Technology and clinical application of ultrasonic dissection.

The implementation of UltraCision (The Harmonic Scalpel) can be seen as a milestone in the development of surgery. Ultrasound is now used for tissue cutting and/or simultaneous vessel sealing and transection. No electric current passes through the patient. Thus the typical complications associated with electrocautery can be avoided. This technology was primarily developed for videoscopic surgery and then successfully transferred to all branches of open surgery. The clean and blood-saving dissection technology and the ability to dissect very close to sensitive structures in oncological surgery are highly beneficial for patients. The development of new blades and multifunctional shears has further enhanced both practicability and ergonomics. UltraCision can now be used for the complete surgical spectrum, both in open and in laparoscopic surgery.

MR imaging for the preoperative planning of sphincter-saving surgery for tumors of the lower third of the rectum: use of intravenous and endorectal contrast materials.

PURPOSE: To evaluate the value of magnetic resonance (MR) imaging with a flexible surface coil in predicting the resectability of tumors in the lower rectum and the feasibility of sphincteral salvage. MATERIALS AND METHODS: In a prospective study, 61 patients with histologically proved primary adenocarcinoma of the lower or middle third of the rectum (<12 cm from the pectinate line) were examined at double-contrast-material-enhanced MR imaging with a circular polarized flexible surface coil. RESULTS: Assessment of anal sphincteral infiltration at MR imaging was excellent, with a specificity of 98% and a sensitivity of 100%. In the determination of tumor infiltration into adjacent organs (T4), the specificity was 100%, and the sensitivity was 90%, with surgical and histologic findings as the standards. While MR imaging showed negative nodes in 40 patients (stage N0 at MR imaging), histologic examination showed negative nodes in 27 patients and positive nodes in 34. At MR imaging, sensitivity was 68%, and specificity was 24%. CONCLUSION: While preoperative staging at MR imaging according to the TNM system still has limited value and accuracy, MR imaging provides the surgeon with valuable information regarding the presence of sphincteral invasion and the surrounding structures in patients with cancers in the lower third of the rectum.

Effects of temperature on the flight activity of Graphocephala atropunctata (Hemiptera: Cicadellidae).

Graphocephala atropunctata (Signoret) is the principal vector of Xylella fastidiosa (Wells, Raju, Hung, Weisberg, Mandelco-Paul and Brenner), the bacterium that causes Pierce's disease of grapevine in coastal California. Monitoring the activity of C. atropunctata in the early spring is important for timing insecticide sprays and assessing the potential for disease spread to adjacent vineyards. Trapping studies with yellow sticky traps over 3 yr in Napa Valley, CA, established a significant correlation between early spring trap catch and temperature. Sticky trap catches of G. atropunctata occurred in the springs of 1996-1998 only when temperature was greater than or equal to 14.5 degrees C. In 1997 and 1998, the degree-hours (> 14.5 degrees C) per day from sunrise to sunset during March and April, but not in May, correlated significantly with trap catches. The temperature threshold of 14.5 degrees C in the early spring can be used to improve the timing of insecticidal applications aimed at reducing C. atropunctata populations in vineyards affected by Pierce's disease.

[Anal sphincter restoration using dynamic graciloplasty--results of 50 patients]. / Analer Schliessmuskelersatz mittels dynamischer Gracilisplastik--Ergebnisse der Anwendung an 50 Patienten.

During the period from 1992 to 1998, 50 patients underwent anal sphincter restoration by dynamic graciloplasty for primary (n = 26) or secondary (n = 6) total anorectal reconstruction (TAR) following abdominoperineal rectal resection (APR) or acquired (n = 9) or congenital (n = 9) fecal incontinence, respectively. Forty-seven patients were operated on by a single-stage procedure using a modified technique for the muscle wrap ("split sling"). Muscle fiber transformation by controlled stimulation was achieved at the beginning of the learning curve within 8 weeks and in the meantime within 4 weeks. Rectal injury (n = 10) turned out to be the most serious postoperative complication and was observed mainly in patients following TAR (n = 8). As the most prominent functional problem constipation in patients following TAR hampered the postoperative functional result; however, this was overcome by regular enemas. An improvement in the continence status was observed in 80% of the patients treated for fecal incontinence, and following APR 66% of the patients had acceptable results without a permanent colostomy.

Bacteroides fragilis toxin 2 damages human colonic mucosa in vitro.

