Distribution Of Gonadotropin-releasing Hormone Research Articles

104 Distribution of Gonadotropin-Releasing Hormone and Kisspeptin Neurons in the Preoptic Area and Hypothalamus During the Estrous Cycle in Cows

Gonadal steroids hormones indirectly regulate gonadotropin-rleasing hormone (GnRH) secretion. Kisspeptin (Kp) co-expresses steroid receptors and modulates GnRH release. The objective of the study was to characterise the number and proportion of GnRH and Kp immunoreactive cells and their association in the preoptic area (POA) and hypothalamus during different phases of the oestrous cycle in cows. Daily ovarian ultrasonography was performed to detect follicle development and ovulation (Day 0) after prostaglandin treatment. On Day 5, cows were assigned randomly to the following groups: proestrus (n = 2), metestrus (n = 2) or diestrus (n = 3). Cows in the diestrus group were killed on Day 8. Cows in the proestrus and metestrus groups were given luteolytic dose of prostaglandin on Day 5.5 and Day 6 and were killed on Day 7 and 24 h after the ensuing ovulation, respectively. Cow heads were perfused with 4% paraformaldehyde via the carotid arteries to fix the brain in situ. The brain-stem (rostral portion of the POA to the mamillary body) was isolated by dissection and placed in 4% paraformaldehyde for 48 h. Following cryoprotection, the tissue block containing the POA and hypothalamus was frozen at –80°C and sectioned serially at a thickness of 50 mm using a cryostat microtome. Every 20th free-floating section was processed for double labelling using 2 sequential immuno-peroxidase reactions and ABC staining; Kp was immuno-labelled with Nickel-DAB at a dilution of 1:10,000 rabbit anti-kisspeptin (AC566, INRA, France), and GnRH was stained with DAB using 1:40,000 rabbit anti-GnRH (LR-5, Dr Benoit). The numbers of neuron cell bodies and fibres were recorded in different areas of the POA and the hypothalamus by brightfield microscopy using 10× and 40× objective lenses. Data were compared among groups by ANOVA. Major aggregations of Kp cells were localised in the mPOA, OVLT, and ARC. Overall, the number of Kp cells was higher in the metestrus v. diestrus group (719 ± 94 v. 378 ± 8; P = 0.01), but was similar to the proestrus group (558 ± 9). The number of Kp cells in the POA (mPOA, OVLT) tended to be higher in the metestrus v. diestrus group (395 ± 56 v. 147 ± 44; P = 0.06), and was intermediate in the proestrus group (206 ± 6). The number of Kp cells in the ARC did not differ among groups (metestrus 310 ± 26, diestrus 206 ± 53, proestrus 321 ± 99; P = 0.4). The number of GnRH cells bodies was not different among groups (metestrus 40 ± 3, diestrus 50 ± 9, proestrus 43 ± 8; combined; P = 0.8), and the distribution was higher in the POA (metestrus 25 ± 2, diestrus 30 ± 3, proestrus 33 ± 2) than hypothalamus. The proportion of GnRH cells in apposition to Kp fibres tended to be highest in the proestrus v. metestrus and diestrus groups (50.5 ± 1% v. 34.1 ± 9% and 31.4 ± 3%; P = 0.09). In conclusion, the number of Kp immunoreactive cells, but not GnRH cells, present in the POA and hypothalamus changed among different phases of the oestrous cycle due primarily to an increase in number of Kp cells in POA during metestrus. The proestrous phase was associated with an increase in apposition between Kp fibres and GnRH cells.

Hypothalamic neuroendocrine functions in rats with dihydrotestosterone-induced polycystic ovary syndrome: effects of low-frequency electro-acupuncture.

