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The Effect of α-Trinositol on Cholera Toxin–Induced Fluid Accumulation in Pig (Sus scrofa domesticus) Jejunum in vivo
Comp. Biochem. Physiol. Vol. 118A, No. 2, pp. 305–307, 1997 Copyright 1997 Elsevier Science Inc.
ISSN 0300-9629/97/$17.00 PII S0300-9629(96)00310-6
The Effect of α-Trinositol on Cholera Toxin–Induced Fluid Accumulation in Pig (Sus scrofa domesticus) Jejunum in vivo Tyge T. Tindholdt,1 Mark B. Hansen,1 Vibeke S. Elbrønd,1 Martin Makinde,2 Eva J. Westerberg,3 and Erik Skadhauge1 1
Department of Anatomy and Physiology, The Royal Veterinary and Agricultural University, Copenhagen, Denmark, 2 Department of Preclinical Veterinary Studies, University of Zimbabwe Mount Pleasant, Harare, Zimbabwe, and 3 Perstorp Pharma, Lund, Sweden ABSTRACT. The effect of α-trinositol (D-myo-inositol-1,2,6-trisphosphate) on cholera toxin–induced fluid accumulation (i.e., net fluid secretion) was studied in the pig jejunum in vivo. Cholera toxin caused a dosedependent fluid accumulation in control experiments. Intravenous injection of α-trinositol produced a reduction of the response to cholera toxin with a significant maximal inhibition of 36%. However, in high concentrations of α-trinositol this inhibition was absent. comp biochem physiol 118A;2:305–307, 1997. 1997 Elsevier Science Inc. KEY WORDS. α-Trinositol, cholera toxin, diarrhoea, intestinal secretion, jejunum, pig
INTRODUCTION The pathogenic Vibrio cholerae causes diarrhoea partly due to the direct action of its enterotoxin, cholera toxin (CT), on the intestinal epithelium (16), and partly due to an inflammatory response (5,7). α-Trinositol (α-T) is an isomer of inositol-1,4,5-trisphosphate and is produced by hydrolysis of phytic acid by the phytase enzyme of yeast. It reduces inflammatory reactions such as edema formation and capillary leakage in several experimental models of inflammation (3,6,8,11,13). In the rat intestinal tract α-T reduces fluid losses caused by choleragenic diarrhoea (14). The morphology and physiology of the gastrointestinal tract in pigs resemble the gastrointestinal tract of humans (12). Therefore the pig was used as an experimental model for humans to evaluate the effect of α-T on CT-induced fluid accumulation (i.e., net fluid secretion) in the jejununal section of the small intestine in vivo. MATERIALS AND METHODS Danish Landrace/Yorkshire crossbred fully weaned 8-weekold female pigs (12–16 kg) were examined. The animals were fasted for 12 hr before surgery, but had free access to sterile drinking water containing D-glucose (55 g/l).
Address reprint requests to: Tyge T. Tindholdt, Department of Anatomy and Physiology, The Royal Veterinary and Agricultural University, 13 Bu¨lowsvej, 1870 Frederiksberg C, Denmark. Fax 145 3528 2525. Received 29 May 1996; accepted 31 May 1996.
