Acta Physiol Scand 1992, 146, 377-383
Pain-induced inhibition of gastric motility is mediated by adrenergic and vagal non-adrenergic reflexes in the rat L. BOJO, J. C A S S U T O and P. N E L L G A R D Department of Internal Medicine and Department of Physiology, University of Goteborg, Department of Clinical Physiology, Central Hospital Karlstad, and Department of Anaesthesiology, Central Hospital, Molndal, Sweden.
BoJO, L., CASSUTO, J. & NELLG~RD, P. 1992. Pain-induced inhibition of gastric motility is mediated by adrenergic and vagal non-adrenergic reflexes in the rat. Acta Physiol Scand 146, 377-383. Received 15 October 1991, accepted 18 May 1992. ISSN 00016772. Department of Internal Medicine and of Physiology, University of Goteborg, Department of Clinical Physiology, Central Hospital, Karlstad, and Department of Anaesthesiology, Central Hospital, Molndal, Sweden. Painful stimuli have been shown to inhibit gastric motility in animal experiments and delay gastric emptying in humans. The aim of the present study was to investigate in detail mechanisms involved in pain-induced gastric inhibition. Pain stimulation by exerting pressure on a testicle induced a prompt gastric relaxation which lasted throughout the period of stimulation. Pain-induced gastric relaxation was significantly reduced by the selective a-1 blocker, prazosin, and by the non-selective /Iblocker, propranolol. Similarly pain-induced inhibition of gastric tone was significantly reduced by bilateral cervical vagotomy. In contrast, gastric relaxation following pain stimulation was significantly potentiated by the selective adrenergic 01-2 blocker, yohimbine. Combined administration of prazosin and propranol followed by bilateral cervical vagotomy abolished gastric relaxation in response to pain stimulation. In conclusion, gastric relaxation in response to painful stimulation was found to be reflex-mediated via sympathetic neurons acting on a-1 and preceptors and possibly also via vagal non-adrenergic fibres. Pain-induced inhibition of gastric tone was significantly increased by yohimbine. It is suggested that yohimbine by blocking presynaptic inhibitory receptors on adrenergic neurons facilitates the release of noradrenaline in response to pain stimulation.
Key words : pain, vagus nerve, adrenergic antagonists, gastrointestinal motility.
Various conditions involving pain such as trauma, intestinal obstruction, appendicitis, myocardial infarction and major surgery are often accompanied by nausea and vomiting (Feldman 1983). A feared complication is aspiration of gastric contents which may cause hypoxia and pneumonitis, particularly in patients with lowered level of consciousness (Naudy & Salducci 1984). Correspondence : Dr Leif Bojo, Department of Clinical Physiology, Central Hospital, S-651 85 Karlstad, Sweden.
Nausea and vomiting are associated with perturbed gastric motility characterized by an inhibition of gastric tone and a reduction of normal motility which, during vomiting, is replaced by antiperistalsis (Smith & Brizzee 1961, Feldman 1983). Nociceptive intra-abdominal stimulation has been shown to induce inhibition of gastric tone both via adrenergic and vagal non-adrenergic reflexes (Glise et al. 1980, Glise & Abrahamsson 1980. T h e aim of the present study was to investigate the neural pathways and receptors mediating gastric relax-
377
378
L. Bo30 et al. Prazosin 150 pg kg' i.v. Blood pressure mm
Hg
200
I
50
50
450
450
350
350
600
600
900
900
Heart rate
(iastric volume
Pf?
Time (min)
Fig. 1. Gastric relaxation induced bl- pain-stimulation is significantly reduced after the administration of 2-1 blocker prazosin compared to control
tion of changes in gastric volume on a Grass polygraph (Grass Instr. Quincy, MA, USA). Changes in the volume of the reservoir were registered as inverse to changes in gastric volume. Calibration was performed b!- adding a known volume of water to the reservoir X I I.:T HOD S after temporaril!- closing the tube connecting the Experiments were performed on 35 male Spragueballoon to the reservoir. Dawley rats weighing 300-350 g. 'The experimental LVhen catheters and the balloon had been introprotocol was approved b>-the Animal Ethics Ctrmduced, the animals were allowed to recmer from mittee. Animals were housed for at least 7 dal-s prior surgerl- for a t least 30 min before starting the to the experiments in a ventilated and temperatureexperiment. Pain stimulation (PS) was performed by controlled room and had water and food ad libitum. exerting a constant pressure of 120-130 mmHg during The day-night c!cle was constants at 12-h periods. 1 min on one testicle by- the use of a clamp. T h e clamp Before the experiments the animals were fasted for \?-asmade of two parallel plastic slides ( 2 x 3 cm), each l l t l 5 h with free access to water. Anaesthesia \$-as with a central excavation approximately the size and induced with methohexital (Brietdl, Lilley, France, shape of the testicle and held together by two 60 mg kg- I ) intraperitoneall!- and maintained b!- a adjustable coils. continuous intravenous infusion of chloralose Each experiment was started by testing the degree (l..&mg kg-' min-I). A tracheostomy w s performed of gastric relaxation induced by PS. This procedure and a tracheal cannula was inserted to secure free Itas repeated u-ith at least 10 min between pain airway. Body temperature was kept constant at 38 "C stimulations. When gastric relaxation differed less by a heating pad coupled to a temperature sensor in than 20°, following two consecutive stimulations and the pharynx. Blood pressure was monitored using a when gastric basal tone had returned to a stable level pressure transducer (Statham P23AC Statham Instr.. and remained there for at least 10 min the experimental Osnard, C.4, USA) connected to a cannula in a drug was administered intravenously. Pain stimulation femoral artery. Heart rate (HR) a a s monitored, using was then repeated 1.5 min after administration of the a heart rate meter. drug. .4fter finishing the experiments, the volume of Gastric volume was recorded at constantl!- loa- the gastric balloon was determined by aspiration. The intragastric pressure ( 2 cmH,O) bj- a aater-filled, animals were killed by an intravenous injection of thin-walled flaccid balloon introduced 1-ia the esophasaturated KC1 and the location of the balloon in the gus. T h e balloon was connected b!- a polyethene stomach w a s controlled by dissection. catheter (PE 90) to a water reservoir mounted on a The drugs used were: prazosin (Peder Inc., New force-transducer which allowed continuous registra'l-ork City, NY, USA), propranolol (JCI Pharma-
ation in response to noxious stimulation in anaesthetized rats.
Pain-induced gastric relaxation
379
Table 1. Effects of adrenergic antagonists and vagotomy on basal gastric volume. Mean$SEM Basal GV (ml)
Treatment
Dose
n
1.1& 0.1 1.3k0.1 1.1 & 0.1 1.okn.i
Prazosin Yohimbine Propranolol BCV
1SO ,ug kg-' 1 mg kg-' 1 mg kg-'
7 6 8 6
__
Effect of treatment on GV (PO
+ 10k30 +200$.50
ns. < 0.05 n.s. < 0.05
-90f40 +290f100
GV = Gastric volume; RCV = Bilateral cervical vagotomy; Reduced gastric volume.
P-value
+ = Increased
gastric volume; and
-
=
Table 2. Relative gastric relaxation induced by pain stimulation after drug administration and vagotomy. Data are given as a percentage of the control. Mean k SEM
Treatment
n
Prazosin Yohimbine Propranolol Prazosin and propranolol BCV Prazosin, propranolol and BCV
7 6 8 6 6 8
Gastric relaxation induced by PS after treatment in percent of control
P-value
< 0.05 < 0.05 < 0.01 < 0.05
f67f16
+ 144f33 +56fll +48_f7
< 0.05
+42i8 -4f5
< 0.01
PS = Pain stimulation; BCV = Bilateral cervical vagotomy; Reduced gastric volume.
ceuticals, Macclesfield, UK) and yohimbine (Merck & Co. Inc., Rahway, NJ, USA). All drugs were
administered intravenously in a femoral vein. Adequate doses of the adrenergic antagonists against the effects of the corresponding agonists on circulation and gastric tone have been tested previously (Bojo et al. 1991). Statistical analysis was performed using the signed rank test. Differences resulting in P-values less than 0.05 were considered significant. Values are given as mean fSEM.
RESULTS Pain stimulation (PS) induced an immediate and pronounced gastric relaxation and a partial or complete inhibition of the spontaneous antral phasic contractions. T h e inhibition of gastric tone lasted throughout the period of stimulation and returned to the pre-stimulation level within 5 min of termination of PS (Fig. 1).
+ = Increased gastric volume; and
-
=
Data regarding basal gastric volumes and the effects of drugs and bilateral cervical vagotomy on gastric volume are presented in Table 1. Pain stimulation before treatment in all 35 experiments increased gastric volume by 227 & 25 pl
( P < 0.01). Prazosin (150 ug kg-l i.v.), an a-1 antagonist, did not cause a significant change in basal gastric volume (Table 1) but significantly reduced paininduced inhibition of gastric tone as compared to the control (n = 7, P < 0.05) (Fig. 1 & Table 2). Propranolol (1 mg kg-' iv.), an unselective pblocker, did not significantly change basal gastric volume (Table 1). After administration of propranolol gastric reflex relaxation following PS was significantly reduced compared to the control ( n = 7, P < 0.05) (Fig. 2 & Table 2). Combined administration of prazosin (150 ug kg-' i.v.) and propranolol (1 mg kg-l) significantly reduced gastric relaxation in te-
.NO
L. BoJO et al. After Prazosin 150 ug kg-1
Fig. 2. After the administration of the 2-1 blocker prazosin pain-induced inhibition of gastric tone is further reduced after i.\. injection of the /l-blocker propranolol. Bilateral cervical vagotomy
I Hlinwt preswre
m m Hg
Heart rate
(;astric volume
800
800
lo00
loo0
1200
1200
UI
'lime (rnin)
Fig. 3. Pain-induced gastric relasation is significantly reduced bl- bilateral cervical vagotomy
sponse to pain stimulation however, a significant response to PS remained in all experiments ( 1 1 = h) (Table 2 ) . Yohimbine (1 mg kg i.v.), an 3 - 2 antagonist, significantlv reduced basal gastric tone and significantly increased pain-induced inhibition of gastric tone (n = 6) (Table 2 ) . Bilateral cerx ical \-agotom>-induced a gastric relaxation in all the experiments (?I = 7 , P < 0.0.5) ('Table 1). The gastric response to PS was
significantly reduced by vagotomy ( a = 7, P < 0.05) (Fig. 3 & Table 2 ) . Combined administration of prazosin and propranolol, at the doses given above, followed by bilateral cervical vagotomy abolished the gastric relaxatory response to pain stimulation (ri = 6, P < 0.05) (Fig. 4 & Table 2). In two out of six experiments in this group pain-stimulation induced a slight gastric contraction following the treatment above.
Pain-induced gastric relaxation
38 1
After Prazosin 150 ug kg-1 and Propranolol 1 mg kg-1 Bilateral cervical vagutumy
I Hlood pressure 111111 Hg
Heart rate 600
900
Time ImhJ
Fig. 4. Pain-induced reflex inhibition of gastric tone is abolished after the combined administration of the a-1 blocker, prazosin, and the a-blocker, propranolol, followed by bilateral cervical vagotomy.
DISCUSSION Painful or otherwise noxious stimuli to the gut or other parts of the body have been shown to inhibit gastric motility. In man, cold pain stimulation has been shown to decrease gastric emptying (Thompson et al. 1983) and reduce the amplitude of the antral phasic pressure in response to a solid meal (Stanghellini et al. 1983). I n the cat, nociceptive stimulation of the gut was shown to inhibit gastric motility via both adrenergic and spinovagal, non-adrenergic reflexes (Glise et al. 1980, Glise & Abrahamson 1980). In the present study a standardized pressure on a testicle was used as a model for pain stimulation. The testis is regarded as a visceral organ since it descends during foetal life to the scrotum from the abdominal cavity (Langman 1975, Kumazawa & Mizumara 1977). The presence of polymodal sensory receptors responsive to noxious stimulation by mechanical, thermal and chemical means have been demonstrated in the testis of the dog by Kumazawa & Mizumura (1977). Pressure applied on the scrotum and testicles is therefore likely to activate both visceral and somatic pain-receptors. In vitro studies have shown adrenergic inhibition of gastric smooth muscle tone to be mediated both by a- and ,&receptors in man
(Bennet & Whitney 1966, Haffner et al. 1969) and in the guinea-pig (Bailey 1971, Guimaraes 1969). Administration of the non-selective /3agonist isoprenaline and the p-2 selective agonist salbutamol significantly prolonged gastric emptying in healthy volunteers (Rees et al. 1980). In vivo experiments in rats showed that gastric relaxation was induced by stimulation of a-1 and /3-2 receptors (Bojo et al. 1991). Concordant results were obtained in the present experiments showing that pain-induced gastric relaxation was significantly reduced both by the a-1 receptor blocker, prazosin, and by the non-selective /3receptor antagonist, propranolol (Table 2). These results suggest that pain-induced gastric relaxation is mediated by activation both of a-1 and p-2 adrenergic receptors. The possibility of pain-induced activation of preganglionic sympathetic efferents to the adrenal glands causing a release of adrenaline which contributes to the gastric relaxation alongside with noradrenaline can not be ruled out. a-2 adrenergic receptors are generally located presynaptically on adrenergic nerve terminals and serve to inhibit noradrenaline release from postganglionic neurons (Burks 1987). I n the present study yohimbine, an a-2 blocker, induced a significant increase in gastric relaxation following pain-stimulation (Table 2). This may suggest that yohimbine, by blocking inhibitory
382
L. Bojii e t al.
presynaptic 2-2 receptors, facilitates the release of noradrenaline from inhibitory gastric efferent neurons in response to pain stimulation thus potentiating gastric relaxation. T h e vagal neri-es carry to thc stomach, besides cholinergic excitatory fibres, non-adrenergic inhibitory fibres. These inhibitory neurons have been shown to mediate receptive relaxation in response to activation of mechanorecptors in the pharynx or the oesophagus (-Abrahanison & Jansson 1969) and to inhibit gastric tone in response to ischaemic or mechanical stimulation of the heart (L4brahamsson & Thoren 1972). \lye have recently presented data, using the present rat model, showing that colonic distension induced gastric relaxation via a vagal nonadrenergic pathway (Bojo & Cassuto 1992). I n the present study bilateral cervical vagotomlsigniticantl!- increased gastric volume resulting in a decreased gastric capacity of further relaxation. T h e gastric response to pain stimulation was reduced following vagotomy. T h e possibilit!- that this reduction was due to the relaxatory effect of vagotomj- itself cannot be ruled out. IIowever, our results show that the pain-induced gastric relaxation remaining after combined z-l and /,’-adrenergic blockade was abolished after vagotomy suggesting a role for the \-agal nerves in pain-induced inhibition of gastric motilitJ-.T h i s vagall!--mediated inhibition of gastric tone could be due to decreased excitatory cholinergic impulse activity and/or increased actirity in the inhibitory \-agal efferent neurons to the stomach. Experimental support for2 reciprocal arrangement in impulse activation was given b!- Grundy &- Scratcherd (1982). T h e y demonstrated in the anaesthetized ferret that intestinal distension induced an increased impulse activity in some vagal efferent fibres and a decreased activity in other vagal efferent neurons. Previous studies in the cat showed that sympathetic inhibition of gastric tone required a spontaneous background discharge in the vagi or artificially induced activitv by electrical stimulation of the vagal nerves (Jansson 1969, Glise 8i .%brahamsson 1980, Glise rt 01. 1980). Electric stimulation of adrenergic fibres had only a weak inhibitor!. effect on gastric tone after acute \-agotom>-or administration of atropine. I t was therefore suggested that the sympathetic fibres exerted their inhibitory action on cholinergic ganglionic cells involved in the excitatory control of gastric motility (Janson 1969). Howei-er, a
direct sympathetic inhibition of gastric smooth muscle has also been proposed by Andrews & LaN-es (1984) in a study on the anaesthetized ferret. T h e y found that the principle mechanism for controlling intragastric pressure following inflation of the stomach was by increased activity in vagal non-adrenergic, non-cholinergic neurons. -4fter vagotomy the most important mechanism for adjusting the intragastric pressure to an increased gastric volume appeared to be by direct sympathetic inhibition of the gastric smooth muscle. O u r results also suggest the presence of a component of sympathetic inhibition of gastric motility independent of vagal nerve function since pain induced inhibition of gastric tone can, to a substantial extent, still be elicited after bilateral cervical vagotomy alone (Fig. 3) while pain-induced gastric relaxation was abolished by vagotomy following a-1 and padrenergic blockade (Fig. 4). I n conclusion we found that gastric relaxation in response to a standardized pain stimulation in the rat is reflexly mediated via sympathetic neurons acting on x-l and Ijj-adrenoreceptors and possibly also via vagal non-adrenergic fibres. Pain-induced inhibition of gastric tone was significantly increased by yohimbine. It is suggested that yohimbine by blocking presj-naptic inhibitory receptors on adrenergic neurons facilitates the release of noradrenaline in response to pain stimulation. This stud!- was supported by grants from Bohuslandstinget, University of Goteborg, the Medical Society of Goteborg and the Swedish Medical Research Council (Grant No. 09072). REFERENCES .IBRAHA\~SSON, H. & JANSSON, G. 1969. Elicitation of reflex vagal relaxation of the stomach from the pharynx and oesophagus in cat. Acta Physiol Scund 77, 172-177. ,4BR,4HAMSSON, H. & THOREN, P. 1972. Reflex relaxation of the stomach elicited from receptors located in the heart. .4n analysis of the receptors and afferents involved. Acta Physid S u n d 84, 197-207. . ~ N D R E W S ,P.L.R. &. LAWES, I.N.C. 1984. Interactions between splanchnic and vagus nerves in the control of mean intragastric pressure in the ferret. 3 Ph.ysiol 351, 4i3490. BAILY, D.M. 1951. Inhibitory and excitatory effects of sympathomimetic amines on muscle strips from the stomach of guinea-pig. B r 3 Pharmac 41, 227-238.
Pain-induced gastric relaxation RENNET,A. & WHITNEY, B. 1966. A pharmacological investigation of human isolated stomach. B r 3 Pharmuc Chemother 27, 286-298. RURKS, T.F. 1987. Actions of drugs on gastrointestinal motility. In: L.R. Johnsson (ed.). Physiology of the gastro-intestinal tract, pp, 723-743. Raven Press, New York. BoJO, L. & CASSUTO, J. 1992. Gastric reflex relaxation by colonic distension. 3 Auton Nerve Syst 38, 57-64. BoJO, L. NELLGKRD, P. & CASSUTO, J. 1991. Effects of selective adrenergic agonists and antagonists on gastric tone in the rat. Acta Physiol Scand 142, 517-522. FELDMAN, M. 1983. Nausea and vomiting. I n : M.H. Sleisenger & J.S. Fordtran (ed.) Gastro-intestinal disease, pp. 160-177. Saunders Company, Philadelphia. GLISE,H., LINDAHL, B.-0. & ABRAHAMSSON, H. 1980. Reflex adrenergic inhibition of gastric motility by nociceptive intestinal stimulation and peritoneal irritation in the cat. Scand 3 Gastroenterol 15, 673-68 1. GLISE,H. & ABRAHAMSSON,H. 1980. Reflex vagal inhibition of gastric motility by intestinal nociceptive stimulation in the cat. ScandJ Gastroenterol 15, 769-774. GRUNDY, D. & SCRATCHERD, T. 1982. A splanchnovagal component of the inhibition of gastric motility by distension of the intestines. In: M. Wienbeck (ed.). Motility of the digestive tract, pp. 3 9 4 3 . Raven Press, New York. GUIMARAES, S.1969. Alpha excitatory, alpha inhibitory
383
and beta inhibitory adrenergic receptors in the guinea-pig stomach. Arch Int Pharmacodyn Ther 179, 188-201. HAFFNER, J.F.W., LIAVAG, I. & SETEKLEIV, J. 1969. Excitatory adrenergic receptors in the human stomach and pylorus. Scand 3 Gastvoenterol 4, 145-1 50. JANSSON, G. 1969. Extrinsic nervous control of gastric motility. Acta Physiol Scand suppl 326, 1-42. T. & MIZUMURA, K. 1977. The polymodaf KUMAZAWA, receptors in the testis of dog. Brain Res 136, 553-558. LANGMAN, J. 1975. Medical embryology. Williams & Wilkins Company, Baltimore. NAUDY,B. & SALDUCCI, J. 1984. Vomiting. I n : L.M.A. Akkermans, A.G. Johnson & N.W. Read (eds). Gastric and Gastroduodenul Motility, pp. 3 4 4 0 . Praeger Publishers, New York. SMITH, C.C. & BRIZZEE, K.R. 1961. Cineradiographic analysis of vomiting in the cat. Gastroenterolo~40, 654-664. STANGHELLINI, V. MALAGELADA, J.-R. ZINMEISTER, A.R., Go, V.L.W. & KAO, P.C. 1983. Stressinduced gastro-duodenal motor disturbances in humans : possible humoral mechanisms. Gastroenterologji 85, 83-91. C.D. 1980. REES,M.R. CLARK, R.A. & HOLDSWORTH, The effect of beta-adrenoceptor agonists and antagonists on gastric emptying in man. B 3 Clin Pharmacol 10, 551-554. THOMPSON, D.G. RICHELSON,E. & MALAGELADA, J.-R. 1983. Perturbation of upper gastrointestinal function by cold stress. Gut 24, 277-283.
Pain-induced inhibition of gastric motility is mediated by adrenergic and vagal non-adrenergic reflexes in the rat L. BOJO, J. C A S S U T O and P. N E L L G A R D Department of Internal Medicine and Department of Physiology, University of Goteborg, Department of Clinical Physiology, Central Hospital Karlstad, and Department of Anaesthesiology, Central Hospital, Molndal, Sweden.
BoJO, L., CASSUTO, J. & NELLG~RD, P. 1992. Pain-induced inhibition of gastric motility is mediated by adrenergic and vagal non-adrenergic reflexes in the rat. Acta Physiol Scand 146, 377-383. Received 15 October 1991, accepted 18 May 1992. ISSN 00016772. Department of Internal Medicine and of Physiology, University of Goteborg, Department of Clinical Physiology, Central Hospital, Karlstad, and Department of Anaesthesiology, Central Hospital, Molndal, Sweden. Painful stimuli have been shown to inhibit gastric motility in animal experiments and delay gastric emptying in humans. The aim of the present study was to investigate in detail mechanisms involved in pain-induced gastric inhibition. Pain stimulation by exerting pressure on a testicle induced a prompt gastric relaxation which lasted throughout the period of stimulation. Pain-induced gastric relaxation was significantly reduced by the selective a-1 blocker, prazosin, and by the non-selective /Iblocker, propranolol. Similarly pain-induced inhibition of gastric tone was significantly reduced by bilateral cervical vagotomy. In contrast, gastric relaxation following pain stimulation was significantly potentiated by the selective adrenergic 01-2 blocker, yohimbine. Combined administration of prazosin and propranol followed by bilateral cervical vagotomy abolished gastric relaxation in response to pain stimulation. In conclusion, gastric relaxation in response to painful stimulation was found to be reflex-mediated via sympathetic neurons acting on a-1 and preceptors and possibly also via vagal non-adrenergic fibres. Pain-induced inhibition of gastric tone was significantly increased by yohimbine. It is suggested that yohimbine by blocking presynaptic inhibitory receptors on adrenergic neurons facilitates the release of noradrenaline in response to pain stimulation.
Key words : pain, vagus nerve, adrenergic antagonists, gastrointestinal motility.
Various conditions involving pain such as trauma, intestinal obstruction, appendicitis, myocardial infarction and major surgery are often accompanied by nausea and vomiting (Feldman 1983). A feared complication is aspiration of gastric contents which may cause hypoxia and pneumonitis, particularly in patients with lowered level of consciousness (Naudy & Salducci 1984). Correspondence : Dr Leif Bojo, Department of Clinical Physiology, Central Hospital, S-651 85 Karlstad, Sweden.
Nausea and vomiting are associated with perturbed gastric motility characterized by an inhibition of gastric tone and a reduction of normal motility which, during vomiting, is replaced by antiperistalsis (Smith & Brizzee 1961, Feldman 1983). Nociceptive intra-abdominal stimulation has been shown to induce inhibition of gastric tone both via adrenergic and vagal non-adrenergic reflexes (Glise et al. 1980, Glise & Abrahamsson 1980. T h e aim of the present study was to investigate the neural pathways and receptors mediating gastric relax-
377
378
L. Bo30 et al. Prazosin 150 pg kg' i.v. Blood pressure mm
Hg
200
I
50
50
450
450
350
350
600
600
900
900
Heart rate
(iastric volume
Pf?