Adult female rats continuously exposed to androgens from prepuberty have reproductive and metabolic features of polycystic ovary syndrome (PCOS). We investigated whether such exposure adversely affects estrous cyclicity and the expression and distribution of gonadotropin-releasing hormone (GnRH), GnRH receptors, and corticotrophin-releasing hormone (CRH) in the hypothalamus and whether the effects are mediated by the androgen receptor (AR). We also assessed the effect of low-frequency electro-acupuncture (EA) on those variables. At 21 days of age, rats were randomly divided into three groups (control, PCOS, and PCOS EA; n = 12/group) and implanted subcutaneously with 90-day continuous-release pellets containing vehicle or 5α-dihydrostestosterone (DHT). From age 70 days, PCOS EA rats received 2-Hz EA (evoking muscle twitches) five times/week for 4–5 weeks. Hypothalamic protein expression was measured by immunohistochemistry and western blot. DHT-treated rats were acyclic, but controls had regular estrous cycles. In PCOS rats, hypothalamic medial preoptic AR protein expression and the number of AR- and GnRH-immunoreactive cells were increased, but CRH was not affected; however, GnRH receptor expression was decreased in both the pituitary and hypothalamus. Low-frequency EA restored estrous cyclicity within 1 week and reduced the elevated hypothalamic GnRH and AR expression levels. EA did not affect GnRH receptor or CRH expression. Interestingly, nuclear AR co-localized with GnRH in the hypothalamus. Thus, rats with DHT-induced PCOS have disrupted estrous cyclicity and an increased number of hypothalamic cells expressing GnRH, most likely mediated by AR activation. Repeated low-frequency EA normalized estrous cyclicity and restored GnRH and AR protein expression. These results may help explain the beneficial neuroendocrine effects of low-frequency EA in women with PCOS.

Distribution of gonadotropin-releasing hormone (GnRH) by in situ hybridization in the tunicate Ciona intestinalis

Gonadotropin releasing hormone (GnRH) is the key hypothalamic neurohormone that is critical in its role of reproduction in all vertebrates. There are currently twenty-four known forms of GnRH that have been identified, 14 in vertebrates and 10 in invertebrates. In tunicates, the primary structure of nine forms have been identified, all of which have been shown to stimulate gamete release. However, the distribution and function of the various GnRH peptides in tunicates have not been fully examined. Therefore, the objective of this study was to determine tissue specific expression of Ci-gnrh-1 and Ci-gnrh-2 in an adult tunicate, Ciona intestinalis, using reverse transcriptase-polymerase chain reaction (RT-PCR) and in situ hybridization. To examine the expression of the two GnRH genes, total RNA and genomic DNA were isolated from whole animals. Total RNA from neural tissue (cerebral ganglion and neural gland), testis, ovary, heart, and hepatic organ were also isolated. Results from RT-PCR indicated both forms are only expressed in the neural tissue. We extended these studies using fluorescent dual label in situ hybridization. GnRH expression was confirmed to be in the cerebral ganglion bordering the neural gland. These current data along with previous studies suggest that GnRH may be involved in reproduction in the protochordate.

GnRH-immunoreactive fiber changes with unilateral amygdala-kindled seizures

Purpose: To assess whether unilateral amygdala seizures are associated with a change in the number and lateral distribution of gonadotropin releasing hormone (GnRH)-staining fibers in the ventromedial hypothalamus of female rats. Methods: The study compared three groups of female rats: (1) amygdala seizures induced by focal injection of kainic acid (KA); (2) saline injected controls; and (3) naı̈ve controls. The animals were sacrificed at 4 weeks in the diestrus phase. GnRH fibers were counted in the ventromedial hypothalamus and compared among groups. Results: GnRH fiber counts were significantly lower in KA than saline and naı̈ve animals ipsilaterally but not contralaterally. Conclusions: This finding may support a potential mechanism by which (1) temporolimbic epilepsy may promote the development of reproductive endocrine disorders and (2) the laterality of the epilepsy may influence the particular nature of the reproductive endocrine disorder.

Ontogeny and distribution of gonadotropin-releasing hormone (GnRH) neuronal systems in the brain of the cichlid fish Cichlasoma dimerus.

