The long term objective of the proposed research is to elucidate both intra and extracellular mechanisms involved in the regulation of active and passive transport properties of the apical and basolateral membranes of ion transporting epithelia. The methods to be used includes conventional and ion specific microelectrodes, capacitance measurements, fluctuation analysis and """"""""patch"""""""" clamping. The data available from these methods allows a determination of apical and basolateral membrane potentials, conductances, areas, area changes, single ionic channel properties and channel density, as well as intracellular ion activities. Using of all of these methods to study ion transport regulation will allow one to differentiate between activation, insertion and modulation of ion transport pathways. The first specific aim is to determine the mechanism and ionic requirements by which known urinary proteases (urokinase, kallikrein, and plasmin) decrease transepithelial Na+ transport, i.e. a decrease in single channel currents or hydrolysis of single channels. In addition, intermediate forms of the Na+ channel will be determined and a kinetic scheme for channel hydrolysis developed. The second specific aim is to determine why reduced plasma Cl- causes a reduction in the basolateral membrane potassium conductance of rabbit urinary bladder epithelium. Both capacitance measurements and patch clamp studies will determine the voltage sensitivity and hormonal and ionic regulations to channel activity. This will allow a determination of channel regulation by insertion, activation or modulation. The last specific aim is to determine some of the factors responsible for the differentiation of primary cultures of the mammalian urinary bladder epithelium and the kidney tissue culture line MDCK. The physiological role of the urinary protease kallikrein, although unknown has been speculated to be important in renal water and electrolyte balance. Since preliminary studies from this laboratory suggest that this enzyme hydrolyses Na+ channels, and others have demonstrated that patients with essential hypertension secrete reduced levels of kallikrein compared to normotensives, this study will determine whether the decrease in kallikrein secretion will cause a significant increase in the reabsorptive capacity of renal tissues in hypertensive patients compared to normotensives.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
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Physiology Study Section (PHY)
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University of Texas Medical Br Galveston
Schools of Medicine
United States
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Lewis, S A; Clausen, C; Wills, N K (1991) Transport-related modulation of the membrane properties of toad urinary bladder epithelium. Biochim Biophys Acta 1070:99-110
Jungwirth, A; Lewis, S; Lang, F (1991) Kallikrein does not modify the transepithelial potential of rat renal distal convoluted tubules. Nephron 58:225-8
Lewis, S A; Clausen, C (1991) Urinary proteases degrade epithelial sodium channels. J Membr Biol 122:77-88
Jovov, B; Wills, N K; Lewis, S A (1991) A spectroscopic method for assessing confluence of epithelial cell cultures. Am J Physiol 261:C1196-203
Jovov, B; Wills, N K; Donaldson, P J et al. (1990) Vectorial secretion of a kallikrein-like enzyme by cultured renal cells. I. General properties. Am J Physiol 259:C869-82
Donaldson, P J; Lewis, S A (1990) Effect of hyperosmotic challenge on basolateral membrane potential in rabbit urinary bladder. Am J Physiol 258:C248-57
Donaldson, P J; Chen, L K; Lewis, S A (1989) Effects of serosal anion composition on the permeability properties of rabbit urinary bladder. Am J Physiol 256:F1125-34
Zweifach, A; Lewis, S A (1988) Characterization of a partially degraded Na+ channel from urinary tract epithelium. J Membr Biol 101:49-56