Time (min)
Fig. 1. Gastric relaxation induced bl- pain-stimulation is significantly reduced after the administration of 2-1 blocker prazosin compared to control
tion of changes in gastric volume on a Grass polygraph (Grass Instr. Quincy, MA, USA). Changes in the volume of the reservoir were registered as inverse to changes in gastric volume. Calibration was performed b!- adding a known volume of water to the reservoir X I I.:T HOD S after temporaril!- closing the tube connecting the Experiments were performed on 35 male Spragueballoon to the reservoir. Dawley rats weighing 300-350 g. 'The experimental LVhen catheters and the balloon had been introprotocol was approved b>-the Animal Ethics Ctrmduced, the animals were allowed to recmer from mittee. Animals were housed for at least 7 dal-s prior surgerl- for a t least 30 min before starting the to the experiments in a ventilated and temperatureexperiment. Pain stimulation (PS) was performed by controlled room and had water and food ad libitum. exerting a constant pressure of 120-130 mmHg during The day-night c!cle was constants at 12-h periods. 1 min on one testicle by- the use of a clamp. T h e clamp Before the experiments the animals were fasted for \?-asmade of two parallel plastic slides ( 2 x 3 cm), each l l t l 5 h with free access to water. Anaesthesia \$-as with a central excavation approximately the size and induced with methohexital (Brietdl, Lilley, France, shape of the testicle and held together by two 60 mg kg- I ) intraperitoneall!- and maintained b!- a adjustable coils. continuous intravenous infusion of chloralose Each experiment was started by testing the degree (l..&mg kg-' min-I). A tracheostomy w s performed of gastric relaxation induced by PS. This procedure and a tracheal cannula was inserted to secure free Itas repeated u-ith at least 10 min between pain airway. Body temperature was kept constant at 38 "C stimulations. When gastric relaxation differed less by a heating pad coupled to a temperature sensor in than 20°, following two consecutive stimulations and the pharynx. Blood pressure was monitored using a when gastric basal tone had returned to a stable level pressure transducer (Statham P23AC Statham Instr.. and remained there for at least 10 min the experimental Osnard, C.4, USA) connected to a cannula in a drug was administered intravenously. Pain stimulation femoral artery. Heart rate (HR) a a s monitored, using was then repeated 1.5 min after administration of the a heart rate meter. drug. .4fter finishing the experiments, the volume of Gastric volume was recorded at constantl!- loa- the gastric balloon was determined by aspiration. The intragastric pressure ( 2 cmH,O) bj- a aater-filled, animals were killed by an intravenous injection of thin-walled flaccid balloon introduced 1-ia the esophasaturated KC1 and the location of the balloon in the gus. T h e balloon was connected b!- a polyethene stomach w a s controlled by dissection. catheter (PE 90) to a water reservoir mounted on a The drugs used were: prazosin (Peder Inc., New force-transducer which allowed continuous registra'l-ork City, NY, USA), propranolol (JCI Pharma-
ation in response to noxious stimulation in anaesthetized rats.
Pain-induced gastric relaxation
379
Table 1. Effects of adrenergic antagonists and vagotomy on basal gastric volume. Mean$SEM Basal GV (ml)
Treatment
Dose
n
1.1& 0.1 1.3k0.1 1.1 & 0.1 1.okn.i
Prazosin Yohimbine Propranolol BCV
1SO ,ug kg-' 1 mg kg-' 1 mg kg-'
7 6 8 6
__
Effect of treatment on GV (PO
+ 10k30 +200$.50
ns. < 0.05 n.s. < 0.05
-90f40 +290f100
GV = Gastric volume; RCV = Bilateral cervical vagotomy; Reduced gastric volume.
P-value
+ = Increased
gastric volume; and
-
=
Table 2. Relative gastric relaxation induced by pain stimulation after drug administration and vagotomy. Data are given as a percentage of the control. Mean k SEM
Treatment
n
Prazosin Yohimbine Propranolol Prazosin and propranolol BCV Prazosin, propranolol and BCV
7 6 8 6 6 8
Gastric relaxation induced by PS after treatment in percent of control
P-value
< 0.05 < 0.05 < 0.01 < 0.05
f67f16
+ 144f33 +56fll +48_f7
< 0.05
+42i8 -4f5
< 0.01
PS = Pain stimulation; BCV = Bilateral cervical vagotomy; Reduced gastric volume.
ceuticals, Macclesfield, UK) and yohimbine (Merck & Co. Inc., Rahway, NJ, USA). All drugs were
administered intravenously in a femoral vein. Adequate doses of the adrenergic antagonists against the effects of the corresponding agonists on circulation and gastric tone have been tested previously (Bojo et al. 1991). Statistical analysis was performed using the signed rank test. Differences resulting in P-values less than 0.05 were considered significant. Values are given as mean fSEM.