BACKGROUND: Strains of Bacteroides fragilis producing a 20 kDa protein toxin (B fragilis toxin (BFT) or fragilysin) are associated with diarrhoea in animals and humans. Although in vitro results indicate that BFT damages intestinal epithelial cells in culture, the effects of BFT on native human colon are not known. AIMS: To examine the electrophysiological and morphological effects of purified BFT-2 on human colonic mucosa in vitro. METHODS: For resistance (R) measurements, colonic mucosa mounted in Ussing chambers was exposed to luminal or serosal BFT-2 (1.25-10 nM) and after four hours morphological damage was measured on haematoxylin and eosin stained sections using morphometry. F actin distribution was assessed using confocal microscopy. RESULTS: Serosal BFT-2 for four hours was four-, two-, seven-, and threefold more potent than luminal BFT-2 in decreasing resistance, increasing epithelial 3H-mannitol permeability, and damaging crypt and surface colonocytes, respectively (p<0.05). Confocal microscopy showed reduced colonocyte F actin staining intensity after exposure to BFT-2. CONCLUSIONS: BFT-2 increases human colonic permeability and damages human colonic epithelial cells in vitro. These effects may be important in the development of diarrhoea and intestinal inflammation caused by B fragilis in vivo.

Restoration of anal sphincter function by single-stage dynamic graciloplasty with a modified (split sling) technique.

BACKGROUND: Controlled muscle fiber conversion by electrostimulation makes transformation of fast twitching type II muscle fibers to slow twitching type I fibers possible, which gives skeletal muscles the capacity for tetanic contraction. This phenomenon has been recently applied in the so-called "dynamic graciloplasty" to restore function of an insufficient or excised anal sphincter. This paper describes our results with this method in patients with fecal incontinence or following an abdomino-perineal resection (APR) of the anorectum. METHODS: From April 1992 through April 1997, 28 patients (12 women and 16 men) were treated by dynamic graciloplasty. The median age was 53.5 years (range 16 to 79). Indications were as follows: APR + synchronous restoration of the excised sphincter by graciloplasty (n = 12); total anorectal reconstruction (TAR) following APR in the past (n = 6); Patients with acquired fecal incontinence (n = 4); and Congenital atresia (n = 6). Muscle transposition, implantation of stimulation electrodes and pulse generator were done as a single-stage procedure, the "neosphincter" was wrapped in a modified technique (split-sling technique). Muscle transformation was performed by controlled neuromuscular stimulation during 8 weeks (from 1992 to 1995) and 4 weeks (since 1996), respectively. RESULTS: No postoperative mortality (90 days) was observed in either group. In our early experience, rectal injury occurred in 4 patients as the most prominent complication. Evaluation of the functional outcome showed the best results in patients operated either for congenital of acquired incontinence who achieved a continence for solids and liquids or solids alone, respectively (1 or 2 according to Williams' score) in 90%, while patients following APR showed a satisfying outcome (continence for solids and liquids, solids alone or with occasional episodes for liquids) in only 55.5%. In patients following APR, defecation disorders turned out to be the most prominent functional problem and had to be treated by enemas. CONCLUSION: In this series, we have been able to perform dynamic graciloplasty as a one-stage procedure using a modified muscle wrap (split-sling-technique) thus reducing the time period until continence could be achieved to 7 weeks. We found the appropriate tension of the muscle wrap essential to prevent direct injury to the rectum as it was seen in our early experience. For this reason, we have introduced a modified device to perform intraoperative anal manometry and to measure pressures created by the neosphincter objectively.

Epidermal growth factor attenuates Clostridium difficile toxin A- and B-induced damage of human colonic mucosa.

Epidermal growth factor (EGF) exhibits a cytoprotective effect on gastrointestinal epithelia via a receptor-mediated mechanism. We investigated the effect of EGF on Clostridium difficile toxin A (TxA)- and toxin B (TxB)-induced damage of human colon. Ussing-chambered colonic mucosa was exposed serosally to EGF before and during luminal exposure to TxA and TxB. Resistance was calculated from potential difference and short-circuit current. Epithelial damage was assessed by light microscopy and alteration of F-actin by fluoresceinated phalloidin. Luminal exposure of colonic strips to TxA and TxB caused a time- and dose-dependent decrease in electrical resistance, necrosis and dehiscence of colonocytes, and disruption and condensation of enterocyte F-actin. These effects were inhibited by prior, but not simultaneous, serosal application of EGF (20 nM). Administration of the tyrosine kinase inhibitor genistein (10(-6) M) inhibited the protective effects of EGF. We conclude that EGF protects against TxA and TxB probably by stabilizing the cytoskeleton, the main target of these toxins.

