Cortistatin modulates the expression and release of corticotrophin releasing hormone in rat brain. Comparison with somatostatin and octreotide
Highlights
► We investigated the effects of cortistatin on CRH production from isolated rat brain regions. ► We also compared the action of cortistatin with those of somatostatin and octreotide. ► Cortistatin inhibited the release and gene expression of CRH from rat hypothalamus. ► Cortistatin inhibited basal and stimulated CRH release from rat hippocampus. ► Somatostatin and octreotide showed a different action profile from cortistatin.
Introduction
Cortistatin (CST) is a 14-aa neuropeptide belonging to the family of somatostatin (SST)-like Cys–Cys loop peptides; it was cloned from human, rat and mouse tissues [38]. In the rat, CST is synthesized as a prepropeptide (preproCST) yielding two biologically active peptides: CST-29 and CST-14 (thereafter referred to as CST), the latter preferentially released via the regulated secretory pathway [18], [34]. CST possesses a remarkable structural similarity to SST-14, although it is the product of a different gene [16], [17]. In fact, CST shares 11 out of 14 amino acids with SST-14, including the amino acid sequence phe-trp-lys-thr required to bind all five cloned SST receptors (SST-R1–SST-R5) [48]. These analogies explain certain common biological activities of CST and SST, including the depression of neuronal activity, the inhibition of cell proliferation and endocrine and metabolic activities [6], [9], [16], [22]. However, the profile of CST is not simply redundant with respect to SST; in fact, several studies showed marked differences between the two hormones in terms of tissues distribution, regulation of synthesis and biological actions, especially in the central nervous system (CNS) and immune system [5], [8], [20], [23], [27]. Based on these differences, specific CST receptors, able to bind selectively CST but not SST, have been postulated. Putative CST receptors are the Mas-related gene X2 (MrgX2) receptor and the GHS-R1a ghrelin receptor [1], [15], [35]. MrgX2 receptor is not expressed in the cerebral cortex, where CST expression is most abundant [35]; MrgX2 appears to be non-specific to CST, since it acts as a receptor for pro-adrenomedullin and related peptides [31] as well. The possible role GHS-R1a as CST receptor has also been questioned. Therefore, the existence of specific CST receptors remains under scrutiny. On this regard, truncated but functional SST5 receptor variants have recently been described in human, rodents and pig; these variants show distinct ligand-selective signaling profiles with respect to CST and SST, respectively [12], [13].
Corticotrophin-releasing hormone (CRH) is a 41-amino acid neuropeptide originally isolated from the ovine hypothalamus [46]. The CRH system currently includes two receptors sub-families (CRH-R1 and -R2), and four ligands (CRH and the related peptides Urocortin I, II, and III) as well a CRH-binding protein [2]. CRH plays a major role in coordinating the behavioral, endocrine, autonomic and immune responses to stress. In fact, CRH produced in the paraventricular nucleus in response to stress modulates ACTH release from the anterior pituitary via CRH-R1 [47], with consequent increases in glucocorticoid secretion from the adrenal cortex. Basal glucocorticoid levels and transient increases during stress are essential for neuronal plasticity and normal brain function. On the contrary, chronic stress causes inappropriate CRH and glucocorticoid levels, with consequent metabolic and immune changes. Extra-hypothalamic CRH seems to play a important role in behavioral adaptive responses to stress as well as in the development of affective disorders [2], [4]. In fact, CRH and its receptors are widely distributed throughout the brain. In particular, the hippocampus displays the highest concentrations of CRH-binding sites and CRH-immunoreactive nerve terminals [10], [19], suggesting that CRH might modulate important hippocampal functions, associated to the control of emotions and response to stress. Indeed, CRH was shown to be secreted directly from nerve terminals within the hippocampus during exposure to stress [10]. A large body of evidence shows that an altered functioning of CRH system may be critical in the development of affective disorders, including anxiety, depression and stress-related disorders [3], [29]. On this line of evidence, it was suggested that the efficacy of anti-depressive agents is associated to the reduction of CRH production and release within the CNS [24]. In this regard, our group showed that different drugs used in mood disorders share the ability to reduce, through different mechanisms, the production and release of CRH from rat hypothalamic and hippocampal explants [21], [42], [44], [45].
The possible role of SST in the control of stress responses has been investigated in the last decades, showing that SST might blunt stress responses both via a direct inhibitory action on hypothalamic CRH [41] and the inhibition of CRH-stimulated ACTH release at pituitary level [39]. Instead, the putative effects of CST in stress control have received so far little attention. Within this framework, in the present study we investigated the effects of CST on the release of CRH from two rat brain areas, namely the hypothalamus and the hippocampus. Moreover, we compared the effects of CST with those of SST and its stable analog Octreotide (OCT) in the same paradigm.
Section snippets
Animals
Male Wistar rats (200–300 g) were used. They were kept four per cage and maintained at a temperature of 23 ± 1.5 °C, with a relative humidity of 65 ± 2%. The animals were exposed to 12-h light (06.00–18.00 h) followed by 12-h dark, and had free access to food and water. The use of animals for this experimental work has been approved by the Italian Ministry of Health (licensed authorization to P. Navarra).
Hypothalamic and hippocampal incubations
On day of the experiment the animals were decapitated between 09.00 and 10.00 a.m. and the brains
Effects of CST, SST and OCT on basal CRH release from rat hypothalamic explants
The first series of experiments was carried out on rat hypothalamic explants; the latter release measurable amounts of CRH under basal and stimulated conditions [45]. In 1 h incubation experiments, CST induced a reduction in CRH release that was statistically significant at 1 μM (Fig. 1A). In the same experimental paradigm, SST given in the range 10–1000 nM showed a trend toward inhibition of basal CRH release which it was not statistically significant (Fig. 1B). On the contrary, OCT, a
Discussion
In the present study we show for the first time that CST negatively modulates the expression and release of CRH from rat hypothalamic and hippocampal explants; these in vitro actions suggest that CST might have a role in the regulation of adaptive responses to stress.
Several studies showed that CST possesses the same actions of SST in different endocrine systems in vivo but especially in vitro, so that some researchers consider CST as a natural duplicate to SST with regard to its endocrine
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