Using immunocytochemistry we have described the distribution and ontogeny of three distinct gonadotropin-releasing hormone (GnRH) neural systems, emphasizing the analysis during the period of sex differentiation in the South American cichlid fish Cichlasoma dimerus. In the forebrain a group of neurones immunoreactive to salmon GnRH that formed clusters in the nucleus olfacto retinalis (NOR), was located at the junction of the olfactory bulb and the telencephalon. These neurones differentiated 3 days after fertilization from the olfactory placodes. GnRH immunoreactive neurones along the olfactory nerves through the rostrobasal olfactory bulb were observed on day 4 and at the NOR on day 10. A group of neurones immunoreactive to chicken GnRH II was seen in the dorsal midbrain tegmentum. They originate from the ventricular ependyma between days 5 and 6. These neurones remained close to blood vessels throughout development. Between days 22 and 30 a group of neurones immunoreactive to seabream GnRH was detected in the anterior basal preoptic area. GnRH innervation of the pituitary was detected after the differentiation of preoptic neurones and in coincidence with gonadal differentiation. We hypothesize that the GnRH neural systems have three distinct embryonic origins. Furthermore, we show that the NOR and the midbrain GnRH neurones might have functions other than gonadal development, whereas the preoptic GnRH neurones in C. dimerus might be associated with gonadal sex differentiation.

Identification and neuroanatomical distribution of immunoreactivity for mammalian gonadotropin-releasing hormone (mGnRH) in the brain and neural hypophyseal lobe of the toad Bufo arenarum.

The presence and distribution of gonadotropin-releasing hormone (GnRH) in sexually mature specimens of Bufo arenarum was studied by reverse phase/high performance liquid chromatography (RP-HPLC) combined with radioimmunoassay and immunocytochemistry. The analysis of brain extracts with RP-HPLC followed by radioimmunoassay with PBL#45 antiserum showed the presence of only one peak with immunoreactivity for GnRH (ir-GnRH) having the chromatographic and immunological characteristics of mammalian GnRH (mGnRH). This peak was further analyzed with two mGnRH-specific antisera, EL-15 and m1076, yielding serial dilution displacement curves parallel to those obtained with the mGnRH synthetic standard. Immunocytochemical results with the monoclonal antibody LRH13 showed the presence of a terminal nerve-septo-preoptic system with neurons and fibers distributed from the olfactory bulb, septal area, and anterior preoptic area toward the hypothalamus and hypophyseal neural lobe. The main group of ir-GnRH fibers and neurons was identified in the anterior preoptic area. These neurons appear to be the origin of fibers that, after surrounding the preoptic recess, border the dorsal surface of the optic chiasma, extend through the infundibulum, traverse the external layer of the median eminence, and end in the pars nervosa.

Investigation of the contribution from peripheral GnRH-like immunoreactive ‘neuroblasts’ to the regenerating central nervous system in the protochordateCiona intestinalis

The neural ganglion of the ascidian Ciona intestinalis regenerates in its entirety within a few weeks of ablation. Here we investigate the role of gonadotropin–releasing hormone–like immunoreactive (GnRH–li) cells in regeneration. Immunocytochemical studies show that in addition to a previously described plexus of GnRH–like neurones located in association with the dorsal strand, the normal adult brain contains GnRH–li neurones. These are predominantly localized to the ventral cortical rind at the posterior of the ganglion. Following ablation, non–process bearing GnRH–li cells appear in the regenerating area within two days. By day 8 post–ablation, process bearing GnRH–li cells are detected close to the regenerating brain. The number of these cells increases at later stages. GnRH–li cells are first detected within the regenerating brain at 14 days post–ablation and their number subsequently increases. These cells are initially concentrated along the ventral surface of the regenerating brain near to the dorsal strand. Double labelling studies with bromodeoxyuridine show that none of the GnRH–li cells are labelled at any stage of regeneration. The data are consistent with a sub–population of the new neurones being derived from GnRH–li neuroblasts born prior to ablation, which migrate from the dorsal strand complex into the regenerating ganglion.

Reproduction in the Mexican Leaf Frog,Pachymedusa dacnicolor: VI. Presence and Distribution of Multiple GnRH Forms in the Brain

The presence and distribution of gonadotropin-releasing hormone (GnRH) has been investigated in the Mexican leaf frog,Pachymedusa dacnicolor,brain during development and in the adult. The ontogenetic pattern of GnRH neurons illustrates their extracranial as well as intracranial sites. Immunohistochemical analysis indicates that GnRH-immunoreactive neurons appear during the metamorphic climax. They are located in the mesencephalon and subsequently other GnRH neurons appear in the peripheral terminal nerve and anterior preoptic area of the brain. Use of specific antisera and homologous combined with heterologous preabsorption tests indicate that mammalian and chicken GnRH-II-like peptide-containing neurons are differentially located within the brain, the former in the anterior preoptic area and peripheral terminal nerve and the latter in the mesencephalon. HPLC and RIA data suggest the presence of three forms of immunoreactive GnRH in theP. dacnicolorbrain. A mammalian GnRH-like molecule and a chicken GnRH-II-like molecule are present. A third form, suspected to be [hydroxyproline9]mGnRH elutes before the mammalian GnRH.

