bluuebunny/crossref_metadata_2025 huggingface dataset:
The Effect of Photoperiod on Plasma Levels of Melanin‐Concentrating Hormone in the Trout
Authors: Lyon V. E. , Baker B. I.
Published: September 2006

The neurohypophysial melanin‐concentrating hormone, MCH, plays a role in adaptive colour change in teleost fishes, inducing pallor when the fish is placed in pale‐coloured surroundings. The present study shows that its plasma concentration, measured in groups of white‐adapted fish, is not uniformly high throughout the day but follows a clear diurnal pattern. Over a 24 h cycle, plasma concentrations rise gradually during the morning to reach peak values around the middle of the photophase, after which they decline significantly before night. Lowest concentrations are observed during the dark period. This pattern was observed under a long photoperiod in summer and a short photoperiod in winter. The peak was shifted within a week of changing the onset of either light or dark. When dawn was delayed by 6 h for fish held on a long photoperiod, the usual morning rise in hormone titres was suppressed but, with the advent of light, hormone concentrations rose more rapidly than usual to reach peak values at about the normal time. If the dawn was advanced by 6 h for fish held under short photoperiod conditions, then peak concentrations were attained 6 h precociously. Fish from a long photoperiod placed in constant light showed a pattern of MCH release which approximated to the normal over the first 24 h period but plasma values then became raised and periodicity was no longer discernible. Plasma hormone concentrations were much reduced in trout kept in black coloured tanks in which nocturnal and daytime values differed, but significant differences during the photophase were not demonstrable. The results suggest that an illuminated white background can initiate the early morning release of MCH, and that an endogenous pacemaker underlies the pattern of MCH secretion.
Regulation of Vasopressin (VP) Gene Expression in the Bed Nucleus of the Stria Terminalis: Gonadal Steroid‐Dependent Changes in VP mRNA Accumulation are Associated with Alterations in mRNA Poly (A) Tail Length but are Independent of the Rate of VP Gene Transcription
Authors: Carter David A. , Murphy David
Published: September 2006

Forebrain vasopressin (VP) neurons of the bed nucleus of the stria terminalis (BNST) contrast with hypothalamic VP neurons in exhibiting nuclear gonadal steroid receptors which may directly effect steroid‐induced changes in VP gene expression observed in BNST cells. A transcription and Northern mRNA analysis has been performed to determine the mechanism through which gonadal steroids regulate VP gene expression in the BNST. In addition to confirming a distinctive, sexually dimorphic pattern of VP mRNA expression in the BNST as compared with the hypothalamic supraoptic nucleus (SON), our results show that the marked decrease in BNST VP mRNA levels observed two weeks after castration is not associated with a change in transcriptional activity of the VP gene. Similarly, VP gene transcription is not increased, relative to castrated animals, in the BNST of castrated rats treated with testosterone which exhibit normal or somewhat elevated levels of VP mRNA in the BNST. A post‐transcriptional mechanism therefore appears to underlie the gonadal steroid‐regulated changes in VP gene expression in the BNST. Since modulation of mRNA size (due to changes in poly (A) tail length) was also observed following castration and testosterone treatment it is apparent that the post‐transcriptional mechanism may involve regulated changes in VP mRNA poly (A) tail length.

The present findings contrast with the osmotic up‐regulation of VP mRNA levels in the SON which is primarily a transcriptional response, and provide a demonstration of the potential physiological importance of post‐transcriptional mechanisms of hormonal gene regulation.
Mating‐lnduced FOS‐Like Immunoreactivity in the Rat Forebrain: A Sex Comparison and a Dimorphic Effect of Pelvic Nerve Transection
Authors: Wersinger S. R. , Baum M. J. , Erskine M. S.
Published: September 2006

Previous research has shown that mating induces the expression of the immediate‐early gene, c‐fos, as detected by the increased presence of nuclear FOS‐like immunoreactivity (FOS‐IR), in specific forebrain regions of both male and female rats. In the male both olfactory/vomeronasal (O/V) and genital/somatosensory (G/S) inputs appear to contribute to the neural FOS response to mating whereas in the female G/S input carried by the pelvic nerves appears to mediate the forebrain FOS response. To date, however, no direct sex comparison of the mating‐induced forebrain FOS response has been made in rats maintained under the same steroidal conditions nor has the contribution of afferent sensory input from the pelvic nerves been assessed in males. We first compared the level of FOS‐IR in brain regions of mated and unpaired gonadectomized male and female rats given 5μg/kg estradiol benzoate (EB) for 7 days and 500 μg progesterone (P) 4 h prior to testing. One h after experiencing 1 ejaculation, both sexes showed increased FOS‐IR in the medial preoptic area (MPOA), bed nucleus of the stria terminalis (BNST), medial amygdala, the ventro‐lateral portion of the ventromedial nucleus of the hypothalamus (VMN), and the midbrain central tegmental field (CTF). This increase was significantly greater in the MPOA and medial amygdala of mated females than of males. Bilateral transection of the pelvic nerves significantly attenuated the increase in FOS‐IR after mating in the CTF of male rats and in the MPOA, BNST, VMN, medial amygdala and CTF of females. Thus, following mating there is no sex difference in the brain regions which express c‐fos, but there is a dimorphism in the contribution of afferent information conveyed by the pelvic nerves to the mating‐induced FOS response. The neural FOS response of the female to mating is heavily dependent upon the G/S afferent inputs carried by the pelvic nerves whereas the male's neural c‐fos response may depend on O/V input plus G/S input conveyed via other afferent pathways such as the pudendal nerves.
Short‐Term Regulation of Gonadotropin Subunit mRNA Levels by Estrogen: Studies in the Hypothalamo‐Pituitary Intact and Hypothalamo‐Pituitary Disconnected Ewe
Authors: Mercer Julie E. , Phillips David J. , Clarke Iain J.
Published: September 2006

In this study the levels of mRNA for the pituitary gonadotropin hormone subunits luteinizing hormone β (LHβ), follicle stimulating hormone β (FSHβ) and the common α‐subunit were assessed during the acute feedback stages of estradiol benzoate (EB) action in ovariectomized (OVX) ewes with and without hypothalamo‐pituitary disconnection (HPD).

