bluuebunny/crossref_metadata_2025 huggingface dataset:
Evidence that Neural Aromatization of Androgen Regulates the Expression of Sexual Behaviour in Female Musk Shrews
Authors: Rissman Emilie F.
Published: September 2006

The experiments reported on here were conducted to test the hypothesis that sexual behaviour in the female musk shrew (Suncus murinus) is regulated by the neural aromatization of testosterone to oestradiol. In the first experiment ovariectomized animals received subcutaneous hormone implants containing either an aromatizable androgen (testosterone or androstenedione), a non‐aromatizable androgen (dihydrotestosterone or methyltrienolone), or cholesterol. Only females that received an aromatizable androgen exhibited significant amounts of sexual behaviour as compared with controls (cholesterol). To examine the role of the oestrogen receptor, the anti‐oestrogen, tamoxifen (200 or 400 μg daily) was given to ovary intact or ovariectomized females treated with testosterone. Tamoxifen treatment had significant negative effects both on female sexual behaviour and on the weights of several peripheral tissues as compared with control treatments. A similar set of experiments was conducted to examine the effect of an anti‐androgen on female sexual behaviour. The androgen receptor blocker, flutamide, had no effect on sexual behaviour or weights of peripheral tissues. To determine whether flutamide can act as an anti‐androgen in this species two final experiments were conducted in male musk shrews. Flutamide treatment in males did affect several measures of sexual behaviour. In summary, these data demonstrate that the oestrogen receptor is involved in the control of female copulatory behaviour. The androgen receptor plays a role in the expression of male, but not female, sexual behaviour. Female musk shrews display copulatory behaviour in advance of follicular development when oestradiol concentrations in plasma are very low. Thus, they may have evolved a strategy of aromatizing peripherally produced androgens in the brain to concentrate the oestrogen required for the expression of sexual behaviour.
Effects of Ethane Dimethane Sulphonate and Orchidectomy on Luteinizing Hormone Secretion
Authors: Dong Qihan , Handelsman David J.
Published: September 2006

The aim of this study was to determine whether testicular products of non‐Leydig cell origin modulate rat luteinizing hormone (LH) secretion in vivo. We therefore compared the effects of ethane dimethane sulphonate (EDS), a toxin regarded as highly selective for Leydig cells, with that of bilateral orchidectomy on LH secretion in mature male Wistar rats. The intention was thereby to compare the effects of selective removal of Leydig cells with that of removing both Leydig cells and seminiferous tubules, respectively. Following a single dose of EDS (75 mg/kg, ip), plasma LH concentrations rose equally with those of castrated rats for the first 3 days. After that time, however, plasma LH concentrations in the EDS‐treated rats fell progressively below those of the orchidectomized rats despite the continuing castrate level of circulating testosterone until Day 17. The effects of EDS treatment or orchidectomy on pulsatile LH secretion were then compared after 11 days to castrate levels of testosterone. EDS‐treated rats demonstrated reduced LH pulse amplitude, mean plasma LH levels and net LH secretion compared with castrate rats, although LH pulse frequency was unaltered. However, a further group of rats treated with EDS and orchidectomized failed to demonstrate that these changes were fully reversed by the castration and therefore EDS may have direct effects upon pituitary LH secretion. In order to determine the mechanism of the reduced LH pulse amplitude after EDS treatment, a further study was conducted to determine whether EDS treatment resulted in reduced pituitary sensitivity to gonadotropin‐releasing hormone (GnRH). Responsiveness of pituitary LH to exogenous GnRH (0.01 to 10 μg/kg body wt) was studied 11 days after removal of testicular testosterone feedback by either EDS or castration. Plasma LH response was linearly related to the log of the GnRH dose. At 10 min after GnRH administration, the plasma LH response in EDS‐treated rats was less sensitive than in castrate rats. We conclude that the lesser augmentation of LH secretion between Days 3 and 17 after EDS treatment compared with castrate rats cannot be explained solely by changes in Leydig cell secretion but may involve direct effects of EDS on pituitary LH secretion or non‐Leydig cell testicular products. Dampening of LH pulse amplitude without change in LH pulse frequency together with the reduced sensitivity to GnRH in EDS‐treated rats suggests that this toxin may have direct effects on pituitary LH secretion independent of its effects on Leydig cell function.
Effects of Pentobarbital on Proopiomelanocortin Opioid Products of Neonatal Piglets During Normoxia and Hypoxia
Authors: Moss Immanuela R. , Inman Jamie D. G.
Published: September 2006

Endogenous opioids have been shown to suppress physiological functions in the neonate. It has been suggested that anesthesia with barbiturates might enhance this suppression by influencing opioid systems directly. To explore this possibility, naive piglets, 2.2±0.8 (x̄±SD) days old, underwent one of five protocols: 1) normoxia (control); 2) 10% 02/90% N2 (hypoxia); 3) saline injection ip during normoxia (sham anesthesia); 4) pentobarbital sodium, 25 mg/kg ip, during normoxia (barbiturate anesthesia); and 5) pentobarbital sodium, 25 mg/kg ip, during hypoxia (combined hypoxia and barbiturate anesthesia). Following the inhalation of either gas mixture for at least 30 min, and precisely 30 min after an injection, blood, cerebrospinal fluid and a dorsal medullary slice containing the nucleus tractus solitarii were collected and processed for measurement by radioimmunoassay of opioid proopiomelanocortin products. These comprised β‐lipotropin (the precursor), β‐endorphin‐like immunoreactivity (containing the active peptide β‐endorphin) and N‐acetyl β‐endorphin (a deactivated peptide).

The most striking result was seen in the cerebrospinal fluid: As compared to barbiturate anesthesia, peptide levels with all other treatments, including combined hypoxia and barbiturate anesthesia, were consistently higher. In the plasma, peptide levels after either combined hypoxia and barbiturate anesthesia or hypoxia alone were generally higher than those of their respective controls (sham anesthesia, control). Plasma levels of β‐endorphin‐like immunoreactivity and estimated β‐endorphin with combined hypoxia and barbiturate anesthesia were also higher than those with barbiturate anesthesia. The latter pattern was reversed in the nucleus tractus solitarii, in which β‐endorphin‐like immunoreactivity and estimated β‐endorphin levels were lower with combined hypoxia and barbiturate anesthesia than with barbiturate anesthesia alone, although no significant differences were achieved.

These results suggest that pentobarbital may decrease the central neuronal release of active endorphins, and thus decrease the quantity of these ligands available for interaction with opioid receptors. Hypoxia, on the other hand, appears to increase such release even in the presence of pentobarbital. Thus, during a hypoxic insult, the suppressive influence of opioids on physiological functions would be enhanced regardless of the presence of barbiturate anesthesia.
Regulation of Melatonin Release and N‐Acetyltransferase Activity in Ovine Pineal Cells
Authors: Van Camp Gilles , Ravault Jean‐Paul , Falcón Jacky , Collin Jean‐Pierre , Voisin Pierre
Published: September 2006

A suspension culture of ovine pineal cells was developed to investigate the regulation of melatonin synthesis and release. Dosedependent stimulation of melatonin release by a series of adrenergic agonists yielded a typical β‐adrenergic profile. Serotonin N‐acetyltransferase (NAT), the rate‐limiting enzyme in melatonin biosynthesis, was also stimulated by a β‐adrenergic mechanism. However, NAT activity appeared less sensitive than melatonin release to β‐adrenergic stimulation. No evidence was obtained for a contribution of α1‐adrenergic receptors to the regulation of NAT activity and melatonin release. Activation of adenylate cyclase by forskolin or addition of a cyclic AMP analogue increased both melatonin release and NAT activity. In contrast, the Ca2+ ionophore A23187 stimulated melatonin release without a detectable increase in NAT activity. Together, the present data argue for a β‐adrenergic regulation of both NAT activity and melatonin release in ovine pinealocytes. The evidence also suggests that two intracellular mechanisms may control melatonin release in ovine pinealocytes: a cyclic AMP‐dependent mechanism, associated with an increase in NAT activity and a Ca2+‐dependent mechanism, independent of NAT activity.
Effects of Various Types of Hypothalamic Deafferentation on Luteinizing Hormone Pulses in Ovariectomized Rats
Authors: Ohkura Satoshi , Tsukamura Hiroko , Maeda Kei‐ichiro
Published: September 2006

The pulsatile luteinizing hormone (LH) secretion in chronically ovariectomized rats bearing various types of hypothalamic deafferentation was examined. Ovariectomized rats were subjected to complete, anterolateral or anterior hypothalamic deafferentation and bled every 6 min for 3 h through an indwelling atrial cannula 5 days after the brain surgery. Another group of ovariectomized animals was subjected to posterior‐anterior hypothalamic deafferentation (PAD), which cut off the anterior part of the arcuate nucleus from the mediobasal hypothalamus, and bled 1, 3 or 5 weeks after the deafferentation. Coronal sections of the brain were immunostained with anti‐LH‐releasing hormone (LHRH) serum. The pulsatile LH secretion was observed in rats bearing anterior, anterolateral or complete hypothalamic deafferentation and these types of deafferentation did not affect the frequency of LH pulses. The mean LH level during the 3‐h sampling period and the amplitude of LH pulses decreased as the incision extended postero‐laterally. Rats bearing PAD showed an irregular fluctuating pattern in plasma LH concentration 1 week after PAD. Parameters of LH pulses were restored with time after PAD. This suggests that the system generating LHRH pulses severed by PAD had been reorganized. LHRH‐immunopositive neuronal fibres were found in the external layer of the median eminence in the rats bearing any type of deafferentation. The present results suggest that the frequency of LH pulses could be controlled by the LHRH pulse generator, which consists of non‐LHRH neurons and is located in the mediobasal hypothalamus.
Phospholipase A2 and Somatostatin Release are Activated in Response to N‐Methyl‐D‐Aspartate Receptor Stimulation in Hypothalamic Neurons in Primary Culture
Authors: Rage Florence , Pin Jean‐Philippe , Tapia‐Arancibia Lucia
Published: September 2006

We have recently shown that glutamate primarily induces somatostatin release in hypothalamic neurons through N‐methyl‐D‐aspartate (NMDA)‐type receptor sites. Here we report that glutamate and NMDA also stimulate the release of [3H]arachidonic acid in a dose‐dependent manner. The NMDA‐induced effects (arachidonic acid release and somatostatin secretion) were both inhibited by MK‐801, an NMDA receptor‐type antagonist, or mepacrine, a phospholipase A2 inhibitor. In addition, mepacrine was able to inhibit A23187‐stimulated arachidonic acid release and somatostatin secretion. p‐Bromophenacylbromide, another phospholipase A2 inhibitor, also blocked NMDA‐induced secretion of somatostatin. However, responses to NMDA were unaffected by H7 (inhibitor of protein kinase C), nordihydroguaiaretic acid or indomethacin (inhibitors of lipoxygenase and cyclooxygenase). Melittin, a phospholipase A2 activator, was found to stimulate both responses, but omission of extracellular Ca2+ from the incubation media strongly reduced melittin‐induced somatostatin release. Six‐h pertussis toxin pretreatment did not significantly reduce the action of NMDA on either of the two parameters studied. High‐performance liquid chromatography analysis of [3H]metabolites released in the medium after NMDA stimulation revealed that [3H]arachidonic acid was the only detectable metabolite. External addition of arachidonic acid increased the release of somatostatin, whereas E2 and F2α prostaglandins had no effect. Our results show a close correlation between arachidonic acid release and somatostatin secretion, the two parameters we investigated.
Inhibitory Actions of Acute Estradiol Treatment on the Activity and Quantity of Tyrosine Hydroxylase in the Median Eminence of Ovariectomized Rats
Authors: Pasqualini Catherine , Leviel Vincent , Guibert Bernard , Faucon‐Biguet Nicole , Kerdelhué Bernard
Published: September 2006

