Cells In Arcuate Nucleus Research Articles

THE MORPHOLOGICAL AND FUNCTIONAL STATE OF THE ARCUATE NUCLEUS OF THE HYPOTHALAMUS OF RATS IN BURN INJURY DYNAMICS

Morphological and morphometric features of cells of arcuate nucleus of hypothalamus in dynamics of thermal burn trauma of skin are considered. The relevance of the study is due to the important role of reactive changes in the neuroendocrine regulatory complex in the pathogenesis of skin burn wounds. Since the hypothalamus is part of neuroendocrine cooperation, it shows the most pronounced signs of neuronal damage. In this regard, the aim of the study was to study qualitative and quantitative structural changes in the arcuate nucleus of the middle hypothalamus of rats in the simulation of thermal burn injury to the skin. Burn exposure was simulated in male nonlinear rats weighing 200-230 g by applying contact thermal trauma in the interblade region of the back. The functional activity of the arcuate nucleus of the hypothalamus was evaluated by quantitative analysis of the morphometric parameters of neurocyte nuclei because these organoids correlate with transcription intensity and can serve as a tool for assessing their functional state. To carry out quantitative analysis of histological preparations of hypothalamus, planimetric properties of neurons and their structures were determined: absolute values of neurons, such as: area of pericaryons, area of neurons nuclei, area of cytoplasm of neurons, nuclear-cytoplasmic ratio of neurons of arcuate nuclei of hypothalamus. On the basis of the obtained data on absolute values of neurons relative values were calculated - median area of pericaryons, median area of nuclei, median area of cytoplasm of pericaryons, median of nuclear-cytoplasmic ratio of neurons of arcuate nuclei of hypothalamus. The study revealed destructive transformations of this hypothalamic brain zone in all stages of inflammatory-regenerative reaction: on 2, 4, 7, 10 days after burn exposure. Comparative analysis of hypothalamus preparations of laboratory animals showed characteristic destructive changes in arcuate nuclei in conditions of thermal skin injury, which reflected the development of processes of reversible and irreversible damage to neurons. The detected features are generally consistent with existing ideas about the disruption of morphofunctional organization of neurons in this zone due to the implementation of adaptive mechanisms characterized by activation of local and systemic compensatory-restorative processes, focal gliosis and development of edema changes of nerve cells.

Endogenous amylin contributes to birth of microglial cells in arcuate nucleus of hypothalamus and area postrema during fetal development.

Amylin acts in the area postrema (AP) and arcuate nucleus (ARC) to control food intake. Amylin also increases axonal fiber outgrowth from the AP→nucleus tractus solitarius and from ARC→hypothalamic paraventricular nucleus. More recently, exogenous amylin infusion for 4 wk was shown to increase neurogenesis in adult rats in the AP. Furthermore, amylin has been shown to enhance leptin signaling in the ARC and ventromedial nucleus of the hypothalamus (VMN). Thus, we hypothesized that endogenous amylin could be a critical factor in regulating cell birth in the ARC and AP and that amylin could also be involved in the birth of leptin-sensitive neurons. Amylin+/- dams were injected with BrdU at embryonic day 12 and at postnatalday 2; BrdU+ cells were quantified in wild-type (WT) and amylin knockout (KO) mice. The number of BrdU+HuC/D+ neurons was similar in ARC and AP, but the number of BrdU+Iba1+ microglia was significantly decreased in both nuclei. Five-week-old WT and KO littermates were injected with leptin to test whether amylin is involved in the birth of leptin-sensitive neurons. Although there was no difference in the number of BrdU+c-Fos+ neurons in the ARC and dorsomedial nucleus, an increase in BrdU+c-Fos+ neurons was seen in VMN and lateral hypothalamus (LH) in amylin KO mice. In conclusion, these data suggest that during fetal development, endogenous amylin favors the birth of microglial cells in the ARC and AP and that it decreases the birth of leptin-sensitive neurons in the VMN and LH.

Switching from high-fat diet to foods containing resveratrol as a calorie restriction mimetic changes the architecture of arcuate nucleus to produce more newborn anorexigenic neurons.