In OVX/HPD ewes maintained on hourly pulses of 250 μg of gonadotropin‐releasing hormone (GnRH) a single i.m. injection of EB in oil caused a biphasic (decrease and then increase) change in plasma LH levels and a monophasic decrease in FSH levels. There was a decrease in pituitary α‐subunit and FSHβ mRNA levels during the acute negative (8 h post EB) and through the positive feedback (20 h post EB) stages of the response. No significant change was seen in LHβ mRNA levels following treatment with EB.

In hypothalamic‐pituitary intact OVX ewes the same EB treatment as above caused a biphasic change in LH secretion with the positive feedback component being much greater than in GnRH‐pulsed OVX‐HPD ewes. The levels of mRNA for all three gonadotropin subunits were reduced by 8 h after EB injection and remained low throughout the positive feedback period.

These data suggest that the LH surge in this experimental model does not require an increase in LHB mRNA levels. Furthermore, the fall in LHβ subunit mRNA seen after estrogen injection to OVX ewes is most likely due to an effect of estrogen to decrease GnRH secretion, since pulsatile GnRH replacement prevents this effect. These data also show that estrogen feedback can effect rapid alterations in pituitary gonadotropin subunit mRNA levels. Short‐term changes in FSHβ mRNA are reflected in changes in FSH secretion; the same is not true for LH.
Differential Seasonal Regulation of Melatonin Receptor Density in the Pars Tuberalis and the Suprachiasmatic Nuclei: A Study in the Hedgehog (Erinaceus europaeus, L.)
Authors: Gauer François , Masson‐Pévet Mireille , Saboureau Michel , George Denise , Pévet Paul
Published: September 2006

Using quantitative autoradiography, we have studied the seasonal changes of high affinity melatonin receptor density in both the SCN and PT of the hedgehog, a seasonal breeder and an hibernator. Animals in 3 different physiological states were studied: sexually active animals, and sexually inactive animals during the hibernation period, being then either euthermic or hypothermic. In sexually active animals, Bmaax were 75.8 ± 7.1 fmol/mg protein in PT and 9.1 ± 0.5 fmol/mg protein in SCN; and Kd values were: 94 ± 22 pM in the PT and 101 ± 15 pM in the SCN. This specific binding was strongly decreased in the PT of sexually inactive animals. Moreover, this decrease was significantly stronger in hypothermic than in euthermic hedgehogs. Saturation studies and Scatchard analysis revealed that the observed decrease in the PT resulted from change in the Bmax but not in the Kd, Bmax values being respectively 56.4 ± 5.9 and 29.5 ± 1.9 fmol/mg protein in euthermic and hypothermic sexually at rest animals. In none of the different physiological states, did the density of melatonin receptors of the SCN show any changes, Bmaax values being respectively 9.8 ± 0.5 and 9.8 ± 0.4 fmol/mg protein in euthermic and hypothermic sexually at rest animals. This shows for the first time a tissue‐specific regulation of melatonin receptor density occurring in the PT but not in the SCN. Furthermore, this decrease of binding in the PT is correlated with both sexual inactivity and hibernation period. This strongly suggests that the mediation of the photoperiodic effect on seasonal functions like seasonal hypothermia and reproduction involves an effect of melatonin on the PT rather than on the SCN.
Antidopaminergic‐lnduced Hypothalamic LHRH Release and Pituitary Gonadotrophin Secretion in 12 Day‐Old Female and Male Rats
Authors: Lacau‐Mengido Isabel M. , Becú‐Villalobos Damasia , Thyssen Sandra M. , Rey Estela B. , Lux‐Lantos Victoria A. R. , Libertun Carlos
Published: September 2006

In previous studies we have shown that the developing rat provides an interesting physiologic model in which the dopaminergic control of both LH and FSH is well defined in contrast to the controversial results obtained in adult rats. We wished to establish the role of testosterone in antidopaminergic induced gonadotrophins release in 12 day‐old male and female rats, and evaluate the effect of antidopaminergic drugs at the hypothalamic level during this developmental stage.

Haloperidol, an antidopaminergic drug, increased both LH and FSH in female 12 day‐old rats but not in male littermates. The effect was blocked by bromocriptine and not by phentolamine indicating that haloperidol acted on the dopaminergic receptor, and that unspecific stimulation of the noradrenergic system was not involved. Haloperidol was ineffective when female rats were previously ovariectomized and injected with testosterone propionate at 9 days of age. If females were treated on the day of birth with testosterone propionate, haloperidol‐induced FSH and LH release was also abolished. In control males haloperidol had no effect on the release of LH or FSH. But if males were orchidectomized at birth or at 9 days of age, haloperidol released both LH and FSH during the infantile period.

In an attempt to establish the site of action of antidopaminergic drugs on gonadotrophin release, hypothalami (mediobasal and preoptic‐suprachiasmatic area) from 12 day‐old infant female rats were perfused with either haloperidol or domperidone (2±10−6 M). Both drugs increased LHRH release into the perifusate. Besides haloperidol did not modify the release of LH or FSH from adenohypophyseal cells incubated in vitro.

