses was most effective for producing high GH peaks. This simulation model suggests that SRIF may play an important role not only in the production of GH troughs, but also in the maintenance of GH surges with distinct peaks in the male rat.
Evidence for Independently Regulated Secretory Pathways of Proopiomelano‐cortin‐Related Peptides in the Mouse Pars Intermedia
Authors: Leenders H. J. , Jenks B. G. , Rêlo A. L. , Roubos E. W.
Published: September 2006

The existence of multiple secretory pathways within endocrine cells has been receiving increasing attention (1–5). In some cases there is evidence for independently regulated pathways, for example in prolactin‐producing cells (6) and parathyroid hormone‐producing cells (7, 8). These pathways concern newly synthesized hormone versus hormone sequestered in mature secretory compartments. In the present study, we have analysed secretion of newly synthesized and mature peptides from proopiomelanocortin (POMC)‐producing cells of the mouse pars intermedia. Evidence has been found for independent regulation and we show that the peptide composition of the two secretory pathways differs with respect to some of the POMC‐derived peptides.
Pituitary Hormone Gene Expression in Male Golden Hamsters: Interactions Between Photoperiod and Testosterone
Authors: Hegarty Catherine M. , Jonassen Julie A. , Bittman Eric L.
Published: September 2006

Daylength regulates neuroendocrine function in male golden hamsters. Exposure to short days triggers gonadal regression and decreases serum luteinizing hormone (LH), prolactin and testosterone concentrations. Inhibitory photoperiods also amplify the negative feedback actions of androgens upon gonadotropin secretion. To examine whether these changes arise from altered adenohypophyseal gene expression, we measured the abundance of the messenger ribonucleic acids (mRNAs) encoding β‐LH, prolactin and proopiomelanocortin in anterior pituitaries of male golden hamsters which were either left intact, castrated, castrated and implanted with testosterone, or pinealectomized and maintained in either long (14 h light/10 h dark) or short (5 h light/19 h dark) days. Short days caused testicular atrophy in intact male hamsters and reduced serum LH in intact and castrated, testosterone‐replaced hamsters. The relative abundance of β‐LH mRNA was suppressed by exposure to short days only in castrated hamsters. Serum prolactin was decreased by short days regardless of circulating testosterone concentrations. Prolactin mRNA abundance was decreased by short days in all pineal‐intact groups. Castration reduced proopiomelanocortin mRNA abundance in long days and testosterone replacement reversed this effect. In the presence of testosterone, photoperiod influenced serum LH concentrations without altering hypophyseal abundance of β‐LH mRNA. In contrast, photoperiodic influences on prolactin secretion were correlated with alterations in steady‐state mRNA abundance.
Ontogenesis and Sexual Dimorphism of Rat Growth Hormone Messenger Ribonucleic Acid as Studied by in situ Hybridization
Authors: Martinoli M. G. , Pelletier G.
Published: September 2006

We have investigated the ontogeny of growth hormone (GH) mRNA in the developing rat foetus and also from birth to adulthood. Using quantitative in situ hybridization, we studied the variations in the levels of GH mRNA during foetal and postnatal life in the pituitary of both male and female rats. A cDNA probe to GH mRNA was used to detect GH transcripts on fixed pituitary sections at different stages of development. Few labelled cells were observed in the lateral wings of the anterior pituitary from the 17th to 19th day of gestation in both sexes. The amounts of GH mRNA significantly increased in both male and female rats from neonatal to adult life, reaching the highest levels after puberty. A clear sexual dimorphism was observed at the 20th day of foetal life, GH mRNA levels being higher in male than in female rats. After birth, no significant differences of GH mRNA levels could be observed between male and female rats until 30 days of age (prepubertal period) when male rats exhibited GH mRNA levels higher than females. This sexual difference in GH mRNA levels remained constant throughout adult life. Moreover, gonadectomy performed at neonatal, prepubertal and adult periods in both male and female rats did not modify GH mRNA levels in either sex. These results indicate that an early synthesis of GH mRNA occurs in foetal pituitary and that the sexual dimorphism of GH mRNA observed from 30 days of age is not related to sex steroids.
Oxytocin Messenger Ribonucleic Acid Levels in the Medial Preoptic Area are Increased During Lactation
Authors: Brooks Philip J. , Lund P. Kay , Stumpf Walter E. , Pedersen Cort A.
Published: September 2006

