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Characterization of the responses of submucosal neurons to tachykinins in the guinea pig ileum
A704
AGA ABSTRACTS
• CHARACTERIZATION OF THE RESPONSES OF SUBMUCOSAL NEURONS TO TACHYKININS IN THE GUINEA PIG ILEUM. S. Vanner. W. K. MacNaughton. GI Diseases Research Unit, Queen's University, Kingston, and Dept. of Physiology, University of Ottawa, Ottawa, Canada. Substance P (SP) or a related tachykinin is thought to be an important excitatory neurotransmitter in the submucosal plexus but the exact tachykinin(s) and receptor(s) involved is unknown. This study examined the properties of tachykinin responses using intracellular and Ussing chamber recording techniques from in vitro preparations from the guinea pig ileum. Intracellular recordings under current clamp conditions were made fi'om neurons i n snbmncosal preparations (n=34); only cells which received slow excitatory post-synaptic potentials (sEPSP's) greater than 5 mV following fiber tract stimulation were studied. All drugs were added to the bath by snpeffusion. SP (3~30nM; ECso=gnM) evoked a dose-dependant depolarization which was associated with increased spike discharge and frequent bursts of fast excitatory post-synaptic potentials (fEPSP's). Neurokinin A and neurokinin B also depolarized these neurons but were less potent than SP. The NK~ receptor agonist (Sarg'MetO2n)SP (I-10nM) mimicked the SP responses. Tlie NKa agonist Sanktide also evoked dose-dependent depolarizations (10-100nM) but the increase in spike discharge and burst of fEPSP's was substantially less than that seen with SP and the NKI agonist. The NK1 non-peptide antagonist CP-99,994 (30nM) caused a parallel shift to the right in the SP dose-response curve. CP+ 99,994 (100nM) completely blocked maximal responses to SP (30 nM) and the NKt agunist (Sarg,MetO2u ) SP (10nM) but had no effect on responses to the NKa agonist Senktide (30nM).' CP-99,994 (100nM) was used to examine whether tachykinins acting at NK~ receptors are released by intrinsic enteric neurons or extrinsic capsaicin-sensitive nerves. The amplitude or duration of the sEPSP evoked by fiber tract stimulation was unaffected by CP-99,994 ( n = 7 ) . In Ussing chamber studies, where we have previously shown that capsaicinsensitive sensory nerves directly activate submncosal secretomotor neurons, the capsaicin (200 nM) evoked net secretory response was inhibited 53% by CP99,994, enmpared to controls (n=5,p<.O2). This study suggests that snbmueosal neurons in the guinea pig ileum are activated by taehykinins acting at NKt and NK3 receptor, NK~ receptors on submneosal secretomotor neurons can be activated by release of tachykinin(s) from capsaiein-sensitive nerves~ Direct evidence that tachykinins released from intrinsic neurons also act at NKt receptors on submucosal neurons remains to be demonstrated.
• SOLITARY RECTAL ULCER SYNDROME (SRUS) AND RECTAL PROLAPSE: MORE THAN BAD LUCK P Velio, C Bertoglio, D. Conte, L. Cermesoni, E. Guffanti Istituto Seienze Mediche, IRCCS, Ospedale Polielinico di Milano-Italy. SRUS is considered a rare condition whose etiology, pathogenesis and management are still debated. It is variably associated with rectal prolapse and characterized clinically by obstructed evacuation and blood per rectum. With the aim of evaluating the prevalence and role of rectal prolapse in SRUS, all patients with histology proven SRUS observed from 1986 to May 1994 underwent full clinical investigation including evacuation proetography, and medical (high-fiber diet, drugs) or surgical treatment. 23 patients (16 F, mean age 47 yr, range 21-74) were studied and followed up for 4 years (range 0.68.6), 2 died and 1 moved. Results are reported in the table. Clinical findings No of cases (%) Defeen~,ravhie findinas No of cases(%) Pain 21 (91.3) Rectal prolapses total 21 (91.3) Constipation 19 (82.6) external 8 (34.8) Blood per rectum 14 (60.8) Anterior reetocele 19 (82.6) Digital evacuation 13 (56.5) Descending permeum 17 (73.9) Bloating 8 (34.8) Anular rectal stenosis 8 (34.8) Anal incontinence 7 (30.4) Puboreetalis dyssynergia 8 (34.8) Mucus per rectum 6 (26.1) Urgency 6 (26.1) Mental disorders 4 (17.4) At endoscopy 1 other patient had rectal prolapse making the total number of patients with prolapses 22 (95.6%); 21 patients had ulcers, multiple in 3, anterior m 17, posterior in 3 and lateral in 1 High-fiber diet improved eunstipation in 67% (8/12), 5-ASA or steroids reduced pain or rectal bleeding in 44% (4/9), laxatives improved constipation in 25% (3/12), surgery of prolapse healed ulcers that persisted in 50%/3/6) with prolapse recurrence Conclusions. Frequency of rectal prolapse was very high (95.6%) and was shown by defecography in 91.3%; it may explain the clinical picture of SRUS. This finding and the clinical outcome after surgery suggest that rectal prolapse has the main etiologic role, and evacuation proctography is a good method to demonstrate it and its characteristics Constipation seems to be less important in some patients. The frequency of anismus (38%) was similar to that observed in the normal population, suggesting a doubtful pathogenetie role. Anterior rectoeele and descending perineum appear ro be mostly the consequences of excessive straining, and anal incontinence +the final event. High-fiber diet is the best trentment for constipation and local 5A S A or steroids may reduce rectal bleeding. Only severe diseases should be treated by surgery as its results are unpredictable.