We therefore conclude that antidopaminergic‐induced gonadotrophins release is modulated by serum testosterone concentrations, and that the site of action is probably the LHRH‐secreting neuron of the hypothalamus.
Immunocytochemical Evidence for Oestrogen Receptors within GABA Neurones Located in the Perinuclear Zone of the Supraoptic Nucleus and GABAA Receptor β2/β3 Subunits on Supraoptic Oxytocin Neurones
Authors: Herbison Allan E.
Published: September 2006

The mechanisms by which oestrogen modulates the biosynthetic and secretory activity of magnocellular oxytocin neurones are poorly understood. Using an antibody directed against the oestrogen receptor (ER), the distribution of ER‐immunoreactive (‐IR) cells in relation to the supraoptic nucleus (SON) was examined. Although no ER‐IR cells were detected within the SON, a small population of immunoreactive cells separate from those in the preoptic area was identified in the perinuclear zone of the SON. Double‐labelling experiments with an antibody specific for glutamic acid decarboxylase (GAD), the neuronal enzyme producing gamma aminobutyric acid (GABA), revealed that approximately 60% of perinuclear zone ER‐IR cells contained GAD. A further set of immunocytochemistry experiments using an antibody raised against the β2. and β3 sub‐units of the GABAA receptor revealed immunoreactivity in the SON. Double‐labelling experiments demonstrated that both oxytocin‐IR and non‐oxytocin‐IR neurones in the SON were immunoreactive for β2 and/or β3 sub‐units of the GABAA receptor. These studies have identified ERs within a GABAergic neural population in the perinuclear zone of the SON and shown that magnocellular oxytocin neurones in the SON possess GABAA receptors comprised of β2 and/or β3 sub‐units. In conjunction with previous evidence that the perinuclear zone GABA neurones are an important source of GABA terminals in the SON, these results provide a morphological basis for the hypothesis that perinuclear zone GABA neurones may be part of a steroid‐sensitive neural circuitry transmitting oestrogen input to oxytocin neurones in the SON.
Characterization of the GABA‐lnduced Current in Frog Pituitary Melanotrophs
Authors: Louiset E. , Mei Y. A. , Valentijn J. A. , Vaudry H. , Cazin L.
Published: September 2006

The molecular mechanisms regulating GABAA receptor activity in cultured frog melanotrophs were studied using the patch‐clamp technique. In the whole‐cell configuration, application of GABA evoked a dose‐related increase of inward chloride currents. The ED50 value, estimated from the sigmoidal dose‐response curve was 2 × 10−6 M and the Hill coefficient was 1.55. The amplitude of the GABA‐induced current decayed with time. Kinetics analysis of the desensitization revealed that the time‐course of the current decrement was fitted by one exponential. Graded doses of GABA or association of GABA with the benzodiazepine receptor agonist flunitrazepam accelerated the desensitization process. In contrast, the time‐course of the current did not significantly vary at different holding potentials. In the outside‐out configuration, GABA was found to activate channels which displayed three unitary conductance levels (8, 15 and 30 pS). The channel openings of the more frequent conductance level (30 pS) exhibited short and long lasting open states (1.2 and 28.3ms at ‐60mV). Altogether these data reveal that frog melanotrophs possess a single population of GABAA receptors which interconvert into a higher affinity state in the presence of benzodiazepine receptor agonists. Two GABA molecules must bind to the receptor to trigger long lasting channel openings. In addition, the activity of the GABAA receptor appears to be independent of the accumulation of intracellular chloride ions.
Characterization of Opioid Binding Sites in the Neural and Intermediate Lobe of the Rat Pituitary Gland by Quantitative Receptor Autoradiography
Authors: Boersma C. J. C. , Pool C. W. , Van Heerikhuize J. J. , Van Leeuwen F. W.
Published: September 2006

Previous studies have suggested an involvement of enkephalins in regulation of oxytocin (OXT) and vasopressin (AVP) release, which seems to disagree with the very low affinities of Met‐ and Leu‐enkephalin for the kappa opioid receptor. As opioid receptors in the neural lobe exclusively exist of kappa receptors, we studied the binding characteristics of larger pro‐enkephalin derived peptides for opioid binding sites in the neural lobe by means of light microscopic receptor autoradiography. In addition, the pharmacological characteristics of opioid binding sites in the neural lobe were compared with those in other parts of the pituitary.

In the neural as well as the intermediate lobe both high and low affinity 3H‐bremazocine binding sites were present. Binding to these sites was completely displaceable by both naloxone and nor‐binaltorphimine, suggesting that these sites represent kappa opioid receptors. Also with regard to selectivity and affinity characteristics to other ligands, opioid binding sites in the neural and intermediate lobe were quite similar. In the anterior lobe a very low level of bremazocine binding was present, which could not be displaced by nor‐binaltorphimine.

Displacement studies with pro‐enkephalin and pro‐dynorphin derived peptides showed that both groups of peptides could bind to opioid binding sites in the neural and intermediate lobe. Especially the relatively large pro‐dynorphin and pro‐enkephalin derived peptides, such as dynorphin 1–17 and BAM22, appeared to be very potent ligands for these opioid binding sites and were much more potent than smaller fragments, such as dynorphin 1–8, and Met‐ and Leu‐enkephalin. These results contradict the existence of a mismatch in the neural (and intermediate) lobe with regard to the local type of opioid peptides and receptors present.
Vasopressin Antisense Oligonucleotide Induces Temporary Diabetes Insipidus in Rats
Authors: Skutella Thomas , Probst Joseph Christopher , Engelmann Mario , Wotjak Garsten T. , Landgraf Rainer , Jirikowski Gustav F.
Published: September 2006