The peptide hormone oxytocin has an important role in parturition, lactation and maternal behavior. The present study employed in situ hybridization histochemistry to determine whether oxytocin messenger ribonucleic acid (mRNA) levels in cells in the medial preoptic area, a brain area known to control maternal behavior, change during pregnancy and lactation in the rat. Female rats were perfused on either Day 18 or 22 of pregnancy or Day 5 of lactation. Ovariectomized female rats were included as an additional control group. Cells expressing oxytocin mRNA were detected by in situ hybridization using an [125 l]‐labeled 38 base synthetic oligodeoxynucleotide probe complementary to the C‐terminal coding region of the preprooxytocin. Relative differences in oxytocin mRNA levels were determined by silver grain counting of labeled cells. A group of oxytocin neurons in the dorsal medial preoptic area, called the lateral subcommissural nucleus, showed elevated oxytocin mRNA levels in lactating animals relative to ail other groups. Oxytocin mRNA levels in the neurons of the periventricular nucleus of the preoptic area did not change across pregnancy and lactation. This result extends the findings of others showing elevated oxytocin mRNA levels in magnocellular nuclei of lactating animals. The results are discussed in terms of the possible role of oxytocin cells in the medial preoptic area in the expression of maternal behavior.
N‐Methyl‐DL‐Aspartate Receptor Antagonism by D‐2‐Amino‐5‐Phosphonovaleric Acid Delays Onset of Puberty in the Female Rat
Authors: Wu F. C. W. , Howe D. C. , Naylor A. M.
Published: September 2006

To investigate the possible role of N‐methyl‐DL‐aspartate (NMDA) receptor activation in the initiation of puberty, we examined the effects of the selective competitive antagonist 2‐amino‐5‐phosphonovaleric acid (AP5) on the timing of vaginal opening. Paired and weight‐matched litter mates of immature female rats were implanted with osmotic minipumps for the intracerebroventricular infusion of DL‐ or D‐AP5 or artificial cerebrospinal fluid from 27 to 30 days of age for 14 days. Each animal was weighed and examined daily for vaginal opening as the indicator of first oestrous. Infusion of 20 or 40 mM DL‐AP5 beginning on Day 30 failed to delay vaginal opening. Administration 50mM of the single enantiomer D‐AP5 beginning on Day 27 significantly delayed the age of vaginal opening to 40.6±1.1 (mean ± SEM) days compared to the cerebrospinal fluid‐infused controls (36.5 ± 0.6 days). Blockade of NMDA receptors in the D‐AP5‐treated animals was confirmed on Day 32 by the suppression of luteinizing hormone response to intravenous NMDA (20 mg/kg) while the response to exogenous luteinizing hormone‐releasing hormone (50 ng/kg) remained intact. AP5‐treated animals had a slower rate of growth (3.1 ± 0.2 g/day) compared to controls (4.2 ± 0.2 g/day). However, a similar degree of growth retardation produced by a 75% restricted diet in untreated juvenile animals did not delay vaginal opening. This suggests that the slower growth rate in the D‐AP5‐treated animals could not account for the delayed onset of puberty. In conclusion, these data suggest that blockade of central NMDA receptors inhibits excitatory mechanisms which may be important in the control of pubertal onset in the female rat.
Regulation of the Rat Oxytocin Gene by Estradiol
Authors: Peter J. , Burbach H. , Adan Roger A. H. , van Tol Hubert H. M. , Verbeeck Marleen A. E. , Axelson John F. , van Leeuwen Fred W. , Beekman Johanna M. , Ab Geert
Published: September 2006

