Salt appetite, along with other primordial forms of awareness such as thirst and hunger, are thought to be evolutionary precursors to more complex forms of consciousness. Our research interests are focused on the role of hypothalamic structures in the behavioural control of salt intake. We have used Affymetrix microarray gene profiling to catalogue gene expression in these brain regions in euhydrated and dehydrated male rats. These gene catalogues were then subject to robust statistical analysis to identify genes that are differentially regulated as a consequence of dehydration. Network reconstruction analysis of our transcriptome data from dehydrated rats revealed a putative node around Gonadotrophin inducible transcription factor 1 (Giot1), which is robustly up-regulated in the dehydrated hypothalamus. Rats are normally averse to 2% (w/v) NaCl. However, this aversion is overcome if 2% (w/v) NaCl is their only fluid source. An initial decline in fluid consumption is followed by a progressive increase in drinking over the course of the 7-day stimulus, concurrent with an increase in the excretion of large volumes of urine. Hypothalamic injection of a lentiviral vector that expresses a Giot1 shRNA completely blocked fluid intake following the onset of salt loading. We thus hypothesis that Giot1 regulates neural circuits in the brain that modulate reward and motivation. We will now use a battery of complementary state-of-the-art technologies to understand the mechanisms responsible for the profound behavioural actions of Giot1. Disorders of fluid balance are evident in a substantial proportion of elderly patients admitted to hospital, and dehydration and hypo-nateremia are frequent cause of morbidity and mortality in old people. This is partly due to decreased thirst perception and altered salt appetite. Thus, a major question to be addressed is how are the brain mechanisms controlling salt intake are altered by ageing.
1. What are the molecular mechanisms that mediate the regulation of Giot1 by physiological cues?
2. What are the molecular mechanisms of Giot1 action?
3. How does Giot1 alter neural function?
4. What are the downstream neural mechanisms of Giot1 action?
5. How are brain mechanisms controlling salt intake altered by ageing?
The student will:
1. Use ChiP-Seq to identify transcriptional targets of Giot1.
2. Use yeast-two hybrid analysis to identify protein partners.
3. Validate the genomic results.
4. Engage in functional studies using lentiviral vector transduction of shRNAs to target neurones, assessing salt appetite, thirst, and the electrophysiological properties of transduced cells.
When applying please select ’Medicine PhD’ within the Faculty of Health Sciences.
Greenwood M, Mecawi A, Hoe SZ, Mustafa MR, Johnson K, Al-Mahmoud G, Elias L, Paton J, Antunes-Rodrigues J, Gainer H, Murphy D and Hindmarch C (2015) A comparison of physiological and transcriptome responses to water deprivation and salt loading in the rat supraoptic nucleus. Am J Physiol 308:R559-568
Konopacka A, Qiu J, Yao ST, Greenwood MP, Greenwood M, TLancaster T, Inoue W, Mecawi A, Vechiato FMV, de Lima JBM, Coletti R, Hoe SZ, Martin A, Lee J, Joseph M, Hindmarch C, Paton J, Antunes-Rodrigues J, Bains J and Murphy D (2015) Osmoregulation requires brain expression of the renal Na-K-2Cl co-transporter NKCC2. J Neurosci 35:5144-5155
Greenwood M, Greenwood MP, Paton JFR and Murphy D (2015) Transcription Factor CREB3L1 Regulates Endoplasmic Reticulum Stress Response Genes in the Osmotically Challenged Rat Hypothalamus. PLoS One 10:e0124956
Johnson KR, Hindmarch CCT, Salinas YD, Shi YJ , Greenwood M, Ziau HS, Murphy D and Gainer H (2015) A RNA-Seq Analysis of the Rat Supraoptic Nucleus Transcriptome: Effects of Salt Loading on Gene Expression. PLoS One 10:e0124523
Greenwood MP, Greenwood M, Paton JFR and Murphy D (2015) Control of Polyamine Biosynthesis by Antizyme Inhibitor 1 is Important for Transcriptional Regulation of Arginine Vasopressin in the Male Rat Hypothalamus. Endocrinology 156:2905–2917
This paper was the subject of a commentary in the same journal by Yoichi Ueta (AZ Street in Vasopressin Synthesis. Endocrinology 2015 156:2749-2750)
Konopacka A, Greenwood M, Loh S-Y, Paton J and Murphy D (2015) RNA binding protein Caprin-2 is a pivotal regulator of the central osmotic defense response. eLife (In Press)
Greenwood M, Greenwood MP, Mecawi AS, Loh S-Y, Antunes Rodrigues J, JPaton JFR and Murphy D (2015) Transcription factor CREB3L1 mediates cAMP and Glucocorticoid Regulation of Arginine Vasopressin Gene Transcription in the Rat Hypothalamus. Molecular Brain (In Press)