The transcriptome of the medullary area postrema: The thirsty rat, the hungry rat and the hypertensive rat
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ABSTRACT: The area postrema (AP) is a sensory circumventricular organ characterised by extensive fenestrated vasculature and neurons which are capable of detecting circulating signals of osmotic, cardiovascular, immune and metabolic status. The AP can communicate these messages via efferent projections to brainstem and hypothalamic structures that are able to orchestrate an appropriate response. We have used microarrays to profile the transcriptome of the AP in the Sprague Dawley (SD) and Wistar Kyoto (WKY) rat and present here a comprehensive catalogue of gene expression, focussing specifically on the population of ion channels, receptors and G protein-coupled receptors (GPCRs) expressed in this sensory tissue; of the GPCRs expressed in the rat AP we identified ~36% that are orphans having no established ligand. We have also looked at the ways in which the AP transcriptome responds to the physiological stressors of 72-hours dehydration (DSD) and 48-hours fasting (FSD) and have performed microarrays under these conditions. Comparison between the DSD and SD or between FSD and SD revealed only a modest number of AP genes that are regulated by these homeostatic challenges. The expression levels of a much larger number of genes are altered in the spontaneously hypertensive rat (SHR) AP compared to the normotensive WKY controls however. Finally, analysis of these ‘hypertension-related’ elements revealed genes that are involved in both the regulation of blood pressure and immune function and as such are excellent targets for further study. Using Affymetrix microarrays (Rat 230:2.0) we anaysed the transcriptome of the area postrema. Each microarray in the study represents an independent biological replicate of 5 animals.
Project description:We have used microarrays to comprehensively describe the transcriptomes of the supraoptic nucleus (SON), the paraventricular nucleus (PVN) and the neurointermediate lobe (NIL) of adult male Sprague-Dawley (SD) and Wistar-Kyoto (WKY) rats, as well as the paraventricular nucleus of Wistar (WIST) rats. Comparison of these gene lists has enabled us to identify surprisingly large differences in hypothalamo-neurohypophyseal system gene expression patterns in these three strains. We have also shown that different transcript populations are enriched in the PVN and the SON of SD and WKY rats. The transcriptome differences catalogued here may be molecular substrates for the neuro-humoral phenotypic differences exhibited by different strains of rats. Experiment Overall Design: For each experimental group (SON-WKY, PVN-WKY, NIL-WKY, SON-SD, PVN-SD, NIL-SD, PVN-WISTAR) five chips were hybridised with independantly pooled RNA from a biological n=5.
Project description:We have used Affymetrix microarray-driven gene profiling to comprehensively describe the expression of mRNAs in the nucleus tractus solitarii (NTS) in the adult male spontaneously hypertensive rat (SHR) as compared to its normotensive parental Wistar-Kyoto (WKY) strain. Keywords: Gene expression NTS was dissected from hypertensive (SHR) and normotensive (WKY) strains of rat. For each structure, 10 independent Affymetrix Genechip 230 2.0 experiments were performed (5 SHR, 5 WKY), with each chip representing an independent biological replication (n=5).
Project description:In these studies we have for the first time described the transcriptome of the rat SFO, and have in addition identified genes the expression of which is significantly modified by either water or food deprivation. Experiment Overall Design: For each condition (food deprivation, water deprivation, untreated), 25 animals were denied food for 48 hours prior to sacrifice. The Subfornical organ was removed under a dissecting microscope from a 1mm slice of brain removed from an ice cold brain matrix. SFO's from 5 animals were pooled for each sample prior to RNA extraction to give n=5.
Project description:The hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei are important integrative structures that regulate co-ordinated responses to perturbations in cardiovascular homeostasis. Through descending projections from parvocellular neurons to the brainstem, the PVN acts as an autonomic 'premotor nucleus', regulating reflex changes in sympathetic nerve activity that are involved in blood pressure and blood volume regulation. Endocrine responses are mediated through axonal projections from SON and PVN magnocellular neurons (MCNs) to the capillaries of the posterior pituitary neural lobe. In response to dehydration, a massive release of the antidiuretic peptide hormone vasopressin (VP) into the circulation is accompanied by a dramatic functional remodelling of the HNS. We have used microarrays to comprehensively catalogue the genes expressed in the PVN, the SON and the neurointermediate lobe (NIL) of the pituitary gland. Further, we have identified transcripts that are regulated as a consequence of dehydration, as well as RNAs that are enriched in either the PVN or the SON. We suggest that these differentially expressed genes represent candidate regulators and effectors of HNS activity and remodelling. Experiment Overall Design: In total, 10 Affymetrix Genechip Rat Genome 230 2.0 were used. The experiment compared hypothalamic supraoptic nucleus from 3 day dehydrated and control male Sprague Dawley rats (10-12 weeks). For each chip, tissue from 5 animals was pooled prior to extraction of total RNA and in total 5 chips were used for each condition using independently prepared RNA samples from separate groups of animals.