Nature and Distribution of Gonadotropin-Releasing Hormone (GnRH) in the Brain, and GnRH and GnRH Binding Activity in Serum of the Spotted Dogfish Scyliorhinus canicula

The distribution of different molecular forms of gonadotropin-releasing hormone (GnRH) in the brain and serum of the spotted dogfish, Scyliorhinus canicula, was investigated by an indirect immunofluorescence method, using antisera against salmon (s-), chicken-II (cII-) and mammalian (m-) GnRHs, and by reverse-phase high-performance liquid chromatography (RP-HPLC) coupled with radioimmunoassays. Five GnRH molecular forms were demonstrated on the basis of the retention time in the RP-HPLC system. The characterestics of four of these GnRH peptides are consistent with those of m-, cII-, dogfish (df-), and sGnRH. The fifth form appears to be novel. Immunoreactive sGnRH structures were confined to the diencephalon; whereas cIIGnRH and mGnRH were found in the telecephalon and diencephalon, cIIGnRH-and dfGnRH-like molecules were detected in the serum. Moreover, specific, low-affinity GnRH binding protein (GnRH-BP) was found in the serum of the spotted dogfish. The binding of [ 125I]sGnRHA to the serum GnRH-BP was dependent on incubation time, equilibrium being reached within 1 hr at 4° binding was rapid and incompletely reversible. Scatchard analysis yielded a linear plot with a K d of 7.9 × 10 -7 M. The presence of a Gnrh-BP in spotted dogfish serum suggests a probable action of GnRH via the general circulation.

Distribution of gonadotropin‐releasinghormone immunoreactivity in the brain of the pacific hagfish, Eptatretus stouti (craniata: Myxinoidea)

The distribution of gonadotropin-releasing hormone (GnRH)-like immunoreactivity in the brain of a myxinoid, the Pacific hagfish (Eptatretus stouti), was investigated via immunohistochemistry, including the use of six different antisera. In the diencephalon, immunoreactive cell bodies were found in two systems: the infundibular hypothalamus, a neuromodulatory nucleus with diffuse projections of varicose fibers to most areas of the brain, and a primarily preoptic system of putatively hypophysiotropic neurons that projects to the neurohypophysis. Some potential neurovascular and CSF contacts were also identified. These findings are consistent with those of similar studies in other craniates and suggest that a preoptic hypophysiotropic system may be present in all craniates. We therefore tentatively accept the homology of this system in hagfish and vertebrates. The homology of the distributed hypothalamic system is more dubious. It may be homologous to a caudal GnRH system of modulatory neurons found in many vertebrates. Antiserum PBL-49 displays a differential affinity for the two systems, indicating that the two systems differ in the amount or identity of the immunoreactive substance. We suggest that the two systems have distinct functions in hagfish. The primitive function of GnRH-like molecules in craniates may have thus been both neuromodulatory and hypophysiotropic. These findings also indicate that the brain-pituitary axis of hagfish is more similar to that of vertebrates than has been previously suggested.

Immunohistochemical double-labeling study of gonadotropin-releasing hormone (GnRH)-immunoreactive cells and oxytocin-immunoreactive cells in the preoptic area of the dwarf gourami, Colisa lalia

The distribution of gonadotropin-releasing hormone (GnRH)-immunoreactive (ir) cells in the preoptic area (POA) of the dwarf gourami, Colisa lalia, was immunohistochemically studied. These neurons form cell columns on both sides of the common ventricle, and their axons project to the pituitary gland by forming distinct bundles. Also examined was the distribution of isotocin (IST) cells in the POA by using an anti-oxytocin (OXT) serum which has been proven to crossreact with IST. These two kinds of immunoreactive cells were distributed quite similarly in the POA. However, by using an immunofluorescence double-labeling method on thinner sections we found that a population of small IST cells in the ventral POA were also immunoreactive to GnRH, but that large IST cells in the dorsal POA were not immunoreactive to GnRH, and small GnRH-ir cells in the most ventral POA were not immunoreactive to the OXT antiserum. In the pituitary gland, GnRH-ir fibers were found in both the neurohypophysis and proximal pars distalis, but IST fibers were found only in the neurohypophysis.

