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Inhibition by met-enkephalin of peristaltic activity in the guinea pig ileum, and its reversal by naloxone
European Journal of Pharmacology, 41 (1977)341--342
341
© Elsevier/North-Holland Biomedical Press, Amsterdam -- Printed in The Netherlands
Rapid communication INHIBITION BY MET-ENKEPHALIN OF PERISTALTIC ACTIVITY IN THE GUINEA PIG ILEUM, AND ITS REVERSAL BY NALOXONE JAN M. VAN NUETEN, JAN M. VAN REE* and PAUL M. VANHOUTTE**
Department of Pharmacology, Research Laboratories, Janssen Pharmaceutica, B-2340 Beerse, Belgium Received 7 December 1976, accepted 8 December 1976
When the guinea pig ileum is distended for prolonged periods, its peristaltic reflex activity decreases gradually ('fatigue'); this phenomenon is reversed by naloxone (Van Nueten et al., 1976). Parallel to the fatigue, an unknown endogenous substance is liberated which produces a similar pattern of inhibition of peristaltic reflex activity in the nonfatigued ileum. The effect of the endogenous substance is also inhibited by naloxone, which prompted the conclusion that it is responsible for the decreased peristalsis in fatigued preparations (Van Nueten et al., 1976). The reversal by naloxone suggests that the endogenous substance is acting, directly or indirectly, on opiate receptors. There is a high degree of similarity between opiate receptors in the brain and in the ileum (Terenius, 1975). Enkephalin, described as an endogenous ligand for opiate receptors, has been found in both the brain and the ileum (Hughes, 1975); its most active component on the guinea pig ileum is the pentapeptide methionine-enkephalin (met-enkephalin) (Hughes et al., 1975). The present experiments demonstrate that the inhibitory effect of met-enkephalin on the peristaltic reflex activity of the guinea pig ileum is reversed by naloxone. *Rudolf Magnus Institute for Pharmacology, University of Utrecht, Medical Faculty, Utrecht, The Netherlands. **Department of Internal Medicine, Universitaire Instelling Antwerpen, Wilrijk, Belgium.
The peristaltic reflex was studied on the isolated guinea pig ileum by means of a modified Trendelenburg method (Van Nueten et al., 1973). Briefly, an ileum segment is suspended in Tyrode solution of 37°C at pH 7.4 with a gas mixture 95 % 02--5 % CO2. Pressure is increased in the lumen of the segment from --5 to 20 mm H2 O, and is maintained at this level for the duration of the experiment. This pressure elicits rhythmic peristaltic reflex activity, i.e. coordinated contractions of both muscle layers, leading to peristaltic waves and the expulsion of the intraluminal fluid. Parameters recorded are the activity of the longitudinal muscle, measured as tension changes, the activity of the circular muscle, measured as development of intraluminal pressure, and the fluid volume displacement. In the guinea pig ileum segment, met-enkephalin causes a dose-dependent inhibition of the rhythmic peristaltic reflex activity induced by increasing the distending pressure. At the threshold concentration (10 ng/ml) the inhibitory effect of met-enkephalin is characterized by decreased activity of both the longitudinal and circular muscle layers resulting in increased filling; the frequency of the peristaltic movements is decreased. At the higher concentrations, intermittent (at 40 ng/ml; fig. 1A) or prolonged (at 160, fig. 1B, and 630 ng/ml) interruptions of the peristaltic activity are observed. At all concentrations tested, the inhibitory effect of
342 A r
NALOXONE li~ nn/rnl r - -
MET- ENKEPHALIN ~,L0 n g / m l
+
B MET-'ENKEPHALIN ,1160 ng ~'ml
+ NALOXONE 10ng/ml, r
a
'
b
'
'"'~
1 I' t i! I,'
C
observed with met-enkephalin resembles both the spontaneous decrease in peristaltic activity seen in fatigued preparations (fig. 1C) and the depressant effect of the unidentified endogenous substance liberated by such preparations. The latter two effects are also reversed by naloxone (Van Nueten et al., 1975). On prolonged exposure to met-enkephalin, a partial recovery of the peristaltic activity is noted which can be attributed to the rapid enzymatic destruction of the pentapeptide in this preparation. This may also explain, in the case of fatigue, the alternation of periods of complete interruption of peristalsis with bursts of peristaltic waves. The present experiments are thus consistent with the possible role of enkephalin as a mediator or a modulator of fatigue in the guinea pig ileum. T h e y do, however, not rule out that other endogenous substances, including polypeptides or adenine nucleotides, may be involved.
i115 min.