GASTROENTEROLOGY, Vol. 108, No. 4
DIAGNOSTIC VALUE OF DUMPING PROVOCATION IN PATIENTS AFTER GASTRIC SURGERY. J. Vecht F.G.M. van der Kley, A.A.M. Masclee, C.B.H.W. Lamers. Dept. G astroenterology-Hepatology, University Hospital Leiden, The Nethedands. Dumping syndrome is a combination of distressing postprandial systemic symptoms in patients with previous gastric surgery. Based on medical history it is often difficult to discern early and late dumping symptoms from other postcibal complaints. Therefore, we designed a dumping provocation test (DPT) in order to select patients with severe symptoms and objective signs of dumping for octreotide treatment. T h e diagnostic value of the DPT was evaluated in 36 symptomatic patients with previous gastdc surgery: 16 patients (age 30-74 yr) had eady dumping symptoms, 8 patients (age 31-74 yr) had late dumping symptoms; 12 patients (age 31-70 yr) after gastric surgery with abdominal complaints but without dumping symptoms served as controls. The DPT consisted of oral ingestion of a 50 g glucose load with evaluation of symptoms and objective parameters (heart rate, hematocrit, breath hydrogen excretion, blood glucose) scored every 15 rain for 360 min during standardized test conditions. Results (mean±SEM) eady dumping late dumping controls heart rate, increment (n/rain) hematocrit, increment breath H2> 10 ppm (%) nadir glucose (mmol/I)
18+9"
10+_2
4+9
0.015:0.006
0.01±0.005
0.01t-0.004
94%* 3,6±-0.3
85%*
8%
2.8±-0,2*
4.4£-0.3
* 13<0.05 compared to controls. An increase in heart rate of > 12/min (cardiovascular sign) and a rise in breath H2 excretion _> 10 ppm (indicative for accelerated transit) o had high sensitivity (both 94'/,) and specificity (100oVo and 92 o~ resp) for early dumping. A nadir blood glucose _< 3.3 mmoVI was present in 75% of late dumpers and in 17% of controls, When DPT was repeated with octreotide (50 I.tg s.c.) 100% of eady dumpers, 75% of late dumpers and only 25% of the controls had complete relief of symptoms and signs. It is concluded that dumping provocation is of clinical value in diagnosing eady and late dumping after gastric surgery. A positive test result is indicative for the response to octreotide.