The purpose of this study was to downregulate the transcriptiona message of arginine vasopressin (AVP) by antisense treatment. A complete phosphorothioate antisense oligodesoxynucleotide corresponding to the beginning of the coding region of rat AVP mRNA was constructed and injected into the lateral ventricle of rats. Within 3–6 h animals exhibited a temporary diabetes insipidus, which lasted up to 9 h. Accordingly, vasopressin immunoreactivity in the hypothalamic nuclei was reduced. Our results demonstrate that a specific and reversible inhibition of neuropeptide expression can be accomplished in the intact hypothalamo‐neurohypophysial system by antisense treatment, thus providing a novel tool for studies on stimulus‐secretion coupling in vivo.
Similar Age Related Increase of Vasopressin Colocalization in Paraventricular Corticotropin‐Releasing Hormone Neurons in Controls and Alzheimer Patients
Authors: Raadsheer Frederik C. , Tilders Fred J. H. , Swaab Dick F.
Published: September 2006

Recent studies on experimental animals showed that long term activation of the hypothalamo‐pituitary‐adrenal axis is associated with increased vasopressin (AVP) colocalization in paraventricular corticotropin‐releasing hormone (CRH) neurons. In the present study we estimated the fraction of CRH neurons in which AVP is colocalized by double label immunocytochemistry in hypothalami of 10 control subjects of 21 ‐91 years of age and 10 age‐matched Alzheimer patients. CRH neurons in the paraventricular nucleus (PVN) of Alzheimer patients and control subjects showed similar age dependent increases in AVP colocalization. Based on this parameter, it seems that CRH neurons of Alzheimer patients are not overactivated as compared to age‐matched controls, but e.g. changes in m‐RNA for CRH should still be established.
Hypothalamic Dopaminergic Neurons in Prolactin‐Deficient Ames Dwarf Mice: Localization and Quantification of Deficit by Tyrosine Hydroxylase Immunocytochemistry
Authors: Phelps Carol J. , Carlson Susan W. , Vaccarella Myra Y.
Published: September 2006

Hypothalamic tuberoinfundibular prolactin‐inhibiting neurons show decreased levels and synthesis of dopamine in two types of genetically prolactin‐deficient dwarf mice (Snell, Ames) which arise from separate mutations. A reduction to 2% of normal in this neuronal population has been quantified for Snell dwarfs. The present study was undertaken in order to quantify morphometrically the deficit and its distribution in Ames dwarf mice, including comparisons of sex and adult age. The brains of dwarf (df/df) and normal phenotypic (DF/?) sibling mice of both sexes from 4 to 16 months of age were immunostained for tyrosine hydroxylase, the rate‐limiting enzyme in dopamine synthesis; neuronal perikarya were counted in coronal sections of tuberoinfundibular arcuate nucleus (area A12), medial zona incerta (A13) and anterior periventricular (A14) hypothalamic areas at 180 μm rostral‐to‐caudal intervals. Normal (DF/?) mice exhibited no differences in neuron numbers, with regard to age or sex, in any of the three dopaminergic areas. In dwarf mice, a tendency toward decreased neuron numbers with age was statistically significant for area A14 only, and the size of the neuronal population in A12 was reduced in males compared with females. Total A12 neuron number in dwarfs was 48% of that in normal mice (P< 0.001). Periventricular (A14) perikaryal numbers were reduced slightly (P<0.05) in dwarfs compared with normals. Numbers of A13 neurons were comparable for DF/? and df/df. The morphometric distribution of tyrosine hydroxylase‐immunoreactive neurons in A12 showed that the decrease in neuronal number in dwarfs was distributed throughout the rostral‐to‐caudal length of the nucleus, with significant decrease of total perikarya (P<0.05) at each 180 μm sampling interval. Thus, lifelong absence of prolactin in Ames dwarf mice is accompanied by a significant decrease in hypothalamic tyrosine hydroxylase‐immunoreactive neurons, which is restricted to hypophysiotropic areas, uniformly distributed within A12, and is less severe than the reduction in the phenotypically similar Snell dwarf.
Immunohistochemical Localization of GnRH in the Crested Newt (Triturus carnifex) Brain and Terminal Nerve: a Developmental Study
Authors: D'Aniello B. , Masucci M. , di Meglio M. , Lela L , Rastogi R. K.
Published: September 2006

Localization of GnRH‐immunoreactive neuronal system was studied by immunohistochemistry in the nasal‐brain area of the crested newt, Triturus carnifex. Besides adults, developmental stages were those from hatchlings up to complete metamorphosis. Neurons containing immunoreactive GnRH were first detected in the nasal area of larvae with yet undifferentiated gonads. Subsequently, in prometamorphic stages, GnRH‐immunoreactive cell bodies and fibers were detected in the proximal part of the terminal nerve as well as along the ventromedial surface of the olfactory bulbs. In older larvae with sexually differentiated gonads and up to the metamorphic climax GnRH‐neurons were detected, as a rostral to caudal continuum, along the ventromedial surface of the olfactory bulbs and midtelencephalon. This is exactly the route followed by the terminal nerve. In the adult brain, besides the presence of occasional GnRH‐neurons and fibers in the terminal nerve proximal to olfactory bulbs, olfactory bulbs and the mid‐basal telencephalon, another aggregate of immunoreactive neurons was present in the anterior preoptic area, and a greater number of fibers in the habenular area as well as in the infundibular floor, median eminence and pars nervosa. These data suggest the nasal area to forebrain migration (along the course of the terminal nerve) of GnRH‐neurons during development in the crested newt.
Calcium Waves and Dynamics Visualized by Confocal Microscopy in Xenopus Oocytes Expressing Cloned TRH Receptors
Authors: Eidne Karin A. , Zabavnik Jelka , Allan W. T. Gordon , Trewavas Anthony J. , Read Nick D. , Anderson Lorraine
Published: September 2006