Oxytocin (OT) plays a role in reproduction at the level of the pituitary and mammary glands and uterus. This OT is synthesized in the hypothalamo‐neurohypophyseal system (HNS). A number of observations have suggested that estrogens regulate the production of OT in the HNS. In this study the effect of 17β‐estradiol on the activity of the OT gene promoter was examined as well as the effect of 17β‐estradiol in vivo on OT messenger ribonucleic acid (mRNA) and peptide revels in the rat HNS. Vasopressin (VP) and its mRNA were also determined in the in vivo studies. The direct transcriptional stimulation of OT gene expression by 17β‐estradiol was studied in two different heterologous expression systems. When a plasmid having nucleotides −363 to +16 of the rat OT gene fused to the firefly luciferase reporter gene was co‐transfected with an estrogen receptor expression vector in P19 embryonal carcinoma cells, luciferase activity was stimulated 80‐fold by 17β‐estradiol. In estrogen receptor containing MCF‐7 cells transfected with a plasmid having nucleotides −188 to +16 of the rat OT gene fused to the chloramphenicol acetyl transferase gene, 17β‐estradiol induced the expression of the chloramphenicol acetyl transferase gene through the cloned promoter element. After in vivo treatment of ovariectomized rats with 17β‐estradiol, levels of OT mRNA and VP mRNA were measured in microdissected supraoptic and paraventricular nuclei as well as VP and OT levels in these nuclei and the pituitary gland. As compared to non‐treated ovariectomized rats there was no difference in contents of OT mRNA and VP mRNA in these hypothalamic nuclei and in levels of the peptides in paraventricular nuclei and the pituitary gland. A 30% reduction of the OT content of the supraoptic nuclei only was found, while the VP content did not change. To explain the results immunocytochemical analyses of the hypothalamus were performed, showing that the estrogen receptor was absent in the magnocellular neurons of the supraoptic and paraventricular nuclei. The results demonstrate that the 5’flanking region of the OT gene confers estrogen‐sensitivity to transcription of the OT gene. This potential to respond to estrogens is not used in the OT‐producing neurons of supraoptic and paraventricular nuclei probably due to the absence of the estrogen receptor.
Immunocytochemical Localization of Glucagon‐Related Peptides in the Cerebral Ganglion of the Protochordate Ascidian, Styela plicata
Authors: Pestarino Mario
Published: September 2006

The central nervous system of the protochordate ascidian Styela plicata has been investigated by means of immunocytochemical techniques in order to localize peptides of the glucagon/secretin family. In particular, glucagon‐ and peptide histidine isoleucine‐containing neurons were localized in the cortex of the cerebral ganglion, and numerous nerve cell bodies containing vasoactive intestinal polypeptide occurred mainly in the medullary zone. Moreover, coexistence of glucagon/peptide histidine isoleucine and glucagon/vasoactive intestinal polypeptide was detected in a few cortical neurons. The localization of peptide histidine isoleucine‐like peptides in the central nervous system and alimentary tract supports the occurrence of a ‘brain‐gut axis’ in protochordates as well as in vertebrates and some invertebrates, while glucagon‐ and vasoactive intestinal polypeptide‐like substances can be considered exclusively as neuropeptides.
Stress‐Induced Oxytocin Release in the Rat After Lesion of the Paraventricular Nuclei; Possible Deficiency of Corticotrophin‐Releasing Factor
Authors: Higuchi Takashi , Honda Kazumasa , Takano Shin , Negoro Hideo
Published: September 2006