Muscle transformation of the sartorius muscle in a canine model: clinical impact for electrodynamic graciloplasty as a "neosphincter".

PURPOSE: Transformation of fast-twitching skeletal muscles to slow-twitching, slowly fatigable muscles has become of clinical interest in the recent past. Transposition and transformation of the gracilis muscle to use it as a substitute for a resected or defected anal sphincter (graciloplasty) have been reported as achieving promising results in the treatment of fecal incontinence caused by sphincter defects or following abdominoperineal anorectal excision for cancer. METHOD: This experimental study used a canine model and the sartorius muscle to evaluate the functional efficiency of two different configurations of the muscle loop to compare the presently applied transformation program (8 weeks) with a shorter (5 weeks) protocol. In six beagle dogs, both sartorius muscles were wrapped around two stomas, either in an alpha fashion or in the so-called split-sling technique. Muscle transformation was achieved by controlled neuromuscular stimulation either during eight (Program A) or five weeks (Program B). After completion of the transformation period, the function of the muscle slings was evaluated by manometry, and histomorphologic evaluation of the sartorius muscles was performed. RESULTS: It was shown that muscle transformation led to a slowly fatigable muscle that made it possible to perform continuos (tetanic) contraction, regardless of the configuration or the duration of the transformation. Median pressures created by these muscles also did not differ significantly. In accordance with these functional findings, the histologic evaluation showed the typical, significant increase of Type I fibers in both muscle slings and following both transformation protocols. Although the decrease of fast-twitching Type II fibers was more pronounced following the conventional (8 weeks) program, this finding did not influence the functional results. CONCLUSIONS: Results of our experiment indicate the possibility for using a shorter transformation protocol for transformation of the gracilis muscle during graciloplasty in the clinical setting. Furthermore, the efficacy and safety of the modified (split-sling) wrap technique was demonstrated.

Effect of growth factors on epithelial restitution of human colonic mucosa in vitro.

BACKGROUND: Epithelial restitution enables resurfacing of epithelial discontinuities by enterocyte migration. This study investigated the effect of basic fibroblast growth factor (bFGF), insulin-like growth factor (IGF-1), and epidermal growth factor (EGF) on restitution of human colonic mucosa in vitro. METHODS: After base-line incubation human colonic mucosal strips, mounted in Ussing chambers, were luminally exposed to 0.5 mM sodium deoxycholate (NaDOC) for 10 min. Thereafter tissues were incubated with buffer alone or luminal buffer containing various concentrations of bFGF, IGF-1, and EGF for 3 h. Resistance (R) was calculated from potential difference (PD) and short-circuit current (Isc). All tissues were processed for light microscopy. Extent of damage was measured by morphometry. RESULTS: Luminal 0.5 mM NaDOC for 10 min caused R to drop by 43% (n = 4; P < 0.05). Compared with controls 50 ng/ml EGF induced an approximately 30% R increase until the end of the experiments (P < 0.05, n = 4, paired). Ten minutes after injury 50.2 +/- 4% of the mucosa was damaged (n = 6), and after 3 h damage was significantly reduced by EGF (17.2 +/- 3% versus 31.7 +/- 4%, 50 ng/ml EGF versus controls) (P < 0.05, n = 6 per group). Histology showed that EGF stimulated enterocyte migration over the basal lamina. Various doses of bFGF and IGF-1 did not impair restitution when compared with controls. CONCLUSION: In contrast to bFGF and IGF-1, EGF was shown to promote epithelial restitution of human colonic mucosa in vitro.

Laminin stimulates rapid epithelial restitution of rabbit duodenal mucosa in vitro.