RESULTS Pain stimulation (PS) induced an immediate and pronounced gastric relaxation and a partial or complete inhibition of the spontaneous antral phasic contractions. T h e inhibition of gastric tone lasted throughout the period of stimulation and returned to the pre-stimulation level within 5 min of termination of PS (Fig. 1).
+ = Increased gastric volume; and
-
=
Data regarding basal gastric volumes and the effects of drugs and bilateral cervical vagotomy on gastric volume are presented in Table 1. Pain stimulation before treatment in all 35 experiments increased gastric volume by 227 & 25 pl
( P < 0.01). Prazosin (150 ug kg-l i.v.), an a-1 antagonist, did not cause a significant change in basal gastric volume (Table 1) but significantly reduced paininduced inhibition of gastric tone as compared to the control (n = 7, P < 0.05) (Fig. 1 & Table 2). Propranolol (1 mg kg-' iv.), an unselective pblocker, did not significantly change basal gastric volume (Table 1). After administration of propranolol gastric reflex relaxation following PS was significantly reduced compared to the control ( n = 7, P < 0.05) (Fig. 2 & Table 2). Combined administration of prazosin (150 ug kg-' i.v.) and propranolol (1 mg kg-l) significantly reduced gastric relaxation in te-
.NO
L. BoJO et al. After Prazosin 150 ug kg-1
Fig. 2. After the administration of the 2-1 blocker prazosin pain-induced inhibition of gastric tone is further reduced after i.\. injection of the /l-blocker propranolol. Bilateral cervical vagotomy
I Hlinwt preswre
m m Hg
Heart rate
(;astric volume
800
800
lo00
loo0
1200
1200
UI
'lime (rnin)
Fig. 3. Pain-induced gastric relasation is significantly reduced bl- bilateral cervical vagotomy
sponse to pain stimulation however, a significant response to PS remained in all experiments ( 1 1 = h) (Table 2 ) . Yohimbine (1 mg kg i.v.), an 3 - 2 antagonist, significantlv reduced basal gastric tone and significantly increased pain-induced inhibition of gastric tone (n = 6) (Table 2 ) . Bilateral cerx ical \-agotom>-induced a gastric relaxation in all the experiments (?I = 7 , P < 0.0.5) ('Table 1). The gastric response to PS was
significantly reduced by vagotomy ( a = 7, P < 0.05) (Fig. 3 & Table 2 ) . Combined administration of prazosin and propranolol, at the doses given above, followed by bilateral cervical vagotomy abolished the gastric relaxatory response to pain stimulation (ri = 6, P < 0.05) (Fig. 4 & Table 2). In two out of six experiments in this group pain-stimulation induced a slight gastric contraction following the treatment above.
Pain-induced gastric relaxation
38 1
After Prazosin 150 ug kg-1 and Propranolol 1 mg kg-1 Bilateral cervical vagutumy
I Hlood pressure 111111 Hg
Heart rate 600
900
Time ImhJ
Fig. 4. Pain-induced reflex inhibition of gastric tone is abolished after the combined administration of the a-1 blocker, prazosin, and the a-blocker, propranolol, followed by bilateral cervical vagotomy.
DISCUSSION Painful or otherwise noxious stimuli to the gut or other parts of the body have been shown to inhibit gastric motility. In man, cold pain stimulation has been shown to decrease gastric emptying (Thompson et al. 1983) and reduce the amplitude of the antral phasic pressure in response to a solid meal (Stanghellini et al. 1983). I n the cat, nociceptive stimulation of the gut was shown to inhibit gastric motility via both adrenergic and spinovagal, non-adrenergic reflexes (Glise et al. 1980, Glise & Abrahamson 1980). In the present study a standardized pressure on a testicle was used as a model for pain stimulation. The testis is regarded as a visceral organ since it descends during foetal life to the scrotum from the abdominal cavity (Langman 1975, Kumazawa & Mizumara 1977). The presence of polymodal sensory receptors responsive to noxious stimulation by mechanical, thermal and chemical means have been demonstrated in the testis of the dog by Kumazawa & Mizumura (1977). Pressure applied on the scrotum and testicles is therefore likely to activate both visceral and somatic pain-receptors. In vitro studies have shown adrenergic inhibition of gastric smooth muscle tone to be mediated both by a- and ,&receptors in man
(Bennet & Whitney 1966, Haffner et al. 1969) and in the guinea-pig (Bailey 1971, Guimaraes 1969). Administration of the non-selective /3agonist isoprenaline and the p-2 selective agonist salbutamol significantly prolonged gastric emptying in healthy volunteers (Rees et al. 1980). In vivo experiments in rats showed that gastric relaxation was induced by stimulation of a-1 and /3-2 receptors (Bojo et al. 1991). Concordant results were obtained in the present experiments showing that pain-induced gastric relaxation was significantly reduced both by the a-1 receptor blocker, prazosin, and by the non-selective /3receptor antagonist, propranolol (Table 2). These results suggest that pain-induced gastric relaxation is mediated by activation both of a-1 and p-2 adrenergic receptors. The possibility of pain-induced activation of preganglionic sympathetic efferents to the adrenal glands causing a release of adrenaline which contributes to the gastric relaxation alongside with noradrenaline can not be ruled out. a-2 adrenergic receptors are generally located presynaptically on adrenergic nerve terminals and serve to inhibit noradrenaline release from postganglionic neurons (Burks 1987). I n the present study yohimbine, an a-2 blocker, induced a significant increase in gastric relaxation following pain-stimulation (Table 2). This may suggest that yohimbine, by blocking inhibitory