Twenty min before general anaesthesia was commenced the pigs were sedated by intramuscular injection (5 mg/kg) of azaperone (Sedaperone, Janssen, Denmark). Following intravenous injection (10 mg/kg) of pentobarbital (Mebumal, University Pharmacy, Denmark) anaesthesia was maintained with halothane inhalation (2% in oxygen) on a closed curcuit. A catheter was placed in the femoral vein, and infusion of α-T [4; 8; 16; 32 (mg/kg)/hr] (Perstorp Pharma, Lund, Sweden) dissolved in saline was commenced following an intravenous bolus injection of α-T (2 mg/kg in 20 ml saline). Control experiments were performed with infusion of saline following a bolus injection of 20 ml saline. The infusion rate in both control and α-T experiments was the same [0.15 (ml/kg)/min]. A midline abdominal incision was made and the jejunum was located 0.6 m aborally to the Treitz ligament. Seventeen jejunal loops were prepared by ligaturing between the mesenteric arcades. Each loop was ,12 cm in length and the space between loops was ,2 cm. Fifteen loops were injected with 3 ml (3040 mg) buffered solution [(in mM) 10 Na1, 7 Cl2, 3 HCO 32 , 80 glucose; pH 7.4] (9) containing graded doses of CT (1; 5; 10; 20; 40 µg/loop). One loop was filled with buffer and one loop was left unfilled. The loops were filled randomly to eliminate regional differences. After filling, the exteriorized loops were returned to the peritoneal cavity and the abdomen was closed using forceps. The bolus was administered and the infusion of α-T was initialized 10 min before the loops were filled. Two hundred thirty min after the abdomen was closed, a blood sample was collected to determine the plasma level
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of α-trinositol, ensuring that the infusion mechanism was intact throughout the experiment. The abdomen was reopened 240 min after closure. The loops were removed and the animals were killed by intracardiac injection of pentobarbital. The loops were weighed with and without their luminal contents, and the fluid accumulation was calculated by subtracting the two values. Results were presented as mean 6 SEM and were analysed for significance with the unpaired Student’s t-test. N and n represented the total number of pigs and loops respectively that were prepared for every corresponding dose of CT and concentration of α-T. Data were considered to be significant at P values less than 0.05. The ED50 (CT dose eliciting 50% of the maximal fluid accumulation) and Emax (the maximal fluid accumulation) values were calculated using non-linear curve fitting applied to Michaelis-Menten kinetics. The computer program used for all data analysis was Sigmaplot for Windows version 1.0. RESULTS Plasma levels of α-T increased linearily with the amount of α-T being infused. Thus, control experiments reflected a low level of 5 6 2 nM (N 5 5) and levels in α-T experiments using 32 (mg/kg)/hr increased up to 117 6 12 µM (N 5 6). In both control and α-T experiments the unfilled control loops were empty, and thus there was no unstimulated net fluid secretion. This means that α-T did not alter the physiological net fluid transport. Also the filled control loops were empty in all experiments, meaning that a net fluid absorption had taken place at the termination point of the individual experiments. CT caused a dose-dependent fluid accumulation (ED50 5 1.4 6 0.2 µg/loop; Emax 5 7057 6 143 mg/loop) in control pigs (Fig. 1). Treatment with α-T [except for 32 (mg/kg)/hr] caused an inhibition of the CT response (Fig. 1). In the CT dose range of 5–40 µg/loop fluid accumulation data in pigs treated with 8 and 16 (mg/kg)/hr of α-T were significantly different to control experiments. Infusion of 16 (mg/kg)/hr produced a maximal inhibition of 36% (ED50 5 1.4 6 0.4 µg/loop; Emax 5 4847 6 241 mg/loop) corresponding to a CT dose of 10 µg/loop. Infusion of 32 (mg/kg)/hr did not alter the CT-induced fluid accumulation. DISCUSSION The present study demonstrates that α-T reduces fluid accumulation caused by the pathogenic mechanisms of the enterotoxin CT. The results do not reveal how α-T interdigitates with these mechanisms, and thus we can only speculate about this, taking previous studies on α-T into consideration. CT causes distinct morphological changes in human and rat jejunum (1,18), which is reflected in numerous inflam-
FIG. 1. Dose-dependent effect of intravenous a-T injection
[0 (control); 4; 8; 16; 32 (mg/kg)/hr] on intraluminal CTinduced (1; 5; 10; 20; 40 mg/loop) fluid accumulation. Each point represents the mean fluid accumulation 6 SEM (n 5 11–24, N 5 5–6). *P , 0.05; **P , 0.01.