The effects of acute estradiol (E2) treatment on both the activity of tyrosine hydroxylase (TH) in the median eminence and the serum level of prolactin (PRL) were investigated. Twelve‐day‐ovariectomized rats were injected with 17β‐E2 (25μg sc) at 1100 h and sacrificed hourly from 1200 to 2300 h. TH activity was quantified by measuring the amount of exogenous tyrosine converted to L‐DOPA in vitro by aliquots of median eminence homogenates. Serum PRL levels were evaluated by radioimmunoassay. A biphasic response of TH activity to treatment was observed: an immediate decrease occurred—preceding and accompanying a rise in serum PRL—followed by an increase beyond control levels 2 h after the maximal release of PRL. The increase in TH activity could be prevented by the pretreatment of rats with a specific rat PRL antiserum, suggesting it was not due to E2 per se but rather mediated by the E2‐induced PRL elevation. To pin‐point the process underlying the E2‐induced decrease in TH activity, we evaluated the kinetic parameters of TH in the median eminence as well as its quantity (by Western blot analysis) in the median eminence and arcuate nucleus. Finally, we used a sensitive dot‐blot assay to quantify specific TH messenger ribonucleic acid in the arcuate nucleus. The decrease in TH activity after E2 treatment paralleled an immediate decrease in the affinity of TH for its pterin cofactor (6‐MPH4), while Vmax remained unchanged. A decrease in the amount of TH protein in the arcuate nucleus and median eminence as well as in the TH messenger ribonucleic acid level in the arcuate nucleus was also observed, but the latency of these effects precluded a major involvement in the immediate decline of TH activity. Therefore, when observed separately from those of PRL, E2 effects on TH in tuberoinfundibular dopaminergic neurons are clearly inhibitory consisting of a ‘deactivation’ of the enzyme together with a reduction of its synthesis.
Neurotensin‐lmmunoreactive Neurons in the Rat Medial Preoptic Area are Oestrogen‐Receptive
Authors: Herbison Allan E. , Theodosis Dionysia T.
Published: September 2006

The identity of neurons influenced by oestrogen is critical for the understanding of ovarian steroid actions in the brain. The medial preoptic area (MPOA) contains one of the largest oestrogen‐receptive cell populations in the rat brain and participates in several oestrogen‐dependent functions, including the regulation of luteinizing hormone (LH) secretion and sexual behaviour. Using double immunostaining procedures with antibodies specific for the oestrogen receptor and neurotensin, a neuropeptide implicated in the regulation of LH secretion within this area, we found that approximately half of the neurotensin‐immunoreactive neurons in the MPOA also displayed immunoreactivity for the oestrogen receptor. We estimate that oestrogen‐receptive neurotensin neurons represent 5% to 12% of all oestrogen receptor‐positive cells in the MPOA. Our results provide morphological evidence that neurotensin mediates oestrogen‐dependent mechanisms within the brain and suggest that oestrogen may act through preoptic neurotensin neurons to aid in the generation of the LH surge.
Stimulation of Luteinizing Hormone‐Beta Messenger Ribonucleic Acid and Post‐Translational Modification of Luteinizing Hormone Isoforms by Second Messengers Mediating the Action of Gonadotrophin‐Releasing Hormone
Authors: Leigh A. J. , Wilson C. A. , Edger M. J. , Tipping K. E. , Patel M. , Chapman A. J. , Whitehead S. A.
Published: September 2006

Several second messenger systems have been implicated in mediating the action of gonadotrophin‐releasing hormone on the pituitary gonadotrophs and numerous studies have shown that activation of these systems induces luteinizing hormone (LH) secretion. However, it is not known how gonadotrophin‐releasing hormone or the second messenger systems induce de novo LH biosynthesis and post‐translational modification of the hormone. In these experiments hemipituitary glands have been perifused with drugs which activate second messengers or stimulate protein kinase C directly. The LH secretory responses have been correlated with measurements of common a and LHβ mRNA and the molecular species of LH which were present in the pituitary perifusate after exposure to the drugs.