Acknowledgements
c ~I~LU2SJJ~'~LIJ~-JJ3'J.~JJ~--UJ---~ H I rain.
Fig.1. Peristaltic reflex activity of the guinea pig ileum induced by increasing distension pressure. A en B: inhibition of the activity by met-enkephalin At 40 ng/ml: intermittent interruptions (A); at 160 ng/ml: complete inhibition (B). Partial recovery occurs after 5--6 rain; naloxone restores the normal activity. • C: fatigue caused by continuous distension of the lumen ( II : interruption of the tracing for a period of 15 rain), a: longitudinal muscle tension; b: volume displacement; c: circular muscle activity, measured as intraluminal pressure.
met-enkephalin is completely reversed by naloxone 10 ng/ml (fig. 1A and B). Similar results have been described for the electrically driven guinea pig ileum (Hughes et al., 1975}. The pattern of inhibition
We thank Mr. L. Helsen for his skilful technical assistance and Dr. Paul A.J. Janssen for his continued interest.
References Hughes, J., 1975, Isolation of an endogenous compound from the brain with pharmacological properties similar to morphine, Brain Res. 88, 295. Hughes, J., T.W. Smith, H.W. Kosterlitz, L.A. Fothergill, B.A. Morgan and H.R. Morris, 1975, Identification of two related pentapentides from the brain with potent opiate agonist activity, Nature 258,577. Terenius, L., 1975, Comparison between narcotic 'receptors' in the guinea-pig ileum and the rat brain, Acta Pharmacol. Toxicol. 3 7 , 2 1 1 . Van Nueten, J.M., H. Geivers, J. Fontaine and P.A.J. Janssen, 1973, An improved method for studying peristalsis in the isolated guinea-pig ileum, Arch. Intern. Pharmacodyn. Therap. 203,411. Van Nueten, J.M., P.A.J. Janssen and J. Fontaine, 1976, Unexpected reversal effects of naloxone on the guinea pig ileum, Life Sci. 18, 803.
341
© Elsevier/North-Holland Biomedical Press, Amsterdam -- Printed in The Netherlands
Rapid communication INHIBITION BY MET-ENKEPHALIN OF PERISTALTIC ACTIVITY IN THE GUINEA PIG ILEUM, AND ITS REVERSAL BY NALOXONE JAN M. VAN NUETEN, JAN M. VAN REE* and PAUL M. VANHOUTTE**
Department of Pharmacology, Research Laboratories, Janssen Pharmaceutica, B-2340 Beerse, Belgium Received 7 December 1976, accepted 8 December 1976
When the guinea pig ileum is distended for prolonged periods, its peristaltic reflex activity decreases gradually ('fatigue'); this phenomenon is reversed by naloxone (Van Nueten et al., 1976). Parallel to the fatigue, an unknown endogenous substance is liberated which produces a similar pattern of inhibition of peristaltic reflex activity in the nonfatigued ileum. The effect of the endogenous substance is also inhibited by naloxone, which prompted the conclusion that it is responsible for the decreased peristalsis in fatigued preparations (Van Nueten et al., 1976). The reversal by naloxone suggests that the endogenous substance is acting, directly or indirectly, on opiate receptors. There is a high degree of similarity between opiate receptors in the brain and in the ileum (Terenius, 1975). Enkephalin, described as an endogenous ligand for opiate receptors, has been found in both the brain and the ileum (Hughes, 1975); its most active component on the guinea pig ileum is the pentapeptide methionine-enkephalin (met-enkephalin) (Hughes et al., 1975). The present experiments demonstrate that the inhibitory effect of met-enkephalin on the peristaltic reflex activity of the guinea pig ileum is reversed by naloxone. *Rudolf Magnus Institute for Pharmacology, University of Utrecht, Medical Faculty, Utrecht, The Netherlands. **Department of Internal Medicine, Universitaire Instelling Antwerpen, Wilrijk, Belgium.