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presynaptic 2-2 receptors, facilitates the release of noradrenaline from inhibitory gastric efferent neurons in response to pain stimulation thus potentiating gastric relaxation. T h e vagal neri-es carry to thc stomach, besides cholinergic excitatory fibres, non-adrenergic inhibitory fibres. These inhibitory neurons have been shown to mediate receptive relaxation in response to activation of mechanorecptors in the pharynx or the oesophagus (-Abrahanison & Jansson 1969) and to inhibit gastric tone in response to ischaemic or mechanical stimulation of the heart (L4brahamsson & Thoren 1972). \lye have recently presented data, using the present rat model, showing that colonic distension induced gastric relaxation via a vagal nonadrenergic pathway (Bojo & Cassuto 1992). I n the present study bilateral cervical vagotomlsigniticantl!- increased gastric volume resulting in a decreased gastric capacity of further relaxation. T h e gastric response to pain stimulation was reduced following vagotomy. T h e possibilit!- that this reduction was due to the relaxatory effect of vagotomj- itself cannot be ruled out. IIowever, our results show that the pain-induced gastric relaxation remaining after combined z-l and /,’-adrenergic blockade was abolished after vagotomy suggesting a role for the \-agal nerves in pain-induced inhibition of gastric motilitJ-.T h i s vagall!--mediated inhibition of gastric tone could be due to decreased excitatory cholinergic impulse activity and/or increased actirity in the inhibitory \-agal efferent neurons to the stomach. Experimental support for2 reciprocal arrangement in impulse activation was given b!- Grundy &- Scratcherd (1982). T h e y demonstrated in the anaesthetized ferret that intestinal distension induced an increased impulse activity in some vagal efferent fibres and a decreased activity in other vagal efferent neurons. Previous studies in the cat showed that sympathetic inhibition of gastric tone required a spontaneous background discharge in the vagi or artificially induced activitv by electrical stimulation of the vagal nerves (Jansson 1969, Glise 8i .%brahamsson 1980, Glise rt 01. 1980). Electric stimulation of adrenergic fibres had only a weak inhibitor!. effect on gastric tone after acute \-agotom>-or administration of atropine. I t was therefore suggested that the sympathetic fibres exerted their inhibitory action on cholinergic ganglionic cells involved in the excitatory control of gastric motility (Janson 1969). Howei-er, a
direct sympathetic inhibition of gastric smooth muscle has also been proposed by Andrews & LaN-es (1984) in a study on the anaesthetized ferret. T h e y found that the principle mechanism for controlling intragastric pressure following inflation of the stomach was by increased activity in vagal non-adrenergic, non-cholinergic neurons. -4fter vagotomy the most important mechanism for adjusting the intragastric pressure to an increased gastric volume appeared to be by direct sympathetic inhibition of the gastric smooth muscle. O u r results also suggest the presence of a component of sympathetic inhibition of gastric motility independent of vagal nerve function since pain induced inhibition of gastric tone can, to a substantial extent, still be elicited after bilateral cervical vagotomy alone (Fig. 3) while pain-induced gastric relaxation was abolished by vagotomy following a-1 and padrenergic blockade (Fig. 4). I n conclusion we found that gastric relaxation in response to a standardized pain stimulation in the rat is reflexly mediated via sympathetic neurons acting on x-l and Ijj-adrenoreceptors and possibly also via vagal non-adrenergic fibres. Pain-induced inhibition of gastric tone was significantly increased by yohimbine. It is suggested that yohimbine by blocking presj-naptic inhibitory receptors on adrenergic neurons facilitates the release of noradrenaline in response to pain stimulation. This stud!- was supported by grants from Bohuslandstinget, University of Goteborg, the Medical Society of Goteborg and the Swedish Medical Research Council (Grant No. 09072). REFERENCES .IBRAHA\~SSON, H. & JANSSON, G. 1969. Elicitation of reflex vagal relaxation of the stomach from the pharynx and oesophagus in cat. Acta Physiol Scund 77, 172-177. ,4BR,4HAMSSON, H. & THOREN, P. 1972. Reflex relaxation of the stomach elicited from receptors located in the heart. .4n analysis of the receptors and afferents involved. Acta Physid S u n d 84, 197-207. . ~ N D R E W S ,P.L.R. &. LAWES, I.N.C. 1984. Interactions between splanchnic and vagus nerves in the control of mean intragastric pressure in the ferret. 3 Ph.ysiol 351, 4i3490. BAILY, D.M. 1951. Inhibitory and excitatory effects of sympathomimetic amines on muscle strips from the stomach of guinea-pig. B r 3 Pharmac 41, 227-238.
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