Laser scanning confocal microscopy was used to analyse changes in free cytosolic calcium ([Ca2+]i) in Xenopus laevis oocytes expressing the cloned rat TRH receptor in response to TRH. In oocytes expressing TRH receptors, TRH invariably evoked a dose‐dependent, biphasic calcium response. This response consisted of an initial transient planar wave of calcium propagating just below the surface of the membrane followed by a slower, secondary calcium phase. The TRH antagonist, chlordiazepoxide, markedly inhibited this calcium wave. The origins of calcium involved in this biphasic response were investigated using a variety of intra‐ and extra‐cellular calcium antagonists. The intracellular calcium antagonists thapsigargin and TMB‐8 reduced the initial and to a lesser extent the secondary phase of the planar calcium wave. In contrast, EGTA and the calcium channel blocker nifedipine produced a profound inhibition of the secondary phase while the initial phase was only slightly reduced. These results indicate that the release of intracellular calcium is predominantly responsible for the initial phase of the calcium wave while the influx of extracellular calcium is mainly involved in the secondary phase. Qualitative changes in the patterns of calcium release induced by TRH were observed following pretreatment with intracellular calcium antagonists. Following pretreatment with these compounds, TRH induced spiral or regenerative calcium waves. Addition of EGTA to the extracellular medium did not alter these responses confirming the importance of intracellular calcium in the generation of these spiral calcium waves. This study demonstrates the nature and multiplicity of regulating mechanisms of [Ca2+]i following activation of TRH receptors expressed in Xenopus oocytes.
Oxytocin Administered Centrally Facilitates Formation of a Partner Preference in Female Prairie Voles (Microtus ochrogaster)
Authors: Williams Jessie R. , Insel Thomas R. , Harbaugh Carroll R. , Carter C. Sue
Published: September 2006

Prairie voles (Microtus ochrogaster) are monogamous mammals that form male‐female pair bonds. Partner preference formation, one component of the pair bond in prairie voles, occurs following male‐female cohabitation and is facilitated by mating. The peptide hormone oxytocin is released during physical contact and particularly following vaginal stimulation. Oxytocin has been implicated in mother‐infant bond formation. The present study tested the hypothesis that oxytocin participates in the partner preference component of pair bond formation in adult prairie voles. Ovariectomized female prairie voles were implanted with osmotic mini‐pumps releasing oxytocin (1–100 ng/h) or artificial cerebrospinal fluid (CSF). Pumps were implanted intracerebroventricularly or subcutaneously and females then were housed for 6 h with a male partner, followed by a preference test in which females could elect to spend time with either the partner or an unfamiliar male. Females in groups that received centrally‐administered oxytocin (10 or 100 ng/h), but not CSF, exhibited a significant preference for the partner present during infusion. The induction of a partner preference after oxytocin administration appeared specific for central oxytocin pathways as peripheral oxytocin administration was ineffective. Moreover, central administration of a selective oxytocin receptor antagonist inhibited the behavioral effect of exogenous oxytocin. These results suggest that oxytocin may be one factor contributing to the development of partner preferences in this monogamous rodent.
Luteinizing Hormone‐Releasing Hormone Receptor Messenger Ribonucleic Acid Expression in the Rat Pituitary during Lactation and the Estrous Cycle
Authors: Funabashi Toshiya , Brooks Philip J. , Weesner Gary D. , Pfaff Donald W.
Published: September 2006

To study mechanisms underlying the modulation of luteinizing hormone‐releasing hormone receptor (LHRH‐R) during lactation and the estrous cycle, we used a reverse transcriptase‐polymerase chain reaction (RT‐PCR) procedure to generate a probe for rat LHRH‐R messenger RNA (mRNA). Using primers based on the mouse sequence, we amplified an approximately 300 bp fragment from rat pituitary complementary DNA. This PCR product was shown to be part of LHRH‐R cDNA by direct sequencing and by comparing to the rat LHRH‐R cDNA reported recently. Then, this PCR fragment was used as a probe for northern blotting analysis.

The level of LHRH‐R mRNA in the pituitary was significantly decreased during lactation, by approximately 80%, compared to that of ovariectomized and intact (diestrous and metestrous cycling) rats while no statistical difference in glyceraldehyde‐3‐phosphate‐dehydrogenase (GAPDH) mRNA level was observed between groups. During the estrous cycle, the level of LHRH‐R mRNA in the pituitary was about two‐fold higher on diestrous day 2 and the morning of proestrus than that on diestrous day 1 and quickly returned toward control level by noon of proestrus. In addition, we found that GAPDH mRNA levels from a so‐called housekeeping gene often thought to be unchanged under different conditions, were significantly higher on proestrus while levels of 18S rRNA were not significantly changed.

The large decrease in LHRH‐R mRNA during lactation could account for the changes in LHRH binding previously reported.
A Critical Period for Glutamate Receptor‐Mediated Induction of Precocious Puberty in Female Rats
Authors: Smyth Catherine , Wilkinson M.
Published: September 2006