matory characteristics, such as accumulation of mononuclear cells in the lamina propria, mucosal edema and desquamation, and necrosis of the mucosal epithelium. These studies suggest that this inflammatory reaction plays an essential role in the secretory response to CT. Considering the anti-inflammatory properties of α-T (3,6,8,11,13), the partial reduction in net fluid secretion demonstrated in our setup, therefore, could be due to the inhibition of the inflammatory response caused by CT. Unfortunately the antiinflammatory mechanisms of α-T remain unknown and further investigation will be required. Isomers of inositol phosphates bind with high affinity to specific sites on different cell types, eliciting different cellular responses (17). Inositol-1,3,4,5-phosphate [Ins(1,3, 4,5)P4 ] shows binding characteristics similar to those of α-T (20). Furthermore, Ins(1,3,4,5)P4 is believed to activate Ca21 permeable channels (20), raising the possibility that α-T affects these same channels, and thereby regulates transmural fluid transport by altering the Ca21 influx. In the intestine 5-hydroxytryptamine (5-HT) originates in enterochromaffin (EC) cells (15), mast cells (4) and in the enteric neurons (10,19). The EC cells release 5-HT following exposure to CT in vivo, and the decrease in intracellular 5-HT contents correlates closely to the increased rate of intraluminal fluid accumulation (15). This proposes that CT-induced fluid accumulation is due, in part, to the release of 5-HT (2,15). Different phosphorylated inositol analogs were demonstrated to bind to bovine adrenal chromaffin cell membranes (17). This suggests that α-T might bind to
Cholera Toxin–Induced Diarrhoea and α-Trinositol
the EC cell membrane, inhibiting the discharge of 5-HT by unknown mechanisms, and thereby result in the observed reduction in the fluid accumulation. In summary, the present study showed that α-T is a weak anti-secretory agent, reducing CT-induced fluid accumulation in the pig jejunum in vivo. Surprisingly, high concentrations of α-T did not affect the secretory response elicited by CT. We can only explain this by an unspecific action of α-T when reaching high concentrations.
307
9. 10. 11. 12.
The authors wish to express their sincere thanks to Jens E. Thorbøll, Gerda M. Jensen, Lars Thomsen, Marie L. Grøndahl, and Jens Randrup for their assistance. This investigation was supported by Perstorp Pharma, Lund, Sweden. This article is based on a study first reported in Pharmacology & Toxicology, 78, 104–110;1996.
13.
14.
References 1. Asakura, H.; Morita, A.; Morishita, T.; Tsuchiya, M.; Fukumi, H.; Ohashi, M.; Uylangco, C.; Castro, A. Pathologic findings from intestinal biopsy specimens in human cholera. Digest. Dis. 18:271–278;1973. 2. Beubler, E.; Kollar, G.; Saria, A.; Bukhave, K.; Rask-Madsen, J. Involvement of 5-hydroxytryptamine, prostaglandin E2 , and cyclic adenosine monophosphate in cholera toxin-induced fluid secretion in the small intestine of the rat in vivo. Gastroenterology 96:368–376;1989. 3. Cassuto, J.; Jo¨nsson, A.; Tarnow, P.; Nellga˚rd, P.; Rimba¨ck, G. Anti-inflammation mediated effects in several acute conditions by PP56 (D-myo-inositol-1,2,6-triphosphate). Circ. Shock. Suppl. 1:51;1993. (Abstract 12.20) 4. Castro, G.A.; Harari, Y.; Russel, D. Mediators of anaphylaxisinduced ion transport changes in small intestine. Am. J. Physiol. 253:G540–G548;1987. 5. Ciancio, M.J.; Chang, E.B. Epithelial secretory response to inflammation. Ann. N. Y. Acad. Sci. 664:210–221;1992. 6. Claxson, A.; Morris, C.; Blake, D.; Sire´n, M.; Halliwell, B.; Gustafsson, T.; Lo¨fkvist, B. Bergelin, I. The anti-inflammatory effects of D-myo-inositol-1.2.6-triphosphate (PP56) on animal models of inflammation. Agents Actions 29:68–70;1990. 7. Crosby, W.H.; Kugler, H.W. Intraluminal biopsy of the small intestine. Am. J. Digest. Dis. 2:236–241;1957. 8. Gunther, R.A.; Gustafsson, T.; Brofeldt, B.T. α-Trinositol (α-
15.
16. 17.
18. 19.
20.