BACKGROUND: This study investigated the effect of the basal lamina constituents fibronectin, collagen IV, and laminin on epithelial restitution of rabbit duodenum in vitro. METHODS: Rabbit duodenal mucosal sheets were mounted in Ussing chambers, luminally exposed to 10 mM HCI for 10 min, and incubated with buffer or luminal buffer containing 25-100 micrograms/ml of collagen IV, fibronectin, laminin, or polyclonal antisera directed against these proteins (diluted 1:50-1:20) for 3 h. Resistance was calculated from potential difference and short-circuit current. Mucosal damage was assessed by morphometry on hematoxylin- and eosin-stained sections. RESULTS: Acid exposure caused a 40% drop in resistance (119 +/- 5 versus 71 +/- 5 Ohm.cm2 before versus after injury; P < 0.05, n = 6) and mucosal damage of 58 +/- 4% (n = 6). Three hours after injury resistance was 102 +/- 6, 117 +/- 4, and 48 +/- 5 Ohm.cm2 in the control, laminin, and anti-laminin groups, respectively. Furthermore, 36 +/- 2%, 16 +/- 2%, and 64 +/- 5% of the mucosa was damaged in the control, laminin, and anti-laminin groups, respectively, 3 h after injury (P < 0.05 versus controls). Laminin promoted epithelial wound closure by stimulation of enterocyte migration, which was inhibited by anti-laminin. Fibronectin, collagen IV, anti-fibronectin, and anti-collagen IV did not impair restitution. CONCLUSION: Our results show that laminin promotes electrophysiologic restoration and epithelial restitution of rabbit duodenum in vitro. We therefore suggest that laminin plays an important part in the orchestration of epithelial integrity and barrier function.

Epidermal growth factor promotes rapid response to epithelial injury in rabbit duodenum in vitro.

BACKGROUND & AIMS: Growth factors are mainly involved in the regulation of intestinal epithelial barrier function. This study investigated the effect of epidermal growth factor (EGF) and insulin-like growth factor 1 (IGF-1) on epithelial restitution of rabbit duodenum in vitro. METHODS: Rabbit duodenal mucosal strips mounted in an Ussing chamber were luminally exposed to 10 mmol/L HCl for 10 minutes and then incubated with buffer alone or luminal buffer containing various concentrations of EGF and IGF-1 for 3 hours. Resistance was calculated from potential difference and short-circuit current. Damage was assessed by morphometry on H&E-stained sections. RESULTS: HCl caused resistance to decrease from 112 +/- 2 to 51 +/- 4 ohms x cm2 10 minutes after injury (n = 6; P < 0.05). Postinjury treatment with 25 or 50 ng/mL luminal EGF for 3 hours stimulated resistance to recover to 94 +/- 3 and 104 +/- 3 ohms x cm2, respectively, vs. 81 +/- 3 omega x cm2 in controls (P < 0.05). Ten minutes after injury, 62% of the mucosa was damaged; 3 hours after injury, damage was reduced to 24% +/- 1.09% and 10% +/- 1.42% in the 25 and 50 ng/mL EGF group, respectively, vs. 38% +/- 0.93% in controls (n = 6 per group). EGF stimulated enterocyte migration. IGF-1 did not impair epithelial restitution. CONCLUSIONS: EGF, but not IGF-1, promoted epithelial restitution of rabbit duodenum in vitro.

Effects of extracellular pH on intracellular pH-regulation and growth in a human colon carcinoma cell-line.

Mechanisms of intracellular pH (pHi) regulation seem to be involved in cellular growth and cell division. Little is known about how extracellular acidosis, known to occur in central regions of solid tumors, or alkaline conditions affect pHi regulation in colonic tumors. pHi changes in the colonic adenocarcinoma cell-line SW-620 were recorded by spectrofluorimetric monitoring of the pH-sensitive, fluorescent dye BCECF, and proliferative activity was assessed by [3H]thymidine uptake. Resting pHi in Hepes-buffered solution was 7.53 +/- 0.01 (n = 36). Both 1 mM amiloride and Na(+)-free solution inhibited pHi recovery from acidification and decreased pHi in resting cells. In HCO3-/CO2-buffered media resting pH1 was 7.42 +/- 0.01 (n = 36). Recovery from acidification was Na(+)-dependent, CI(-)-independent, and only partially blocked by 1 mM amiloride. In the presence of amiloride and 200 microM H2DIDS pHi recovery was completely inhibited. In Na(+)-free solution pHi decreased from 7.44 +/- 0.04 to 7.29 +/- 0.03 (n = 6) and no alkalinization was observed in CI(-)-free medium. Addition of 5 microM tributyltin bromide (an anion/OH-exchange ionophore) caused pHi to decrease from 7.43 +/- 0.05 to 7.17 +/- 0.08 (n = 5). The effects of pH0 on steady-state pHi, pHi recovery from acidification and proliferative activity after 48 h were investigated by changing buffer [CO2] and [HCO3-]. In general, increases in pH0 between 6.7 and 7.4 increased pHi recovery, steady-state pHi and growth rates. In summary, SW-620 cells have a resting pHi > 7.4 at 25 degrees C, which is higher than other intestinal cells. Acid extrusion in physiological bicarbonate media is accomplished by a pHi-sensitive Na+/H+ exchanger and a pHi-insensitive Na(+)-HCO3-cotransporter, both of which are operational in control cells at the resting pHi. No evidence for activity of a CI-/HCO3- exchanger was found in these cells, which could account for the high pHi observed and may explain why the cells continue to grow in acidic tumor environments.

