Journal of Surgical Research 161, 278–281 (2010) doi:10.1016/j.jss.2009.02.009
Esophageal and Gastric Smooth Muscle Activity After Carbon Dioxide Pneumoperitoneum1 Tutku Soyer, M.D.,*,2 Zuhal Aktuna, M.D.,† Tolga Resxat Aydos, M.D.,† Go¨khan Osmanog˘lu, M.D.,‡ Og˘uzhan Korkut, M.D.,† Hu¨lya Akman, M.D.,* and Murat C ¸ akmak, M.D.* *Department of Pediatric Surgery; †Department of Pharmacology; and ‡Department of General Surgery, Kırıkkale University, Faculty of Medicine Kırıkkale, Turkey Submitted for publication November 8, 2008
Background. The metabolic changes associated with carbon dioxide (CO2) pneumoperitoneum include metabolic acidosis and lowered intra-abdominal pH values. An experimental study was performed to evaluate the effect of CO2 pneumoperitoneum on esophageal and gastric smooth muscle sensitivity in response to several agonists. Methods. Wistar albino rats, weighing 200–250 g, were allocated into three groups. After anesthetization with ketamine hydrochloride and xylazine, abdominal esophagus, gastroesophageal junction, and gastric fundus were removed via median laparotomy in the control group. In the oxygen (O2) group, a 16G catheter was inserted into the abdomen above the umbilicus and insufflated with 95% O2 and 5% CO2 with a pressure of 10 mm Hg. In the CO2 group, CO2 was insufflated at the same pressure within the same time and the tissues were removed at the end of a 60 min period of pneumoperitoneum. Abdominal esophageal segment (n:6), gastroesophageal junction (n:6) and gastric fundus (n:12) were suspended under 0.5 to 2 g resting tension in Tyrode solution in organ baths. Contraction responses were obtained by carbachol and serotonin and relaxation responses were evaluated by isoproterenol in each group. All the responses were compared by nonparametric Kruskal Wallis test. Results. Carbachol and serotonin induced contractile responses of abdominal segments, gastroesophageal junction, and gastric fundus showed no difference between the control, O2, and CO2 groups (P > 0.05). Isoproterenol relaxation responses of the three groups 1 This study was presented at 9th Annual Congress of British Association of Paediatric Endoscopic Surgeons, October 8–10, 2007, _ Istanbul, Turkey. 2 To whom correspondence and reprint requests should be addressed at Department of Pediatric Surgery, Kırıkkale University, Faculty of Medicine, Fabrikalar mah. 71100, Kırıkkale, Turkey. E-mail:
[email protected].
0022-4804/$36.00 Ó 2010 Elsevier Inc. All rights reserved.
were also not statistically different from each other (P > 0.05). Conclusion. CO2 pneumoperitoneum of 60 min has no influence on esophageal and gastric smooth muscle responses to different agonists in rats. Ó 2010 Elsevier Inc. All rights reserved.
Key Words: esophagus; gastric; smooth muscle; carbon dioxide; pneumoperitoneum.
INTRODUCTION
Laparoscopic surgery has become widely accepted within pediatric surgical practice over the last decade [1]. Laparoscopic techniques were introduced with their superiority to classical surgical procedures and are supposed to lead to reduction in overall morbidity. The minimally invasive surgery generally requires a pneumoperitoneum for adequate visualization and exposure of the structures to be operated upon [2]. Carbon dioxide (CO2) is the most common gas used for pneumoperitoneum due to its high diffusion capacity with subsequent rapid absorption and excretion [2]. It is evident from several clinical and experimental studies that CO2 pneumoperitoneum during laparoscopy is associated with adverse physiologic effects like changes in ventilation, and it may also lead to alterations in cellular, hormonal, and immunological parameters [2]. The most common side effects of CO2 pneumoperitoneum are hypercapnia, acidosis, and pulmonary hypertension [3]. Although it is believed that laparoscopic surgery contributes to earlier recovery of postoperative gastrointestinal motility compared with conventional
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approach, several studies have demonstrated that hypercapnia and acidosis inhibit the activity of gastrointestinal smooth muscle [4]. Our aim was thus to evaluate the effect of CO2 pneumoperitoneum on esophageal and gastric smooth muscle activity. For this purpose, an in vitro study was performed to investigate the sensitivity of rat esophagus and gastric fundus to contracting and relaxing agonists after CO2 pneumoperitoneum. MATERIALS AND METHODS Eighteen Wistar albino rats, weighing 200–250 g, were allocated into three groups (control, oxygen [O2], and CO2 groups). Care and use of the animals in this study conformed to the recommendations of the Declaration of Helsinki, and internationally and locally accepted principles were taken into consideration.
