Preparation and some properties of a suspension of fragmented tubules from rat kidney

Abstract

A method is described whereby short fragments of rat kidney tubule were obtained when kidney slices were gently dispersed by exposure to collagenase and hyaluronidase. When suspended in buffered saline the fragmented tubules respired actively over a period of several hours, the rate of oxygen consumption being proportional to the amount of cell protein. Oxygen uptake was stimulated by the addition of glucose, lactate, butyrate, alpha-oxoglutarate and other substrates and was decreased by the omission of Ca(2+) from the suspending medium. With alpha-oxoglutarate as the added substrate, dinitrophenol strongly stimulated oxygen uptake. Dinitrophenol had a less-marked stimulatory effect when glucose was the added substrate, and inhibited respiration in the absence of added substrate. Oligomycin inhibited respiration and this inhibition was partially reversed by dinitrophenol. Fragmented tubules synthesized glucose from lactate at a high rate but this capacity for gluconeogenesis was abolished by dinitrophenol and by physically damaging the cells.

A rapid enzymatic method for the isolation of defined kidney tubule fragments from mouse

“…[7,14,15,18,22,28,36,38,45,49]). Currently, several techniques are employed for digesting kidneys to obtain single cells of defined segmental origin for cell culture, e.g.…”

“…Complex organs such as the kidney, with a variety of functionally and morphologically different compartments and cell types, impose additional difficulties. Numerous techniques, such as whole isolated kidney perfusion [25], in situ microperfusion of single nephron segments in the intact kidney [39], biochemical and transport studies in tubule suspensions [15,45,49] or isolated membrane vesicles [29,35], isolation of kidney tubules [14,15,31,36,38], kidney slices [2,3,5] and preparations of cell cultures [4,21,23,41,44] have been developed and used to investigate kidney organ function as well as segment-or cell-specific functions. The preparation of isolated kidney tubule fragments allowed for the first time access to all nephron segments and the controlled examination of transport processes.…”

The fate of oxalic acid in the Wistar rat

“…Isolated rat kidney tubules were prepared according to the method of Dawson (1972) in phosphatebuffered saline at pH7.4 (Dawson 1977). Aliquots (1 ml) of this cell suspension, containing 4-15mg protein/ml, were incubated at 37'C in conventional Warburg flasks containing either ~-[ U -~~C ] g l u c o s e (1 mni) or ~-[U-'~C]lactate ( 2 m n ) with and without oxalic acid (1 mhl).…”

Gluconeogenesis in the Newborn Rat: the Substrates and their Quantitative Significance

“…However, it is the mobilization of tissue glycogen stores, built up in times of plentiful carbohy drate supply, that leads to glucose production and lactate formation; the latter can then serve as a gluconeogenic substrate in times of carbohydrate deprivation. Lactate is an efficient precursor in vitro giving rates of glucose synthesis compa rable to pyruvate in rat liver and kidney slices (104,136), in perfused liver and kidney (44,143,161), in isolated liver cells (85,91), and in isolated kidney tubule preparations (32,62,63,136). Hepatic gluconeogenesis from lactate in vivo occurs at a high rate and increases proportionally with the circulating lac tate concentration (2), as is observed also in the perfused liver (44) and in isolated liver cells (85).…”