• Corticotropin-releasing factor (CRF) acts in the brain to inhibit thyrotropin-releasing hormone (TRH) analogue, RX-77368-induced vagal stimulation of gastric motility. We investigated CRF receptor-mediated actions of rat urocortin (rUcn) injected intracisternally (ic) on gastric motor function.
• Urethane-anaesthetized rats with strain gauges on the gastric corpus were injected i.c. with rUcn and 20min later, with i.c. RX-77368. CRF antagonists were injected i.c. 10min before rUcn.
• RX-77368 (1.5, 3, 10, 30 and 100ng, i.c.) dose-dependently increased corpus contractions, expressed as total area under the curve (AUC, mVmin⁻¹) to 2.6±2.5, 6.1±5.9, 9.8±2.6, 69.7±21.7 and 74.9±28.7 respectively vs 0.2±0.1 after i.c. saline. Ucn (1, 3 or 10μg) inhibited RX-77368 (30ng)-induced increase in total AUC by 28, 62 and 93% respectively vs i.c. saline+RX-77368.
• The CRF₁/CRF₂ antagonist, astressin-B (60μg, i.c.) completely blocked i.c. rUcn (3μg, i.c.)-induced inhibition of gastric motility stimulated by RX-77368 (30ng).
• The selective CRF₂ antagonist, astressin₂-B (30, 60 or 100μg, i.c. ) dose-dependently prevented i.c. rUCn action while the CRF₁ antagonist, NBI-27914 did not.
• In conscious rats, rUcn (0.6 or 1μg, i.c.) inhibited gastric emptying of an ingested chow meal by 61 and 92% respectively. rUcn action was antagonized by astressin₂-B.
• These data show that i.c. rUcn acts through CRF₂ receptors to inhibit central vagal gastric contractile response and postoprandial emptying.

Corticototropin-releasing factor (CRF) is well established to act centrally to inhibit gastric motor function (Taché et al., 2001, review). CRF injected into the cerebrospinal fluid at the level of the lateral ventricle (i.c.v.), third ventricle or cisterna magna (i.c.) or microinjected into specific hypothalamic or medullary nuclei delays gastric emptying of acaloric (methycellulose) and caloric (glucose) liquids as well as gastric transit of a solid chow meal in conscious rats, and dogs (Taché et al., 1987; Lenz et al., 1988a; Sheldon et al., 1990; Mönnikes et al., 1992; 1994; Smedh et al., 1995; Lee & Sarna, 1997; Martínez et al., 1998a). In addition, CRF injected i.c. or into the dorsal vagal complex (DVC) inhibits vagally mediated high amplitude gastric contractions induced by the stable thyrotropin-releasing hormone (TRH) analogue, RX-77368 co-injected with CRF into the DVC in urethane anaesthestized fasted rats (Garrick et al., 1988; Heymann-Mönnikes et al., 1991). In conscious fasted dogs, i.c.v. injection of CRF abolished the phase III of the migrating motor complex cyclic activity front in the antrum, and induced a long-lasting disruption of interdigestive gastric cyclic contractions (Buéno & Fioramonti, 1986; Buéno et al., 1986). Recently, rat urocortin (rUcn) has been characterized as a 40 amino-acid peptide which shares 45% homology with rat/human (r/h)CRF. However, so far only one study investigated the central action of rUcn on gastric motor function (Kihara et al., 2001) and showed that the i.c.v. injection of rUcn decreased antrum motility under conditions of fed state while disrupting the fasted motor patterns of gastroduodenal activity to induce a fed-like motor pattern in conscious rats.