• EFFECTS OF NOREPINEPHRINE ON ELECTRICAL SLOW WAVES AND PHASIC CONTRACTIONS IN CAT COLON CIRCULAR MUSCLE K. Venkova a n d J. Krier, Department of Physiology, Michigan State University, East Lansing, MI 48824 The postjunctional effects of norepinephrine and selective adrenoceptor agonists were tested on electrical slow Waves and phasic contractions of cat colon circular muscle. Resting membrane potential and electrical slow waves were recorded near the submucosal border. Myogenic activity was studied in the presence o f tetredotoxin (0.5 gM) or atropine (0.5 gM). Smooth muscle cells had a resting membrane potential of -76.1 4. 1.2 InV. They exhibited electrical slow waves (ESW) at a frequency of 4.2 +- 0.1 cycles/rain. ESW consist of an upstroke depolarization of resting membrane potential to -43.7 4- 1.3 mV, a plateau potential of -45.7 4- 0.8 mV and a repolaiization phase. The duration of ESW was 5.3 -+ 0.3 s. Norepinephrine (t I.tM) significantly increased ESW duration, plateau potential and the magnitude of phasic contractions. These effects were potentiated by propranotol and abolished by prazoslo. In strips pretreated with phenoxybenzamiue, an irreversible Ctl-adrenoceptor antagonist, norepinephrine reduced the duration of ESW. Phenylephrine (1 gM), a selective tx 1-edrenoeeptor agonist, caused an increase of ESW duration (7.6 4- 0.5 s), depolarized the plateau potential and decreased the rate of repolarization. In contrast, isoproterenol (1 gM), a selective ~-adreuoceptor agonist, decreased the duration of electrical slow waves (2.8 4- 0.2 s), hyperpolarized the plateau potential and increased the rate of repolarization. There was a linear concentration-response relation between the effect of phenylephrine (0.01;tM t o 5 gM) On area o f E S W (r = 0.97) anti phasic contractions (r = 0.95). Within this concentration range, resting membrane potential and resting tension did not significantly change. Higher concentrations of phenylephrine (> 5 JaM) depolarized the resting membrane potential and increased smooth muscle tone. Nitrendipine (1 ~tM for 10 rain) and verapamil (1 pM for 10 rain) did not alter resting membrane potential. In contrast, they nearly abolished the ESW plateau, reduced the upstroke potential, decreased the rate of depolarization and repolarization and reduced phasic contractions. In the presence of nitrendipine or verapamil the effects of phenylephrine and isoproterenol on electrical slow waves and phasic contractions were abolished. The results indicate that norepinephrine directly regulates ESW and phasic contractions of the circular muscle in non-spkincteric regions of cat colon. The myogenic effects of norepinephrine involve Ctl-adrenoceptors that potentiate ESW and phasic contractions and 13-adrenocept0rs that reduce ESW and phasic contractions. Both, the ct- and [~-adrenoceptor effects were sensitive to dihydropyridines that block L-type Ca 2+ channels. (NIH-DK-29920)
AGA ABSTRACTS
• CHARACTERIZATION OF THE RESPONSES OF SUBMUCOSAL NEURONS TO TACHYKININS IN THE GUINEA PIG ILEUM. S. Vanner. W. K. MacNaughton. GI Diseases Research Unit, Queen's University, Kingston, and Dept. of Physiology, University of Ottawa, Ottawa, Canada. Substance P (SP) or a related tachykinin is thought to be an important excitatory neurotransmitter in the submucosal plexus but the exact tachykinin(s) and receptor(s) involved is unknown. This study examined the properties of tachykinin responses using intracellular and Ussing chamber recording techniques from in vitro preparations from the guinea pig ileum. Intracellular recordings under current clamp conditions were made fi'om neurons i n snbmncosal preparations (n=34); only cells which received slow excitatory post-synaptic potentials (sEPSP's) greater than 5 mV following fiber tract stimulation were studied. All drugs were added to the bath by snpeffusion. SP (3~30nM; ECso=gnM) evoked a dose-dependant depolarization which was associated with increased spike discharge and frequent bursts of fast excitatory post-synaptic potentials (fEPSP's). Neurokinin A and neurokinin B also depolarized these neurons but were less potent than SP. The NK~ receptor agonist (Sarg'MetO2n)SP (I-10nM) mimicked the SP responses. Tlie NKa agonist Sanktide also evoked dose-dependent depolarizations (10-100nM) but the increase in spike discharge and burst of fEPSP's was substantially less than that seen with SP and the NKI agonist. The NK1 non-peptide antagonist CP-99,994 (30nM) caused a parallel shift to the right in the SP dose-response curve. CP+ 99,994 (100nM) completely blocked maximal responses to SP (30 nM) and the NKt agunist (Sarg,MetO2u ) SP (10nM) but had no effect on responses to the NKa agonist Senktide (30nM).' CP-99,994 (100nM) was used to examine whether tachykinins acting at NK~ receptors are released by intrinsic enteric neurons or extrinsic capsaicin-sensitive nerves. The amplitude or duration of the sEPSP evoked by fiber tract stimulation was unaffected by CP-99,994 ( n = 7 ) . In Ussing chamber studies, where we have previously shown that capsaicinsensitive sensory nerves directly activate submncosal secretomotor neurons, the capsaicin (200 nM) evoked net secretory response was inhibited 53% by CP99,994, enmpared to controls (n=5,p<.O2). This study suggests that snbmueosal neurons in the guinea pig ileum are activated by taehykinins acting at NKt and NK3 receptor, NK~ receptors on submneosal secretomotor neurons can be activated by release of tachykinin(s) from capsaiein-sensitive nerves~ Direct evidence that tachykinins released from intrinsic neurons also act at NKt receptors on submucosal neurons remains to be demonstrated.