The Nature and Distribution of Gonadotropin-Releasing Hormones in Brains and Plasma of Ranid Frogs

Highly specific antisera and reversed-phase HPLC were used to examine the nature and distribution of gonadotropin-releasing hormone (GnRH) in the brains and plasma of three species of Rana frog previously reported to vary in the types and brain distributions of GnRH. The three species examined, Rana pipiens, Rana esculenta, and Rana ridibunda, exhibit differences in total concentration and relative concentrations of GnRH in brain areas, but all three contain the mammalian and chicken-II forms of GnRH (mGnRH and cGnRH-II). Both forms were found in the telencephalon and diencephalon, but mGnRH is consistently the most abundant form in the preoptic-hypothalamic area (e.g., ratios of mGnRH:cGnRH-II > 3:1 in hypothalamus-median eminence), whereas, cGnRH-II is the most abundant in the telencephalon and the only form measured in the cerebellum and medulla. The total content of cGnRH-II in the whole brain is about 1.5-2 times higher than that of mGnRH, due to the larger size of the areas outside the preoptic-hypothalamic area. These general patterns were the same for adults and juveniles. We found no evidence of a third form of GnRH corresponding to salmon GnRH in hypothalamus or other brain areas of R. esculenta as previously reported. These analyses also revealed the presence of both forms of GnRH in plasma draining the hypothalamic area, but the concentration of cGnRH-II is relatively higher than that in the corresponding hypothalamic tissue, suggesting differentially greater release or slower degradation of this form. These results do not support the conclusion that the ranid frogs are highly variable in the nature and distribution of GnRH in the brain, although they suggest that both forms of GnRH are potentially involved in the direct regulation of pituitary gonadotropes.

The distribution of gonadotropin-releasing hormone (GnRH) neurons and fibers throughout the chick brain (Gallus domesticus)

Nerve fibers and perikarya containing gonadotropin-releasing hormone (GnRH-like) immunoreactivity were investigated in the brain of the three-week-old chick. Gallus domesticus using the technique of immunocytochemistry. Six major groups of perikarya were found to include the olfactory bulb, olfactory tubercle/lobus parolfactorius, nucleus accumbens, septal preoptic hypothalamic region (three sub-nuclei), lateral anterior thalamic nucleus and in and about the oculomotor complex. The immunostaining was unusual in the latter group, suggesting that the neurons may contain a GnRH-II like material. Immunoreactive fibers for GnRH were found throughout the entire brain extending from the olfactory bulbs to the caudal brainstem. Two anatomical areas, not emphasized in the past literature, which had distinct GnRH-like immunoreactivity, included the lateral anterior thalamic nucleus and the preoptic recess. The former included a group of GnRH perikarya that is also known to be a retino-recipient area while the latter contained neuronal terminals some of which appeared to be contacting the cerebrospinal fluid of the preoptic recess. An attempt was made to list all anatomical structures that contained or were juxta-positioned to sites that displayed immunoreactive perikarya and fibers including circumventricular organs.

Immunocytochemical Localization of Gonadotropin and Gonadotropin Releasing Hormone in Pituitary and Brain of the Gilthead Seabream, Sparusaurata L. (Teleost)