Oxytocin is released under stressful conditions and corticotrophin‐releasing factor (CRF) is known to be involved in mediating general ‘stress responses’. We therefore examined whether CRF neurons in the paraventricular nucleus participate in the stress‐induced oxytocin release in the rat. CRF (0.02 to 2 nmol) injected into the third ventricle produced a dose‐dependent rise in the plasma oxytocin concentration. The oxytocin release induced by CRF occurred without a change in blood pressure, and was not affected by dexamethasone pretreatment, which prevents adrenocorticotrophin release following CRF injection. Lesioning of the paraventricular nucleus reduced oxytocin release by immobilization stress, but did not alter the release of oxytocin in response to osmotic stimulation induced by intraperitoneal injection of hypertonic saline. Anti‐CRF serum injection into the third ventricle reduced delayed oxytocin response to immobilization stress. These results are consistent with the hypothesis that CRF neurons in the paraventricular nucleus are involved in the oxytocin release during immobilization stress in the rat.
Vasotocin Target Sites in the Capsular Region Surrounding the Nucleus Robustus Archistriatalis of the Canary Brain
Authors: Voorhuis T. A. M. , Elands J. P. M. , De Kloet E. R.
Published: September 2006

The aim of this study was to characterize neurohypophyseal hormone receptors in the brain of the canary (Serinus canarius) by using autoradiographical and biochemical procedures with a radioiodinated vasotocin analogue, [125 l]d(CH2)5[Tyr(Me)2, Thr4, Orn8, Tyr‐NH29]vasotocin ([125 I]‐OTA). This synthetic analogue was used previously to identify a population of oxytocin‐like receptors in the rat brain that have high affinity for vasotocin.

In vitro autoradiography revealed high affinity binding sites for [125 I]‐OTA in the canary brain in the archistriatum surrounding the nucleus robustus archistriatalis. Scatchard analysis of [125 I]‐OTA binding to a synaptic membrane fraction prepared from the archistriatal region including the nucleus robustus archistriatalis indicated the presence of a single population of binding sites (Kd= 0.05 nM; Bmax= 4 fmol/mg protein).

Displacement studies revealed that the order of potency of certain peptides to inhibit [125 I]‐OTA binding was as follows: vasotocin (Ki= 0.4 nM) > oxytocin = vasopressin > mesotocin (Ki= 8.0 nM). The administration of testosterone to female canaries did not affect [125 I]‐OTA labelling in the archistriatum detected by autoradiography and did not influence [125 I]‐OTA binding constants in the membrane binding assay. In conclusion, this study provides morphological and biochemical evidence of a vasotocin‐target site in the archistriatum in close association with the central vocal control circuit in the canary brain.
Non‐Phasic Discharge and Osmoresponsiveness of Cat Magnocellular Neuroendocrine Cells
Authors: Fagan M. , Andrew R. D.
Published: September 2006

Repetitive bursting (phasic firing) generated endogenously by magnocellular neuroendocrine cells (MNCs) in the rat facilitates systemic release of vasopressin from axon terminals in the neurophypophysis. However, little is known of how MNCs function in other mammals. Using coronal slices of hypothalamus we studied the firing behaviour and intrinsic membrane properties of homologous neurons in the cat supraoptic nucleus where vasopressinergic MNCs outnumber oxytocinergic cells. Less than 1% of units recorded in cat supraoptic nuclei (2 of 270) spontaneously fired in a phasic mode compared to 39% in the rat (90 of 230). A discrete level of steady current across the extracellular recording micropipette promoted phasic firing in 66 of 152 non‐phasic units tested in rat supraoptic nuclei, but no phasic activity in 189 units from the cat. One or several stimuli applied dorsal to supraoptic nuclei triggered a single burst (afterdischarge) in 115 of 180 MNC units from the rat, whereas none of 173 MNC units tested in the cat fired an afterdischarge. Intracellular recordings from 56 feline MNCs revealed that unlike the rat, spike depolarizing afterpotentials were absent in all cells. This explains both the absence of phasic firing and the inability to trigger regenerative bursts in the intact cat. The possible Osmoresponsiveness of cat MNCs was examined using unit recording. These units reversibly increased their firing rate as osmolality was elevated with mannitol or NaCl (10 to 100 mOsm/kg), comparable to rat units. However, in no case did hyperosmotic conditions elicit phasic firing. We conclude that cat MNCs lack a regenerative burst capability but that unit Osmoresponsiveness is comparable to rat MNC units. We hypothesize that since the kidney of the cat normally functions at high efficiency in terms of water resorption, there may be little need for the rapid and pronounced elevation in vasopressin release evoked by phasic firing.
Both Pertussis Toxin‐Sensitive and Insensitive G‐Proteins Link Melatonin Receptor to Inhibition of Adenylate Cyclase in the Ovine Pars Tuberalis
Authors: Morgan P. J. , Davidson G. , Lawson W. , Barrett P.
Published: September 2006