CRF and Ucn biological actions are mediated through binding to two high-affinity membrane-bound receptors, the subtype 1 (CRF₁) and the subtype 2 (CRF₂) which are coupled to G_s to stimulate cyclic AMP formation (Chalmers et al., 1996; Perrin & Vale, 1999). In rats, two CRF₂ splice variants, α and β, differing in their N-terminal domains, have been identified (Perrin & Vale, 1999). CRF₁ and CRF₂ receptors display distinct affinities for the mammalian members of the CRF family of peptides. In cells transfected with CRF receptors, rUcn binds with equal high affinity to both CRF receptor subtypes and their variants while r/hCRF displays high binding affinity to CRF₁ receptor, almost equal to that of rUcn, but is 40 fold less potent at the CRF_2α and CRF_2β receptors (Vaughan et al., 1996; Perrin et al., 1999). Recently two new peptide members of the CRF family, urocortin II and urocortin III, have been identified by sequence homology from the human genome data base and the mouse orthologs cloned (Reyes et al., 2001; Lewis et al., 2001). Both mouse and human urocortin II and urocortin III are highly selective for the type 2 CRF receptors and exhibit low affinities for the CRF₁ receptor (Reyes et al., 2001; Lewis et al., 2001).

Studies with the specific, non-selective CRF₁/CRF₂ receptor peptide antagonists, α-helical r/h CRF_9–41, and the more potent [D-Phe¹²,Nle^21,38,C^'γ MeLeu³⁷]r/hCRF_12–41 and astressin (Rivier et al., 1984; Hernandez et al., 1993; Gulyas et al., 1995) established that central CRF-induced inhibition of gastric motor function is CRF receptor mediated in rats and dogs (Lenz et al., 1988b; Sheldon et al., 1990; Taché et al., 1991; Sütó et al., 1994; 1996; Smedh et al., 1995; Lee & Sarna, 1997; Martínez et al., 1997; 1998a; 1999; Rivier et al., 1998). The central action of rUcn to alter gastric motility in fed or fasted state was also prevented by the i.c.v. injection of α-helicalr/hCRF_9–41 (Kihara et al., 2001). However, the CRF receptor subtype involved in the central action of CRF or Ucn to inhibit gastric motor function is still to be characterized. Indirect pharmacological evidence based on the use of a selective CRF₁ receptor antagonist, and non-mammalian CRF-related peptides with higher affinity than r/hCRF to the CRF₂ receptor pointed to a role of CRF₂ receptor in mediating i.c. r/hCRF-induced delayed gastric emptying of Purina chow ingested by rats previously food deprived (Martínez et al., 1998a). A major breakthrough has been the development of selective CRF₂ receptor antagonists, namely, antisauvagine-30 and more recently, the long-acting derivative, astressin₂-B which provided new tools to assess the role of CRF₂ receptor in the biological actions of CRF and related peptides (Ruhmann et al., 1998; Rivier et al., 2001; Higelin et al., 2001; Million et al., 2002).

In the present study, we investigated the actions of rUcn injected i.c. on gastric motor functions in rats. First, we assessed the dose-related effect of i.c. rUcn on RX-77368 injected i.c. at a dose inducing a plateau stimulation of corpus contractility in urethane-anaesthetized rats. To gain insight into the CRF receptor subtype involved in rUcn inhibitory action under these conditions, we used the recently developed long acting and potent CRF₁/CRF₂ receptor antagonist, astressin B, cyclo(30–33) [D-Phe¹²,Nle^21,38,Glu³⁰,Lys³³,C^αMeLeu^27,40]Ac-r/h CRF_9–41 (Rivier et al., 1999), the non-peptide selective CRF₁ receptor antagonist NBI-27914 (Maciejewski-Lenoir et al., 2000) and the CRF₂ antagonist, astressin₂-B, cyclo(31-34)[D-Phe¹¹,His¹²Nle¹⁷,C^dMeLeu^13,39,Glu³¹,Lys³⁴]Ac-Sauvagine_(8–40) which has similar affinity to the CRF₂ variants (Rivier et al., 2001). Lastly, we investigated in conscious rats whether the i.c. injection of rUcn influences gastric emptying of a solid meal and the mediation of rUcn action through the CRF₂ receptor.

