Abstract

Studies have shown that glucose is able to decrease the pH of the surface epithelium jejunal preparations when added in vitro and the existence of a high concentration of protons in the immediate area of the mucosa could be of considerable significance for absorption of electrolytes. The aim of this study is to assess whether the change in pH of Tyrode (solution used for perfusion of the jejunum) interferes with the absorption of glucose and electrolytes. Male Wistar rats weighing 200 to 220 g (n = 6) were utilized. Jejunal absorption of glucose and electrolytes was investigated in rats. A Tyrode solution containing twice glucose, sodium and potassium concentration (pH 7.0, 7.4, 8.0 and 8.5) was infused through the jejunal loops during 40 minutes. The glucose absorption was not significantly affected by Tyrode. However, there was significantly decrease in sodium absorption at pH 7.0 and 8.5 (41.13 ± 2.79 and 41.37 ± 1.71, respectively, P < 0.05) when compared with the uptake at pH 7.4 and 8.0 (61.06 ± 6.50 and 56.28 ± 7.03, respectively, P < 0.05). Moreover, potassium absorption increased at pH 8.0 (1.04 ± 0.07) when compared with the uptake at pH 7.0 (0.59 ± 0.04), 7.4 (0.78 ± 0.08) and 8.5 (0.54 ± 0.05) (P < 0.05). These data indicate that the pH of Tyrode has no influence on glucose absorption. However, the major potassium uptake occurs at pH 8.0, while the absorption of sodium is impaired at pH 7.0 and 8.5.

Keywords

Absorption; pH; Glucose; Sodium; Potassium; Jejunum

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1. INTRODUCTION

Studies have shown that glucose is able to decrease the pH of the surface epithelium of duodenal and jejunal preparations when added in vitro [1,2], although the acidification is not caused by glucose transport alone [3]. This effect is reproduced when the substrate is administered in vivo [1]. It is plausible that the generation of protons, which are secreted into this region, is closely related to the intracellular metabolism of glucose and lactate production. The existence of a high concentration of protons in the immediate surface of the mucosa could be of considerable significance for the absorption of electrolytes [1].

Recent studies suggest that changes in extracellular pH affect the ionization state of membrane proteins includeing transporters and ion channels with specific functions in transporting epithelia [4]. The surface pH of rat intestine had been measured in vivo, confirming the phenomenon of acidification in the jejunum followed by an alkalinization process in the ileum [5]. However, no difference in the pH of the intestinal surface was detected when the bicarbonate buffer was used or when glucose was included in the buffer. Several factors can affect the intestinal absorption of glucose, electrolytes and water, such as nutritional deficiency [6] or absorption modulation by hormones [7] and peptides [8-12] via the autonomic nervous system. However, the different experimental models used in absorption studies have employed different physiological solutions for infusion, such as Ringer’s solution [13], Krebs’ solution [14], Tyrode solution [8-11,14,15] and phosphate buffer [16,17], with a broad range of variation in the pH of these solutions (pH 7 to pH 8).

The purpose of the present study was to determine whether a change in the pH of Tyrode solution (used for jejunal perfusion) affects the absorption of glucose and electrolytes. These significant new data concerning the dependence of glucose or electrolytes absorption on the pH of Tyrode solution may help to improve methodological approach in future studies of intestinal transport.

2. MATERIAL AND METHODS

2.1. Animals

Adult male Wistar rats weighing 200 to 220 g were housed under standard laboratory conditions of a 12:12-h light/dark cycle and controlled temperature (23˚C ± 3˚C). The animals were fasted for 12 h prior to the experimenttal procedures, but water was offered ad libitum. All experiments complied with the International Principles of Animal Care and the study received approval from the local Ethics Committee on Animal Experimentation (CETEA/UFMG process n˚ 230/2010).

2.2. General Procedures

The rats were anesthetized with thiopental (Cristália, Brazil) (40 mg/kg i.p.). Following the procedures of median xypho-pubic laparotomy, a 20-cm segment of jejunum after the duodenojejunal ligament was isolated, preserving the nerves and the vascular pedicle. Two cannulae were then inserted into the extremities of the jejunal loop—one for perfusion and the other for fluid drainage. The abdominal wall was then closed in order to prevent tissue dehydration. Both cannulae were exteriorized through the extremities of the abdominal suture. Tyrode solution (137 mM NaCl, 2.7 mM KCl, 1.36 mM CaCl₂, 0.49 mM MgCl₂, 11.9 mM NaHCO₃, and 5 mM D-glucose) was maintained in a bottle connected to the catheter infusion pump at 37˚C in a bath. Tyrode solution pH 8.0 (buffered by) was perfused at a rate of 0.5 ml·msin^–1 for 15 min in order to equilibrate the fluids to reach steady state within the jejunal lumen [8].

The rats were divided into four groups: Group 1 (n = 6) received Tyrode solution pH 7.0; Group 2 (n = 6) received Tyrode solution pH 7.4; Group 3 (n = 6) received Tyrode solution pH 8.0; Group 4 (n = 6) received Tyrode solution pH 8.5 The rats in all four groups were submitted to the infusion of a Tyrode solution containing twice the usual concentrations of glucose, sodium and potassium during the 40-min experiment, under the same conditions described above for Tyrode solution. The effluents were collected separately in test tubes at 10-min intervals and maintained in ice, then stored in a freezer at –20˚C until the biochemical analysis.

2.3. Biochemical Determinations

The glucose concentration of the effluent was determined by an enzymatic method based on glucose oxidase (Glucose PAP Liquiform, Labtest, Brazil), estimated from a standard glucose concentration curve. Effluent potassium and sodium ion concentrations were measured by flame photometry. The results were expressed by the difference between influx and efflux.

2.4. Statistical Analysis

Split-plot analysis of variance (ANOVA), followed by the Student-Newman-Keuls method, was used for the statistical analysis, with the level of significance set at 5% (P < 0.05).

3. RESULTS AND DISCUSSION

Figure 1 shows the effect of Tyrode pH on jejunal glucose absorption. There were no differences in glucose absorption among the four groups.

Conflicts of Interest

The authors declare no conflicts of interest.

References