Bordetella pertussis toxin (islet activating protein, IAP) has been used to investigate the G‐proteins involved in mediating the action of the melatonin receptor. Melatonin inhibits iorskolin‐stimulated cyclic AMP production in ovine pars tuberalis (PT) cells. In cells treated with IAP for 16 h this response is attenuated in a dose‐dependent manner, but not abolished. IAP catalyses the incorporation of [32 P‐ADP]ribose into a 41 kd protein present in PT membranes, but this labelling can be reduced if PT cells are preincubated with IAP for 16 h. Treatment of crude membrane preparations with IAP (20 /ig/ml) suppresses the binding of 2‐[125 l]iodomelatonin by 20%, whereas 1 mM GTP alone reduces binding by 40%, and in combination with IAP its effect is additive (60% inhibition). Therefore, these results indicate that the melatonin receptor acts via two G‐proteins, one pertussis toxin‐sensitive and the other pertussis toxin‐insensitive.
Control of Phospholipid Turnover and Prolactin Release in a Dopamine‐Sensitive, Prolactin‐Secreting Rat Pituitary Adenoma and in Two Dopamine‐Resistant, Prolactin‐Secreting Rat Pituitary Tumors
Authors: Forget H. , Huot C. , Collu R.
Published: September 2006

The secretion of prolactin by the pituitary gland is under a tonic inhibitory control exerted by tubero‐infundibular dopamine. Recently, it has been suggested that dopamine may exert its action by inhibiting production of inositol phosphates and mobilization of intracellular Ca2+. To study the effects of dopamine on the production of inositol phosphates and prolactin release, we have utilized an estrone‐induced, dopamine‐sensitive rat pituitary adenoma and two transplantable and dopamine‐resistant rat pituitary tumors, 7315a and MtTW15. Purified cells, obtained from the three tissues, were incubated for 30 min in media with drugs (thyrotropin‐releasing hormone or angiotensin II) stimulating inositol phosphates and prolactin release, in the presence or the absence of dopamine. Basal production of inositol phosphates and prolactin release by adenomatous lactotrophs were inhibited by dopamine. Thyrotropin‐releasing hormone and angiotensin II stimulated inositol phosphates by adenomatous and 7315a cells. This effect was antagonized by dopamine in adenomatous cells. Prolactin release by adenomatous cells only was stimulated by thyrotropin‐releasing hormone and angiotensin II. This stimulation was inhibited by dopamine. The results show differences, in the mechanisms of regulation of prolactin release, between adenoma and transplantable pituitary tumors as well as between the two tumors themselves. These differences may be responsible, in part, for the resistance of the two transplantable pituitary tumors to the inhibitory effects of dopamine on prolactin release and tumor size. Our results obtained both with adenoma and tumoral cells also suggest that inositol phosphates probably intervene only in the late phases of dopamine inhibition of prolactin release and only in the presence of a normal Ca2+ signaling system.
Vasopressin Expression in Cultured Neurons is Stimulated by Cyclic AMP
Authors: Ceding Petra , Schilling Karl , Schmale Hartwig
Published: September 2006