T) and decreased fluid balance following thermal injury. Circ. Shock. Suppl. 1:51;1993. (Abstract 12.19) Hansen, M.B.; Skadhauge, E. Ketanserin and granisetron reduce cholera toxin-induced hypersecretion in pig jejunum. Scand. J. Gastroenterol. 29:908–915;1994. Hardcastle, J.; Hardcastle, P.T.; Redfern, J.S. Action of 5-hydroxytryptamine on intestinal ion transport in the rat. J. Physiol. 320:41–55;1981. Lund, T.; Reed, R.K. α-Trinositol inhibits edema generation and albumin extravasation in thermally injured skin. J. Trauma 36(6):761–765;1994. Miller, E.R.; Ullrey, D.E. The pig as a model for human nutrition. Ann. Rev. Nutr. 7:361–382;1987. Nakazawa, H.; Gustafsson, T.O.; Traber, L.D.; Herndon, D.N.; Traber, D.L. α-Trinositol decrease lung edema formation following smoke inhalation in an ovine model. J. Appl. Physiol. 76(1):278–282;1994. Nellga˚rd, P.; Jo¨nsson, A.; Cassuto, J. Effects of D-myo-inositol-1,2,6-trisphosphate (PP56) on cholera toxin-induced secretion. Acta Physiol. Scand. 146(Suppl. 608):2.50;1992. Nilsson, O.; Cassuto, J.; Larsson, P.-A.; Jodal, M.; Lidberg, P.; Ahlman, H.; Dahlstro¨m, A.; Lundgren, O. 5-Hydroxytryptamine and cholera secretion: A histochemical physiological study in cats. Gut 24:542–548;1983. Norris, H.T.; Curran, P.F.; Schultz, S.G. Modification of intestinal secretion experimental cholera. J. Infec. Dis. 119: 117–125;1969. Regunathan, S.; Reis, D.J.; Wahlestedt, C. Specific binding of in hexakisphosphate (phytic acid) to adrenal chromaffin cell membranes and effects on calcium dependent catecholamine release. Biochem. Pharmacol. 43:1331–1336;1992. Shirgi-Degen, A.; Pabst, M.A.; Klimpfinger, M.; Beubler, E. Histopathological effects of cholera toxin in the rat jejunum in vivo. Gastroenterology 104:A279;1993. Timmermans, J.-P.; Scheuermann, D.W.; Stach, W.; Adriaensen, D.; De Groodt-Lasseel, M.H.A. Distinct distribution of CGRP-, enkephalin-, galanin-, neuromedin U-, neuropeptide Y-, somatostatin-, substance P-, VIP- and serotonin-containing neurons two submucosal ganglionic neural networks of the porcine small intestine. Cell Tissue Res. 260:367–379; 1990. Yoo, H.; Lindahl, A.; Veznedaroglu, E.; Wahlestedt, C. Studies on 3H-ins[1,2,6]P3 and other 3H-inositol phosphate binding sites in rat brain. Soc. Neurosci. 18:477;1992. (Abstract)
ISSN 0300-9629/97/$17.00 PII S0300-9629(96)00310-6
The Effect of α-Trinositol on Cholera Toxin–Induced Fluid Accumulation in Pig (Sus scrofa domesticus) Jejunum in vivo Tyge T. Tindholdt,1 Mark B. Hansen,1 Vibeke S. Elbrønd,1 Martin Makinde,2 Eva J. Westerberg,3 and Erik Skadhauge1 1
Department of Anatomy and Physiology, The Royal Veterinary and Agricultural University, Copenhagen, Denmark, 2 Department of Preclinical Veterinary Studies, University of Zimbabwe Mount Pleasant, Harare, Zimbabwe, and 3 Perstorp Pharma, Lund, Sweden ABSTRACT. The effect of α-trinositol (D-myo-inositol-1,2,6-trisphosphate) on cholera toxin–induced fluid accumulation (i.e., net fluid secretion) was studied in the pig jejunum in vivo. Cholera toxin caused a dosedependent fluid accumulation in control experiments. Intravenous injection of α-trinositol produced a reduction of the response to cholera toxin with a significant maximal inhibition of 36%. However, in high concentrations of α-trinositol this inhibition was absent. comp biochem physiol 118A;2:305–307, 1997. 1997 Elsevier Science Inc. KEY WORDS. α-Trinositol, cholera toxin, diarrhoea, intestinal secretion, jejunum, pig
INTRODUCTION The pathogenic Vibrio cholerae causes diarrhoea partly due to the direct action of its enterotoxin, cholera toxin (CT), on the intestinal epithelium (16), and partly due to an inflammatory response (5,7). α-Trinositol (α-T) is an isomer of inositol-1,4,5-trisphosphate and is produced by hydrolysis of phytic acid by the phytase enzyme of yeast. It reduces inflammatory reactions such as edema formation and capillary leakage in several experimental models of inflammation (3,6,8,11,13). In the rat intestinal tract α-T reduces fluid losses caused by choleragenic diarrhoea (14). The morphology and physiology of the gastrointestinal tract in pigs resemble the gastrointestinal tract of humans (12). Therefore the pig was used as an experimental model for humans to evaluate the effect of α-T on CT-induced fluid accumulation (i.e., net fluid secretion) in the jejununal section of the small intestine in vivo. MATERIALS AND METHODS Danish Landrace/Yorkshire crossbred fully weaned 8-weekold female pigs (12–16 kg) were examined. The animals were fasted for 12 hr before surgery, but had free access to sterile drinking water containing D-glucose (55 g/l).
