A preparation of perfused small intestine for the study of absorption in amphibia

Abstract

1. A preparation of amphibian small intestine perfused through its vascular system is described. Vascular perfusion with a bicarbonate Ringer solution containing a colloid is used to control the composition of the environment of the submucosal faces of the absorbing cells and to carry away for collection any material extruded from these cells. Oxygenation of the mucosal cells is derived primarily from fluid circulated through the intestinal lumen. The preparation exhibits physiological properties of transport for periods of up to 5 hr. After 5 hr perfusion the epithelial cells show no signs of gross cellular damage when examined either by light or by electron microscopy.2. The relationship between the hydrostatic pressure at the mesenteric artery and the rate of perfusion through the vascular bed is substantially linear. The pressure-flow relationships in the mesenteric bed, including an apparent ;critical closing pressure', are primarily determined by the hydrostatic pressure in the intestinal lumen. Alterations in the hydrostatic pressure in the intestinal lumen also change the relative proportions of the vascular infusate which appear in the portal venous effluent and in the fluid exuded from the serosal surface of the preparation (;sweat'). Hydrostatic distension pressures above about 10 cm H(2)O reduce the rate of collection of fluid from the portal vein and increase the rate of collection of ;sweat'.3. An increase in the rate of vascular perfusion increases the total rate of glucose appearance although the glucose concentrations in both the portal effluent and the ;sweat' are reduced.4. The glucose translocation rate is related in an alinear saturable fashion to the luminal concentration of glucose. By making a correction for metabolic loss of glucose during its passage through the intestinal cell, the relationship existing between the lumen concentration and the uptake of the sugar by the mucosal cells has been calculated. This relationship is found to fit Michaelis-Menten type kinetics. The K(m) of the intestinal translocation process for glucose in Rana pipiens was 0.45 +/- 0.13 (4) muM. The mean V(max) was 137.5 +/- 35.3 (4) muM/hr/g fat-free dry wt.5. When phlorrhizin (10(-5)M) is added to the vascular perfusate, no inhibition of glucose transport is seen for at least 60 min. When strophanthin is added to the vascular perfusate (5 x 10(-5)M), a markedly greater inhibition of glucose transport is observed than when it is introduced to the luminal circulation.6. Earlier studies of the vascular perfusion of isolated small intestine are tabulated. The experimental findings are discussed in relation to a model of the mode of action of the epithelial cell for glucose transport.