These days, obesity threatens the health for which one of the main interventions is calorie restriction (CR). Due to the difficulty of compliance with this treatment, CR mimetics such as resveratrol (RSV) have been considered. The present study compared the effects of RSV and CR on hypothalamic remodeling in a diet-switching experiment. C57BL/6 male mice received high-fat diet (HFD) for 4weeks, subsequently their diet switched to chow diet, HFD + RSV, chow diet + RSV or CR diet for a further 6weeks. Body weight, fat accumulation, hypothalamic apoptosis and expression of trophic factors as well as generation and fate specification of newborn cells in arcuate nucleus (ARC) were evaluated. Switching diet to RSV-containing foods leading to weight and fat loss after 6weeks. In addition, not only a significant reduction in apoptosis but also a considerable increase in production of newborn cells in ARC occurred following consumption of RSV-enriched diets. These were in line with augmentation of hypothalamic ciliary neurotrophic factor and leukemia inhibitory factor expression. Interestingly, RSV-containing diets changed the fate of newborn neurons toward generation of more proopiomelanocortin than neuropeptide Y neurons. The CR had effects similar to those of RSV-containing diets in the all-evaluated aspects besides neurogenesis in ARC. Although both RSV-containing and CR diets changed the fate of newborn neurons to create an anorexigenic architecture for ARC, newborn neurons were more available after switching to RSV-enriched diets. It can be consider as a promising mechanism for future investigations.

Expression and colocalization patterns of calbindin-D28k, calretinin and parvalbumin in the rat hypothalamic arcuate nucleus

Calcium binding proteins (CaBPs) form a diverse group of molecules that function as signal transducers or as intracellular buffers of Ca2+ concentration. They have been extensively used to histochemically categorize cell types throughout the brain. One region which has not yet been characterized with regard to CaBP expression is the hypothalamic arcuate nucleus, which plays a vital role in neuroendocrine control and the central regulation of energy metabolism. Using in situ hybridization and immunofluorescence, we have investigated the cellular distribution of the three CaBPs, calbindin-D28k (CB), calretinin (CR) and parvalbumin (PV) in the rat arcuate nucleus. Both mRNA and immunoreactivity was detected in the arcuate nucleus for CB – located in the medial aspects – and CR – located ventrolaterally. No PV mRNA was detected in the arcuate nucleus. Immunofluorescence results for PV were ambiguous; while one antibody detected a group of cell somata, a different antibody failed to visualize any arcuate nucleus cell profiles. Using double-labeling, neither of the examined CaBPs were observed in cells immunoreactive for the signaling molecules agouti gene-related protein, tyrosine hydroxylase, neurotensin, growth hormone-releasing hormone, somatostatin, enkephalin, dynorphin or galanin. We did, however, observe CB- and CR-immunoreactivity, in two distinct populations of neurons immunoreactive for the melanocortin peptide α-melanocyte-stimulating hormone. These data identify distinct subpopulations of arcuate neurons defined by their expression of CaBPs and provide further support for differentiation between subpopulations of anorexigenic melanocortin neurons.

The Kisspeptin/Neurokinin B/Dynorphin (KNDy) Cell Population of the Arcuate Nucleus: Sex Differences and Effects of Prenatal Testosterone in Sheep

Recent work in sheep has identified a neuronal subpopulation in the arcuate nucleus that coexpresses kisspeptin, neurokinin B, and dynorphin (referred to here as KNDy cells) and that mediate the negative feedback influence of progesterone on GnRH secretion. We hypothesized that sex differences in progesterone negative feedback are due to sexual dimorphism of KNDy cells and compared neuropeptide and progesterone receptor immunoreactivity in this subpopulation between male and female sheep. In addition, because sex differences in progesterone negative feedback and neurokinin B are due to the influence of testosterone (T) during fetal life, we determined whether prenatal T exposure would mimic sex differences in KNDy cells. Adult rams had nearly half the number of kisspeptin, neurokinin B, dynorphin, and progesterone receptor-positive cells in the arcuate nucleus as did females, but the percentage of KNDy cells colocalizing progesterone receptors remained high in both sexes. Prenatal T treatment also reduced the number of dynorphin, neurokinin B, and progesterone receptor-positive cells in the female arcuate nucleus; however, the number of kisspeptin cells remained high and at levels comparable to control females. Thus, sex differences in kisspeptin in the arcuate nucleus, unlike that of dynorphin and neurokinin B, are not due solely to exposure to prenatal T, suggesting the existence of different critical periods for multiple peptides coexpressed within the same neuron. In addition, the imbalance between inhibitory (dynorphin) and stimulatory (kisspeptin) neuropeptides in this subpopulation provides a potential explanation for the decreased ability of progesterone to inhibit GnRH neurons in prenatal T-treated ewes.