Address reprint requests to: Tyge T. Tindholdt, Department of Anatomy and Physiology, The Royal Veterinary and Agricultural University, 13 Bu¨lowsvej, 1870 Frederiksberg C, Denmark. Fax 145 3528 2525. Received 29 May 1996; accepted 31 May 1996.
Twenty min before general anaesthesia was commenced the pigs were sedated by intramuscular injection (5 mg/kg) of azaperone (Sedaperone, Janssen, Denmark). Following intravenous injection (10 mg/kg) of pentobarbital (Mebumal, University Pharmacy, Denmark) anaesthesia was maintained with halothane inhalation (2% in oxygen) on a closed curcuit. A catheter was placed in the femoral vein, and infusion of α-T [4; 8; 16; 32 (mg/kg)/hr] (Perstorp Pharma, Lund, Sweden) dissolved in saline was commenced following an intravenous bolus injection of α-T (2 mg/kg in 20 ml saline). Control experiments were performed with infusion of saline following a bolus injection of 20 ml saline. The infusion rate in both control and α-T experiments was the same [0.15 (ml/kg)/min]. A midline abdominal incision was made and the jejunum was located 0.6 m aborally to the Treitz ligament. Seventeen jejunal loops were prepared by ligaturing between the mesenteric arcades. Each loop was ,12 cm in length and the space between loops was ,2 cm. Fifteen loops were injected with 3 ml (3040 mg) buffered solution [(in mM) 10 Na1, 7 Cl2, 3 HCO 32 , 80 glucose; pH 7.4] (9) containing graded doses of CT (1; 5; 10; 20; 40 µg/loop). One loop was filled with buffer and one loop was left unfilled. The loops were filled randomly to eliminate regional differences. After filling, the exteriorized loops were returned to the peritoneal cavity and the abdomen was closed using forceps. The bolus was administered and the infusion of α-T was initialized 10 min before the loops were filled. Two hundred thirty min after the abdomen was closed, a blood sample was collected to determine the plasma level
306
T. T. Tindholdt et al.
of α-trinositol, ensuring that the infusion mechanism was intact throughout the experiment. The abdomen was reopened 240 min after closure. The loops were removed and the animals were killed by intracardiac injection of pentobarbital. The loops were weighed with and without their luminal contents, and the fluid accumulation was calculated by subtracting the two values. Results were presented as mean 6 SEM and were analysed for significance with the unpaired Student’s t-test. N and n represented the total number of pigs and loops respectively that were prepared for every corresponding dose of CT and concentration of α-T. Data were considered to be significant at P values less than 0.05. The ED50 (CT dose eliciting 50% of the maximal fluid accumulation) and Emax (the maximal fluid accumulation) values were calculated using non-linear curve fitting applied to Michaelis-Menten kinetics. The computer program used for all data analysis was Sigmaplot for Windows version 1.0. RESULTS Plasma levels of α-T increased linearily with the amount of α-T being infused. Thus, control experiments reflected a low level of 5 6 2 nM (N 5 5) and levels in α-T experiments using 32 (mg/kg)/hr increased up to 117 6 12 µM (N 5 6). In both control and α-T experiments the unfilled control loops were empty, and thus there was no unstimulated net fluid secretion. This means that α-T did not alter the physiological net fluid transport. Also the filled control loops were empty in all experiments, meaning that a net fluid absorption had taken place at the termination point of the individual experiments. CT caused a dose-dependent fluid accumulation (ED50 5 1.4 6 0.2 µg/loop; Emax 5 7057 6 143 mg/loop) in control pigs (Fig. 1). Treatment with α-T [except for 32 (mg/kg)/hr] caused an inhibition of the CT response (Fig. 1). In the CT dose range of 5–40 µg/loop fluid accumulation data in pigs treated with 8 and 16 (mg/kg)/hr of α-T were significantly different to control experiments. Infusion of 16 (mg/kg)/hr produced a maximal inhibition of 36% (ED50 5 1.4 6 0.4 µg/loop; Emax 5 4847 6 241 mg/loop) corresponding to a CT dose of 10 µg/loop. Infusion of 32 (mg/kg)/hr did not alter the CT-induced fluid accumulation. DISCUSSION The present study demonstrates that α-T reduces fluid accumulation caused by the pathogenic mechanisms of the enterotoxin CT. The results do not reveal how α-T interdigitates with these mechanisms, and thus we can only speculate about this, taking previous studies on α-T into consideration. CT causes distinct morphological changes in human and rat jejunum (1,18), which is reflected in numerous inflam-
FIG. 1. Dose-dependent effect of intravenous a-T injection
[0 (control); 4; 8; 16; 32 (mg/kg)/hr] on intraluminal CTinduced (1; 5; 10; 20; 40 mg/loop) fluid accumulation. Each point represents the mean fluid accumulation 6 SEM (n 5 11–24, N 5 5–6). *P , 0.05; **P , 0.01.