Gonadotrophin‐releasing hormone (50 ng/ml, 42 nM), with and without the presence of extracellular Ca2+, the Ca2+ ionophore, A23187 (10 μM), and phorbol 12‐myristate (1 μM) all stimulated an increase in LHβ mRNA compared with controls and the appearance of a different isoform of LH to that found stored in and released from the unstimulated pituitary gland. Phospholipase C was without effect on LHβ mRNA levels and showed minimal efficacy in inducing the appearance of the different LH isoform.
Autoradiographic Detection of Oxytocin‐ and Vasopressin‐Binding Sites in Various Subnuclei of the Bed Nucleus of the Stria Terminalis in the Rat. Effects of Functional and Experimental Sexual Steroid Variations
Authors: Kremarik P. , Freund‐Mercier M. J. , Stoeckel M. E.
Published: September 2006

Oxytocin‐ and vasopressin‐binding sites were detected by autoradiography on films and on emulsion‐coated sections of rat brains using highly selective [125|]‐labelled oxytocin and vasopressin antagonists. Two distinct areas with high concentrations of oxytocin‐binding sites were detected in the bed nucleus of the stria terminalis: 1) the principal encapsulated nucleus and the associated cell‐sparse zone in the posterior medial part, and 2) the oval nucleus in the anterior lateral part. A weak diffuse labelling was, in addition, detected around the oval nucleus in the anterior lateral and anterior dorsal areas. The vasopressin‐binding sites were restricted to the anterior lateral part of the bed nucleus of the stria terminalis where they were highly concentrated in the juxtacapsular nucleus and present with lower density in a discrete cell group dorsal to the oval nucleus.

Autoradiographic analyses of the bed nucleus of the stria terminalis from pregnant, lactating and ovariectomized rats (oestradiol treated or not) indicated that only the oxytocin‐binding sites in the principal encapsulated nucleus and the associated cell‐sparse zone were oestrogen‐dependent. These observations are in agreement with earlier data suggesting that the two major divisions of the bed nucleus of the stria terminalis are involved in distinct regulations. The anterior lateral part, including the oval nucleus in which oxytocin receptors are not oestrogen‐dependent, is, rather, involved in central autonomie regulations. The posterior medial part, where oestrogen‐dependent oxytocin receptors are concentrated in the principal encapsulated nucleus and the associated cell‐sparse zone, is implicated in neuroendocrine regulations and in reproductive behaviour.
Lack of Glucocorticoids Enhances the Early Activation of the Medullary Catecholaminergic Cell Groups Triggered by Restraint Stress
Authors: Lachuer Joël , Buda Michel , Tappaz Marcel
Published: September 2006

We have investigated whether the stress‐induced activation of the medullary catecholaminergic neurons, that was shown previously to provide the main central activation input to the hypothalamo‐pituitary‐adrenocortical axis during an immobilization stress, is sensitive to circulating corticosteroids. Experiments were carried out on adrenalectomized rats that were first maintained on corticosterone in the drinking water for 5 days following surgery and then switched to corticosterone‐free water 15 h before stress application. Some of the latter animals were injected with dexamethasone. Activation of the brainstem Catecholaminergic neurons was estimated by assaying 3,4‐dihydroxyphenylacetic acid (DOPAC), a side metabolite of the noradrenaline and adrenaline biosynthesis pathway that was established previously as a reliable index of the activity of these neurons. In the so‐called A1C1 and A2C2 Catecholaminergic groups of the medulla, lack of corticosterone under these experimental conditions did not modify the basal level of DOPAC but led to a further enhancement (+ 35% to 40%) of the approximately 2‐fold increase in DOPAC content observed 15 min after the onset of the 5‐min immobilization stress. No significant further enhancement of the stress‐induced DOPAC increase was observed in the locus coeruleus. In these adrenalectomized rats, dexamethasone pretreatment prevented the enhancement of the stress‐induced increase in DOPAC level observed in the medullary cell groups but did not abolish the response to stress. Lack of endogenous corticosteroids led to a 10‐fold enhancement of the adrenocorticotropin increase following immobilization stress. Pretreatment with dexamethasone fully abolished the stress‐induced increase in adrenocorticotropin plasma level.

Our results show that circulating corticosteroids reduce the stress‐induced activation of the medullary A1 and A2C2 groups i.e. those that contribute mainly to the catecholaminergic innervation of the hypothalamo‐pituitary‐adrenocortical axis within the hypothalamic paraventricular nuclei. However, since the feedback regulation of the central catecholaminergic systems is much less efficient than the feedback actually observed on the adrenocorticotropin secretion, we suggest that it is likely to play a minor physiological role in the overall feedback regulation exerted by circulating corticosteroids on the hypothalamo‐pituitary‐adrenocortical axis.
cFos Activity Identifies Recruitment of Luteinizing Hormone‐Releasing Hormone Neurons During the Ascending Phase of the Proestrous Luteinizing Hormone Surge
Authors: Lee Wen‐Sen , Smith M. Susan , Hoffman Gloria E.
Published: September 2006

The proto‐oncogene product of the c‐fos gene, cFos, is a useful marker for luteinizing hormone‐releasing hormone (LHRH) neuronal activation. While recent data indicate that in the rat, an LHRH surge plays an active role in stimulating the proestrous luteinizing hormone (LH) surge, the mechanics of the LHRH surge remain unknown. The aim of this study was to determine whether LHRH neuronal activation occurs entirely at the beginning of the LH surge or whether the number of LHRH neurons activated increases during the ascending phase of the surge. To accomplish this aim, we determined the relationship between the number of LHRH neurons expressing cFos and LH concentrations during the ascending limb of the proestrous LH surge. During the estrous cycle in the rat, on the afternoon of proestrus, the number of LHRH neurons expressing cFos increased as plasma LH levels increased to reach peak concentrations. The regression line describing these two variables had a very highly significant correlation coefficient, indicating a linear relationship. Treatment with RU486 to block progesterone's action on the afternoon of proestrus significantly reduced both the number of LHRH neurons expressing cFos and the magnitude of LH secretion during the entire ascending phase of the LH surge. An analysis of covariance with comparison of regression lines from untreated and RU486‐treated animals revealed that while both sets of data established significant linear relationships between the degree of activation of LHRH neurons and plasma LH values, the slopes of the two lines were different (P = 0.031) with no statistical difference in the two intercepts. These data, together with the demonstration of an overall reduction of cFos intensity following removal of progesterone's actions, suggest progesterone alters the dynamics of LHRH neuronal activation by significantly reducing the recruitment of LHRH neurons and suppressing the level of activation of individual LHRH neurons. The results of our study support the hypothesis that the ascending phase of the LH surge results from the gradual recruitment of LHRH neurons into the active state.
Acute Luteinizing Hormone and Prolactin Responses to Paced Mating Stimulation in the Estrous Female Rat
Authors: Erskine Mary S. , Kornberg Eva
Published: September 2006

