FUNCTIONAL EXPRESSION OF CAMP-DEPENDENT AND INDEPENDENT UREA TRANSPORTERS IN XENOPUS-OOCYTES

Facilitated transport of urea by the inner medullary collecting duct in kidney is important for the urinary concentrating mechanism. To examine the nature and tissue distribution of urea transporters, mRNA was isolated from different tissues and expressed in Xenopus oocytes. [C-14]urea and [H-3]methylglucose uptake were measured at 21-degrees-C at 64 h after microinjection of mRNA. Relative urea uptake in oocytes injected with 50 ng of unfractionated mRNA was (n = 6-42): 1.0 (water-injected control), 1.0 +/- 0.3 (human kidney cortex), 2.9 +/-0.5 (rat kidney papilla), 2.5 +/- 0.5 (human kidney papilla), 2.7 +/-0.3 (rat liver), 1.1 +/- 0.3 (rat brain), 1.2 +/- 0.3 (rat muscle), and 2.6 +/- 0.3 (rabbit reticulocyte). Urea uptake was inhibited to near control values by 0.2 mM phloretin and 0.2 mM p-chloromercuribenzenesulfonate (pCMBS) in oocytes injected with mRNA from kidney medulla, liver, and reticulocyte; phloretin and pCMBS had no effect in control oocytes and oocytes injected with mRNA from kidney cortex, brain, and muscle. Urea uptake was strongly increased in oocytes injected with kidney medulla mRNA (4.4-fold over control) by a 5-min preincubation with the adenosine 3',5'-cyclic monophosphate (cAMP) agonist adenosine-3',5'-cyclic monophosphorothioate (Sp-cAMPS) or a mixture of CPT-cAMP, forskolin, and 3-isobutyl-1-methylxanthine; cAMP agonists did not affect urea uptake in oocytes expressing the reticulocyte and liver urea transporters. As an internal control, (phloretin inhibitable) glucose uptake was enhanced in all oocytes (up to 5-fold greater than control), and was not affected by pCMBS and the cAMP agonists. Size-fractionated mRNA from human kidney papilla gave a 2- to 3-kb size for the cAMP-stimulated urea transporter. These findings 1) suggest that the existence of a facilitated urea transporting pathway in liver, 2) suggest that the cAMP-dependent urea transporter in renal medulla is different from the cAMP-independent urea transporters in erythrocyte or liver, and 3) require that cAMP stimulation of the kidney type urea transporter does not involves vesicular trafficking.