Project description:This SuperSeries is composed of the following subset Series:; GSE3110: Comprehensive description of the transcriptome of hypothalamo-neurohypophyseal system in euhydrated and dehydrated rat; GSE3111: Comprehensive description of the transcriptome of PVN in euhydrated and dehydrated rat; GSE3125: Comprehensive description of the transcriptome of neurointermediate lobe in euhydrated and dehydrated rat Experiment Overall Design: Refer to individual Series
Project description:The hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei are important integrative structures that regulate co-ordinated responses to perturbations in cardiovascular homeostasis. Through descending projections from parvocellular neurons to the brainstem, the PVN acts as an autonomic 'premotor nucleus', regulating reflex changes in sympathetic nerve activity that are involved in blood pressure and blood volume regulation. Endocrine responses are mediated through axonal projections from SON and PVN magnocellular neurons (MCNs) to the capillaries of the posterior pituitary neural lobe. In response to dehydration, a massive release of the antidiuretic peptide hormone vasopressin (VP) into the circulation is accompanied by a dramatic functional remodelling of the HNS. We have used microarrays to comprehensively catalogue the genes expressed in the PVN, the SON and the neurointermediate lobe (NIL) of the pituitary gland. Further, we have identified transcripts that are regulated as a consequence of dehydration, as well as RNAs that are enriched in either the PVN or the SON. We suggest that these differentially expressed genes represent candidate regulators and effectors of HNS activity and remodelling. Experiment Overall Design: In total, 10 Affymetrix Genechip Rat Genome 230 2.0 were used. The experiment compared hypothalamic supraoptic nucleus from 3 day dehydrated and control male Sprague Dawley rats (10-12 weeks). For each chip, tissue from 5 animals was pooled prior to extraction of total RNA and in total 5 chips were used for each condition using independently prepared RNA samples from separate groups of animals.
Project description:The hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei are important integrative structures that regulate co-ordinated responses to perturbations in cardiovascular homeostasis. Through descending projections from parvocellular neurons to the brainstem, the PVN acts as an autonomic 'premotor nucleus', regulating reflex changes in sympathetic nerve activity that are involved in blood pressure and blood volume regulation. Endocrine responses are mediated through axonal projections from SON and PVN magnocellular neurons (MCNs) to the capillaries of the posterior pituitary neural lobe. In response to dehydration, a massive release of the antidiuretic peptide hormone vasopressin (VP) into the circulation is accompanied by a dramatic functional remodelling of the HNS. We have used microarrays to comprehensively catalogue the genes expressed in the PVN, the SON and the neurointermediate lobe (NIL) of the pituitary gland. Further, we have identified transcripts that are regulated as a consequence of dehydration, as well as RNAs that are enriched in either the PVN or the SON. We suggest that these differentially expressed genes represent candidate regulators and effectors of HNS activity and remodelling. Experiment Overall Design: In total, 10 Affymetrix Genechip Rat Genome 230 2.0 were used. The experiment compared hypothalamic supraoptic nucleus from 3 day dehydrated and control male Sprague Dawley rats (10-12 weeks). For each chip, tissue from 5 animals was pooled prior to extraction of total RNA and in total 5 chips were used for each condition using independently prepared RNA samples from separate groups of animals.
Project description:The area postrema (AP) is a sensory circumventricular organ characterised by extensive fenestrated vasculature and neurons which are capable of detecting circulating signals of osmotic, cardiovascular, immune and metabolic status. The AP can communicate these messages via efferent projections to brainstem and hypothalamic structures that are able to orchestrate an appropriate response. We have used microarrays to profile the transcriptome of the AP in the Sprague Dawley (SD) and Wistar Kyoto (WKY) rat and present here a comprehensive catalogue of gene expression, focussing specifically on the population of ion channels, receptors and G protein-coupled receptors (GPCRs) expressed in this sensory tissue; of the GPCRs expressed in the rat AP we identified ~36% that are orphans having no established ligand. We have also looked at the ways in which the AP transcriptome responds to the physiological stressors of 72-hours dehydration (DSD) and 48-hours fasting (FSD) and have performed microarrays under these conditions. Comparison between the DSD and SD or between FSD and SD revealed only a modest number of AP genes that are regulated by these homeostatic challenges. The expression levels of a much larger number of genes are altered in the spontaneously hypertensive rat (SHR) AP compared to the normotensive WKY controls however. Finally, analysis of these ‘hypertension-related’ elements revealed genes that are involved in both the regulation of blood pressure and immune function and as such are excellent targets for further study.