Effect of apoptotic proteins on function of rat vasopressin-and dopaminergic hypothalamic neurons

To study character of effect of apoptosis signal proteins on activities of neurosecretory cells and neurons of rat hypothalamus, pharmacologic inhibitors of proapoptotic protein p53 Pifithrin-alpha and antiapoptotic protein Bcl-2 HA14-1 were injected into the hypothalamus. Activation of vasopressinergic neurosecretory cells at administration of the blocker Bcl-2 HA14-1 was shown: there were observed an increase of vasopressin mRNA in neurons of hypothalamus supraoptical and paraventricular nuclei, a decrease of the immunoreactive vasopressin content in posterior pituitary, and reduction of diuresis. Inactivation of p53 inhibited release of vasopressin from hypothalamus cell bodies, which is indicated by an elevated content of immunoreactive vasopressin in neurosecretory cell bodies with its unchanged synthesis, a decrease of the neurohormone content in the posterior pituitary, and an increase of diuresis rate. Activation of vasopressinergic neurons of the suprachiasmatic nucleus was also shown. Administration of the blocker Bcl-2 has been revealed to decrease functional activity both of dopaminergic neurons (Zona Incerta) and of dopaminergic neurosecretory cells (arcuate nucleus), in which a decrease of the tyrosine hydroxylase content was observed. The p53 inactivation also led to a decrease of activity of dopaminergic neurosecretory cells of arcuate nucleus, whereas activity of the proteins Zone Incerta did not change. Thus, it has been shown that a change of the apoptotic protein content in vasopressinergic and dopaminergic neurons and neurosecretory cells leads to a change of their functional activity, the character and possibly mechanisms of effects of apoptotic proteins on activities of vasopressin- and dopaminergic cells being different.

Homologous Upregulation of sst2 Somatostatin Receptor Expression in the Rat Arcuate Nucleus in vivo

In vitro studies using various cell systems have provided conflicting results regarding homologous regulation of somatostatin (SRIH) receptors, and information on whether SRIH regulates the expression of its own receptors in vivo is lacking. In the present study we examined, by in situ hybridization, the effects of pretreatment with the sst2-preferring SRIH analog, octreotide, in vivo, on mRNA levels of two SRIH receptor subtypes, sst1 and sst2, in rat brain and pituitary. ¹²⁵I-[DTrp⁸]-SRIH binding was also measured in these regions. Three hours after the iv injection of 50 µg octreotide to conscious adult male rats, there was a 46% increase (p < 0.01) in the labeling density of sst2 mRNA-expressing cells in the hypothalamic arcuate nucleus compared to normal saline-pretreated controls, but not in any of the other brain regions examined. Computer-assisted image analysis revealed that 3 h exposure to octreotide significantly (p < 0.01) augmented both the number and labeling density of sst2 mRNA-expressing cells in the arcuate nucleus, compared to those in saline-treated controls. By contrast, within the anterior pituitary gland, in vivo exposure to octreotide did not affect the expression of sst2 mRNA. No changes in sst1 mRNA-expressing cells were observed after octreotide treatment in any of the regions measured, indicating that the observed effects were homologous, i.e. specific of the receptor subtype stimulated. Octreotide pretreatment was also without effect on the density of ¹²⁵I-[DTrp⁸]-SRIH binding in either the arcuate nucleus or pituitary. These results demonstrate, for the first time, that SRIH preexposure in vivo upregulates the expression of a subtype of its own receptors, sst2, within the central nervous system. They further suggest that pretreatment with SRIH in vivo does not cause sst2 receptor desensitization in arcuate nucleus and pituitary. Such homologous regulatory mechanisms may play an important role in the neuroendocrine control of growth hormone (GH) secretion by the arcuate nucleus.

GHRP-6-induced changes in electrical activity of single cells in the arcuate, ventromedial and periventricular nucleus neurones [correction of nuclei] of a hypothalamic slice preparation in vitro.