Metabolic base production and mucosal vulnerability during acid inhibition in a mammalian stomach in vitro.

Acid inhibition increases gastric mucosal susceptibility to damage by luminal acid. This might be due to reduced metabolic CO2 and bicarbonate whereas, during normal acid, secretion cytoprotective CO2/HCO3- production parallels acid production. Metabolic activity and mucosal damage caused by luminal acid perfusion was determined in an in vitro mouse stomach, with and without acid inhibition, and at 0%, 1%, or 5% serosal CO2 supply. Without acid inhibition there was no mucosal damage at any level of serosal CO2/HCO3- supply. Acid inhibition reduced metabolic CO2 production by 29% (P < 0.004) and resulted in microscopic damage to 55% of the mucosal area and perforation in four of five stomachs (P < 0.05). Although, 1% CO2 supply completely replaced the reduction in metabolic CO2, it did not protect against mucosal damage. Overreplacement by 5% serosal CO2/HCO3- was required to prevent damage. There was no correlation between luminal CO2/HCO3- output and mucosal damage. The protection by endogenous or exogenous CO2/HCO3- appears to act intracellularly rather than by intragastric or intercellular neutralization.

Peptic esophageal stricture: is surgery still necessary?

Gastroesophageal reflux disease is frequently complicated by peptic esophageal stricture formation. Treatment of choice over the past 25 years has changed from resection of the stenotic esophagus towards fundoplication, or conservative treatment combined with dilatation. Reports on the long-term results of the clinical course of such patients are still rare. Between 1965 and 1990 200 patients were treated for peptic esophageal stricture by surgery or bougienage with antisecretory medication. Retrospective analysis of the clinical outcome according to the primary therapeutic strategy was performed after a follow-up period of 1.5 to 267 months. 139 patients (group A) primarily received bougienage and medical treatment. After 71 months 36% of the patients were symptom-free, 52% had received further dilatation and 11% had undergone surgery. One fatal complication occurred. 61 patients (group B) underwent primary surgical treatment. Fundoplication was performed in 72% of the cases, resection in 18% and other procedures in 10%. After a median period of 84 months following standard fundoplication (n = 43) 44% were free of symptoms, 39% had received further dilatations and 12% had to be reoperated. Fatal complications occurred in 2 patients (5%). The risk of undergoing surgery after primary dilatation was 16% after 2 years, remaining on this level throughout follow-up time. We conclude that resection of peptic strictures of the esophagus is rarely indicated any more. Treatment of choice consists of primary bougienage combined with antisecretory medication. If conservative treatment fails or patient compliance is low we recommend fundoplication with intraoperative dilatation within the first 2 years after diagnosis of symptomatic stricture.

[Dilatation treatment of achalasia as initial therapy. Results of a prospective study]. / Dilatationsbehandlung der Achalasie als Initialtherapie. Ergebnisse einer prospektiven Studie.

From 1972 to 1991 primary dilatation therapy was investigated in a prospective study in 77 patients suffering from achalasia of the gastroesophageal sphincter. The patients were followed up continuously and the median follow-up period was ten years. 70% of patients were symptom-free after dilatation therapy at the time of the last follow-up examination, whereby 50% were cured of symptoms after a single dilatation. 27% required surgery after unsuccessful dilatation therapy. The probability of avoiding operation after dilatation therapy within 15 years of diagnosis of achalasia was 73%. The clinical symptomatology at diagnosis graded according to a scheme from I to IV, was significantly worse in patients requiring an operation (Wilcoxon test, p < 0.0005). The data of esophageal manometry were of no prognostic relevance.

Clostridium difficile toxin B is more potent than toxin A in damaging human colonic epithelium in vitro.

