Central tachykinin NK3 receptors in the inhibitory action on the rat colonic propulsion of a new tachykinin, PG-KII

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Abstract

The inhibitory action of the natural selective tachykinin NK3 receptor agonist, PG-KII, (pGlu-Pro-Asn-Pro-Asp-Glu-Phe-Val-Gly-Leu-Met-NH2), on colonic propulsion was studied in rats after central administration. Intracerebroventricular injection of PG-KII (0.1, 1, 10 and 100 ng/rat) produced a dose-related inhibition of colonic propulsion, measured as the increase in the mean expulsion time of a 5-mm glass bead placed in the distal colon. At the same doses as PG-KII, the selective tachykinin NK3 receptor agonist, senktide, (succ-[Asp6-MePhe8] substance P-(6–11)), induced a similar dose-related inhibition. Conversely, substance P (0.1, 1 and 10 μg/rat), a tachykinin NK1-preferring receptor agonist, had weaker antipropulsive effects, neurokinin A (0.1, 1 and 10 μg/rat), a tachykinin NK2-preferring receptor agonist, at the highest dose used only slightly inhibited colonic propulsion and neurokinin B (0.1, 1 and 10 μg/rat), a tachykinin NK3-preferring receptor agonist, left propulsion unchanged. Pretreatment with the selective tachykinin NK3 receptor antagonist, 3-indolycarbonyl-Hyp-Phg-N(me)-Bzl, referred as to R820 (6.2 μg/rat), prevented PG-KII-induced colonic antipropulsion, whereas the tachykinin NK1 receptor antagonist, (S)-1-(2-[3-(3,4-dichlorophenyl)-1-(3-isopropoxyphenylacetyl)piperidin-3-yl] ethyl)-4-phenyl-1-azoniabicyclo[2.2.2] octane chloride, referred to as SR 140,333 (1 μg/rat), and the tachykinin NK2 receptor antagonist, ([Tyr5,d-Trp6,8,9, Arg10] neurokinin A-(4–10)), referred to as Men 10,376 (5 μg/rat), left it unchanged. These findings show that of the tachykinins tested, PG-KII and senktide are the most potent central inhibitors of colonic propulsion in the rat, suggesting that the central tachykinin NK3 receptor system plays an inhibitory role in modulating colonic transit. As well as confirming the selectivity of PG-KII for tachykinin NK3 receptors, we show that PG-KII provides useful information about the physiological role of central tachykinin NK3 receptors and that glass bead expulsion test is a reliable non-invasive in vivo method for evaluating the tachykinin NK3 receptor selectivity of new synthetic or natural tachykinins.

Introduction

Tachykinins are widely distributed in the central nervous system (Saffroy et al., 1988) and in the gastrointestinal tract (Mussap et al., 1993). Although they participate in the control of many gastrointestinal functions, their central and peripheral physiological role in the control of peristalsis remains unclear.

The most investigated action of tachykinins in the gut relates to their role as neuromuscular excitatory transmitters through the activation of tachykinin NK1 (substance P-preferring) or tachykinin NK2 (neurokinin A-preferring) receptors (Barthó and Holzer, 1985; Holzer-Petsche, 1995; Shuttleworth and Keef, 1995; Maggi et al., 1997). Tachykinins can also affect intestinal motility by releasing inhibitory transmitters via tachykinin NK1 and NK3 receptors (Holzer and Holzer-Petsche, 1997). Evidence for tachykinin NK1 and NK3 receptor agonist inhibition of motor activity comes mainly from in vitro studies in the canine colon (Hou et al., 1989), the rat distal colon (Scheurer et al., 1994) and the guinea pig small and large intestine (Maggi et al., 1993, Maggi et al., 1994a, Maggi et al., 1994b, Maggi et al., 1994c, Maggi et al., 1994d; Giuliani and Maggi, 1995). The role of tachykinin NK3 receptors in intestinal motility under in vivo conditions remains less clear, because of the lack of potent and selective tachykinin NK3 receptor agonists and antagonists. One study only has reported preliminary in vivo evidence that tonic activation of peripheral tachykinin NK3 receptors by endogenous tachykinins stimulates inhibitory reflexes during prolonged localised distension of the rat colon (Lecci et al., 1996). Nothing is known about the role of central tachykinin NK3 receptors in the regulation of colonic motility.

