OSMOLARITY-STIMULATED UREA TRANSPORT IN RAT TERMINAL IMCD - ROLE OF INTRACELLULAR CALCIUM

We showed previously that both increasing osmolality by adding NaCl or mannitol (hyperosmolarity) or adding vasopressin can stimulate urea p ermeability in rat terminal inner medullary collecting ducts (IMCD). V asopressin acts via adenosine 3',5'-cyclic monophosphate (cAMP), but t he mechanism by which hyperosmolarity acts is unknown. To study the me chanism, we determined the effect of varying osmolality (with NaCl) on two potential second messenger systems, i.e., cAMP and intracellular calcium. There was no significant difference in cAMP production among tubules incubated at 290, 490, 690, or 890 mosmol/kg. In contrast, cAM P did increase significantly after vasopressin (10(-8) M) addition. In tracellular calcium increased significantly when osmolality was increa sed from 290 to 490 mosmol/kg in the absence of vasopressin. To examin e whether changes in intracellular calcium affect urea permeability, w e added thapsiprgin (and removed bath calcium) while maintaining osmol ality at 290 mosmol/kg. Both intracellular calcium and urea permeabili ty increased significantly. Next, we buffered intracellular calcium by pretreatment with the acetoxymethyl ester of ,2-bis(2-aminophenoxy)et hane-N,N,N',N'-tetraacetic acid (BAPTA, 50 muM). BAPTA completely bloc ked the increase in urea permeability occurring when osmolality was in creased from 290 to 490 mosmol/kg, but did not block the increase in u rea permeability occurring when vasopressin (10(-8) M) was added. In s ummary, 1) hyperosmolarity increases intracellular calcium, but has no effect on cAMP accumulation; 2) thapsigargin increases intracellular calcium and urea permeability; and 3) BAPTA blocks the hyperosmolarity -stimulated increase in urea permeability, but not vasopressin-stimula ted urea permeability. We conclude that hyperosmolarity and vasopressi n stimulate urea permeability in rat terminal IMCD via two different s econd messenger pathways, i.e., hyperosmolarity via changes in intrace llular calcium and vasopressin via cAMP.