Anaesthesia All surgical procedures and experimental protocols except otherwise stated, were performed in rats under urethane anaesthesia. Urethane (1.5gkg⁻¹) was injected intraperitoneally (i.p.) 30min before the start of surgery with an additional i.p. injection (0.1–0.25gkg⁻¹) immediately before surgery if, as was usually the case, the corneal reflex was present. Adequate depth of anaesthesia was monitored at regular intervals by the absence of response to ear-pinch and corneal reflex. A temperature-controlled heating pad was used to maintain rectal temperature within 35.5–37.5°C throughout the experiment.

Cannulation of the cisterna magna The cisterna magna was cannulated as previously described (Kiraly et al., 1994). After the tracheotomy performed to facilitate ventilation, the head of the rat was positioned in a stereotaxic apparatus (Kopf, Model 900), and the atlanto-occipital membrane was exposed. A 25-gauge needle was inserted perpendicularly into the membrane to two-thirds of the depth of its bevel, at approximately 1.5mm caudal to the edge of the occipital bone, then removed. A PE-10 polyethylene catheter (Intramedic, Becton Dickinson, Parsippany, NJ, U.S.A.; length: 6cm) was inserted 2mm in length through the hole into the cisterna magna. The PE-10 catheter was connected to an injection port comprising a short length of stainless steel tubing joined to a PE-20 polyethylene tubing (total dead space: 5μl). Successful cannulation was verified by leakage of clear cerebrospinal fluid from the distal catheter. If blood was observed, the experimental procedure was not further pursued. A drop of cyanoacrylate glue (Duro® SuperGlue, Rocky Hill, CT, U.S.A.) was used to hold the catheter in place, and when secured, it was covered with cotton and the incision was sutured. The cannula was capped to prevent leakage. In all experiments, we reconfirmed the correctness of the intracisternal cannula placement immediately after euthanasia. This was performed by injecting methylene blue (Sigma) under similar conditions of administration as i.c. injection of tested compounds and assessing the distribution of the dye inside the cisterna magna by visual inspection after euthanasia.

Intravenous catheter A PE-50 catheter was inserted into the right external jugular vein, and sterile, pyrogen-free 0.9% saline (Sigma) was continuously infused at a rate of 1.2mlh⁻¹ with a syringe pump to maintain hydration.

Implantation of gastric strain gauge A strain gauge (Micro-Measurement Inc; Raleigh, NC, U.S.A.; size: 10×7×1mm) was sutured to the serosa side of gastric corpus, parallel to circular smooth muscles in the greater curvature side, using Tevdek II 7-0 suture (Deknatel; Fall River, MA, U.S.A.). Then, the abdomen was closed with a suture using 3-0 silk.

Gastric motility in urethane-anaesthetized rats After the completion of surgical procedures (average 60min), and the stabilization period (30min), gastric corpus contractions were monitored for 30min before and 120min after i.c. injection of RX-77368 in urethane-anaesthetized rats fasted for 18-h.

Dose-related effects of RX-77368 and urocortin injected i.c. In the first study, after a 30min basal period, RX-77368 (1.5, 3, 10, 30, and 100ng) or saline was injected i.c. in separate groups of rats. Based on the dose-response curve, RX-77368 at 30ng, which induces a plateau gastric contractile response, was selected to stimulate gastric contractility in all further experiments. In the second study, separate groups of rats were injected i.c. with rUcn at 1, 3, or 10μg or double-distilled water and 20min later, with RX-77368 (30ng, i.c.).

Effects of CRF receptor antagonists on urocortin action The non-selective CRF₁/CRF₂ receptor antagonist, astressin B (30 or 60μg), the selective CRF₁ receptor antagonist, NBI-27914 (50 or 100μg), the selective CRF₂ receptor antagonist, astressin₂-B (30, 60 or 100μg), or respective vehicles was injected i.c. 10min before rUcn (3μg, i.c.) or i.c. water, and RX-77368 (30ng) was injected i.c. 20min later. The dose of rUcn was selected based on the above dose-response study. The dose of NBI-27914 was based on previous studies (Martínez & Taché, 2001).