The present experiments sought to characterize the particular stimuli received during mating in the female rat which induce acute increases in luteinizing hormone (LH) and prolactin (PRL) following copulation. Comparisons were made between cycling females mated on the evening of proestrus in partitioned chambers in which spontaneous patterns of approach/withdrawal toward the male served to pace copulatory stimulation (paced), in non‐partitioned chambers in which female regulation of intervals between copulatory mounts was prevented (non‐paced), or under conditions in which they received mounts‐without‐intromission (mounts‐only). Frequent blood samples were withdrawn via surgically implanted intra‐atrial catheters. In experiment 1, blood samples for LH determinations were taken at 15‐min intervals for 1 h prior to and for 2 h after mating on the evening of proestrus. In experiment 2, samples for PRL determinations were taken at 10‐min intervals for 30 min prior to and for 90 min after mating on proestrus and at 0300, 0400 and 0500 h on the day of estrus (reported times corrected for reversed light cycle). LH levels were significantly higher in paced animals 15 min after initiation of mating than in non‐paced and mounts‐only females; no differences in LH were seen between females who subsequently became or did not become pregnant/pseudopregnant (P/PSP). PRL values were not different between groups receiving the different mating treatments at any time; however, P/PSP animals showed significantly higher levels of PRL between 20 to 60 min after mating than did non‐P/PSP females. No differences in PRL were seen between mating treatments or pseudopregnancy condition at 0300 to 0500 h on estrus. Paced females in both experiments received intromissions at a significantly slower rate than did non‐paced females. There was a significant positive correlation (r = 0.619, P<0.001) between LH concentration at 15 min and the inter‐intromission interval (in seconds) in paced and non‐paced groups of females. These data suggest that an LH response to mating is dependent upon the particular characteristics of mating stimulation received. In addition, they demonstrate that PRL increases acutely after mating stimulation in animals destined to become P/PSP but does not increase in response to those characteristics of mating stimulation which induced increases in LH.
Characterization of Melatonin Binding Sites in the Pars Tuberalis of the European Hamster
Authors: Skene D. J. , Masson‐Pévet M. , Pévet P.
Published: September 2006

Melatonin binding sites in the pars tuberalis of the European hamster (Cricetus cricetus) have been characterized using the radioligand 2‐[125I]iodomelatonin. Specific 2‐[125I]iodomelatonin binding was assessed using radioreceptor studies of pars tuberalis membrane preparations. Saturation studies revealed a single, high affinity site (Kd 39.8 (± 7.6 SEM) pM and Bmax 4.1 (± 0.5 SEM) fmol/mg protein, n=4). Kinetic experiments showed the 2‐[125I]iodomelatonin binding to be rapid, saturable and reversible. The Kd calculated from the dissociation and association rate constants was 19.4 pM. The order of potency of different indoles for inhibition of 2‐[125I]iodomelatonin binding was 6‐chloromelatonin > melatonin > 6‐hydroxymelatonin > N‐acetylserotonin > 5‐methoxytryptophol > serotonin > 5‐methoxytryptamine. GTP caused a dose‐dependent inhibition of the 2‐[125I]iodomelatonin binding. A saturation study showed that GTP reduced the number of binding sites by a third without altering their affinity. These results imply the presence of a G‐protein‐coupled melatonin receptor in the pars tuberalis of sexually active European hamsters.
Sexual Behavior Triggers the Appearance of Non‐Neuronal Cells Containing Gonadotropin‐Releasing Hormone‐Like Immunoreactivity
Authors: Silver R. , Ramos C. L. , Silverman A‐J.
Published: September 2006

Gonadotropin‐releasing hormone (GnRH) regulates the secretion of pituitary gonadotropins and facilitates the display of sexual behavior. We report that in doves, following a brief period of courtship, non‐neuronal cells containing GnRH‐like immunoreactivity (ir) are seen in the habenula of both male and female doves. These cells appear to be translocated from either the cerebrospinal fluid or from capillaries in the pia or choroid plexus into the parenchyma of the brain. Immunoreactive cells are virtually absent in the habenula in control animals housed alone. The identity of the ir cells that enter the habenula is unknown but they would appear to be of either the macrophage or the mast cell lineage. Both of these blood‐derived cells have heterochromatic nuclei and irregular cell surfaces with many filamentous processes, as do the GnRH‐ir cells. The vacuolated granules of the ir cells could be indicative of mast cell degranulation or of endocytic vesicles of a phagocytosing cell. These data suggest that there is a population of cells within the habenula that are of similar size and morphology to the GnRH‐ir ceils and are metachromatic when stained with toluidine blue. The latter is a property of heparin‐containing mast cells. Lineage specific markers that permit a double‐label study will be required to determine the exact nature of the GnRH‐ir cells. Whatever their lineage, the translocation of non‐neuronal cells into the undamaged adult central nervous system has not been described previously, and may provide a means of delivering biologically active substances into specific brain regions.
Interactions Between the Hypothalamo‐Pituitary Adrenal Axis and the Thymus in the Rat: A Role for Corticotrophin in the Control of Thymulin Release
Authors: Buckingham J. C. , Safieh B. , Singh S. , Arduino L. A. , Cover P. O. , Kendall M. D.
Published: September 2006

Our recent observations in man suggested that the secretion of the thymic peptide, thymulin, is influenced by hormones of the pituitary‐adrenal axis. In the present study, we have used the rat as a model in order to examine 1) the effects of corticotrophin (ACTH) and glucocorticoids on the release of thymulin in vivo and in vitro, and 2) the influence of an acute rise in plasma thymulin on the secretion of corticosterone and luteinizing hormone.

