Lithium and protein kinase C modulators regulate swelling-activated K-Cl cotransport and reveal a complete phosphatidylinositol cycle in low K sheep erythrocytes

• Published in: The Journal of membrane biology Vol. 177; no. 1; pp. 81 - 93
• Main Authors: Ferrell, C M, Lauf, P K, Wilson, B A, Adragna, N C
• Format: Journal Article
• Language: English
• Published: United States 01.09.2000
• Subjects: Animals; Carrier Proteins - metabolism; Cell Size; Erythrocytes - metabolism; Hydrogen-Ion Concentration; K Cl- Cotransporters; Lithium - metabolism; Phosphatidylinositols - metabolism; Phospholipids - metabolism; Potassium - metabolism; Protein Kinase C - metabolism; Rubidium - metabolism; Sheep; Signal Transduction; Symporters
• ISSN: 0022-2631; 1432-1424
• DOI: 10.1007/s002320001101

K-Cl cotransport (COT), a ouabain-insensitive, Cl-dependent bidirectional K flux, is ubiquitously present in all cells, plays a major role in ion and volume homeostasis, and is activated by cell swelling and a variety of chemical interventions. Lithium modulates several cation transport pathways and inhibits phospholipid turnover in red blood cells (RBCs). Lithium also inhibits K-Cl COT by an unknown mechanism. To test the hypothesis whereby Li inhibits swelling-activated K-Cl COT by altering either its osmotic response, its regulation, or by competing with K for binding sites, low K (LK) sheep (S) RBCs were loaded with Li by Na/Li exchange or the cation ionophore nystatin. K-Cl COT was measured as the Cl-dependent, ouabain-insensitive K efflux or Rb influx. The results show that Li altered the cell morphology, and increased both cell volume and diameter. Internal (Li(i)) but not external (Li(o)) Li inhibited swelling-activated K-Cl COT by 85% with an apparent K(i) of approximately 7 mm. In Cl, Li(i) decreased K efflux at relative cell volumes between 0.9 and 1.2, and at external pHs between 7.2 and 7.4. Li(i) reduced the V(max) and increased the K(m) for K efflux in Cl. Furthermore, Li(i) increased the production of diacylglycerol in a bimodal fashion, without significant effects on the phosphatidylinositol concentration, and revealed the presence of a complete PI cycle in LK SRBCs. Finally, phorbol ester treatment and PD89059, an inhibitor of mitogen-activated protein kinase (ERK2) kinase, caused a time-dependent inhibition of K-Cl COT. Hence, Li(i) appears to inhibit K-Cl COT by acting at an allosteric site on the transporter or its putative regulators, and by modulation of the cellular phospholipid metabolism and a PKC-dependent regulatory pathway, causes an altered response of K-Cl COT to pH and volume.