The excitatory amino acid glutamate and especially its NMDA subtype receptor are important components of the neural system that regulates sexual maturation. It is known that multiple daily injections of immature rats and monkeys with NMDA will induce precocious puberty. We have previously reported that a single daily injection of NMDA administered from 27 days of age to the day of vaginal opening (VO) is sufficient to synchronize and slightly accelerate (1–2 days) first ovulation in female rats. We have now optimized this treatment schedule and show that a higher dose of NMDA (20 mg/kg), or the racemic mixture N‐methyl‐D,L‐aspartate (NMA; 30 mg/kg), initiated earlier in development (24 days to VO) significantly advances first ovulation (4 days). Rats induced to ovulate prematurely had normal estrous cycles. We also report that the same degree of precocity can be obtained when injections are discontinued well before first ovulation occurs. For example, NMA administered from day 21 to 25 or from day 24 to 28 accelerates sexual maturation to the same degree as if injections were continued until VO was observed. It is clear that the hypothalamic‐pituitary‐ovarian (H‐P‐0) axis is stimulated by daily NMDA treatment as shown by the dose‐related luteinizing hormone (LH) release and by an estrogen‐dependent rise in uterine weight. However, stimulation of the P‐O axis with daily injections of GnRH (5 ng/100 g), which elicits an LH response slightly greater than NMDA (20 mg/kg), does not advance puberty. This suggests that NMDA induces some change in hypothalamic control which is not directly related to LH secretion. Interestingly, there also seems to be a critical period of NMDA effectiveness because daily injections of NMA (30 mg/kg) from day 16 to 20 do not induce precocious puberty. Since the ovaries respond with increased estrogen production (increased uterine weight) to gonadotrophin stimulation at this early age (16 days) we conclude that the hypothalamus may be relatively unresponsive to stimulation with NMDA.

Paradoxically the hypothalamus is also hyporesponsive to NMDA in the period preceding spontaneous first ovulation. We now show that an LH dose‐response curve for NMDA at age 28 days demonstrates that in NMDA‐treated rats the LH response to NMDA is less than in the control group. Further, the hyporesponsiveness is not due to pituitary desensitization since an LH dose‐response curve for GnRH at age 28 days is identical in the NMDA‐treated and control groups.

In conclusion we have determined that precocious ovulation can be induced by single, daily injections of NMDA beginning no earlier than postnatal day 21. In the peripubertal period the LH response to NMDA is severely attenuated in both treated and control rats. The evidence therefore suggests that contrary to the prevailing view, the H‐P‐O axis becomes less sensitive with the approach of puberty even though NMDA can induce premature ovulation.
Osmotic Inhibition of Prolactin Secretion in Rats
Authors: Dohanics Janos , Smith M. Susan , Blackburn Ruth E. , Verbalis Joseph G.
Published: September 2006

Chronic hyponatremia is known to cause inhibition of pituitary vasopressin (AVP) and oxytocin (OT) secretion in response to most physiological stimuli, as well as a marked inhibition of synthesis of these peptides. Because many studies have implicated neurohypophyseal peptides in the regulation of pituitary prolactin (PRL) secretion, we investigated the effects of chronic hyponatremia on basal and stimulus‐induced PRL secretion in rats. Hyponatremia was induced by subcutaneous infusion of 1‐deamino‐[8‐D‐arginine]‐vasopressin (dDAVP) (5ng/h) to rats fed a nutritionally balanced liquid diet, and plasma [Na+] was maintained ≤115 mmol/l for 10–12 days. After this period, hyponatremic rats and normonatremic controls fed the same diet without dDAVP were subjected to one of the following stimuli known to stimulate PRL release in rats: 3 min exposure to ether, hemorrhage (20 ml/kg), intravenous injection of 5‐hydroxytryptophane (5‐HTP, 10 mg/kg), or intravenous injection of estradiol (5 μg/kg). A baseline blood sample was collected before each stimulus, and 3–6 additional blood samples were collected at selected intervals after the stimulus. Baseline levels of plasma PRL were not different between normonatremic and hyponatremic rats. However, PRL responses induced by ether or estradiol, but not those induced by hemorrhage or 5‐HTP, were very significantly blunted in the chronically hyponatremic rats. Plasma AVP and OT responses were measured as an index of magnocellular secretion, but did not correlate with the PRL responses for any of the stimuli tested. Our results therefore demonstrate that ether‐ and estradiol‐induced PRL release can be osmotically inhibited, but the mechanisms underlying this inhibition appear to be relatively independent of effects on magnocellular AVP and OT secretion. This allows the possibility that either some parvocellular systems regulating PRL secretion are osmosensitive, or alternatively that other substances released from the neural lobe may selectively modulate pituitary PRL release in response to some, but not all, stimuli.
Somatostatin‐lmmunoreactive Neurones in the Hypothalamic Ventromedial Nucleus Possess Oestrogen Receptors in the Male and Female Rat
Authors: Herbison Allan E.
Published: September 2006

Neurones containing oestrogen receptors (ERs) in the ventrolateral division of the hypothalamic ventromedial nucleus (VMNvl) are believed to play an important role in mediating oestrogen's regulatory influence on reproductive behaviour. As somatostatin (SOM)‐immunoreactive neurones are found exclusively within the ventrolateral division of VMM, this study has used double‐labelling immunocytochemistry techniques to evaluate whether ER‐immunoreactive cells in the VMNvl synthesize SOM in both the male and female rat. Rats gonadecto‐mized 1 week earlier were perfused and brain sections through the mediobasal hypothalamus processed for ER and SOM immunoreactivity using the H222 monoclonal rat ER antibody and a polyclonal rabbit SOM antiserum. Within the VMN, cells immunoreactive for SOM were found predominantly in the rostral portion of the VMNvl while ER‐immunoreactive cells were distributed throughout the VMNvl. Sequential double‐labelling studies revealed that many ER‐containing cells in the rostral VMNvl were also immunoreactive for SOM. A semi‐quantitative analysis of double‐labelled cells in the rostral VMNvl of male and female rats, respectively, estimated that 52 ± 2% and 55 ±8% of SOM‐IR cells possess ERs while 35 ± 1% and 28 ±3% of ER‐IR cells synthesise SOM in the rostral VMNvl. No sex differences were detected at this level. These results show that approximately half of the SOM‐synthesizing neurones in the rostral VMN possess ERs and indicate that SOM should now be considered alongside enkephalin and Substance P as a putative mediator of oestrogen's regulatory influence on reproduction through the VMN of the rat.
Oestrogen Receptors in the Preoptico‐Hypothalamic Continuum: Immunohistochemical Study of the Distribution and Cell Density During Induced Oestrous Cycle in Ovariectomized Ewe
Authors: Blache D. , Batailler M. , Fabre‐Nys C.
Published: September 2006