Dose-related effect of urocortin injected i.c. Rats fasted for 18-h were given ad libitum access to water and Purina chow for a 3-h period, then were injected i.c. with either saline (10μl) or rUcn (0.3, 0.6 or 1μg in 10μl) by puncturing the occipital membrane under short enflurane anaesthesia (2–3min, 5% vapor concentration in O₂; Ethrane-Anaquest, Madison, WI, U.S.A.), as previously described (Martínez et al., 1998a). The selection of rUcn dose range was based on our previous dose response studies in conscious rats in which i.c. injection of r/hCRF or non-mammalian CRF-related peptides (0.1–1μg) inhibited gastric emptying of non-nutrient and nutrient meals (Taché et al., 1987; Martínez et al., 1998a).

Effect of astressin₂-B on urocortin action Rats fasted for 18-h were given ad libitum access to water and Purina chow for a 3-h period, then either water (5μl) or astressin₂-B (10μg in 5μl) followed by that of saline (5μl, i.c.) or rUcn (1μg in 5μl, i.c.) were injected i.c. The dose of astressin₂-B was selected to provide an initial 10:1 antagonist:agonist ratio. In both studies, after the i.c. injections, fed rats were returned to their individual home cages without food and water, and 5-h later, were euthanized to measure gastric emptying of the meal ingested before the i.c. injection.

Figure 1 Dose-related stimulation of gastric contractions induced by intracisternal injection of the TRH analogue, RX-77368 in 18-h fasted urethane-anaesthetized rats. After a 30min basal period, different groups of rats were injected i.c. with either (more ...)

Figure 2 (a) Time course of gastric contractile response to i.c. RX-77368 and dose-related inhibitory effect of i.c. urocortin in 18-h fasted urethane-anaesthetized rats. Each point represents the mean AUCmin−1 of corpus contractions; (b–f) (more ...)

Figure 3 Dose-related inhibitory effect of intracisternal urocortin on RX-77368-induced stimulation of gastric contractions in 18-h fasted urethane-anaesthetized rats. After 30min basal period, different groups of rats were injected i.c. with vehicle (more ...)

Figure 4 Intracisternal astressin B antagonized intracisternal urocortin-induced inhibition of RX-77368 induced stimulation of gastric contractions in 18-h fasted urethane-anaesthetized rats. Intracisternal injection of astressin B or vehicle was followed 10min (more ...)

Figure 5 Intracisternal astressin2-B dose-dependently antagonized intracisternal urocortin-induced inhibition of RX-77368 stimulated gastric contractions in 18h-fasted urethane-anaesthetized rats. Intracisternal injection of astressin2-B or vehicle was (more ...)

Table 1 Effect of the selective CRF1 receptor antagonist, NBI-27914 injected intracisternally on intracisternal urocortin-induced inhibition of gastric corpus contractility stimulated by RX-77368 in urethane-anaesthetized rats

Figure 6 Representative tracing of blockade of intracisternal urocortin-induced inhibition of gastric motor activity stimulated by RX-77368: blockade by the CRF1/CRF2 receptor antagonist, astressin B and the selective CRF2 receptor antagonist, astressin2-B but (more ...)

Figure 7 Dose-related inhibition of gastric emptying of a solid meal by intracisternal injection of urocortin in conscious fed rats (a) and blockade of urocortin effects by pretreatment with the selective CRF2 receptor antagonist astressin2-B (b). Fasted rats (more ...)

In animals injected with vehicle+vehicle, 54.0±5.2% of the meal had emptied from the stomach 5h after the end of the feeding period, while in vehicle+rUcn (1μg), gastric emptying was reduced to 11.2±7.0% (P<0.05 vs vehicle+vehicle; Figure 7b). Pretreatment with astressin₂-B (10μg) antagonized the inhibitory effect of i.c. rUcn on gastric emptying of solid meal and values were not significantly different from those of rats injected with vehicles (Figure 7b). Astressin₂-B (10μg) followed by the injection of vehicle had no effect on basal gastric emptying of a solid meal (53.3±5.2%).