Surgical Procedure The rats were anaesthetized with ketamine hydrochloride (Ketalar; Parke-Davis, Pfizer, Istanbul, Turkey) and xylazine (Rompun; Bayer, Leverkusen, Germany) in all the groups. The abdominal segment of esophagus, gastroesophageal junction and gastric fundus were removed via median laparotomy after an hour of anesthesia without any other procedure in the control group (n:6). In the O2 (n:6) and CO2 (n:6) groups, after the anesthesia, a 16G catheter was inserted into the abdomen above the umbilicus and gas insufflation was performed with an insufflator until the intra-abdominal pressure reached 10 mmHg. In the O2 group, 95% O2 and 5% CO2 was insufflated and intra-abdominal pressure was kept at 10 mm Hg for 60 min. In the CO2 group, CO2 was insufflated for 60 min and intra-abdominal pressure was also maintained at 10 mm Hg during the experiment. CO2 insufflation method is similar to a model that is already known to induce local and systemic acidosis [2]. At the end of the experiments, after exsufflation (in O2 and CO2 groups) and median laparotomy, the tissues were removed and placed in petri dishes containing Tyrode solution.
In vitro Studies After 60 min of carbon dioxide pneumoperitoneum, abdominal esophageal segment, gastroesophageal junction, and gastric fundus were dissected of fat and connective tissues. Shortly after the dissection, all the tissues were suspended under an optimal resting tension in organ baths containing 10 mL of Tyrode solution (pH: 7.4) at 37 C, and they were continuously aerated with 95% O2 and 5% CO2. Resting tension was 1 g for abdominal esophagus, 2 g for gastroesophageal junction and 0.5–0.75 g for gastric fundus preparations. Tissues were allowed to equilibrate for 60 min and Tyrode solution was changed in every 15 min. Tyrode solution consisted of NaCl: 137 Mm, KCl: 2.68 Mm, CaCl2: 1.4 Mm, NaHCO3: 11.9 Mm, NaH2PO4: 0.42 Mm, MgCl2: 0.5 Mm, and glucose: 5.5 mM. To determine the variations in the in vitro sensitivity of rat esophageal and gastric smooth muscles to contractile agonists, cumulative concentration response curves of carbachol (108–104 M) and serotonin (108–104 M) were obtained and to determine the relaxation responses after contracting the tissues with a submaximal concentration of carbachol (105 or 3 3 105 M), cumulative concentration response curves of isoproterenol (108 or 3 3 104 M) were studied. TRI 202P/0-25 G (LSI LECTICA, Scientific Instruments, Pan LAB, S.L., Cornella, Spain) force displacement transducers and Powerlab/8P, Quad Bridge amplifier (ML 118) (AD Instruments, NSW, Australia) were used to record the data.
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Agonists Carbachol, serotonin, and isoproterenol were used during the study (Sigma Chemical Co., St. Louis, MO). The stock solutions were prepared by dissolving them in distilled water and the dilutions of different concentrations were made just before use.
Statistical Analysis All data are expressed as mean 6 SEM. Responses of the contracting agents were expressed as g or mg tension and relaxation responses were expressed as the percentage of the submaximal carbachol contractions in each individual preparation. The results were statistically compared by Kruskal Wallis analysis of variance; post hoc Dunn’s test was used when needed. Results were considered to be significant when P values were less than 0.05.
RESULTS
Abdominal esophageal segments, gastroesophageal junction, and gastric fundus displayed concentrationdependent contractions to a muscarinic receptor agonist, carbachol (108–104 M) (Fig. 1). Although not statistically significant, these responses were increased with O2 insufflation, especially at the higher concentrations of carbachol in abdominal esophageal segments (Fig. 1A), but CO2 insufflation had no prominent effect on carbachol-induced responses in this tissue. Carbachol-induced concentration-dependent contractions in the gastric fundus and gastroesophageal junction did not differ with CO2 or O2 insufflation (Fig. 1B and C). In presence of serotonin (108–104 M), gastric fundus and gastroesophageal junction demonstrated concentration-dependent contractions, but it was not possible to obtain any response in the abdominal esophageal segments. When the serotonin-induced contractions of gastric fundus and gastroesophageal junction were compared, there was no significant difference between the three groups (P > 0.05) (Fig 2A and B). Relaxation responses were obtained in abdominal esophageal segments and gastroesophageal junction. Isoproterenol (108 and 3 3 104 M), a b receptor agonist caused concentration-dependent relaxation responses in these tissues after precontraction with carbachol (105 and 3 3 105 M). CO2 and O2 pneumoperitoneum of 60 min did not affect the isoproterenol relaxation responses in abdominal esophagus and gastroesophageal junction (Fig. 3A and B). DISCUSSION
It has been demonstrated in several clinical and experimental studies that laparoscopic surgery contributes to earlier recovery of postoperative gastrointestinal motility than that experienced with conventional surgery [4]. Some authors have stated that gastrointestinal motility showed no difference following laparoscopic and
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FIG. 1. Concentration-response curves of carbachol (108–104 M) in the rat abdominal esophagus (A), gastric fundus (B), and gastroesophageal junction (C), in control (n ¼ 6), CO2 (n ¼ 6), and O2 (n ¼ 6) groups. Responses are expressed as gram (g) contraction. Each point represents mean 6 SEM of (n) experiments. * P < 0.05.
conventional approaches [5], while others suggested that CO2 pneumoperitoneum may inhibit the activity of gastrointestinal motility [4]. While studies regarding gastrointestinal system motility in laparoscopic surgery are inconsistent, pharmacological studies of gastrointestinal smooth muscle activity in laparoscopic models are still a few. In the present study, we aimed to evaluate the in vitro sensitivity of rat esophagus and gastric smooth muscles to different contracting and relaxing agonists in a CO2 pneumoperitoneum model. CO2 is the preferred gas for use in laparoscopic procedures. It is nonflammable, low in cost and has low risk of venous embolism [2]. It is also well tolerated by children. However, insufflated CO2 is absorbed from the peritoneum into the bloodstream and can induce hypercapnia and acidosis [4]. Hypercapnia and metabolic acidosis
due to CO2 can be observed at the very beginning of insufflations and may persist until 6 h after surgery [4]. These complications not only lead to cardiopulmonary disorders but also impair gastrointestinal motility [4]. Schulze-Delrieu et al. reported that acidosis induces an increase in baseline tension and decrease in amplitude of contracting and relaxing response in opossum esophagus and stomach [6]. Tournadre et al. found that metabolic and respiratory acidosis increase fundic tone and alter pyloric and antral contractility, whereas hypercapnia has no effect on lower esophageal sphincter activity [7]. The proposed mechanism of gastrointestinal dysmotility in acidosis is associated with decreased sensitivity of polarized smooth muscle membrane to cholinergic stimulation and reduced rate of release of stored Ca in transmembranous flow into muscle cells [4]. In our study, there were no differences between
FIG. 2. Concentration-response curves of serotonin (108–104 M) in the rat gastric fundus (A) and gastroesophageal junction (B) in control (n ¼ 6), CO2 (n ¼ 6), and O2 (n ¼ 6) groups. Responses are expressed as gram (g) contraction in gastric fundus and in milligram (mg) contraction in gastroesophageal junction. Each point represents mean 6 SEM of (n) experiments.
