EFFECTS OF HYPERTONICITY ON CAMP PRODUCTION IN CULTURED RENAL EPITHELIAL-CELLS (LLC-PK1)

  • 1 May 1987
    • journal article
    • research article
    • Vol. 13  (3) , 165-172
Abstract
The activaton of adenylate cyclase by vasopressin in the renal medulla takes place in a hypertonic environment whose osmolality fluctuates widely under varying physiologic conditions. We utilized the cultured renal epithelial cell line, LLC-PK1 as a model system to study the effects of hypertonic electrolyte and nonelectrolyte solutes on the vasopressin-adenylate cyclase interaction. These cells do not produce prostaglandins, thus permitting separate evaluation of the direct effects of hypertonic solutes on the adenylate cyclase response. In intact cells, 40-400 mM NaCl and 600 mM sucrose and mannitol increased basal and vasopressin-sensitive cAMP production 2 to 4-fold. Urea in a concentration range of 300-1,200 mM blunted the stimulatory effect of hypertonic NaCl in intact cells. In order to distinguish direct effects of solutes on the adenylate cyclase response, from effects related to hypertonic cell shrinkage, the influence of these same electrolyte and nonelectrolyte solutes on adenylate cyclase activity in membrane particulate fractions was also examined. Increasing NaCl in the concentration range of 25-100 mM increased basal and vasopressin-sensitive adenylate cyclase. This effect was not specific to sodium, since similar degrees of stimulation were seen with the additon of KCl. Addition of higher concentrations of NaCl, sucrose, and mannitol directly in the adenylate cyclase assay were inhibitory. These findings suggested that the stimulatory effect of hypertonicity in the intact cells was not due to a direct effect of these solutes on the enzyme, but rather to hypertonic cell shrinkage. Correspondingly, incubation of intact cells in 10 .mu.M ouabain which reverses intracellular [Na+] and [K+] without altering cell volume had no effect on basal or vasopressin-sensitive cAMP production. Addition of the permeant solute urea directly inhibited enzyme activity, corresponding to its effects in the intact cell. We conclude that hypertonicity is associated with a stimulatory effect on cAMP production. This effect is likely mediated by cell shrinkage and cannot be accounted for by altered intracellular cation composition. Urea directly inhibits the adenylate cyclase response and attenuates the effects of hypertonicity.

This publication has 12 references indexed in Scilit: