European Journal of Pharmacology, 191 (1990) 235-238
235
Elsevier EJP 20142
Short communication
The effect of ondansetron on gastric emptying in the conscious rat Eliot R. Forster and Graham J. Dockray MRC Secretov Control Group, PhysiologicalLaboratory. Universityof Liverpool, Liverpool, U.K.
Received 2 October 1990, accepted 9 October 1990
The 5-HT, receptor antagonist ondansetron (GR38032F) enhanced the action of a protein-rich solution in delaying gastric emptying in the conscious gastric fistula rat, but had no effect on the emptying of isotonic or hypertonic saline, acid or FOY-305 which delays emptying by release of cholecystokinin (CCK). The specific CCK-A antagonist
(L-364,718)increased gastric emptying of protein-rich meals. L364,718 also increased emptying in the presence of ondansetron. They indicate that protein-rich meals release both CCK and 5-hydroxytryptamine which act in different ways to control gastric motility. Gastric emptying; 5-HT, receptors; Protein meals; Ondansetron
1. Introduction
Several different pathways are now known to be involved in controlling the gastric emptying of different types of liquid test meal (Green et al., 1988; Forster et al., 1990). In conscious gastric fistula rats, protein-rich meals in particular delay gastric emptying by a mechanism sensitive to the cholecystokinin-A (CCK-A) receptor antagonist L364,718 and dependent on (a) passage of the solution beyond the proximal duodenum and (b) intact visceral afferents; the emptying of this type of meal is not, however, dependent on an intact pylorus. Hypertonic solutions and acid also inhibit gastric emptying by mechanisms that at least in part, require intact visceral afferents, but they are not sensitive to CCK antagonists and differ in their requirement for either an intact pylorus or passage of the solution beyond the duodenum. Although the data clearly suggest a role for CCK
Correspondence to: G.J. Dockray, Physiological Laboratory, University of Liverpool, P.O. Box 147, Liverpool L69 3BX, U.K.
in controlling the gastric emptying of protein the other humoral mediators that might be involved in controlling emptying of protein remain poorly characterized. In the present study we have examined the role of 5-HT, receptors in controlling gastric emptying of various liquid test meals in the consious gastric fistula rat. The data suggest that the S-HT, antagonist ondansetron (Butler et al., 1988) specifically enhances the action of peptone in delaying gastric emptying.
2. Materials and methods
2.I. Animals
Male Wistar rats (200-250 g, n = 8) were anaesthetised with sodium pentobarbitone (Sagatal, 60 mg kg- ’ i.p.) and fitted with stainless steel cannulas in the body of the stomach as previously described (Dimaline et al., 1986). At least two weeks were allowed for recovery before commencing experiments.
0014-2999/90/$03.50 0 1990 Elsevier Science Publishers B.V. (Biomedical Division)
.F
_.a.
~nimaIs were deprived of solid food overnight lowed free access to water. Trained rats were lightly restrained in Rollman-type cages and the stomach flushed with warm saline. Drugs and test solutions were administered directly into the the cannula. Ondansetron (GR38032 .K.) and L-364,718 (Merck, Sharp and ere freshly prepared immediately prior stration in isotonic saline (ondansetron) methyl cellulose in isotonic saline (L18). Doses of ondansetron ranging from loor the vehicle alone, were given via the cannula 2 h prior to commencing the emptying trials. L364.718 was given at a dose of 1 mg kg- ’ via the cannula 30 min prior to commencing the emptying trials. At the end of this period emptying of the following test solutions (3 ml) was studied: 300 mOsm kg-” NaCl, 900 mOsm kg-’ NaCl. 50 mM HCl, 20 mg ml-’ FOY 305 (camostat m&late, Ono Pharmaceuticals) and 4.5% (w/v) meat peptone II (Sigma). Phenol red (60 mg kg-‘) was added t o each solution as a non-absorbable dilution marker. Gastric emptying was determined from the volume and phenol red concentration of fluid recovered from the cannula 5 min after instillation of the test meal into the stomach (Debas et al., 1975; Green et al., 1988).
i
.c
5
2.5-
2 P
B i=
2-
zi Y 3 P
l-5-
T
OJ
,
0.01
/
I
0.03
0.1
1
0.3
1 1
ONDANSETRON (mg.kg”) Fig. 1. The volume of peptone emptied from the stomach after 5 min in a group of conscious gastric fistula rats treated with ondansetron (0.01-1.0 mg kg-‘). Each dose of ondansetron was studied on a separate day. and animals were used no more than twice weekly. The open symbol to the left shows the emptying of peptone in vehicle-treated rats. Points are means i SE., n = 8; * indicates significantly different from control P < 0.05, paired t-test.
effect of ondansetron on peptone was maximal at a dose of 0.1 mg kg-‘; at lower and higher doses it.s effects declined giving a bell-shaped dose-re-
J-3. Statistics The results are expressed as means f SE. Comparisons were made using a paired t-test.