In the present study, the stable TRH analogue, RX-77368 injected i.c. (1.5, 3, 10, 30 and 100ng) dose-dependently stimulated the low basal gastric contractile activity in 18-h fasted rats anaesthetized with urethane. This was assessed by monitoring mechanical activity of the gastric smooth muscles recorded from signals generated by a strain gauge acutely implanted onto the corpus serosa. The increase of gastric motility induced by i.c. RX-77368 plateaued at 30ng. Time course response showed a rapid onset (within 5min) with a peak reached within 10–15min, followed by a return to the low basal activity between 60–70min after i.c. RX-77368 injection. Basal AUC values increased by 17 fold at the peak response to i.c. RX-77368 at 30ng while saline had no effect. The characteristics of the gastric contractile changes analysed using computer software revealed that i.c. injection of RX-77368 increase dose-dependently the mean amplitude and mean duration of individual contractile spike and the duration of the total contractile response. Likewise, other reports indicate that i.c. injection of RX-77368 (10 or 100ng) increased the duration of high amplitude gastric contractions as monitored using a strain gauge chronically implanted on the corpus in conscious fasted rats (Garrick et al., 1987) or intraluminal gastric pressure in urethane-anaesthetized rats (Raybould et al., 1990).

Convergent neuroanatomical, electrophysiological and functional studies established that the gastric contractile response to i.c. injection of TRH or RX-77368 is mediated by central vagal myenteric–cholinergic activation of the smooth muscles (O-Lee et al., 1997; Sivarao et al., 1997; Miampamba et al., 2001). TRH directly activates post-synaptic preganglionic vagal motor neurons (Travagli et al., 1992). We previously found that RX-77368 injected i.c. induced a sustained increase in gastric vagal efferent discharges in urethane-anaesthetized rats with a rapid onset (~3min) and long duration (~60min) (O-Lee et al., 1997) similar to the time course of increased corpus contractility observed in the present study. This supports the view that gastric changes are associated with an increased vagal efferent outflow. In addition, we recently reported that RX-77368 injected i.c. at 30 and 50ng resulted in a plateau stimulation of gastric myenteric neuronal activity in conscious rats as shown by Fos expression in corpus and antrum myenteric neurons (Miampamba et al., 2001). The activation of corpus myenteric ganglia encompasses cholinergic neurons and is prevented by ganglionic blockade (Miampamba et al., 2001). Lastly, the stimulation of gastric contractility and emptying induced by i.c. RX-77368 or TRH is blocked by vagotomy and atropine (Garrick et al., 1987; Martínez et al., 1998b).