• SOLITARY RECTAL ULCER SYNDROME (SRUS) AND RECTAL PROLAPSE: MORE THAN BAD LUCK P Velio, C Bertoglio, D. Conte, L. Cermesoni, E. Guffanti Istituto Seienze Mediche, IRCCS, Ospedale Polielinico di Milano-Italy. SRUS is considered a rare condition whose etiology, pathogenesis and management are still debated. It is variably associated with rectal prolapse and characterized clinically by obstructed evacuation and blood per rectum. With the aim of evaluating the prevalence and role of rectal prolapse in SRUS, all patients with histology proven SRUS observed from 1986 to May 1994 underwent full clinical investigation including evacuation proetography, and medical (high-fiber diet, drugs) or surgical treatment. 23 patients (16 F, mean age 47 yr, range 21-74) were studied and followed up for 4 years (range 0.68.6), 2 died and 1 moved. Results are reported in the table. Clinical findings No of cases (%) Defeen~,ravhie findinas No of cases(%) Pain 21 (91.3) Rectal prolapses total 21 (91.3) Constipation 19 (82.6) external 8 (34.8) Blood per rectum 14 (60.8) Anterior reetocele 19 (82.6) Digital evacuation 13 (56.5) Descending permeum 17 (73.9) Bloating 8 (34.8) Anular rectal stenosis 8 (34.8) Anal incontinence 7 (30.4) Puboreetalis dyssynergia 8 (34.8) Mucus per rectum 6 (26.1) Urgency 6 (26.1) Mental disorders 4 (17.4) At endoscopy 1 other patient had rectal prolapse making the total number of patients with prolapses 22 (95.6%); 21 patients had ulcers, multiple in 3, anterior m 17, posterior in 3 and lateral in 1 High-fiber diet improved eunstipation in 67% (8/12), 5-ASA or steroids reduced pain or rectal bleeding in 44% (4/9), laxatives improved constipation in 25% (3/12), surgery of prolapse healed ulcers that persisted in 50%/3/6) with prolapse recurrence Conclusions. Frequency of rectal prolapse was very high (95.6%) and was shown by defecography in 91.3%; it may explain the clinical picture of SRUS. This finding and the clinical outcome after surgery suggest that rectal prolapse has the main etiologic role, and evacuation proctography is a good method to demonstrate it and its characteristics Constipation seems to be less important in some patients. The frequency of anismus (38%) was similar to that observed in the normal population, suggesting a doubtful pathogenetie role. Anterior rectoeele and descending perineum appear ro be mostly the consequences of excessive straining, and anal incontinence +the final event. High-fiber diet is the best trentment for constipation and local 5A S A or steroids may reduce rectal bleeding. Only severe diseases should be treated by surgery as its results are unpredictable.