Hyperosmolal saline, acid and peptone all delayed gastric emptying compared to isotonic saline, as previously described (Green et al., 1988; Forster et al., 1990). Ondansteron significantly increased the delay in gastric emptying produced by peptone, but had no effect on the emptying of hyperosmolal saline (control, 2.22 f 0.07 VS. ondansetron, 1.89 &-0.19 ml 5 min-*), isotonic saline (3.05 & 0.11 vs. 3.14 * 0.12 ml 5 mm-‘) or acid (1.95 f 0.08 vs. 1.95 + 0.08 ml 5 min-‘). The
Fig. 2. The rate of emptying of peptone in 5 mm in control rats and after L364,718 (1 mg kg-‘), ondansetron (a.1 mg kg-‘) and both L364,718 and ondansetron. Bars are means*S.E., n = 8. See fig. 1 for further information. Points xe means* SE., n = 8; * indicates significantly different from control P < 0.05, paired t-test.
237
sponse curve (fig. 1). The emptying of peptone is known to be increased following administration of the CCK antagonist, L364,718 (Green et al., 1988). In rats treated with the combination of L-364,718 and ondansetron (0.1 mg kg-‘) the rate of emptying of peptone was not si~fic~tly different from that in control rats (fig. 2). In order to test the idea that ondansetron might impair the release or action of CCK we studied its effect on the emptying of FOY-305 which is known to delay gastric emptying solely by release of CCK (Green et al., 1988). Ondansetron had no effect on the in~bition of gastric emp~g produced by FOY305 suggesting that it acts independently of CCK (2.40 rf:0.07 and 2.41+, 0.17 ml 5 mm-‘, pre and post 0.1 mg kg-’ ondansetron, respectively).
4. Discussion Ondansetron delayed the gastric emptying of the protein-rich peptone solution in conscious gastric fistula rats. The dose-response curve was bell-shaped and resembled that for 5-HT, antagonists described previously in other systems (Sanger, 1987). The action of ondansetron in delaying the emptying of prote~-~ch solutions was specific in that the emptying of isotonic saline, hypertonic saline, acid and FOY-305 was unaffected. Both protein and FOY-305 release CCK, but since ondansetron did not affect the emptying of FOY305 it is not acting to impair either the release or action of CCK. Instead, it seems that protein releases both 5-hydroxyt~ta~e (5-HT) and CCK. Thus 5-HT would appear to act on S-HT, receptors to enhance gastric emptying while CCK delays gastric emptying. When both 5-HTj and CCK receptors are blocked, peptone still causes some inhibition of emptying so that still further mechanisms are involved in mediating the action of protein rich solutions on upper gastrointest~al tract motility. The antiemetic properties of 5-HT, receptor antagonists have been widely reported. It is thought that these effects might be mediated by direct actions on vagal afferent fibres. In this context it is of importance to note that stimulation of gastric vagal afferents by CCK causes reflex
slowin& of gastric motility and gastric emptying in the rat (Dockray, 1989). Because the present data indicate that 5-HT and CCK influence gastric emptying in different ways, it will be of interest to determine the site of action of 5-HT, antagonists in changing gastric ~pty~g. Other recent studies have indicated that 5-I-IT3 receptor antagonists (both benzamide and non-benzamide structures) increase gastric emptying in a number of species including rat (Smith et al., 1984; Costall et al., 1987; Buchheit et al., 1989). These findings contrast with those reported here, and consequently it seems possible that 5-HT, receptors are involved in the control of upper gastrointestinal tract motility in several different ways. Taken together with previous data our findings indicate that the factors regulating the gastric emptying of protein-rich solutions are different from those controlling emptying of hyperosmolal solutions, and acid. The mech~s~ regulating gut motility responses to isotonic protein solutions are relatively complex in that several humoral mediators are involved i.e. 5HT, CCK, and at least one other substance. Given that other solutions activate different mechanisms, it is plain that the control of gut motility by a mixed meal must involve a wide diversity of reflex pathways.
Acknowledgements We are grateful to C. Carter for help in preparation of the manuscript. The work is supported by a Medical Research Council grant. Samples of FDY-305, L364.718 and ond~~~on were kindly donated by Gno Pha~~euticals. Merck, Sharp and Dohrne, and Glaxo Laboratories, respectively.
