ION CONTENT AND CELL-VOLUME IN ISOLATED COLLECTING DUCT CELLS - EFFECT OF HYPOTONICITY

On isolated inner medullary collecting duct (IMCD) cells of the rat ki dney the capability of osmoregulatory adaptation was investigated in v itro. IMCD cells were isolated by differential centrifugation at 600 m Osm (268 mm NaCl) and subsequently exposed to hypotonic buffers (300 m Osm, 118 mm NaCl). The alterations of ion content and cell volume foll owing this change in extracellular osmolarity were studied by electron probe microanalysis and determination of intracellular water. After s welling within 40 seconds to 152 +/- 15% of control (P < 0.001; N = 9) cell volume was restored after 15 minutes. This regulatory volume dec rease (RVD) was observed irrespective whether extracellular osmolarity was changed by using NaCl or mannitol as the major osmolyte. During R VD the cells lost sodium (48 +/- 11%) and chloride (14 +/- 5%), and th e potassium content remained nearly unchanged. Correspondingly, sodium and chloride concentrations were progressively lowered, whereas the p otassium concentration changed only transiently. RVD was diminished by 10(-4) m NPPB, 10(-3) m SITS and in the absence of HCO3-. Twenty mill imoles of ouabain or 5 mm barium also inhibited RVD with little additi ve effect. A total of 10(-3) m amiloride and 10(-4) m bumetanide showe d no effect on the hypoosmotic volume response. The experiments show t hat in isolated IMCD cells exposed to hypotonic conditions, rapid reve rsible changes in cell volume and sustained alterations in cell inorga nic ion content occur, and thereby transmembrane sodium and potassium gradients are maintained. Since the loss in inorganic electrolytes doe s not account for RVD, the major part of volume regulation seems to oc cur via changes in organic osmolytes. Fluxes of inorganic osmolytes th rough a variety of pathways and the alterations in organic osmolyte co ntent therefore must be closely coordinated.