The peristaltic reflex was studied on the isolated guinea pig ileum by means of a modified Trendelenburg method (Van Nueten et al., 1973). Briefly, an ileum segment is suspended in Tyrode solution of 37°C at pH 7.4 with a gas mixture 95 % 02--5 % CO2. Pressure is increased in the lumen of the segment from --5 to 20 mm H2 O, and is maintained at this level for the duration of the experiment. This pressure elicits rhythmic peristaltic reflex activity, i.e. coordinated contractions of both muscle layers, leading to peristaltic waves and the expulsion of the intraluminal fluid. Parameters recorded are the activity of the longitudinal muscle, measured as tension changes, the activity of the circular muscle, measured as development of intraluminal pressure, and the fluid volume displacement. In the guinea pig ileum segment, met-enkephalin causes a dose-dependent inhibition of the rhythmic peristaltic reflex activity induced by increasing the distending pressure. At the threshold concentration (10 ng/ml) the inhibitory effect of met-enkephalin is characterized by decreased activity of both the longitudinal and circular muscle layers resulting in increased filling; the frequency of the peristaltic movements is decreased. At the higher concentrations, intermittent (at 40 ng/ml; fig. 1A) or prolonged (at 160, fig. 1B, and 630 ng/ml) interruptions of the peristaltic activity are observed. At all concentrations tested, the inhibitory effect of
342 A r
NALOXONE li~ nn/rnl r - -
MET- ENKEPHALIN ~,L0 n g / m l
+
B MET-'ENKEPHALIN ,1160 ng ~'ml
+ NALOXONE 10ng/ml, r
a
'
b
'
'"'~
1 I' t i! I,'
C
observed with met-enkephalin resembles both the spontaneous decrease in peristaltic activity seen in fatigued preparations (fig. 1C) and the depressant effect of the unidentified endogenous substance liberated by such preparations. The latter two effects are also reversed by naloxone (Van Nueten et al., 1975). On prolonged exposure to met-enkephalin, a partial recovery of the peristaltic activity is noted which can be attributed to the rapid enzymatic destruction of the pentapeptide in this preparation. This may also explain, in the case of fatigue, the alternation of periods of complete interruption of peristalsis with bursts of peristaltic waves. The present experiments are thus consistent with the possible role of enkephalin as a mediator or a modulator of fatigue in the guinea pig ileum. T h e y do, however, not rule out that other endogenous substances, including polypeptides or adenine nucleotides, may be involved.
i115 min.
Acknowledgements
c ~I~LU2SJJ~'~LIJ~-JJ3'J.~JJ~--UJ---~ H I rain.
Fig.1. Peristaltic reflex activity of the guinea pig ileum induced by increasing distension pressure. A en B: inhibition of the activity by met-enkephalin At 40 ng/ml: intermittent interruptions (A); at 160 ng/ml: complete inhibition (B). Partial recovery occurs after 5--6 rain; naloxone restores the normal activity. • C: fatigue caused by continuous distension of the lumen ( II : interruption of the tracing for a period of 15 rain), a: longitudinal muscle tension; b: volume displacement; c: circular muscle activity, measured as intraluminal pressure.
met-enkephalin is completely reversed by naloxone 10 ng/ml (fig. 1A and B). Similar results have been described for the electrically driven guinea pig ileum (Hughes et al., 1975}. The pattern of inhibition
We thank Mr. L. Helsen for his skilful technical assistance and Dr. Paul A.J. Janssen for his continued interest.
References Hughes, J., 1975, Isolation of an endogenous compound from the brain with pharmacological properties similar to morphine, Brain Res. 88, 295. Hughes, J., T.W. Smith, H.W. Kosterlitz, L.A. Fothergill, B.A. Morgan and H.R. Morris, 1975, Identification of two related pentapentides from the brain with potent opiate agonist activity, Nature 258,577. Terenius, L., 1975, Comparison between narcotic 'receptors' in the guinea-pig ileum and the rat brain, Acta Pharmacol. Toxicol. 3 7 , 2 1 1 . Van Nueten, J.M., H. Geivers, J. Fontaine and P.A.J. Janssen, 1973, An improved method for studying peristalsis in the isolated guinea-pig ileum, Arch. Intern. Pharmacodyn. Therap. 203,411. Van Nueten, J.M., P.A.J. Janssen and J. Fontaine, 1976, Unexpected reversal effects of naloxone on the guinea pig ileum, Life Sci. 18, 803.