References Buc~eit, K.H., R. Gamse, A. Bertholet and H.H. Btischer, 1989, An~on~ts at serotonergic 5-HT+ceptors increase gastric emptying of solids and liquids in the rat. Gastroenterology 96. A63. Butler, A., J.M. Hill, S.J. Ireland, C.C. Jordan and M.B. Tyers, 1988, Pharmacological properties of GR38032F, a novel antagonist at 5-HT, receptors, Br. J. Pharmacol. 94, 397. Costall, B., S.J. Gunning, R.J. Naylor and M.B. Tyers, 1987. The effect of GR38032F. a novel 5-HTs-receptor antagonist
on gastric emptying in the guinea-pig. Br. J. Pharmacol. 91. 263. s. HT.. 0. Farooq and M.I. Grossman, 1975, Inhibition tic emptying is a physiological action of cholecys. GastmenteroIogy 68. 1211. R. N. Carter and S. Barnes. 1986. Evidence for adrenergic inhibition of acid secretion in the consious rat. Am. J. Physiol. 251. G615. G.J., 1989, The integrative functions of CCK in the up&; gastrointestinal tract, in: The Neuropeptide Chole_ cystokinitt (CCK), Anatomy, and Biochemistry. Receptors, pharmacology and Physiology, eds. J. Hughes. G. Dockray and G. Woodruff (Ellis Horwod, Chichester) p. 232.
Forster. E.R.. T. Green. M. Eliot, A. Bremner and G.J. Dockray, 1990, Gastric emptying in rats: role of afferent neurons and cholecystokinin, Am. J. Physiol. 258, 552. Green, T., R. Dimaline, S. Peikin and GJ. Dockray, 1988, Action of cholecystokinin antagonist L364.718 on gastric emptying in the rat, Am. J. Physiol. 255, G685. Sanger, G.J., 1987, Increased gut cholinergic activity and antagonism of 5-hydroxytryptamine M-receptors by BRL 24924: potential clinical importance of BRL 24924, Br. J. Phannacol. 91, 77. Smith, W.L., D.A. Droppleman, R.L. Gregory and R.S. Alphin. 1984, Dazopride (AHR-5531): A novel gastrokinetic agent, Gastroenterology 86, 1257.
235
Elsevier EJP 20142
Short communication
The effect of ondansetron on gastric emptying in the conscious rat Eliot R. Forster and Graham J. Dockray MRC Secretov Control Group, PhysiologicalLaboratory. Universityof Liverpool, Liverpool, U.K.
Received 2 October 1990, accepted 9 October 1990
The 5-HT, receptor antagonist ondansetron (GR38032F) enhanced the action of a protein-rich solution in delaying gastric emptying in the conscious gastric fistula rat, but had no effect on the emptying of isotonic or hypertonic saline, acid or FOY-305 which delays emptying by release of cholecystokinin (CCK). The specific CCK-A antagonist
(L-364,718)increased gastric emptying of protein-rich meals. L364,718 also increased emptying in the presence of ondansetron. They indicate that protein-rich meals release both CCK and 5-hydroxytryptamine which act in different ways to control gastric motility. Gastric emptying; 5-HT, receptors; Protein meals; Ondansetron
1. Introduction
Several different pathways are now known to be involved in controlling the gastric emptying of different types of liquid test meal (Green et al., 1988; Forster et al., 1990). In conscious gastric fistula rats, protein-rich meals in particular delay gastric emptying by a mechanism sensitive to the cholecystokinin-A (CCK-A) receptor antagonist L364,718 and dependent on (a) passage of the solution beyond the proximal duodenum and (b) intact visceral afferents; the emptying of this type of meal is not, however, dependent on an intact pylorus. Hypertonic solutions and acid also inhibit gastric emptying by mechanisms that at least in part, require intact visceral afferents, but they are not sensitive to CCK antagonists and differ in their requirement for either an intact pylorus or passage of the solution beyond the duodenum. Although the data clearly suggest a role for CCK
Correspondence to: G.J. Dockray, Physiological Laboratory, University of Liverpool, P.O. Box 147, Liverpool L69 3BX, U.K.
in controlling the gastric emptying of protein the other humoral mediators that might be involved in controlling emptying of protein remain poorly characterized. In the present study we have examined the role of 5-HT, receptors in controlling gastric emptying of various liquid test meals in the consious gastric fistula rat. The data suggest that the S-HT, antagonist ondansetron (Butler et al., 1988) specifically enhances the action of peptone in delaying gastric emptying.