Previously, we demonstrated that systemic injection of the growth hormone secretagogue, growth hormone-releasing peptide (GHRP)-6, selectively activated cells in the hypothalamic arcuate nucleus, as reflected by increased electrical activity and induction of the immediate early gene c-fos. The growth hormone secretagogue receptor distribution is not confined to the arcuate nucleus, suggesting that additional sites of action may exist. In the present study we characterized the electrophysiological responses of cells in the arcuate nucleus, ventromedial nucleus and periventricular nucleus in an in-vitro hypothalamic slice preparation, following bath application of GHRP-6. Additionally, since central somatostatin administration has been shown to attenuate the induction of the c-fos gene by GHRP-6, we sought to determine whether the arcuate cells activated by GHRP-6 are also somatostatin-sensitive. Male Wistar rats (100-150 g body weight (BW)) were anaesthetized (urethane; 1.2 g/kg BW) and the brains removed. Coronal sections (400 microm thickness) were cut through a block of hypothalamus and were transferred to a slice chamber perfused with artificial cerebrospinal fluid. Forty-one arcuate nucleus cells were tested with bath application of 15 microm GHRP-6 for 10 min, 16 of which were tested subsequently (>30 min later) with application of 10 microM somatostatin. Following GHRP-6 administration, 19 cells (46. 3%) showed a significant increase in firing rate during the 15-min period after GHRP-6 application (P<0.001), 17 cells (41.5%) did not respond and the remaining five cells (12.2%) were significantly inhibited. Six of the eight arcuate nucleus cells that were excited by GHRP-6 were significantly inhibited by somatostatin. By contrast, five of the six arcuate nucleus cells that were unresponsive to GHRP-6 were also unresponsive to somatostatin. In the ventromedial nucleus, of 19 cells tested, eight cells (42.1%) were excited by GHRP-6, eight cells (42.1%) were unresponsive and the remaining three cells (15.8%) were significantly inhibited. Of 19 cells recorded in the periventricular nucleus, 13 (68.4%) were unresponsive to GHRP-6 and six (31.6%) were significantly inhibited. Thus, electrophysiological studies in vitro suggest that: (1) neurones in the hypothalamic arcuate nucleus, ventromedial nucleus and periventricular nucleus show changes in electrical activity in response to GHRP-6; and (2) the arcuate nucleus cells excited by GHRP-6 are also subject to inhibitory control by somatostatin.

Kainate glutamate receptors (GluR5-7) in the rat arcuate nucleus: relationship to tanycytes, astrocytes, neurons and gonadal steroid receptors.

Glutamate action, through its ionotropic, kainate receptors, has been implicated in gonadal steroid-dependent mechanisms of the arcuate nucleus. The objective of the present study was to determine the expression of kainate glutamate receptors in neural and glial elements of this area and their potential relationship to gonadal steroid receptors. Single and double label, light and electron microscopic immunocytochemistry for kainate glutamate receptors and estrogen or androgen receptors revealed the existence of glutamate (GluR) 5-7 kainate receptors in tanycytes, astrocytes and neurons of the arcuate nucleus. In the arcuate nucleus, subsets of GluR5-7-containing neurons were also immunopositive for estrogen (20%) and/or androgen receptors (23%). Glial elements, however, lacked labeling for gonadal steroid receptors. The coexistence of gonadal and kainate receptors in the same perikarya of arcuate nucleus cell populations suggests hormone regulation of excitatory neurotransmission through ionotropic glutamate receptors in these regions. It is also indicated that a kainate receptor-mediated glutamate action may participate in neuro-glial interaction in the arcuate nucleus that, in turn, may underlie the morphological synaptic plasticity induced by gonadal steroids.

Estrogen-regulated synthesis of neurotensin in neurosecretory cells of the hypothalamic arcuate nucleus in the female rat.

Neurotensin (NT) is implicated as a neurohormone in mammals, yet the peptide's neuroendocrine role remains to be determined. NT immunoreactivity has been observed in neurosecretory cells of the arcuate nucleus and paraventricular nucleus, and data suggest that NT release into hypophysial portal blood mediates a component of PRL secretion that is female specific and dependent on ovarian steroids. In the present study, in situ hybridization histochemistry and immunohistochemistry were used to investigate the regulation of NT gene expression in hypothalamic neurosecretory regions of adult rats. In ovariectomized females, estradiol induced expression of messenger RNA (mRNA) encoding NT and neuromedin N (NT/N mRNA) in the dorsomedial division of the arcuate nucleus. In contrast, estradiol did not appreciably alter NT/N mRNA expression in the ventrolateral division of the arcuate nucleus, where labeled cells were numerous, or in the paraventricular nucleus, where labeled cells were virtually absent. Estradiol also increased NT immunoreactivity in the external zone of the median eminence, confirming the neuroendocrine phenotype of NT cells in the dorsomedial division, as well as estrogen-regulated synthesis of NT in this system. In the dorsomedial division of cycling females, NT/N mRNA-expressing cells were significantly more numerous at proestrus than at diestrus, consistent with differences in plasma estradiol levels at these stages. In this same region, NT/N mRNA-expressing cells were significantly more numerous in proestrous females than in gonad-intact males. These results imply that estrogen-regulated release of NT at the median eminence subserves one or more sexually differentiated functions and are consistent with the involvement of tuberoinfundibular NT in estrogen-dependent secretion of PRL or GnRH on the afternoon of proestrus.