Vasopressin and oxytocin genes are expressed in mutually exclusive sets of magnocellular neurons in the hypothalamus. Cell specificity and regulation are probably controlled by extra‐ and intracellular signals acting on one or the other gene. In order to identify factors that regulate peptide expression, we have used primary dissociated cultures derived from 14‐day old foetal rats. Vasopressin expression was monitored by combined immunocytochemistry and in situ hybridization. Treatment of cultures with forskolin and/or the phosphodiesterase inhibitor 3‐isobutyl‐1‐methylxanthine (IBMX), both of which result in elevated intracellular cyclic AMP levels, increased the numbers of vasopressin‐expressing cells up to 10‐fold. The specific Vasopressin messenger ribonucleic acid accumulation was verified quantitatively by ribonuclease protection assays. Forskolin and IBMX did not change the levels of the general neuronal markers, neuron‐specific enolase and synaptophysin, suggesting that the effect of these drugs was specific for vasopressin‐expressing cells. The drugs were not mitogenic for magnocellular neurons. Furthermore, their effect was not mediated trans‐synaptically, as the drugs were also effective in cultures grown in low Ca2+/high Mg2+ medium, as well as in cultures treated with either tetanus toxin or tetrodotoxin. The presence of putative response elements for the transcription factor AP‐2 in the 5’promoter regions of all vasopressin genes sequenced so far may provide the molecular basis of the observed cyclic AMP effect. No such elements are present in the genes for oxytocin, the messenger ribonucleic acid levels of which were not measurably affected by forskolin and IBMX in our cultures.
Central 5,7‐Dihydroxytryptamine Lesions Decrease Hippocampal Glucocorticoid and Mineralocorticoid Receptor Messenger Ribonucleic Acid Expression
Authors: Seckl Jonathan R. , Dickson Karen L. , Fink George
Published: September 2006

Corticosteroids exert effects on the hippocampus by binding to intracellular glucocorticoid and/or mineralocorticoid receptors, but the relative importance of each receptor type in mediating corticosteroid effects is poorly understood. There is an extensive serotoninergic (5‐HT) innervation of the hippocampus which interacts with corticosteroid‐sensitive cells. We have investigated the effect of intracerebroventricular 5,7‐dihydroxytryptamine lesions of 5‐HT neurons on glucocorticoid and mineralocorticoid receptor messenger ribonucleic acid (mRNA) expression in the rat hippocampus using in situ hybridization histochemistry. In controls, glucocorticoid receptor mRNA was highly expressed in dentate gyrus granule cell neurons, and in pyramidal cells of CA1 and CA2, but levels in CAS and CA4 were significantly lower. 5,7‐dihydroxytryptamine‐lesioned animals showed significantly less glucocorticoid receptor mRNA in the dentate gyrus (76% decrease), CA1 (42% decrease) and CA2 (52% decrease; all P<0.05 compared with controls). Mineralocorticoid receptor mRNA was expressed at a similar level in all hippocampal subregions in control rats. 5,7‐dihydroxytryptamine lesioning led to a significant decrease in mineralocorticoid receptor mRNA expression in CA3 (56% fall) and CA4 (45% fall; both P<0.05), but not in the other subregions. Thus the 5‐HT innervation regulates hippocampal corticosteroid receptor mRNA expression.
Preproneuropeptide Y Messenger Ribonucleic Acid in the Hypothalamic Arcuate Nucleus of the Rat is Increased by Food Deprivation or Dehydration
Authors: O'Shea Ross D. , Gundlach Andrew L.
Published: September 2006

The distribution of messenger ribonucleic acid (mRNA) encoding preproneuropeptide Y (prepro‐NPY) in the hypothalamus of rats subjected to food deprivation or dehydration has been investigated by quantitative in situ hybridization. Levels of prepro‐NPY mRNA in the arcuate nucleus (ARC) were selectively increased by both treatments. The very high concentration of prepro‐NPY mRNA seen following 96 h of food deprivation had returned towards control levels after 24 h of refeeding. Levels of preprogalanin (prepro‐GAL) mRNA throughout the hypothalamus were essentially unaffected by both regimes. These results demonstrate that hypothalamic NPY gene expression is regulated by peripheral metabolic status (and osmolality), and confirm the key physiological role of NPY in controlling ingestive behaviour.
Vaginocervical Stimulation of Ferrets Induces Release of Luteinizing Hormone‐Releasing Hormone
Authors: Bibeau Carole E. , Tobet Stuart A. , Anthony Edythe L. P. , Carroll Rona S. , Baum Michael J. , King Joan C.
Published: September 2006