Characterization of gonadotropic cells in the pituitary gland of male gilthead seabream (Sparus aurata) was performed at the light- and electron microscopic level with the peroxidase-anti peroxidase and the immunogold technique, respectively. The cells were identified with antibodies raised against three different teleost gonadotropins (carp α, β-GTH, carp β-GTH, catfish α, β-GTH). Immunostained cells were mainly located in the ventral and dorsal areas of the proximal pars distalis (PPD), with a few cells in the pars intermedia (PI). At the light microscopic level the gonadotrops showed a variable granulation and they were often strongly vacuolated. In the electron microscope the latter appeared to be caused by strongly dilated rough endoplasmic reticulum (RER). Immunogold labeling was restricted to all secretory granules and most of the larger globules. Different fixation and embedding procedures were tested: an optimal combination of good ultrastructure and intense immunogold labeling was obtained when osmium tetroxide postfixation was omitted and with embedding in 'Epon. The distribution of gonadotropin-releasing hormone (GnRH) immunoreactivity was studied with antibodies against mammalian LHRH and salmon GnRH. Only with anti-mammalian LHRH immunostained perikarya were observed in the ventral preoptic area. With both antisera immunostained fibers were seen directed towards the pituitary stalk, as well as in the neurohypophysis, whereas no such fibers were present in the PPD. Immunostained fibers were also found in several other regions of the brain.

The nervus terminalis in the mouse: light and electron microscopic immunocytochemical studies.

The distribution of gonadotropin-releasing hormone (GnRH)-containing neurons and fibers in the olfactory bulb was studied with light and electron microscopic immunohistochemistry in combination with retrograde transport of "True Blue" and horseradish peroxidase and lesion experiments. GnRH-positive neurons are found in the septal roots of the nervus terminalis, in the ganglion terminale, intrafascicularly throughout the nervus terminalis, in a dorso-ventral band in the caudal olfactory bulb, in various layers of the main and accessory olfactory bulb, and in the basal aspects of the nasal epithelium. Electron microscopic studies show that the nerve fibers in the nervus terminalis are not myelinated and are not surrounded by Schwann cell sheaths. In the ganglion terminale, "smooth" GnRH neurons are seen in juxtaposition to immunonegative neurons. Occasionally, axosomatic specializations are found in the ganglion terminale, but such synaptic contacts are not seen intrafascicularly in the nervus terminalis. Retrograde transport studies indicate that certain GnRH neurons in the septal roots of the nervus terminalis were linked to the amygdala. In addition, a subpopulation of nervus terminalis-related GnRH neurons has access to fenestrated capillaries whereas other GnRH neurons terminate at the nasal epithelium. Lesions of the nervus terminalis caudal to the ganglion terminale result in sprouting of GnRH fibers at both sites of the knife cut. The results suggest that GnRH in the olfactory system of the mouse can influence a variety of target sites either via the blood stream, via the external cerebrospinal fluid or via synaptic/asynaptic contacts with, for example, the receptor cells in the nasal mucosa.

The effects of orchidectomy and replacement therapy on the ultrastructure and gonadotropin-releasing hormone content of the median eminence of the rat.

The effects of 2 weeks of orchidectomy and replacement therapy with testosterone upon the content and distribution of gonadotropin-releasing hormone (GnRH) in the median eminence were determined by means of radioimmunoassay and electron microscopy. Photographic montages were prepared from electron micrographs of the lateral median eminence at the point of deepest invagination of the tuberoinfundibular sulcus. Morphometric analysis of photographs of tissues immunohistochemically stained for GnRH was performed to determine changes in the volume density of GnRH-containing axon profiles following the experimental treatments. A decrease in GnRH content after orchidectomy was observed both by morphometric analysis of axon volume density and radioimmunoassay of total GnRH content. Testosterone treatment of orchidectomized animals prevented the postorchidectomy loss of GnRH. Morphometric analysis of conventional electron micrographs revealed an increase in the number of axons containing no dense-core vesicles following orchidectomy, but no decrease in volume density of the neuropil. The results indicate that the change in volume density of immunostained axons was related to the loss of immunostainable dense-core vesicles and not to a change in the size or number of axons. The area corresponding to the location of the highest concentration of GnRH-containing axons was observed to be largely avascular and separated from the vessels of the tuberoinfundibular sulcus by a "border zone" composed of glial foot processes. The unique morphology of the GnRH area has suggested the name "compact zone" to distinguish it from the palisade zone with which it is continuous medially. GnRH axons in this region are probably part of a tract extending farther caudally rather than a terminal field.

Gonadotropin-releasing hormone-immunoreactive neuronal structures in the brain and pituitary of the African catfish, Clarias gariepinus (Burchell).