Project description:The supraoptic nucleus (SON) of the hypothalamus is an important integrative brain structure that co-ordinates responses to perturbations in water balance and regulates maternal physiology through the release of the neuropeptide hormones vasopressin and oxytocin into the circulation. Both dehydration and lactation evoke a dramatic morphological remodelling of the SON, a process known as function-related plasticity. We hypothesise that some of the changes seen in SON remodelling are mediated by differential gene expression, and have thus used microarrays to document global changes in transcript abundance that accompany chronic dehydration in female rats, and in lactation. In situ hydridisation analysis has confirmed the differential expression of 3 of these genes, namely Tumour necrosis factor induced protein 6, Gonadotrophin inducible transcription factor 1 and Ornithine decarboxylase antizyme inhibitor 1. Comparison of differential gene expression patterns in male and female rats subjected to dehydration and in lactating rats has enabled the identification of common elements that are significantly enriched in gene classes with particular functions. Two of these are related to the requirement for increased protein synthesis and hormone delivery in the physiologically stimulated SON (translation initiation factor activity and endoplasmic reticulum-Golgi intermediate compartment respectively), whilst others are consistent with concept of SON morphological plasticity (collagen fibril organisation, extracellular matrix organization and biogenesis, extracellular structure organization and biogenesis and homophilic cell adhesion). We suggest that the genes co-ordinately regulated in the SON as a consequence of dehydration and lactation form a network that mediates the plastic processes operational in the physiologically activated SON. Each independent microarray sample represented the SON from either 5 female euhydrated, 5 dehydrated (water restriction for 72hours with food ad libitum) or 11 days lactating. In total, 5 microarrays were performed for the Dehydrated and the Lactation groups and 4 for the euhydrated controls. Samples within each group may be considered to be both biological and technical replicates. The GeneSpring-derived MAS5 data described in the publication is not included in this submission.
Project description:We have addressed the question of how different rodent species cope with the life-threatening homeostatic challenge of dehydration at the level of transcriptome modulation in the supraoptic nucleus (SON), a specialised hypothalamic neurosecretory apparatus responsible for the production of the antidiuretic peptide hormone arginine vasopressin (AVP). AVP maintains water balance by promoting water conservation at the level of the kidney. Dehydration evokes a massive increase in the regulated release of AVP from SON axon terminals located in the posterior pituitary, and this is accompanied by a plethora of changes in the morphology, electrophysiological properties, biosynthetic and secretory activity of this structure. Microarray analysis was used to generate a definitive catalogue of the genes expressed in the mouse SON, and to describe how the gene expression profile changes in response to dehydration. Comparison of the genes differentially expressed in the mouse SON as a consequence of dehydration with those of the rat has revealed many similarities, pointing to common processes underlying the function-related plasticity in this nucleus. In addition we have identified many genes that are differentially expressed in a species-specific manner. However, in many cases, we have found that the hyperosmotic cue can induce species-specific alterations in the expression of different genes in the same pathway. The same functional end can be served by different means, via differential modulation, in different species, of different molecules in the same pathway. We suggest that pathways, rather than specific genes, should be the focus of integrative physiological studies based on transcriptome data. Animals. Adult male C57BL/6 mice (Harlan Sera-Lab, Loughborough, UK) were group housed (4 per cage) under controlled temperature (21+ 2ºC) and diurnal light conditions (14-h light, 10-h dark, lights on at 05.00). Food and water were available ad libitum until the experiment commenced. Complete fluid deprivation was imposed for 48 hours starting at 11.00 a.m. Control animals maintained free access to drinking water, and both groups had access to standard laboratory rodent chow. Experiments on adult male rats described previously (27). All procedures were conducted in strict accordance with the Animal Scientific Procedures Act (1986), UK, and were approved by the local University of Bristol Ethical Review Process. Tissue collection. Mice were killed using cervical dislocation and the brain was carefully removed from the cranium and snap frozen using powdered dry ice and stored at -80oC for no more than 14 days. Sections of brain (14μm) were cut using an RNase free cryostat and mounted onto RNase free membrane coated glass slides (P.A.L.M. Membrane slides; P.A.L.M. Microlaser Technologies). Immediately after sectioning, frozen sections were thawed and fixed (30s; in 95% [v/v] EtOH), rehydrated (30s in each of 75% [v/v] and 50% [v/v] EtOH) before being stained (60s 1% [v/v] cresyl violet). Sections were then dehydrated in a graded EtOH series (30s in each of 50% [v/v], 75% [v/v] and 95% [v/v]. then 2x 30s in 100% [v/v]). Laser microdissection was performed using a P.A.L.M. MicrolaserSystem (P.A.L.M. Microlaser Technologies). The SON was identified with reference to Franklin and Paxinos (28) and the tissue from each animal was independently pooled into collection vials containing RNAlater® (Ambion, Huntingdon, UK). A single operative carried out all dissections. Total RNA was isolated without delay (within 24h) according to standard procedures that accompany the Ambion RNAqueous MicroKit (Ambion). Microarray analysis. Separate microarrays (n=4) were probed using independently generated target. For each completely independent replicate, tissue from 1 mouse was used for RNA extraction.