Immunoreactive thymulin was readily detectable in plasma from male Sprague‐Dawley rats(≃200 g). Chronic bilateral adrenalec‐tomy, which effectively removed endogenous corticosterone, produced highly significant (P<0.01) increases in the plasma concentrations of both ACTH and thymulin. Treatment of the adrenalectomized rats with dexamethasone, in a dose sufficient to suppress the hypersecretion of ACTH, maintained the plasma thymulin at a low level which did not differ significantly (P > 0.2) from that in sham‐operated controls.

In vitro, two non‐specific depolarizing agents, K+ (56 mM) and veratridine (10 ≃M), caused significant (P<0.01) Ca2+‐dependent increases in thymulin release from segments of rat thymic tissue. Their effects were mimicked by ACTH1–39. The secretory responses to ACTH (0.025 to 1 ng/ml) were concentration‐dependent but a very high concentration (2 ng/ml) of the peptide was without effect. Dexamethasone (0.1 μM) reduced (P<0.05) the spontaneous release of thymulin in vitro but potentiated markedly (P<0.01) the secretory responses to ACTH (0.5 to 1.0 ng/ml).

Administration of thymulin (0.1 and 10 μg/kg ip) produced, within 10 min, striking increases in the plasma thymulin concentration which were still evident at 30 min. The peptide concentration then declined rapidly and, within 24 h, was lower than that in the corresponding vehicle‐treated controls. The serum concentrations of corticosterone and luteinizing hormone were unaffected by the thymulin treatment. The saline vehicle (2.0 ml/kg ip) also produced a small increase in plasma thymulin concentration which was maximal at 10 min; a further small rise was evident 6 h after the injection but thereafter the thymulin values were indistinguishable from those in uninjected controls. A similar biphasic profile of serum corticosterone was apparent after the saline injection but the serum luteinizing hormone was unaffected.

The results suggest that ACTH is a physiological enhancer of thymulin release and that, in certain circumstances, its effects may be potentiated by glucocorticoids.
Central Noradrenergic Lesion Impairs the Adrenocorticotrophin Response to Release of Endogenous Catecholamines
Authors: Al‐Damluji Saad , White Anne
Published: September 2006

Activation of hypothalamic α1‐adrenoceptors stimulates the secretion of corticotrophin‐releasing factors which in turn stimulate pituitary adrenocorticotrophin (ACTH). This mechanism is important in the physiological control of ACTH secretion. This study assesses the feasibility of using the ACTH response to release of endogenous catecholamines as a means of detecting a hypothalamic noradrenergic lesion in vivo. Intracerebroventricular infusion of the catecholamine neurotoxin, 6‐hydroxydopamine, was used to destroy noradrenergic nerve endings in rats, with the purpose of producing a model that could be used to study alterations in ACTH responses that may result from a lesion involving central noradrenergic neurons. 6‐Hydroxydopamine (250 μg icv) significantly reduced hypothalamic noradrenaline content, indicating damage to noradrenergic nerve endings, without affecting postsynaptic receptor function, as judged by preservation of the effect of a selective α1‐adrenergic agonist. Pharmacological release of endogenous catecholamines, effected by combined administration of a catecholamine precursor and an α2‐adrenergic antagonist, stimulated the secretion of ACTH in control, but not in 6‐hydroxydopamine‐treated rats. Degeneration of hypothalamic noradrenergic nerve endings is not followed by denervation hypersensitivity, and is therefore accompanied by impairment of the ACTH response to release of endogenous catecholamines.
Arginine Vasotocin Gene Expression During Osmotic Challenge in the Chicken
Authors: Miihlbauer E. , Hamann D. , Xu B. , Ivell R. , Ellendorff F. , Grossmann R.
Published: September 2006

The avian hypothalamic nonapeptide arginine vasotocin (AVT) is released from axon terminals in the neural lobe upon the application of osmotic stimuli. We have investigated whether, and to what extent, hormone secretion from the neurohypophysis is related to gene expression in the hypothalamus. Results from hybridization experiments with an AVT‐specific cDNA probe indicate that in adult chickens stimulated by water deprivation or by hypertonic saline (2% w/v) drinking water, an upregulation of the AVT mRNA pool takes place, since consistently higher AVT mRNA levels compared to controls were monitored in osmotically challenged birds. This stimulatory effect was even visible at the transcriptional level after 19 h of water deprivation when osmolality was still near the basal value. In hens osmotically challenged by hypertonic saline drinking water for 5 days, a dissociation between osmolality and AVT plasma concentration was visible: extremely high plasma osmolality was accompanied by only moderately increased plasma AVT concentration. This might be caused either by exhaustion of stored hormone, or by downregulation of the system after chronic challenge. The latter suggestion is supported by the fact that the AVT mRNA concentration after 5 days of hypertonic saline challenge was well below the AVT mRNA levels of the groups with the more short‐term stimuli of water deprivation for 19 or 48 h. In 30‐day‐old chicks the hypothalamic AVT mRNA concentration hardly reached 70% of the adult value, although AVT plasma concentrations were similar to those in the mature bird.