In the present study, intracisternal injection of rUcn (1, 3 and 10μg) inhibited dose-dependently i.c. RX-77368-induced maximal stimulation of gastric corpus contractility in urethane-anaesthetized rats as assessed by the reduction in the total AUC, the mean spike amplitude and mean spike duration. Intracisternal injection of rUcn (0.6 and 1μg) also inhibited gastric propulsive motor function in conscious rats, as shown by the dose-related suppression of postprandial gastric emptying. At 1μg, only 9% of the food had emptied from the stomach after 5h, indicative of the potency and long-lasting effect of i.c. rUcn inhibitory action. The observed decreased gastric emptying of the ingested meal is not confounded by a possible reduction of food ingestion reported to occur after central injection of rUcn at similar doses (Spina et al., 1996). In our studies, rUcn was injected i.c. after the 3-h refeeding period and the amount of Purina chow eaten by rats in all the experimental groups was similar before i.c. injection of rUcn. Moreover, animals had no access to food or water after the i.c. treatment. r/hCRF injected i.c. inhibits gastric acid secretion in conscious fasted rats (Taché et al., 1983). Such a possible decrease in gastric fluid secretion would reduce the weight content of the stomach, while we found an increase after i.c. rUcn, suggesting that the gastric emptying measurements reflect a delayed gastric emptying. Consistent with these observations, the non-mammalian CRF-related 40 amino-acid peptide, sauvagine which has high affinity to CRF₂ receptors, injected i.c. also completely suppressed gastric emptying of a solid meal at similar dose range (Martínez et al., 1998a). The i.c.v. injection of rUcn at 0.1–5μg was also reported to suppress dose-dependently the antrum motor activity induced by refeeding after a fast in conscious rats (Kihara et al., 2001). In the present study, rUcn injected i.c. at 0.3–1μg suppressed postprandial gastric emptying by 61–91%, while doses ranging from 1–10μg are required to inhibit centrally stimulated gastric contractility by 29–95%. Such a difference in rUcn dose range to suppress gastric motor function may be related to the different experimental models used: conscious preparation with no surgery in gastric emptying studies vs urethane-anaesthetized rats with acute surgery in the gastric motility experiments. However, irrespective of animal preparations, rUcn injected into the cerebrospinal fluid inhibits gastric motor function as shown by the dose-related suppression of corpus contractions induced by central vagal stimulation in anaesthetized fasted rats (present study), decreases antral motility index associated with refeeding in conscious rats (Kihara et al., 2001), and slows postprandial gastric emptying in conscious rats (present study).

Additional studies using CRF receptor antagonists provided convergent evidence that i.c. rUcn inhibitory action is CRF receptor mediated and that the receptor subtype 2 is primarily involved. First, astressin B injected i.c. completely prevented i.c. rUcn-induced decrease of corpus contractility induced by i.c. RX-77368. Astressin B is a recently developed long-acting antagonist at both CRF₁ and CRF₂ receptors (Rivier et al., 1999). Likewise, i.c.v. rUcn-induced antral motility alteration of the fed and fasted patterns was blocked by the non-selective CRF receptor antagonist, α-helical CRF_9–41 in conscious rats (Kihara et al., 2001). Second, in the present study, the selective CRF₂ antagonist, astressin₂-B (Rivier et al., 2001) injected i.c. dose-dependently abolished i.c. rUcn inhibitory action on vagally stimulated gastric motility in urethane-anaesthetized rats. The choice of astressin₂-B was based on the stability and selectivity of this new CRF₂-receptor antagonist in in vitro binding studies on transfected CRF receptors and in vivo studies (Rivier et al., 2001; Martínez et al., unpublished observations). Moreover, we showed that i.c. rUcn-induced inhibition of gastric emptying of a solid meal was completely antagonized by astressin₂-B in conscious rats. The higher antagonist:agonist ratio (33:1) at which i.c. astressin₂-B completely blocked i.c. rUcn inhibitory action on gastric motility compared with gastric emptying (10:1) may be related to difference in experimental conditions (anaesthetized vs conscious, as well as the nature of the stimuli to be inhibited by rUcn: exogenously induced maximal central vagal stimulation vs physiological postprandial vagal tone). By contrast, the selective CRF₁ receptor antagonist, NBI-27914 (Chen et al., 1996; Maciejewski-Lenoir et al., 2000) did not influence rUcn inhibitory action on stimulated gastric motility. Consistent with these findings, NBI-27914 administered i.c. did not influence i.c. CRF, sauvagine and urotensin I (0.3 or 1μg)-induced delayed gastric emptying of a solid meal in conscious rats (Martínez et al., 1998a). However, the CRF₁ receptor antagonist NBI-27914, injected i.c.v. at similar doses was biologically active to block i.c.v. CRF-induced colonic motor activation (Martínez & Taché, 2001). The inhibitory effect of NBI-27914 per se on total AUC may indicate that the compound acts as an inverse agonist under these conditions. Collectively, these observations provide strong support for the involvement of CRF₂ receptor in mediating i.c. rUcn-induced inhibition of gastric motor function.