Oestrogen plays a key role in the regulation of the endocrine and behavioural events associated with the oestrous cycle. It is important, therefore, to know the location of neurones receptive to this steroid and to know whether their distribution varies with the oestrous cycle. We have undertaken experiments to identify the location of oestrogen receptors (ER) within the preoptico‐hypothalamic continuum of ovariectomized ewes submitted to a variety of different hormone replacement regimes which mimic the different stages of the oestrous cycle. We used a monoclonal antibody to ER and detected receptors with immunohistological methods in the non‐vascular part of the organum vasculosum of the lamina terminalis, the lateral septum, the medial preoptic area, the supraoptic, suprachiasmatic and arcuate (ARC) nuclei, the ventromedial hypothalamus (HVM) and in the region close to the mamillari recess. ER neurones were scarce or absent from the anterior hypothalamus and the paraventricular nucleus. The density of ER staining in the HVM, but in no other localization, was found to be higher, and in a more lateral position, during the induced luteal phase (progesterone treatment) than during the follicular phase (7 days of progesterone treatment followed by oestradiol) or in the ovariectomized female. In all areas studied, except for the ARC, the apparent surface area of the nucleus in ER immunoreactive cells varied with hormonal treatment. These data, and especially those in the HVM, contribute towards our understanding of how steroids may act in the ovine to control sexual behaviour.
Expression and Down‐Regulation by Retinoic Acid of IGF Binding Protein‐2 and ‐4 in Medium from Human Neuroblastoma Cells
Authors: Bernardini Sergio , Cianfarani Stefano , Spagnoli Anna , Annicchiarico‐Petruzzelli Margherita , Melino Gerry , Massoud Renato , Boscherini Brunetto , Finazzi‐Agró Alessandro , Rosenfeld Ron G. , Federici Giorgio
Published: September 2006

Insulin‐like growth factors (IGFs) regulate the autocrine/paracrine growth of neuroblastomas. The IGFs bind to specific binding proteins (IGFBPs) which modulate their biological activity. We investigated, by Western ligand blotting (WLB), the presence of IGFBPs and their possible modulation by retinoic acid (RA), IGF‐I, IGF‐II and truncated Des(1‐3)IGF‐l in conditioned medium (CM) of the human neuroblastoma SK‐N‐BE(2) cell line. We demonstrated the presence of two IGFBPs, with MW 37 kDa and 25 kDa. Following immunoprecipitation, they turned out to be IGFBP‐2 and ‐4, respectively. The RA‐induced differentiation in SK‐N‐BE(2) cells was accompanied by a marked reduction of the intensity of both IGFBP bands after 48 h (32% and 24% of control, respectively) and 72 h (2% and 0% of control, respectively) incubation. The addition of exogenous IGFs, which did not induce cell differentiation, did not change the IGFBP pattern significantly, except for the truncated form of IGF‐I, which induced a marked decrease in both the 37 kDa and 25kDa bands after 72 h incubation (45% and 18% of control, respectively). These findings suggest that IGFBPs have a role in RA‐induced differentiation in human neuroblastoma cells.
[Arg8]Vasotocin Excites Neurones in the Dorsal Vagal Complex in vitro: Evidence for an Action through Novel Class(es) of CNS Receptors
Authors: Ingram C. D. , Tolchard S.
Published: September 2006

Using extracellular recordings from brainstem slices in vitro, it was demonstrated that a high proportion (38/56) of neurones in the dorsal vagal complex of dioestrus, virgin female rats exhibit an excitatory response to [Arg8]‐vasotocin (AVT). Pharmacological characterization suggests that these responses cannot be entirely explained by interaction with either of the currently known classes of central receptors for oxytocin (OT) and vasopressin (V1a). Comparison of the responses with those to the OT receptor‐specific agonist [Thr4,Gly7]‐OT (TGOT), showed that not all neurones that responded to TGOT also responded to AVT (3/27). Furthermore, while the effects of 10−7 M TGOT could be blocked either by the broad‐spectrum antagonist d(CH2)5[d‐Tyr(OEt)2,Val4,Cit8]‐vasopressin or by the selective OT receptor antagonist d(CH2)5[Tyr(Me)2,Thr4,Orn8,Tyr‐NH29]‐vasotocin, these peptides did not completely block the responses to AVT, indicating that AVT is unlikely to act through the central OT receptor. The responses to AVT and [Arg8]‐vasopressin (AVP) indicated the presence of at least 2 classes of receptor with which these agonists could act. Of 42 neurones tested with both AVP and AVT, none responded to AVP in the absence of a response to AVT, while 7/42 responded to AVT without a response to AVP. This might be explained by AVP acting through only the V1 receptor, while AVT acts through both the V1 and its own novel class of receptor. This was substantiated by the fact that two OT/V1 receptor antagonists, d(CH2)5[d‐Tyr(OEt)2,Val4,Cit8]‐VP and d(CH2)5|Tyr(Me)2,Tyr‐NH29]‐AVP, were unable to block completely all the responses to AVT at a dose which suppressed responses to both AVP and TGOT. These data suggest that, in addition to activation by OT and AVP, neurones in the mammalian central nervous system are able to be excited by AVT, possibly involving its own distinct class of receptor.
Cellular Distribution of G Protein Goa in Pituitary Lactotrophs: Effects of Dopamine
Authors: Painson Jean‐Claude , Wenger Tibor , Lagacé Ginette , Masson Nicole D. , Collu Robert
Published: September 2006