2. Materials and methods
2.I. Animals
Male Wistar rats (200-250 g, n = 8) were anaesthetised with sodium pentobarbitone (Sagatal, 60 mg kg- ’ i.p.) and fitted with stainless steel cannulas in the body of the stomach as previously described (Dimaline et al., 1986). At least two weeks were allowed for recovery before commencing experiments.
0014-2999/90/$03.50 0 1990 Elsevier Science Publishers B.V. (Biomedical Division)
.F
_.a.
~nimaIs were deprived of solid food overnight lowed free access to water. Trained rats were lightly restrained in Rollman-type cages and the stomach flushed with warm saline. Drugs and test solutions were administered directly into the the cannula. Ondansetron (GR38032 .K.) and L-364,718 (Merck, Sharp and ere freshly prepared immediately prior stration in isotonic saline (ondansetron) methyl cellulose in isotonic saline (L18). Doses of ondansetron ranging from loor the vehicle alone, were given via the cannula 2 h prior to commencing the emptying trials. L364.718 was given at a dose of 1 mg kg- ’ via the cannula 30 min prior to commencing the emptying trials. At the end of this period emptying of the following test solutions (3 ml) was studied: 300 mOsm kg-” NaCl, 900 mOsm kg-’ NaCl. 50 mM HCl, 20 mg ml-’ FOY 305 (camostat m&late, Ono Pharmaceuticals) and 4.5% (w/v) meat peptone II (Sigma). Phenol red (60 mg kg-‘) was added t o each solution as a non-absorbable dilution marker. Gastric emptying was determined from the volume and phenol red concentration of fluid recovered from the cannula 5 min after instillation of the test meal into the stomach (Debas et al., 1975; Green et al., 1988).
i
.c
5
2.5-
2 P
B i=
2-
zi Y 3 P
l-5-
T
OJ
,
0.01
/
I
0.03
0.1
1
0.3
1 1
ONDANSETRON (mg.kg”) Fig. 1. The volume of peptone emptied from the stomach after 5 min in a group of conscious gastric fistula rats treated with ondansetron (0.01-1.0 mg kg-‘). Each dose of ondansetron was studied on a separate day. and animals were used no more than twice weekly. The open symbol to the left shows the emptying of peptone in vehicle-treated rats. Points are means i SE., n = 8; * indicates significantly different from control P < 0.05, paired t-test.
effect of ondansetron on peptone was maximal at a dose of 0.1 mg kg-‘; at lower and higher doses it.s effects declined giving a bell-shaped dose-re-
J-3. Statistics The results are expressed as means f SE. Comparisons were made using a paired t-test.
Hyperosmolal saline, acid and peptone all delayed gastric emptying compared to isotonic saline, as previously described (Green et al., 1988; Forster et al., 1990). Ondansteron significantly increased the delay in gastric emptying produced by peptone, but had no effect on the emptying of hyperosmolal saline (control, 2.22 f 0.07 VS. ondansetron, 1.89 &-0.19 ml 5 min-*), isotonic saline (3.05 & 0.11 vs. 3.14 * 0.12 ml 5 mm-‘) or acid (1.95 f 0.08 vs. 1.95 + 0.08 ml 5 min-‘). The
Fig. 2. The rate of emptying of peptone in 5 mm in control rats and after L364,718 (1 mg kg-‘), ondansetron (a.1 mg kg-‘) and both L364,718 and ondansetron. Bars are means*S.E., n = 8. See fig. 1 for further information. Points xe means* SE., n = 8; * indicates significantly different from control P < 0.05, paired t-test.
237
sponse curve (fig. 1). The emptying of peptone is known to be increased following administration of the CCK antagonist, L364,718 (Green et al., 1988). In rats treated with the combination of L-364,718 and ondansetron (0.1 mg kg-‘) the rate of emptying of peptone was not si~fic~tly different from that in control rats (fig. 2). In order to test the idea that ondansetron might impair the release or action of CCK we studied its effect on the emptying of FOY-305 which is known to delay gastric emptying solely by release of CCK (Green et al., 1988). Ondansetron had no effect on the in~bition of gastric emp~g produced by FOY305 suggesting that it acts independently of CCK (2.40 rf:0.07 and 2.41+, 0.17 ml 5 mm-‘, pre and post 0.1 mg kg-’ ondansetron, respectively).