Convergent evidence indicates that neuronal pathways, through which i.c. rUcn inhibits i.c. RX-77368 effect via CRF₂ receptor interaction, may take place within the DVC, involving modulation of vagal outflow to the stomach. Direct co-microinjection of CRF and RX-77368 into the DVC inhibited the stimulated gastric motility in urethane-anaesthetized rats (Heymann-Mönnikes et al., 1991), indicative of an interaction between CRF and the stable TRH analogue within medullary nuclei regulating gastric vagal outflow (Berthoud et al., 1991). Cell groups expressing the CRF₂ receptor have been localized in the area postrema and medial nucleus tractus solitarius (NTS) (Bittencourt & Sawchenko, 2000; Van Pett et al., 2000). Urocortin injected i.c.v. at 1μg activates neurons in the area postrema and NTS as shown by Fos expression (Bittencourt & Sawchenko, 2000). The activation of these medullary nuclei has been associated with inhibitory influence on preganglionic vagal neurons as established by electrophysiological and functional studies (Olson et al., 1993; Rogers & McCann, 1993). Moreover, we reported that i.c.v. injection of CRF induced Fos expression in NTS neurons while inhibiting brain medullary TRH circuitry-induced Fos expression in the dorsal motor nucleus of the vagus (DMN) (Wang et al., 1996). Lastly, electrophysiological studies showed that i.c. injection of sauvagine, which has higher affinity for CRF₂ receptors than r/hCRF (Perrin & Vale, 1999), is more potent than i.c. CRF to inhibit gastric vagal efferent discharges in urethane-anaesthetized rats (Kosoyan et al., 1999). Functional reports in conscious rats also established that truncal vagotomy abolished the changes in antrum motility index induced by rUcn injected i.c.v. (Kihara et al., 2001) and the delayed gastric emptying induced by i.c. CRF or sauvagine (Taché et al., 1987; Broccardo & Improta, 1990).

In summary, the present studies demonstrate that i.c. injection of RX-77368 induced a dose-related stimulation of gastric corpus contractility with a plateau response observed at 30 and 100ng in urethane-anaesthetized rats. The i.c. injection of rUcn dose-dependently inhibited corpus contractility induced by i.c. RX-77368 at 30ng in urethane-anaesthetized rats and gastric emptying of a solid meal in conscious rats. The inhibitory action of i.c. rUcn on gastric motor function is mediated through interaction with CRF₂ receptors. This is supported by the blockade of i.c. rUcn-induced suppression of RX-77368-stimulated gastric contractility by i.c. pretreatment with the CRF₁/CRF₂ receptor antagonist, astressin B and the selective CRF₂ receptor antagonist, astressin₂-B while the selective CRF₁ receptor antagonist, NBI-27914 had no effect. Likewise, the selective CRF₂ receptor antagonist blocked i.c. rUcn-induced inhibition of gastric emptying. These data along with previous pharmacological evidence that non-selective CRF receptor antagonists injected centrally prevented the delayed gastric emptying induced by various stressors (Taché et al., 2001, review), suggest that selective CRF₂ receptor antagonist acting in the brain medulla may have relevance to alleviate stress-related inhibition of gastric motor function.

This work was supported by the National Institute of Diabetes and Digestive and Kidney Diseases, R01 Grants DK-30110, DK 33061 and DK 26741 and Center grant DK-41301 (Animal Core). Chih-Yen Chen, is supported by Taiwan NHRI Physician Scientist Fellowship. The authors thank Dr Dimitri Grigoriadis (Neurocrine Bioscience, La Jolla, CA, U.S.A.) for the generous supply of NBI-27914. Dr Alfred Bayati (AstraZeneca, Mölndal, Sweden) is acknowledged for provision of a new version of Data Acquisition® software program, and Mr Paul Kirsch for his assistance in the preparation of the manuscript.