Recently, a novel kassinin-like peptide, PG-KII, (pGlu-Pro-Asn-Pro-Asp-Glu-Phe-Val-Gly-Leu-Met-NH2), has been isolated from the skin of the Australian myobatrachid frog, Pseudophryne güntheri (Simmaco et al., 1990). Our previous reports (Improta et al., 1996; Polidori et al., 1997) indicated that in the rat, after central administration, PG-KII modulates inhibition of gastric acid secretion and alcohol intake, two functions thought to be mediated mainly by tachykinin NK3 receptors. Furthermore, after peripheral administration, PG-KII stimulates saliva secretion through cholinergic pathways, as does neurokinin B (Broccardo et al., 1996). All these data provide evidence confirming PG-KII as a potent and selective tachykinin NK3 receptor agonist.

This newly available highly selective tachykinin NK3 receptor agonist seemed an important tool for investigating the role of central tachykinin NK3 receptors in the regulation of in vivo colonic motility. Continuing research into the possible physiological role of the central tachykinin NK3 receptor system in the mediation of gut motility in the rat, our main aim in this study was to characterize the actions of this novel natural tachykinin on colonic transit. Hence, we compared the effects induced on rat colonic propulsion by intracerebroventricular (i.c.v.) injection of PG-KII with those induced by various tachykinins including substance P (a tachykinin NK1-preferring receptor agonist), neurokinin A (a tachykinin NK2-preferring receptor agonist), neurokinin B (a tachykinin NK3-preferring receptor agonist) and senktide (a synthetic and selective tachykinin NK3 receptor agonist). We also evaluated the activity of PG-KII at central tachykinin receptors in rats pretreated with selective tachykinin NK1, NK2 and NK3 receptor antagonists. The fundamental role of the central opioid receptor system in modulating colonic motility (Broccardo and Improta, 1992, Broccardo and Improta, 1998) prompted us also to investigate a possible central tachykinin–opioid receptor system interaction.

Section snippets

Animals

All animal experiments complied with the Italian D.L. no. 116 of 27 January 1992 and associated guidelines in the European Communities Council Directive of 24 November 1986 (86/609/EEC). Male Wistar rats (Morini, S. Polo D'Enza, Italy) weighing 200–250 g were used for the experiments. All rats were examined at 09:00 h. Rats were placed individually in plastic cages under standard temperature conditions (22°), with 12-h light/dark cycles and food and water ad libitum.

Surgery

At least 3 days before the

Results

The mean colonic bead expulsion time for vehicle-treated rats was 2.26±0.34 min. All the tachykinins tested inhibited colonic propulsion: the rank order of potency was PG-KII=senktide≫substance P⋙neurokinin A≥neurokinin B. PG-KII (0.1, 1, 10 and 100 ng/rat) increased the mean expulsion time significantly and in a dose-related manner; the highest dose tested (100 ng/rat) increased the colonic propulsion time to 37.32±6.4 min and even a very small dose (100 pg/rat) significantly inhibited colonic

Discussion

First of all, in this study, i.c.v. injection of the natural tachykinin PG-KII potently inhibited distal colonic propulsion increasing, in a significant and dose-dependent way, the mean expulsion time of a glass bead placed in the distal colon of rats. Because R820, the most suitable tachykinin NK3 receptor antagonist currently available for studies of the rat central nervous system (Cellier et al., 1997), prevented the PG-KII-induced inhibitory effect, whereas the tachykinin NK1 and NK2

Acknowledgements

We are grateful to Dr. D. Regoli (University of Sherbrooke, Quebec, Canada) for the donation of R820. We also acknowledge Dr. A. Maggi (Menarini, Florence) for the kind gift of Men 10,376. This study was supported by grants from the Italian Ministry of University and Scientific and Technological Research.

References (40)

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