Cortistatin modulates calcitonin gene-related peptide release from neuronal tissues of rat. Comparison with somatostatin
Research highlights
▶ The effects of cortistatin and somatostatin on CGRP release were investigated. ▶ Cortistatin inhibited stimulated CGRP release from rat trigeminal ganglia neurons. ▶ Cortistatin inhibited stimulated CGRP release from rat brainstem explants. ▶ Somatostatin showed a profile of action similar to cortistatin in ganglia neurons. ▶ Cortistatin was more effective than somatostatin in brainstem explants.
Introduction
Cortistatin (CST) is a cyclic neuropeptide cloned from human, rat and mouse tissues [30]. CST expression was initially reported to be restricted to the cerebral cortex and hippocampus, whereas it is currently known to have wide distribution in many organs [14]. In the rat, CST is synthesized as a prepropeptide (preproCST) form that yields two biologically active peptides: CST-29 and CST-14 (thereafter referred to as CST), the latter preferentially released via the regulated secretory pathway [22], [12]. CST exhibits a remarkable structural and functional resemblance to somatostatin (SST), although it is the product of a different gene [10], [11], [30]. CST shares with SST the ability to bind and activate all five cloned SST receptors (SSTR1-5), with similar efficacy and potency [29]. This finding may well explain the considerable overlapping between CST and SST on several biological actions, including the depression of neuronal activity and the inhibition of cell proliferation [10], [7].
However, the profile of CST biological activities is not simply redundant with respect to SST; in fact, recent studies showed that CST has certain proprieties distinct from, and even opposite to those of SST [4], [15], [13]. Indeed, evidence exists that CST might signal through receptors distinct from those of SST, namely the putative CST receptor MrgX2 and the GHSR-1a ghrelin receptor [1], [9], [25]. Moreover, four truncated but functional SST5 receptor variants (SST5TMD4, SST5TMD2 and SST5TMD1 in the mouse; SST5TMD1 in the rat) have been described; these variants show distinct ligand-selective signaling profiles for CST and SST, respectively [8].
Pain perception involves several anatomical structures within both the central and peripheral nervous system, as well as various agents such as neurotransmitters, neuropeptides and cytokines. CST and SST show different activities on the modulation of pain perception. In fact, intracerebroventricular injections of CST elicit a significant increase in pain threshold, assessed through the hot-plate test in the rat; in this paradigm, SST displays no significant activity [20]. Thus, the effect of CTS on pain threshold may not involve SST receptors. MrgX2 receptors, which bind with high affinity CTS but not SST, are expressed in human principally in the small sensory neurons of the dorsal root ganglia (DRG), which include primary sensory fibers associated with acute and neuropatich pain [25]. Human DRG also express CST mRNA. Therefore, a role for this ligand-receptor pair can be postulated in the control of nociception [25].
Within this framework, in the present study we have investigated the effects of CTS on the release of calcitonin gene-related peptide (CGRP, the main neuromediator of trigeminal signaling) from primary cultures of rat trigeminal ganglia neurons as well as from rat brainstem explants containing trigeminal nucleus caudalis. Moreover the effects of CST and SST in this experimental paradigm were also compared. CGRP secretagogues were 56 mM KCl solutions, veratridine and capsaicin. The first two should be considered as non-specific depolarizing agents, with veratridine acting primarily through the opening of Na+ channels, whereas 56 mM KCl solutions elicit directly Ca2+ ion influx within the cells. In contrast, the C-fiber stimulating agent capsaicin is considered a specific stimulus, acting via binding to and activation of vanilloid receptors [5].
Section snippets
Trigeminal neuronal cultures
Trigeminal neuronal cultures were prepared from 6- to 7-day-old Wistar rats as previously described [5]. The use of animals for this experimental work has been approved by the Italian Ministry of Health (licensed authorization to P. Navarra). In brief, animals were decapitated and, after removal of the brain, basal skulls were exposed and trigeminal ganglia from both sides were aseptically removed. Tissues were collected in a Petri dish containing 3–5 ml of ice-cold phosphate buffer saline
Effects of CST and SST on basal and stimulated CGRP release from primary cultures of rat trigeminal neurons
The first series of experiments was carried out on primary cultures of rat trigeminal neurons; the latter release CGRP under basal and stimulated conditions [6].
CST did not modify basal release of CGRP in the concentration range 0.1 nM–1 μM (Fig. 1). A significant and consistent increase in CGRP secretion was elicited by non-specific (56 mM KCl or veratridine) or specific (capsaicin) depolarizing stimuli after 30 min of incubation. Under these conditions, CTS was able to reduce the secretion of
Discussion
In the present study we investigated the effects of CST and SST on CGRP release from primary cultures of rat trigeminal ganglia neurons and from brainstem trigeminal nuclei. We found that both CST and SST were able to inhibit CGRP release from trigeminal neurons evoked by different depolarizing stimuli. Moreover, in the present study we have developed and characterized CGRP release from brainstem explants containing trigeminal nucleus caudalis, which mainly receives afferent C-fibers from
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