Neuronostatin, a recently discovered peptide derived from the somatostatin preprohormone, significantly inhibited both food and water intake when administered centrally in adult male rats. Because neuronostatin is highly produced in the hypothalamus, an area of the brain through which important feeding circuits, including the central melanocortin system, communicate, we sought to determine if the anorexigenic and antidipsogenic effects of neuronostatin would be reversed by pretreatment with the melanocortin 3/4 receptor antagonist, SHU9119. SHU9119 pretreatment reversed the effect of neuronostatin on both food and water intake. We have shown recently that the central oxytocin system is a potential downstream mediator of the anorexignic action of alpha-MSH. We therefore tested whether the effects of neuronostatin also were dependent upon central oxytocin receptors. Neuronostatin-induced anorexia was not reversed by pretreatment with the oxytocin receptor antagonist, OVT, suggesting that neuronostatin acts through a unique subset of POMC neurons that do not signal via central oxytocin receptors.
Yosten et al. Peptides. 2010 Jun 25. [Epub ahead of print]
Neuronostatin (NST) is a newly identified peptide of 13-amino acids encoded by the somatostatin (SST) gene. Using a rabbit polyclonal antiserum against the human NST, neuronostatin-immunoreactive (irNST) cells comparable in number and intensity to somatostatin immunoreactive (irSST) cells were detected in the hypothalamic periventricular nucleus. Fewer and/or less intensely labeled irNST cells were noted in other regions such as the hippocampus, cortex, amygdala, and cerebellum. Double-labeling hypothalamic sections with NST- and SST-antiserum revealed an extensive overlapping of irNST and irSST cells in the periventricular nucleus. Pre-absorption of the NST-antiserum with NST (1 microg/ml) but not with SST (1 microg/ml) abrogated irNST and vice versa. The activity of NST on dissociated and cultured hypothalamic neurons was assessed by the Ca(2+) imaging method. NST (10, 100, 1000 nM) concentration-dependently elevated intracellular Ca(2+) concentrations [Ca(2+)](i) in a population of hypothalamic neurons with two distinct profiles: (1) a fast and transitory increase in [Ca(2+)](i), and (2) an oscillatory response. Whereas, SST (100 nM) reduced the basal [Ca(2+)](i) in 21 of 61 hypothalamic neurons examined; an increase was not observed in any of the cells. Optical imaging with a slow-responding voltage sensitive dye DiBAC(4)(3) showed that NST (100 nM) depolarized or hyperpolarized; whereas, SST (100 nM) hyperpolarized a population of hypothalamic neurons. The result shows that NST and SST, though derived from the same precursor protein, exert different calcium mobilizing effects on cultured rat hypothalamic neurons, resulting in diverse cellular activities.
Dun et al. Neuroscience. 2010 Mar 17;166(2):455-63.
Neuronostatin, a newly identified peptide hormone sharing the same precursor with somatostatin, exerts multiple pharmacological effects in gastrointestinal tract, hypothalamus, and cerebellum. However, the cardiovascular effect of neuronostatin is unknown. The aim of this study was to elucidate the impact of neuronostatin on cardiac contractile function in murine hearts and isolated cardiomyocytes. Short-term exposure of neuronostatin depressed left ventricular developed pressure (LVDP), maximal velocity of pressure development (+/-dP/dt), and heart rate in Langendorff heart preparation. Consistently, neuronostatin inhibited peak shortening (PS) and maximal velocity of shortening/relengthening (+/-dL/dt) without affecting time-to-PS (TPS) and time-to-90% relengthening (TR(90)) in cardiomyocytes. The neuronostatin-elicited cardiomyocyte mechanical responses were mimicked by somatostatin, the other posttranslational product of preprosomatostatin. Furthermore, the neuronostatin-induced cardiomyocyte mechanical effects were ablated by the PKA inhibitor H89 (1 microM) and the Jun N-terminal kinase (JNK) inhibitor SP600125 (20 microM). The PKC inhibitor chelerythrine (1 microM) failed to alter neuronostatin-induced cardiomyocyte mechanical responses. To the contrary, chelerythrine, but not H89, abrogated somatostatin-induced cardiomyocyte contractile responses. Our results also showed enhanced c-fos and c-jun expression in response to neuronostatin exposure (0.5 to 2 h). Taken together, our data suggest that neuronostatin is a peptide hormone with overt cardiac depressant action. The neuronostatin-elicited cardiac contractile response appears to be mediated, at least in part, through a PKA- and/or JNK-dependent mechanism.
Hua et al. Am J Physiol Regul Integr Comp Physiol. 2009 Sep;297(3):R682-9.
Somatostatin is important in the regulation of diverse neuroendocrine functions. Based on bioinformatic analyses of evolutionarily conserved sequences, we predicted another peptide hormone in pro-somatostatin and named it neuronostatin. Immuno-affinity purification allowed the sequencing of an amidated neuronostatin peptide of 13 residues from porcine tissues. In vivo treatment with neuronostatin induced c-fos expression in gastrointestinal tissues, anterior pituitary, cerebellum, and hippocampus. In vitro treatment with neuronostatin promoted the migration of cerebellar granule cells and elicited direct depolarizing actions on paraventricular neurons in hypothalamic slices. In a gastric tumor cell line, neuronostatin induced c-fos expression, stimulated SRE-reporter activity, and promoted cell proliferation. Furthermore, intracerebroventricular treatment with neuronostatin increased blood pressure but suppressed food intake and water drinking. Our findings demonstrate diverse neuronal, neuroendocrine, and cardiovascular actions of a somatostatin gene-encoded hormone and provide the basis to investigate the physiological roles of this endogenously produced brain/gut peptide.
Samson et al. J Biol Chem. 2008 Nov 14;283(46):31949-59.
Understanding how a small brain region, the suprachiasmatic nucleus (SCN) can synchronize the body's circadian rhythms is an ongoing research area. This important time-keeping system requires a complex suite of peptide hormones and transmitters that remain incompletely characterized. Here, capillary liquid chromatography and Fourier-transform mass spectrometry (FTMS) have been coupled with tailored software for the analysis of endogenous peptides present in the SCN of the rat brain. After ex vivo processing of brain slices, peptide extraction, identification and characterization from tandem FTMS data with <5 ppm mass accuracy produced a hyper-confident list of 102 endogenous peptides, including 33 previously unidentified peptides, and 12 peptides that were post-translationally modified with amidation, phosphorylation, pyroglutamination, or acetylation. This characterization of endogenous peptides from the SCN will aid in understanding the molecular mechanisms that mediate rhythmic behaviors in mammals.
Lee et al. Mol Cell Proteomics. 2009 Nov 10.
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|
Fixative |
10% formalin |
Embedding |
Paraffin |
Negative Control |
No primary antibody |
Pretreatment |
N/A |
Blocking |
2% Normal Goat Serum |
Primary Antibody |
Rabbit Anti-Neuronostatin-13 (Human, Porcine) Antibody (Catalog No.:H-060-50) |
Optimal Dilution |
1:200, 1 hour at RT |
Secondary Antibody |
Goat Anti-Rabbit IgG, Biotinylated (1:400), 30 min |
Amplification |
ABC (Vector) (1:400, 30 min) |
Detection System |
HRP |
Substrate |
DAB (Sigma), 3 min |
Counterstained |
Hematoxylin, 30 sec |
Samson et al. J Biol Chem. 2008 Nov 14;283(46):31949-59.