Vaginocervical stimulation of ovariectomized estradiol‐primed ferrets (which are reflex ovulators) with a glass rod in the presence of a neck‐gripping male induced an increase in plasma luteinizing hormone (LH) from undetectable levels (≤0.50 ng/ml) before stimulation, to 2.4 ± 0.43 ng/ml 75 min after stimulation (stimulated females). Forty‐eight h after stimulation plasma LH returned to baseline levels (post‐stimulated females). A significant decrease in the number of perikarya containing LH‐releasing hormone (LHRH), detected by immunocytochemistry, was associated with the increase in plasma LH following stimulation. Approximately one half of the number of immunoreactive LHRH neurons (243±27) were detected in the forebrain of stimulated females, compared to those detected in the forebrain of post‐stimulated animals (436 ± 88) using antiserum AR 744. Equivalent results were obtained with a different antiserum (RM 1076) capable of detecting the extended decapeptide, or precursor, as well as partially or fully processed decapeptide.

We conclude that controlled Vaginocervical stimulation of female ferrets evokes the release of LHRH as well as LH, depleting approximately 50% of the LHRH perikarya of detectable LHRH. Additionally, electron microscopy of LHRH perikarya of stimulated females revealed more Golgi complexes/cell compared to baseline females. We propose that Vaginocervical stimulation also augments the processing of extended precursor forms of LHRH to generate the decapeptide.
Diurnal Differences and Adrenal Involvement in Calmodulin Stimulation of Hippocampal Adenylate Cyclase Activity
Authors: Gannon Maureen N. , Brinton Roberta E. , Sakai Randall R. , McEwen Bruce S.
Published: September 2006

Calciam/calmodulin‐dependent processes are altered by manipulations of the hypothalamic‐pituitary‐adrenal axis, and are associated with changes in synaptic efficacy in the hippocampus, such as long‐term potentiation. Recent evidence indicates that there are diurnal variations in the threshold for long‐term potentiation, as well as diverse effects of the adrenals and of adrenal steroids on electrical activity related to long‐term potentiation. In order to probe possible mechanisms underlying these observations, we investigated the effects of the diurnal cycle, as well as adrenalectomy (ADX) and adrenal demedullation on adenylate cyclase activity. In hippocampal, but not cortical, membranes the adenylate cyclase response to calmodulin was higher during the beginning of the dark phase of the cycle, when endogenous corticosterone levels are high. Basal and forskolin‐stimulated adenylate cyclase activity did not exhibit diurnal variation in either brain region. ADX (6 and 14 days) depressed the adenylate cyclase response to calmodulin in hippocampal membranes, and abolished the diurnal difference. ADX had smaller effects on this response in cortical membranes. ADX also attenuated basal and forskolin‐stimulated adenylate cyclase activity, but these changes were less striking than effects on calmodulin‐stimulated activity. Demedullation (14 days), generating corticosterone levels in the low physiological range, mirrored the effects of ADX on hippocampal adenylate cyclase activity. Corticosterone (20 to 25 μg/ml in the drinking water) did not consistently prevent ADX effects on adenylate cyclase activity. These results demonstrate that adrenal effects on adenylate case activity are regionally specific within the brain, and they suggest that other adrenal secretions besides glucocorticoids may be involved in the feedback of the diurnal rhythm on the hippocampus. Taken together with our recent finding that chronic stress or corticosterone injection selectively attenuated the adenylate cyclase response to calmodulin in cortical, but not hippocampal membranes our findings provide further support for a role of the pituitary‐adrenal axis in modulating neural calmodulin‐dependent adenylate cyclase activity.