matory characteristics, such as accumulation of mononuclear cells in the lamina propria, mucosal edema and desquamation, and necrosis of the mucosal epithelium. These studies suggest that this inflammatory reaction plays an essential role in the secretory response to CT. Considering the anti-inflammatory properties of α-T (3,6,8,11,13), the partial reduction in net fluid secretion demonstrated in our setup, therefore, could be due to the inhibition of the inflammatory response caused by CT. Unfortunately the antiinflammatory mechanisms of α-T remain unknown and further investigation will be required. Isomers of inositol phosphates bind with high affinity to specific sites on different cell types, eliciting different cellular responses (17). Inositol-1,3,4,5-phosphate [Ins(1,3, 4,5)P4 ] shows binding characteristics similar to those of α-T (20). Furthermore, Ins(1,3,4,5)P4 is believed to activate Ca21 permeable channels (20), raising the possibility that α-T affects these same channels, and thereby regulates transmural fluid transport by altering the Ca21 influx. In the intestine 5-hydroxytryptamine (5-HT) originates in enterochromaffin (EC) cells (15), mast cells (4) and in the enteric neurons (10,19). The EC cells release 5-HT following exposure to CT in vivo, and the decrease in intracellular 5-HT contents correlates closely to the increased rate of intraluminal fluid accumulation (15). This proposes that CT-induced fluid accumulation is due, in part, to the release of 5-HT (2,15). Different phosphorylated inositol analogs were demonstrated to bind to bovine adrenal chromaffin cell membranes (17). This suggests that α-T might bind to
Cholera Toxin–Induced Diarrhoea and α-Trinositol
the EC cell membrane, inhibiting the discharge of 5-HT by unknown mechanisms, and thereby result in the observed reduction in the fluid accumulation. In summary, the present study showed that α-T is a weak anti-secretory agent, reducing CT-induced fluid accumulation in the pig jejunum in vivo. Surprisingly, high concentrations of α-T did not affect the secretory response elicited by CT. We can only explain this by an unspecific action of α-T when reaching high concentrations.
307
9. 10. 11. 12.
The authors wish to express their sincere thanks to Jens E. Thorbøll, Gerda M. Jensen, Lars Thomsen, Marie L. Grøndahl, and Jens Randrup for their assistance. This investigation was supported by Perstorp Pharma, Lund, Sweden. This article is based on a study first reported in Pharmacology & Toxicology, 78, 104–110;1996.
13.
14.