Alpha-Melanocyte stimulating hormone: immunohistochemical identification and mapping in neurons of rat brain.

alpha-Melanocyte stimulating hormone (alpha-melanotropin) immunofluorescence was observed in rat brain by means of a highly specific and well-characterized antibody. The hormone was contained in arcuate nucleus cell bodies and in varicose fibers. Dense populations of hormone-containing fibers were present in the septum, the nucleus interstitialis stria terminalis, and the medial preoptic, anterior hypothalamic, dorsomedial, and periventricular nuclei. Moderate numbers of fibers were seen in the paraventricular and arcuate nuclei, the amygdala, the region of the tractus diagonalis, the mammillary body, the central gray, the cuneiform nucleus, and the nucleus of the solitary tract. There is an interesting correlation of alpha-melanocyte stimulating hormone fibers with regions of noradrenergic axonal projections and terminal fields.

Direct evidence that the arcuate nucleus-median eminence tuberoinfundibular system is not of primary importance in the feedback regulation of luteinizing hormone and follicle-stimulating hormone secretion in the castrated rat.

Adult rats which have received monosodium-L-glutamate (MSG, 4 mg/g BW) on alternate days for the first 10 days of life manifest central nervous system lesions largely restricted to the retina and the arcuate nucleus of the hypothalamus in which nearly 90% of the perikarya are destroyed, leaving axons in passage intact. In animals so treated, concentrations of dopamine within the arcuate nucleus and median eminence of the hypothalamus are reduced 52% and 57%, respectively, in males and 45% and 61% in females, whereas concentrations of norepinephrine in these same two areas are normal. Concentrations of both norepinephrine and dopamine in five other hypothalamic nuclei (dorsal septal, medial preoptic, suprachiasmatic, periventricular, and dorsomedial nuclei) are unchanged. Nevertheless, despite the destruction of the arcuate nucleus cell bodies of MSG-treated rats, postcastration levels of serum FSH and LH in males, and FSH in females were not significantly different from FSH and LH values in castrated controls. Serum LH in castrated, MSG-treated females was slightly but significantly lower than in castrated controls. It is concluded that the arcuate nucleus-median eminence tuberoinfundibular neurons are not of primary importance in the tonic, negative feedback regulation of gonadotropin secretion.

A microprobe analysis of Gomori-positive glial cells in the rat arcuate nucleus.

Astrocytes and microglial cells in the arcuate nucleus of the rat hypothalamus contain lipofuscin-like granules which react with chrome alum gallocyanin and exhibit endogenous peroxidase activity. These granules were assessed with energy dispersive X-ray microanalysis and compared to neuronal dense bodies and glial cytoplasm. The granules are distinguished by a consistent content of sulphur and a frequent presence of calcium. The localization of other elements such as iron, copper, potassium and chlorine is impaired by methodical difficulties. The sulphur content as well as the endogenous peroxidase activity is interpreted as indicating a special variant of lipofuscin. The presence of calcium is discussed with respect to recent concepts of glia as a regulator of the ionic environment of the CNS.

Electron microscopic observation on the hypothalamo-hypophyseal system in the rat. II. Ultrafine structure of the median eminence and of the nerve cells of the arcuate nucleus.

Ultrafine structure of the median eminence was examined with the electron microscope in adult female normal and castrated rats. In the internal layer of the median eminence there are many un-myelinated and a few myelinated nerve fibers. The un-myelinated nerve fibers contain in their axoplasma numerous granules and vesicles, the diameter of which varies from 40 to 340mμ. The diameter of vesicles with the largest population measures 60-80mμ. Electron dense granules show two peaks in their size distribution histogram, a major one at 100-140mμ and a minor one at 220-240mp. In the external layer, too, both in its middle portion and in its peripheral contact region, a number of nerve fibers containing granules and vesicles are found. Nerve fibers in this layer are exclusively un-myelinated. The granules and vesicles found in this layer are much more in number and distributed in a narrower size range than those in the internal layer. The largest population have the dense granules and the vesicles the diameter of which measure 120mμ-160mμ and 80-120mμ respectively. Small vesicles resembling synaptic vesicles are also abundantly found. Two weeks after castration, small vesicles in the internal layer are slightly decreased in number, while in the external layer small dense granules (less than 120mμ in diameter) and small vesicles (60, -120mμ) are markedly increased. The number of larger dense granules remained almost unchanged both in the internal and in the external layer. Furthermore, the fine structure of the cells of arcuate nucleus was studied in normal rats. Besides well developed endoplasmic reticula and Golgi apparatus, small electron dense granules measuring about 100mμ in diameter are not infrequently found in the cytoplasm, in particular, in the vicinity of the Golgi apparatus. Possible significance of the findings observed was discussed.