We conclude that osmotic challenge of the hypothalamo‐neurohypophysial system not only causes secretion of AVT from stores in the neural lobe but is accompanied by upregulation of AVT gene expression. Upregulation already occurs after marginal increase in plasma osmolality, as seen after 19 h of water deprivation in hens. In 30‐day‐old chicks gene expression is only slightly upregulated after short‐term water deprivation while increase in plasma AVT is even greater compared to hens.
Localization of Insulin‐Like Growth Factor‐II Receptors in Rat Brain by in vitro Autoradiography and Immunohistochemistry
Authors: Batch Jennifer A. , Werther George A. , Hogg Annette , Mendelsohn Frederick A. O. , Scott Carolyn , Oldfield Brian J.
Published: September 2006

Insulin‐like growth factor‐ll (IGF‐II) and its receptor, which is homologous with the mannose‐6‐phosphate (M6P) receptor, are found in high levels in adult rat and human brain, though their role remains unclear. In order to point to possible regional functions, we have mapped and quantified IGF‐II/M6P receptors in sagittal sections of adult rat brain by in vitro autoradiography/computerized densitometry and immunohistochemistry. While in vitro autoradiography allowed mapping and quantitation, immunohistochemistry both confirmed mapping and allowed more detailed determination of cellular distribution of receptors. The two methods were generally in agreement with few areas of mismatching. By in vitro autoradiography, a discrete and characteristic distribution of IGF‐II receptor binding was demonstrated, with specific binding representing 85% of total binding. Displacement and specificity competition curves in arcuate nucleus and choroid plexus were typical for authentic IGF‐II receptors with half maximal displacement at 1 nM cold IGF‐II. IGF‐II receptor density, estimated by in vitro autoradiography, was very high in circumventricular organs, especially the median eminence, which had the highest binding in the brain. In the remainder of the brain there was concordance between the distribution of receptors identified by the two techniques, with greatest densities in the olfactory bulb and olfactory pathways, the hippocampus and discrete regions of the cerebral cortex, cerebellum, hypothalamus, thalamus and brainstem. There were however, some notable mismatches. Autoradiographic binding was high to very high in the median eminence, arcuate nucleus, suprachiasmatic nucleus and anterodorsal thalamic nucleus, whereas these areas were only poorly immunostained. Conversely, the septum showed moderate autoradiographic binding, but very prominent immunostaining of neurons in its dorsolat‐eral aspect. Using the immunohistochemical technique IGF‐II receptors were localized to specific neuronal groups such as the mitral cells of the olfactory bulb, Purkinje cells of the cerebellum and neurons in the red nucleus. Fibre pathways were not labelled by either technique.

We conclude that IGF‐II/M6P receptors are widespread throughout rat brain, specifically in neurons and blood vessels, with a similar, but distinct distribution to IGF‐I and insulin receptors. Many of these regions have in common high rates of metabolic and synthetic activity, which may be mediated by IGF‐II/M6P and their receptors.
Norepinephrine Inhibits Vasopressin‐Stimulated Flank Marking in the Syrian Hamster by Acting Within the Medial Preoptic‐Anterior Hypothalamus
Authors: Whitman D. Carol , Hennessey Ann C. , Albers H. Elliott
Published: September 2006

Syrian hamsters exhibit a form of scent marking behavior called flank marking. Flank marking, which is elicited during social contact with other hamsters and by the odors of other hamsters, communicates socially important information such as mate choice and dominance status. Vasopressinergic activity within the medial preoptic‐anterior hypothalamic continuum (MPOA‐AH) is essential for the expression of flank marking. In female hamsters, an inverse relationship exists between the expression of flank marking and sexual receptivity during the 4‐day estrous cycle. Since norepinephrine (NE) appears to facilitate sexual receptivity, the present study investigated whether NE might have an inhibitory effect on flank marking by acting on Vasopressinergic activity within the MPOA‐AH. Microinjection of 9.0 μM arginine vasopressin (AVP) into the MPOA‐AH stimulated high levels of flank marking. Microinjection of 9.0 μM AVP combined with NE in concentrations of 4.0, 0.4 or 0.2 nM, drastically reduced or eliminated flank marking. In contrast, AVP in combination with 0.09, 0.04 or 0.004 nM NE produced no significant reductions in flank marking. In addition, microinjection of 9.0 μM AVP, in combination with epinephrine (4.0 nM), but not dopamine (4.0 nM), serotonin (4.0 nM) or neuropeptide Y (900 μM), significantly reduced AVP‐induced flank marking. In male hamsters, microinjection of NE (4.0 nM) combined with AVP (9.0 μM) into the MPOA‐AH was not found to inhibit AVP‐stimulated flank marking. These results suggest that NE is involved in regulating the expression of flank marking during the estrous cycle by acting on Vasopressinergic activity within the MPOA‐AH.
Colocalization of Fos‐ and Glucocorticoid Receptor‐Like Immunoreactivities in the Rat Amygdaloid Complex After Immobilization Stress
Authors: Honkaniemi Jari , Fuxe Kjell , Rechard Leena , Koistinaho Jari , Isola Jorma , Gustafsson Jan‐Åke , Okret Sam , Pelto‐Huikko Markku
Published: September 2006

