AJP - Cell Physiology, Vol 262, Issue 2 C436-C444, Copyright © 1992 by American Physiological Society

ARTICLES

Regulation of vascular endothelial cell volume by Na-K-2Cl cotransport

W. C. O'Neill and J. D. Klein
Department of Medicine, Emory University School of Medicine, Atlanta, Georgia 30322.

The relationship between cell volume and Na-K-2Cl cotransport was studied in cultured bovine aortic endothelial cells. Hypertonic cell shrinkage increased bumetanide-sensitive, Na- or Cl-dependent K influx without altering bumetanide-insensitive influx. Greater stimulation of cotransport was observed in cells shrunken isosmotically either by preincubation in K-free and Na-free medium or by preincubation in hypotonic medium. Cell swelling, produced by preincubation in isotonic high-K medium, inhibited bumetanide-sensitive K influx. Simultaneous measurements of [3H]bumetanide binding and K influx revealed an increased number of binding sites without an increased influx per binding site in shrunken cells. Bumetanide did not alter the volume or ion content of cells in isotonic or hypertonic medium, indicating that no net influx of ions occurs through cotransport under these conditions. In isosmotically shrunken cells, there was greater stimulation of bumetanide-sensitive influx than of bumetanide-sensitive efflux, resulting in net bumetanide-sensitive influx. Rapid recovery of cell K, Na, and water occurred over 10-20 min and was inhibited by bumetanide or by the removal of external Na or Cl. These data demonstrate that Na-K-2Cl cotransport in aortic endothelial cells is regulated by cell volume, possibly through changes in the number of functional cotransporters, and mediates a brisk regulatory volume increase in isosmotically shrunken cells. Although thermodynamically favored, no net influx occurs through Na-K-2Cl cotransport in cells of normal volume or in hypertonically shrunken cells. This suggests additional regulation of cotransport, perhaps through trans-inhibition by intracellular Cl. Regulation of cell volume by Na-K-2Cl cotransport may be important in maintaining endothelial integrity.

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