References 1. Asakura, H.; Morita, A.; Morishita, T.; Tsuchiya, M.; Fukumi, H.; Ohashi, M.; Uylangco, C.; Castro, A. Pathologic findings from intestinal biopsy specimens in human cholera. Digest. Dis. 18:271–278;1973. 2. Beubler, E.; Kollar, G.; Saria, A.; Bukhave, K.; Rask-Madsen, J. Involvement of 5-hydroxytryptamine, prostaglandin E2 , and cyclic adenosine monophosphate in cholera toxin-induced fluid secretion in the small intestine of the rat in vivo. Gastroenterology 96:368–376;1989. 3. Cassuto, J.; Jo¨nsson, A.; Tarnow, P.; Nellga˚rd, P.; Rimba¨ck, G. Anti-inflammation mediated effects in several acute conditions by PP56 (D-myo-inositol-1,2,6-triphosphate). Circ. Shock. Suppl. 1:51;1993. (Abstract 12.20) 4. Castro, G.A.; Harari, Y.; Russel, D. Mediators of anaphylaxisinduced ion transport changes in small intestine. Am. J. Physiol. 253:G540–G548;1987. 5. Ciancio, M.J.; Chang, E.B. Epithelial secretory response to inflammation. Ann. N. Y. Acad. Sci. 664:210–221;1992. 6. Claxson, A.; Morris, C.; Blake, D.; Sire´n, M.; Halliwell, B.; Gustafsson, T.; Lo¨fkvist, B. Bergelin, I. The anti-inflammatory effects of D-myo-inositol-1.2.6-triphosphate (PP56) on animal models of inflammation. Agents Actions 29:68–70;1990. 7. Crosby, W.H.; Kugler, H.W. Intraluminal biopsy of the small intestine. Am. J. Digest. Dis. 2:236–241;1957. 8. Gunther, R.A.; Gustafsson, T.; Brofeldt, B.T. α-Trinositol (α-
15.
16. 17.
18. 19.
20.
T) and decreased fluid balance following thermal injury. Circ. Shock. Suppl. 1:51;1993. (Abstract 12.19) Hansen, M.B.; Skadhauge, E. Ketanserin and granisetron reduce cholera toxin-induced hypersecretion in pig jejunum. Scand. J. Gastroenterol. 29:908–915;1994. Hardcastle, J.; Hardcastle, P.T.; Redfern, J.S. Action of 5-hydroxytryptamine on intestinal ion transport in the rat. J. Physiol. 320:41–55;1981. Lund, T.; Reed, R.K. α-Trinositol inhibits edema generation and albumin extravasation in thermally injured skin. J. Trauma 36(6):761–765;1994. Miller, E.R.; Ullrey, D.E. The pig as a model for human nutrition. Ann. Rev. Nutr. 7:361–382;1987. Nakazawa, H.; Gustafsson, T.O.; Traber, L.D.; Herndon, D.N.; Traber, D.L. α-Trinositol decrease lung edema formation following smoke inhalation in an ovine model. J. Appl. Physiol. 76(1):278–282;1994. Nellga˚rd, P.; Jo¨nsson, A.; Cassuto, J. Effects of D-myo-inositol-1,2,6-trisphosphate (PP56) on cholera toxin-induced secretion. Acta Physiol. Scand. 146(Suppl. 608):2.50;1992. Nilsson, O.; Cassuto, J.; Larsson, P.-A.; Jodal, M.; Lidberg, P.; Ahlman, H.; Dahlstro¨m, A.; Lundgren, O. 5-Hydroxytryptamine and cholera secretion: A histochemical physiological study in cats. Gut 24:542–548;1983. Norris, H.T.; Curran, P.F.; Schultz, S.G. Modification of intestinal secretion experimental cholera. J. Infec. Dis. 119: 117–125;1969. Regunathan, S.; Reis, D.J.; Wahlestedt, C. Specific binding of in hexakisphosphate (phytic acid) to adrenal chromaffin cell membranes and effects on calcium dependent catecholamine release. Biochem. Pharmacol. 43:1331–1336;1992. Shirgi-Degen, A.; Pabst, M.A.; Klimpfinger, M.; Beubler, E. Histopathological effects of cholera toxin in the rat jejunum in vivo. Gastroenterology 104:A279;1993. Timmermans, J.-P.; Scheuermann, D.W.; Stach, W.; Adriaensen, D.; De Groodt-Lasseel, M.H.A. Distinct distribution of CGRP-, enkephalin-, galanin-, neuromedin U-, neuropeptide Y-, somatostatin-, substance P-, VIP- and serotonin-containing neurons two submucosal ganglionic neural networks of the porcine small intestine. Cell Tissue Res. 260:367–379; 1990. Yoo, H.; Lindahl, A.; Veznedaroglu, E.; Wahlestedt, C. Studies on 3H-ins[1,2,6]P3 and other 3H-inositol phosphate binding sites in rat brain. Soc. Neurosci. 18:477;1992. (Abstract)