Toxin A but not toxin B, appears to mediate intestinal damage in animal models of Clostridium difficile enteritis. The purpose of this study was to investigate the electrophysiologic and morphologic effects of purified C. difficile toxins A and B on human colonic mucosa in Ussing chambers. Luminal exposure of tissues to 16-65 nM of toxin A and 0.2-29 nM of toxin B for 5 h caused dose-dependent epithelial damage. Potential difference, short-circuit current and resistance decreased by 76, 58, and 46%, respectively, with 32 nM of toxin A and by 76, 55, and 47%, respectively, with 3 nM of toxin B, when compared with baseline (P < 0.05). 3 nM of toxin A did not cause electrophysiologic changes. Permeability to [3H]mannitol increased 16-fold after exposure to 32 nM of toxin A and to 3 nM of toxin B when compared with controls (P < 0.05). Light and scanning electron microscopy after exposure to either toxin revealed patchy damage and exfoliation of superficial epithelial cells, while crypt epithelium remained intact. Fluorescent microscopy of phalloidin-stained sections showed that both toxins caused disruption and condensation of cellular F-actin. Our results demonstrate that the human colon is approximately 10 times more sensitive to the damaging effects of toxin B than toxin A, suggesting that toxin B may be more important than toxin A in the pathogenesis of C. difficile colitis in man.

[Laparoscopic cholecystectomy--a meta-analysis of 23,700 cases and status of a personal patient sample]. / Laparoskopische Cholezystektomie--eine Metaanalyse von 23,700 Fällen und der Stellenwert im eigenen Patientenkollektiv.

Meta-analysis of 23,700 laparoscopic cholecystectomies of 27 authors shows an intraoperative complication rate of 1.35% and a postoperative complication rate of 3%, including injuries of the bile duct in 0.48% and of the intestine in 0.19%. A second operation was necessary in 1.26%. In 0.43% retained common bile duct stones were observed. The conversion rate was 4.1% and the mortality rate was 0.08%. Our own experience in laparoscopic cholecystectomy as a standard procedure on 455 out of 574 unselected patients (50 acute, 405 elective) resulted in 9 (2.2%) intraoperative and 5 (1.2%) postoperative complications. In 4 cases (0.9%) the complications necessitated a second operation. The conversion rate was 11.6% and the mortality rate was zero. It is concluded that the results of laparoscopic cholecystectomy appear equivalent the "gold standard" of open cholecystectomy up to certain pathomorphological limits. Using an exact preoperative diagnostic regimen (sonography, cholangiography, ERCP if indicated) the place of routine intraoperative cholaniography is discussed critically.

Intracellular pH regulation of human colonic crypt cells.

In order to investigate the regulation of intracellular pH (pHi) in freshly isolated human colonocytes, we have used a newly developed technique for the rapid isolation and covalent attachment of these cells to glass surfaces and microspectrofluorimetric measurement of the pH-sensitive fluorescence of 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF)-loaded specimens in a perfusion chamber (37 degrees C). In N-2-hydroxyethylpiperazine-N'-2-ethanesulphonic-acid-(HEPES)-buffered Ringer solution (HBS) a baseline pHi of 7.35 +/- 0.03 (mean +/- SD; n = 42) was found for human colonocytes and in HBS, NH4Cl-prepulse-induced intracellular acidification in colonocytes is reversed rapidly by the ubiquitous amiloride-sensitive (1 mmol/l) Na+/H+ exchanger. Switching from HBS to HCO(3-)-buffered solution (BBS) led to a transient intracellular acification (7.29 +/- 0.09), followed by a recovery to a final resting pHi of 7.43 +/- 0.03. One-third of the acid extrusion in BBS is amiloridesensitive; the remaining two-thirds are caused by the dihydroderivative of 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (H2DIDS)-sensitive HCO(3-)-dependent mechanisms. The functional activity of an acid-extruding Na+/HCO3- cotransporter in human colonocytes was observed in response to the reintroduction of Na+ into amiloride-containing Na+/Cl(-)-free BBS. In addition, the mechanism leading to alkalinization (7.56 +/- 0.05) in Cl(-)-free BBS was identified as Na(+)-dependent Cl-/HCO3- exchange, by its H2DIDS sensitivity and the specific requirement for Cl- and Na+. The intrinsic buffering capacity (beta i) of the human colonocytes was calculated from pH changes induced by sequential NH4Cl-loading steps during blockage of acid/base transporters.(ABSTRACT TRUNCATED AT 250 WORDS)