The distribution of gonadotropin-releasing hormone (GnRH) immunoreactivity was studied in the African catfish, Clarias gariepinus, by means of immunofluorescence and immunoperoxidase techniques. Immunoreactive neurons were found throughout the preoptic nucleus (NPO). However, only a portion of the secretory perikarya in the NPO showed a positive reaction by use of an anti-LHRH serum. Numerous immunoreactive fibres were found to enter the pituitary and to terminate in its proximal pars distalis, the site of concentration of the gonadotropic cells. Since GnRH is present in the brain and pituitary of the African catfish, the lack of spontaneous ovulation in captivity is apparently due to an insufficient release of GnRH.

Immunohistochemical localization of gonadotropin-releasing hormone (GnRH) in the brain and infundibulum of the sheep.

The distribution of gonadotropin-releasing hormone (GnRH) was studied in the brain and infundibulum (INF) or median eminence of sheep utilizing a peroxidase-antiperoxidase immunohistochemical method. This procedure utilized a specific antiserum generated against GnRH conjugated to bovine serum albumin. In the rostral INF, the greatest concentration of GnRH positive axons was found in the medial region, mostly in the external layer dorsal to the hypophysial portal plexus. In the intermediate portion of the INF, the hormone was mainly observed in the external layer at the more dorsolateral areas ventral to the tuberoinfundibular sulcus. GnRH was generally located medially in the caudal portion of the INF and dorsomedially in the rostral infundibular stalk. Substantial amounts of reaction product were also noted in the internal layer throughout the entire rostrocaudal extent of the INF. The hormone was localized in axons throughout the brain from the septal and medial preoptic areas to the mammillary bodies. GnRH-positive perikarya were scattered in various regions of the infundibular (arcuate) and for the first time in the ventromedial nuclei of sheep hypothalamus. Preabsorption of the specific antiserum with synthetic GnRH abolished staining in both axons and perikarya, whereas preabsorption with thyrotropin-releasing hormone, oxytocin, arginine-vasopressin, and adrenocorticotrophic hormone did not affect staining intensity.

Distribution of gonadotropin-releasing hormone in the mouse brain as revealed by immunohistochemistry.

The distribution of gonadotropin-releasing hormone (GnRH) was studied immunohistochemically in the brain of the adult mouse with the peroxidase-antiperoxidase (PAP) method. Primary antisera were prepared against unconjugated synthetic GnRH and GnRH conjugated to limpet hemocyanin or bovine serum albumin. GnRH was localized in the organum vasculosum of the lamina terminalis (OVLT), with the greatest amount being found ventral to the cephalic end of the third ventricle. In the cephalic region of the median eminence, it was concentrated bilaterally in longitudinal bands located dorsal to the tuberoinfundibular sulci. More caudally, near the junction of the infundibulum with the brain, GnRH accumulated over the apex of the tuberoinfundibular sulci, with several foci being scattered from this region medially to the ependyma of the third ventricle. The greatest aggregation of GnRH occurred in the psotinfundibular median eminence in an area extending from the floor of the third ventricle to the ventral surface of the brain. In the caudal median eminence, GnRH was arranged in a narrow ventral band that crossed the midline. GnRH appeared to be located in axonal processes and terminals of the OVLT and median eminence; the structures observed were granular. GnRH was not localized within the neuronal cell bodies of any hypothalamic nuclei. When one antiserum to conjugated GnRH was used at high concentration, the cytoplasm of ependymal cell bodies and tanycyte processes was stained more intensely than the general background, but, with absorption and/or dilution of the antiseru, this staining was shown to be nonspecific.

Brain immunoreactive gonadotropin-releasing hormone in Huntington's chorea and in non-choreic subjects.

THERE is evidence which suggests that the release of gonadotropin-releasing hormone (GRH) is modulated by dopaminergic neurones1–3. Dopamine receptors seem to be hyper-responsive in Huntington's chorea4,5, and consequently the synthesis and secretion of GRH may be altered in this condition. For this reason we have compared the concentration of immunoreactive GRH in hypothalamic tissues taken postmortem from choreic and non-choreic (‘control’) subjects. The distribution of GRH in the human brain, and changes in concentration which occur with age have also been examined.