In the present paper we demonstrate the effect of immobilization stress on c‐fos‐like immunoreactivity (Fos‐LI) in the rat amygdaloid complex. Furthermore, since the subnuclei of the amygdaloid complex contain numerous glucocorticoid receptor‐immunoreactive (GR‐IR) neurons, we also studied the possible colocalization of GR‐ and Fos‐LI ie. Fos‐Lls and the action of a synthetic glucocorticoid, dexamethasone, and an anti‐glucocorticoid, RU 38486, on Fos‐LI. Immobilization stress caused a remarkable increase in the number of the Fos‐IR neurons in all the subnuclei of the amygdaloid complex except in the lateral nucleus. The majority of Fos‐IR neurons also contained GR‐LI, with the highest colocalization in the central amygdaloid nucleus. A similar induction of Fos‐LI after immobilization was seen in the hypothalamic paraventricular nucleus and almost all the Fos‐IR neurons in this nucleus also exhibited GR‐LI. Treatments with dexamethasone or RU 38486 prior to stress did not have any marked effect on Fos‐LI when compared to stress alone. The present findings suggest that Fos may function as a transcriptional regulator in the amygdaloid complex after stress and affect the synthesis of neurotransmitters and receptors in the amygdaloid neurons. Since we did not observe any effect of dexamethasone or RU 38486 on Fos‐LI, it is likely that glucocorticoids do not directly regulate the expression of the c‐fos gene or the formation of Fos protein. In view of the fact that Fos is capable of forming a stabile complex with GR and repress the transactivational capacity of GR, Fos may inhibit the negative feedback effect of circulating glucocorticoids and thus maintain elevated plasma glucocorticoid levels in stress.
Desensitization of Pituitary Cells by Gonadotrophin‐Releasing Hormone or its Analogues in the Superfusion System: Different Pattern for Males and Females
Authors: Horvath Judit E. , Kéri† György , Seprödi Attila , Teplán István , Flerkó Béla
Published: September 2006

The effect of a 6‐h infusion of gonadotrophin‐releasing hormone (GnRH) or its analogues on dispersed anterior pituitary cells from male or female rats was investigated. The cells were stimulated with 3‐min pulses of K+ and GnRH. Thereafter GnRH (1 nM) or GnRH analogues ([D‐Trp6]GnRH‐ethylamide ([D‐Trp6]GnRH, 50 pM), [D‐Phe6, Gln8]GnRH‐ethylamide (Folligen, 100 pM) and [Asu6]GnRH‐ethylamide ([Asu6]GnRH, 33 pM)) were applied for 6 h. In cells from female rats this treatment resulted in a 20‐fold increase in luteinizing hormone (LH) secretion during the first 90‐min period of the 6‐h incubation. Following this a gradual decrease in LH release occurred, and during the fourth 90‐min period the amount of LH secreted was only one‐third or less of the initial value. The pituitary cells of male rats responded to the same treatment with only a 7‐fold rise of LH secretion during the first period. In the second 90‐min of the 6‐h incubation a 20% to 30% increase was observed. Even in the fourth 90‐min period the amount of LH secreted was two‐thirds or more greater than that of the first 90‐min period. When using 10‐fold greater concentrations of the same peptides in males, the increase in hormone secretion in the second 90‐min was not seen and the hormone release decreased to around 50%. We found definite differences in the responses of male and female rat pituitary cells to the 6‐h infusion of GnRH or its analogues: the initial amplitude of the response in females was higher but desensitization was stronger. In males, the initial response was weaker; however, even using doses one magnitude greater, the level of desensitization did not reach the values obtained in females. The results were similar both with GnRH and the analogues. The responses to 3‐min K+ and GnRH stimuli given after the 6‐h incubation were strongly reduced in cells from female rats compared to the initial responses; however, in cells from male rats the reaction was higher or unchanged. The ratio of LH released by the final K+ stimulus relative to the actual LH content of the cells decreased in females but increased in males. Our data show that the differences between the pattern of desensitization in cells from male and female rats may be caused by the differences in the amount and ratio of immediately releasable hormone and the hormone replenishment into these pools.
Galanin Message‐Associated Peptide‐Like Immunoreactivity in the Nervous System of the Blowfly: Distribution and Chromatographic Characterization
Authors: Lundquist C. Tomas , Rökaeus Åke , Nässel Dick R.
Published: September 2006

Galanin message‐associated peptide (GMAP) is a flanking peptide in mammalian preprogalanin located C‐terminally of galanin (GAL). GMAP‐like immunoreactive (LI) material in the brain of the blowfly Phormia terraenovae was analysed by radioimmunoassay combined with reversed‐phase high‐performance liquid chromatography and immunocytochemistry and compared to GAL‐LI material. A sensitive radioimmunoassay, developed against a species‐conserved portion of mammalian GMAP (synthetic porcine GMAP(19–41)amide), was applied to serially diluted blowfly head extracts. High‐performance liquid chromatography combined with radioimmunoassay showed that the GMAP‐LI material eluted as several different components with one major component coeluting with the synthetic GMAP fragment. One GMAP‐LI peak co‐eluted with a GAL‐LI component of the extract. By immunocytochemistry it was shown that a distinct set of GMAP‐LI neurons and neurosecretory cells is present in the blowfly brain and thoracico‐abdominal ganglion. About 150 GMAP‐LI cell bodies were found in the brain, distributed in the protocerebrum, tritocerebrum and suboesophageal ganglion. Several hundred GMAP‐LI cell bodies were detected in the medulla of the optic lobe. In the fused thoracico‐abdominal ganglion there are about 70 GMAP‐LI cell bodies distributed in a segmental fashion. Several of the GMAP‐LI neurons also contain GAL‐LI material whereas some do not. In addition, there are GAL‐LI neurons that do not react with the GMAP antiserum. Some of the GMAP‐LI interneurons and neurosecretory cells could be traced in detail enabling a resolution of putative sites of action of the peptide.