GASTROENTEROLOGY, Vol. 108, No. 4
DIAGNOSTIC VALUE OF DUMPING PROVOCATION IN PATIENTS AFTER GASTRIC SURGERY. J. Vecht F.G.M. van der Kley, A.A.M. Masclee, C.B.H.W. Lamers. Dept. G astroenterology-Hepatology, University Hospital Leiden, The Nethedands. Dumping syndrome is a combination of distressing postprandial systemic symptoms in patients with previous gastric surgery. Based on medical history it is often difficult to discern early and late dumping symptoms from other postcibal complaints. Therefore, we designed a dumping provocation test (DPT) in order to select patients with severe symptoms and objective signs of dumping for octreotide treatment. T h e diagnostic value of the DPT was evaluated in 36 symptomatic patients with previous gastdc surgery: 16 patients (age 30-74 yr) had eady dumping symptoms, 8 patients (age 31-74 yr) had late dumping symptoms; 12 patients (age 31-70 yr) after gastric surgery with abdominal complaints but without dumping symptoms served as controls. The DPT consisted of oral ingestion of a 50 g glucose load with evaluation of symptoms and objective parameters (heart rate, hematocrit, breath hydrogen excretion, blood glucose) scored every 15 rain for 360 min during standardized test conditions. Results (mean±SEM) eady dumping late dumping controls heart rate, increment (n/rain) hematocrit, increment breath H2> 10 ppm (%) nadir glucose (mmol/I)
18+9"
10+_2
4+9
0.015:0.006
0.01±0.005
0.01t-0.004
94%* 3,6±-0.3
85%*
8%
2.8±-0,2*
4.4£-0.3
* 13<0.05 compared to controls. An increase in heart rate of > 12/min (cardiovascular sign) and a rise in breath H2 excretion _> 10 ppm (indicative for accelerated transit) o had high sensitivity (both 94'/,) and specificity (100oVo and 92 o~ resp) for early dumping. A nadir blood glucose _< 3.3 mmoVI was present in 75% of late dumpers and in 17% of controls, When DPT was repeated with octreotide (50 I.tg s.c.) 100% of eady dumpers, 75% of late dumpers and only 25% of the controls had complete relief of symptoms and signs. It is concluded that dumping provocation is of clinical value in diagnosing eady and late dumping after gastric surgery. A positive test result is indicative for the response to octreotide.
• EFFECTS OF NOREPINEPHRINE ON ELECTRICAL SLOW WAVES AND PHASIC CONTRACTIONS IN CAT COLON CIRCULAR MUSCLE K. Venkova a n d J. Krier, Department of Physiology, Michigan State University, East Lansing, MI 48824 The postjunctional effects of norepinephrine and selective adrenoceptor agonists were tested on electrical slow Waves and phasic contractions of cat colon circular muscle. Resting membrane potential and electrical slow waves were recorded near the submucosal border. Myogenic activity was studied in the presence o f tetredotoxin (0.5 gM) or atropine (0.5 gM). Smooth muscle cells had a resting membrane potential of -76.1 4. 1.2 InV. They exhibited electrical slow waves (ESW) at a frequency of 4.2 +- 0.1 cycles/rain. ESW consist of an upstroke depolarization of resting membrane potential to -43.7 4- 1.3 mV, a plateau potential of -45.7 4- 0.8 mV and a repolaiization phase. The duration of ESW was 5.3 -+ 0.3 s. Norepinephrine (t I.tM) significantly increased ESW duration, plateau potential and the magnitude of phasic contractions. These effects were potentiated by propranotol and abolished by prazoslo. In strips pretreated with phenoxybenzamiue, an irreversible Ctl-adrenoceptor antagonist, norepinephrine reduced the duration of ESW. Phenylephrine (1 gM), a selective tx 1-edrenoeeptor agonist, caused an increase of ESW duration (7.6 4- 0.5 s), depolarized the plateau potential and decreased the rate of repolarization. In contrast, isoproterenol (1 gM), a selective ~-adreuoceptor agonist, decreased the duration of electrical slow waves (2.8 4- 0.2 s), hyperpolarized the plateau potential and increased the rate of repolarization. There was a linear concentration-response relation between the effect of phenylephrine (0.01;tM t o 5 gM) On area o f E S W (r = 0.97) anti phasic contractions (r = 0.95). Within this concentration range, resting membrane potential and resting tension did not significantly change. Higher concentrations of phenylephrine (> 5 JaM) depolarized the resting membrane potential and increased smooth muscle tone. Nitrendipine (1 ~tM for 10 rain) and verapamil (1 pM for 10 rain) did not alter resting membrane potential. In contrast, they nearly abolished the ESW plateau, reduced the upstroke potential, decreased the rate of depolarization and repolarization and reduced phasic contractions. In the presence of nitrendipine or verapamil the effects of phenylephrine and isoproterenol on electrical slow waves and phasic contractions were abolished. The results indicate that norepinephrine directly regulates ESW and phasic contractions of the circular muscle in non-spkincteric regions of cat colon. The myogenic effects of norepinephrine involve Ctl-adrenoceptors that potentiate ESW and phasic contractions and 13-adrenocept0rs that reduce ESW and phasic contractions. Both, the ct- and [~-adrenoceptor effects were sensitive to dihydropyridines that block L-type Ca 2+ channels. (NIH-DK-29920)