Membrane‐bound GTP‐binding (G) proteins mediate signal transduction in a variety of cell systems. The exact mechanisms of G proteins action are still under investigation but they appear to involve effectors located in the plasma membrane as well as in other parts of the cell. With this study, we investigated the cellular and ultrastructural localization of G protein subunits, and particularly of Goa, in normal rat anterior pituitaries and in estrone‐induced rat adenomatous lactotrophs. We also evaluated the effects of Goα cellular redistribution in rat adenomatous lactotrophs following short‐term exposure to dopamine (DA). Using the Protein A‐gold (PAG) methodology, Goα was found to be present in the cysternae of the endoplasmic reticulum of normal pituitary cells and of adenomatous lactotrophs. In the latter, Goα could be co‐localized with prolactin (PRL). By immunoblots, using specific antisera, significant amounts of Goα and Gs42α, together with smaller amounts of Giα, Gs47α and Gβ were found to be present in the uncontaminated supernatant fraction of adenomatous lactotrophs. Unexpectedly, exposure of the cells to DA induced a rapid and short‐lived decrease in the cytosolic fraction of Goα and Gβ associated with a decrease of PRL release. Since cytosolic Goα can be ADP‐ribosylated by pertussis toxin (PT) and is therefore in a heterotrimeric form, our data suggest that the soluble Go protein may play a role during lactotrophs' exposure to an inhibitor of PRL release, perhaps through its relocalization after being internalized with the D2 receptor or by being used for interaction with intracellular and/or membrane‐bound effectors.
Neuroactive Steroids Modulate GABAA Receptors in Peptidergic Nerve Terminals
Authors: Zhang Shuanglin J. , Jackson Meyer B.
Published: September 2006

Neuroactive steroid modulation of GABAA receptors was investigated in the peptidergic nerve terminals of the posterior pituitary using patch clamp techniques. In common with GABAA receptors in cell bodies, the nerve terminal GABAA receptor was potentiated by the synthetic steroid alphaxalone and by physiological concentrations of the progesterone metabolite allopregnanolone. Both of these agents enhanced Cl− currents elicited by GABA. Estradiol‐17β had a weak inhibitory effect on GABA responses of nerve terminals, but only at high concentrations. The potentiating action was manifest as an increase in the probability of channel opening, with no effect on the rate of desensitization of the GABAA receptor. Neuroactive steroids enhanced GABA‐gated Cl∼ channel activity in cell‐free membrane patches, thus demonstrating a membrane delimited response. These results indicated that with regard to allosteric modulation by neuroactive steroids, the nerve terminal GABAA receptor is similar to the GABAA receptors of nerve cell bodies and endocrine cells. Neuroactive steroids are thus capable of altering the chemosensitivity of nerve terminal membranes by enhancing GABA inhibition at this location. The neuroactive steroid sensitivity of nerve terminal GABAA receptors provides a pathway by which gonadal steroid derivatives could regulate peptide secretion from neurosecretory neurons. Such a pathway could participate in the coordination of neuropeptide secretion during complex neuroendocrine functions. With specific regard to the neurohypophysis, neuroactive steroid‐induced changes in the sensitivity of nerve terminal GABAA receptors could play a role in the initiation of oxytocin secretion during the transition between pregnancy and parturition.
Effects of Pituitary Adenylate Cyclase‐Activating Polypeptide (PACAP) and Vasoactive Intestinal Polypeptide (VIP) on Prolactin, Luteinizing Hormone and Growth Hormone Secretion in the Ewe
Authors: Sawangjaroen K. , Curlewis J. D.
Published: September 2006

This study was undertaken to investigate the roles of PACAP and VIP in the control of pituitary hormone secretion in the ewe. The first experiment was designed to identify any direct effects at the level of the pituitary and was conducted during the luteal phase of a prostaglandin‐synchronized oestrous cycle. PACAP (0.008, 0.04, 0.2 and 1.0 nmol/min) or VIP (0.06, 0.2, 0.6 and 1.8 nmol/min) was infused into the carotid artery over a 10 min period. Blood samples were taken before and after the infusions so that plasma PRL, LH and GH concentrations could be measured. Blood pressure was also monitored to determine if the doses used were biologically active. In no case was an effect on hormone secretion observed. In contrast, the highest dose of each peptide induced an increase in heart rate to almost three‐fold the resting value. Although both peptides are active in vivo, this result suggests that neither peptide has a direct effect on hormone release from the pituitary of prostaglandin‐synchronized ewes. In a second experiment, we investigated whether the peptides had central effects on hormone secretion. Intracerebroventricular (ICV) injection of PACAP or VIP at the dose 10nmol was tested in ovariectomized ewes. After injection, PACAP suppressed PRL and GH secretion so that plasma hormone concentrations from 1–3 h after injection were significantly different from the control (P<0.05 for PRL, P<0.01 for GH). In addition, PACAP significantly reduced mean LH concentration (P<0.05) and LH pulse frequency (P<0.01). A similar suppressive effect on LH secretion was also observed after ICV injection of VIP (P<0.05 for both parameters), although PRL and GH release were not affected. These results suggest a possible role for PACAP in the neuroendocrine control of PRL, GH and LH secretion in sheep. In addition, VIP may be involved in the control of LH secretion. In contrast, there is no evidence to suggest that either peptide is a hypophysiotropic factor for PRL, LH or GH in prostaglandin‐synchronized ewes.