4. Discussion Ondansetron delayed the gastric emptying of the protein-rich peptone solution in conscious gastric fistula rats. The dose-response curve was bell-shaped and resembled that for 5-HT, antagonists described previously in other systems (Sanger, 1987). The action of ondansetron in delaying the emptying of prote~-~ch solutions was specific in that the emptying of isotonic saline, hypertonic saline, acid and FOY-305 was unaffected. Both protein and FOY-305 release CCK, but since ondansetron did not affect the emptying of FOY305 it is not acting to impair either the release or action of CCK. Instead, it seems that protein releases both 5-hydroxyt~ta~e (5-HT) and CCK. Thus 5-HT would appear to act on S-HT, receptors to enhance gastric emptying while CCK delays gastric emptying. When both 5-HTj and CCK receptors are blocked, peptone still causes some inhibition of emptying so that still further mechanisms are involved in mediating the action of protein rich solutions on upper gastrointest~al tract motility. The antiemetic properties of 5-HT, receptor antagonists have been widely reported. It is thought that these effects might be mediated by direct actions on vagal afferent fibres. In this context it is of importance to note that stimulation of gastric vagal afferents by CCK causes reflex
slowin& of gastric motility and gastric emptying in the rat (Dockray, 1989). Because the present data indicate that 5-HT and CCK influence gastric emptying in different ways, it will be of interest to determine the site of action of 5-HT, antagonists in changing gastric ~pty~g. Other recent studies have indicated that 5-I-IT3 receptor antagonists (both benzamide and non-benzamide structures) increase gastric emptying in a number of species including rat (Smith et al., 1984; Costall et al., 1987; Buchheit et al., 1989). These findings contrast with those reported here, and consequently it seems possible that 5-HT, receptors are involved in the control of upper gastrointestinal tract motility in several different ways. Taken together with previous data our findings indicate that the factors regulating the gastric emptying of protein-rich solutions are different from those controlling emptying of hyperosmolal solutions, and acid. The mech~s~ regulating gut motility responses to isotonic protein solutions are relatively complex in that several humoral mediators are involved i.e. 5HT, CCK, and at least one other substance. Given that other solutions activate different mechanisms, it is plain that the control of gut motility by a mixed meal must involve a wide diversity of reflex pathways.
Acknowledgements We are grateful to C. Carter for help in preparation of the manuscript. The work is supported by a Medical Research Council grant. Samples of FDY-305, L364.718 and ond~~~on were kindly donated by Gno Pha~~euticals. Merck, Sharp and Dohrne, and Glaxo Laboratories, respectively.
References Buc~eit, K.H., R. Gamse, A. Bertholet and H.H. Btischer, 1989, An~on~ts at serotonergic 5-HT+ceptors increase gastric emptying of solids and liquids in the rat. Gastroenterology 96. A63. Butler, A., J.M. Hill, S.J. Ireland, C.C. Jordan and M.B. Tyers, 1988, Pharmacological properties of GR38032F, a novel antagonist at 5-HT, receptors, Br. J. Pharmacol. 94, 397. Costall, B., S.J. Gunning, R.J. Naylor and M.B. Tyers, 1987. The effect of GR38032F. a novel 5-HTs-receptor antagonist
on gastric emptying in the guinea-pig. Br. J. Pharmacol. 91. 263. s. HT.. 0. Farooq and M.I. Grossman, 1975, Inhibition tic emptying is a physiological action of cholecys. GastmenteroIogy 68. 1211. R. N. Carter and S. Barnes. 1986. Evidence for adrenergic inhibition of acid secretion in the consious rat. Am. J. Physiol. 251. G615. G.J., 1989, The integrative functions of CCK in the up&; gastrointestinal tract, in: The Neuropeptide Chole_ cystokinitt (CCK), Anatomy, and Biochemistry. Receptors, pharmacology and Physiology, eds. J. Hughes. G. Dockray and G. Woodruff (Ellis Horwod, Chichester) p. 232.
Forster. E.R.. T. Green. M. Eliot, A. Bremner and G.J. Dockray, 1990, Gastric emptying in rats: role of afferent neurons and cholecystokinin, Am. J. Physiol. 258, 552. Green, T., R. Dimaline, S. Peikin and GJ. Dockray, 1988, Action of cholecystokinin antagonist L364.718 on gastric emptying in the rat, Am. J. Physiol. 255, G685. Sanger, G.J., 1987, Increased gut cholinergic activity and antagonism of 5-hydroxytryptamine M-receptors by BRL 24924: potential clinical importance of BRL 24924, Br. J. Phannacol. 91, 77. Smith, W.L., D.A. Droppleman, R.L. Gregory and R.S. Alphin. 1984, Dazopride (AHR-5531): A novel gastrokinetic agent, Gastroenterology 86, 1257.
