The Feldberg Lecture 1976. Solute transport across epithelia: what can we learn from micropuncture studies in kidney tubules?

Epithelial transport covers an enormously wide field of research on tissues such as skin, intestine, salivary or sweat glands and kidney tubules which, on first view, seem to have little in common. However, despite the vast number of transport functions which these tissues perform, it appears that all operate on a relatively small number of general principles and it is my intention to describe some of those principles which we can discern. I will do so not by screening the literature for comparative aspects but by focussing mainly on one single epithelium, the rat kidney proximal tubule, and probing further and further into its properties. Our interest in this epithelium was twofold: (1) we knew that the proximal tubule plays a paramount role in the absorption of the glomerular filtrate and hence in the maintenance of the water and electrolyte balance of the body in man the proximal tubules absorb approximately 140 1. of tubular urine per day and (2) we have found that, with respect to its transport functions, renal proximal tubule may serve as an ideal model tissue for a group of epithelia (comprising among others, small intestine, gall-bladder, and choroid plexus), as well as possibly some endothelia, to which the well known frog skin model of transepithelial transport cannot be applied. These epithelia we have classified (Fromter & Diamond, 1972) as 'leaky epithelia' in contrast to the frog skin type 'tight epithelia' which have different transport properties and serve different functions in the body. I will come back to the distinction between tight and leaky epithelia below. A considerable disadvantage of the kidney tubules in transport studies is their small size. Rat proximal tubule has an outer diameter of 45 jtm and a lumen diameter of only 20 ,um (compare Fig. 1). The wall is formed of one layer of uniform cuboidal cells, with nuclei, vacuoles and a dense packing of mitochondria. The luminal cell membrane surface (brush border) and the basal cell membrane surface (basal labyrinth) are greatly amplified by microvilli or basal infoldings respectively. The gaps between neighbouring cells (lateral spaces) are closed near the luminal end by terminal bars (so-called tight junctions; see Fig. 9 below). To study solute and water transport across such tiny structures as renal tubules requires appropriate micropuncture and microanalytical techniques. Such techniques were initially developed between 1920 and 1930 for work with the bigger tubules of frog and Necturus kidney (Richards, 1929) and since then have been more and mor

A simple method for constructing shielded, low-capacitance glass microelectrodes

A new simple method is presented to produce shielded low-capacitance microelectrodes. A metal-shield is vapour-deposited on the inner surface of a glass pipette which is slid over the microelectrode proper and insulated at the tip by dipping in polystyrene. The unshielded protruding tip can be as small as 10 micrometers. A special advantage is the low capacitance between electrode and shield of approximately 0.16 microF/cm shield length

Introduction to electrophysiology and epithelial transport: the use of fast concentration step experiments in the electrical analysis of tubular transport

Publisher Summary: This chapter discusses that the classical electrophysiological technique is a powerful tool in the hands of renal physiologists. The chapter describes the specific advantages of electrophysiological approach. (1) It directly determines the membrane potential, which is one of the conjugate driving forces for ion flow. (2) It can pass current across the membrane, which is not possible in vesicle preparations. (3) One can use ion selective electrodes to measure ion activities and to follow the time course of ion concentration changes in response to changes of system parameters from which net ion fluxes can be calculated. It also explains that electrophysiological techniques offer a unique tool, the microelectrode, which enables to monitor transport events of single cells under their normal living conditions and in their normal environment

Progress in microelectrode techniques for kidney tubules

Although microelectrode measurements have caused a good deal of confusion in the investigation of renal electrolyte transport (for literature see Ref.4), we cannot do without them. Knowledge of the electrical potential steps, which the ions have to pass during their reabsorption, is indispensable for understanding the transport mechanisms involved. Hence the development of more reliable methods to measure the electrical properties of kidney tubules was an important as well as challenging problem. This paper deals with some of the advances effected in our laboratory in the past 5 years. The properties of microelectrodes that present the major problems are: 1. The invisibility of the electrode tip, 2. The instability of the tip potential, and 3. The instability of the tip resistance. Since a brief description of resistance measurement problems has been published elsewhere recently (14), only the first two points will be discussed here. Simple microscopic observation alone can never tell whether the invisible microelectrode tip has penetrated the invisible cell membrane. In order to decide whether the tip sticks within a cell or whether it has reached the tubular lumen a number of localization methods have been developed

Electrophysiological analysis of rat renal sugar and amino acid transport. I. Basic phenomena

The electrical events associated with the absorption ofd-glucose orl-amino acids in renal proximal tubules were studied in microperfusion experiments on rat kidneys in vivo. Intratubular application of these substrates led concomittantly to: 1) a shift of the transepithelial potential into lumen negative direction, 2) a partial depolarization of the tubular cell membranes and 3) a reduction of the electrical resistance of the brushborder membrane. By means of rapid perfusion experiments it was possible to discern two phases in the potential response to substrate perfusion, a fast initial response which reflects a substrate-induced Na+ ion current from lumen to cell, and a slower secondary response which reflects the relaxation of the intracellular ion and substrate concentrations towards new steady states. A quantitative analysis of the data yielded estimates of 1) the apical (Ra) and basal (Rb) cell membrane resistances and of the shunt resistance, Rs, of rat proximal tubule of approximately Ra=255 Ω cm2, Rb=92 Ω cm2 and Rs=5 Ω cm2 (all referred to the quasi macroscopic surface area of the tubular lumen), 2) the conductance of the Na+ and glucose cotransport pathway and 3) the driving forces acting on the cotransport mechanism in the brushborder membrane. The latter were found to be a) the electrical cell membrane potential of −74mV, b) the Na- ion concentration gradient between the tubular lumen (clumNa =145 mmol/l) and the cytoplasm (ccellNa ≈22mmol/l) which corresponds to an additional equivalent potential of 51 mV and c) the substrate concentration gradient, which varies according to the experimental conditions. Moreover the analysis provided a quantitative estimate of the relationship between the substrate-induced changes in transepithelial potential or short circuit current and the actual cotransport current in the brushborder membrane. Based on this analysis it is concluded that the stoichiometry of Na+ and glucose flux coupling in the brushborder membrane of rat proximal tubule is close to 1.0