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FIG. 3. Concentration-response curves of isoproterenol (108 or 3 3 104 M) in the rat abdominal esophagus (A) and gastroesophageal junction (B) in control (n ¼ 6), CO2 (n ¼ 6), and O2 (n ¼ 6) groups. Each point represents mean 6 SEM of (n) experiments. Isoproterenol relaxation responses are expressed as the percentage of the precontraction obtained with carbachol (105 or 3 3 105 M).
CO2, O2, and control groups with respect to esophagus and gastric smooth muscle responses to contracting and relaxing agonists. We suggest that although metabolic changes may occur in the early phase of insufflations, the effect on smooth muscle sensitivity for several agonists may be observed in prolonged exposures. The role of metabolic changes in motility disorders has been clearly identified in several studies; however, CO2 pneumoperitoneum models in organ bath experiments are limited. The first study concerning the effects of CO2 pneumoperitoneum in the rat ileum was reported by Unsal et al. [8]. They found inhibited contractions in ileum segments of rats under high intraabdominal pressures. Their results showed no statistical difference in 10 mmHg pressure, which is the level used in most pediatric and experimental laparoscopic procedures [7]. However, decreased contractile responses were obtained with 20 mm Hg pressure [8]. We found that in CO2 pneumoperitoneum, different esophageal segments and gastric muscle strips had similar responses to both carbachol and serotonin compared with control and O2 groups. Furthermore, isoproterenol relaxation responses showed no difference between the groups. Our results are similar with ileal responses reported by Unsal et al. under 10 mmHg of intraabdominal pressure. In light of these two studies, we suggest that 60 min of CO2 pneumoperitoneum has no major effect on gastrointestinal smooth muscle activity under standard intra-abdominal pressure. In order to identify the difference between ileal, esophageal, and gastric smooth muscle responses, further studies with high intra-abdominal pressures or
with longer duration of pneumoperitoneum should be performed in our experimental model. In conclusion, 60 min of CO2 pneumoperitoneum had no influence on the responses of the esophageal and gastric smooth muscles to different agonists in rats if laparoscopic approaches are performed under standard intra-abdominal pressure and duration. REFERENCES 1. Neuhaus SJ, Watson DI, Ellis T, et al. Metabolic and immunologic consequences of laparoscopy with helium or carbon dioxide insufflation: A randomized clinical study. ANZ J Surg 2001;71:447. 2. Kuntz C, Wunsch A, Bo¨deker C, et al. Effect of pressure and gas type on intra-abdominal, subcutaneous, and blood pH in laparoscopy. Surg Endosc 2000;14:367. 3. McLaughlin JG, Scheeres DE, Dean RJ, et al. The adverse hemodynamic effects of laparoscopic cholecystectomy. Surg Endosc 1995;9:121. 4. Waseda M, Murakami M, Kato T, et al. Helium gas pneumoperitoneum can improve the recovery of gastrointestinal motility after a laparoscopic operation. Min Inv Ther Allied Technol 2005;14:14. 5. Hotokezaka M, Dix J, Mentis EP, et al. Gastrointestinal recovery following laparoscopic versus open colon surgery. Surg Endosc 1996;10:485. 6. Schulze-Delrieu K, Lepsien G. Depression of mechanical and electrical activity in muscle strips of opossum stomach and esophagus by acidosis. Gastroenterology 1982;82:720. 7. Tournadre JP, Allaouchiche B, Malbert CH, et al. Metabolic acidosis and respiratory acidosis impair gastro-pyloric motility in anesthetized pigs. Anesth Analg 2000;90:74. 8. Unsal MA, Imamoglu M, Kadioglu M, et al. The acute alterations in biochemistry, morphology, and contractility of rat-isolated terminal ileum via increased intra-abdominal pressure. Pharmacol Res 2006;53:135.
