| 16875859 |
Ivanova TI, Agalakova NI, Gusev GP: Activation of sodium transport in rat erythrocytes by inhibition of protein phosphatases 1 and 2A. Comp Biochem Physiol B Biochem Mol Biol. 2006 Sep;145(1):60-7. Epub 2006 Jun 21.
Four structurally different protein phosphatases (PPs) inhibitors - fluoride, calyculin A, okadaic acid and cantharidin--were tested for their ability to modulate unidirectional Na (+) influx in rat red blood cells. Erythrocytes were incubated at 37 degrees C in isotonic and hypertonic media containing 1 mM ouabain and (22) Na in the absence or presence of PP inhibitors. Exposure of the cells to 20 mM fluoride or 50 nM calyculin A for 1 h under isosmotic conditions caused a significant stimulation of Na (+) influx, whereas addition of 200 microM cantharidin or 100 nM okadaic acid had no effect. After 2 h of treatment, however, all these PPs blockers significantly enhanced Na (+) transport in rat erythrocytes. Selective inhibitors of PP-1 and PP-2A types, calyculin A, cantharidin and okadaic acid, produced similar ( approximately 1.2-1.4-fold) stimulatory effects on Na (+) influx in the cells. Activation of Na (+) influx was unchanged with increasing calyculin A concentration from 50 to 200 nM. No additive stimulation of Na (+) influx was observed when the cells were treated with combination of 20 mM fluoride and 50 nM calyculin A. Na (+) influx induced by PPs blockers was inhibited by 1 mM amiloride and 200 muM bumetanide approximately in the equal extent, indicating the involvement of Na (+)/H (+) exchange and Na-K-2Cl cotransport in sodium transport through rat erythrocytes membrane. Activation of Na (+) transport in the cells induced by calyculin A and fluoride was associated with increase of intracellular Na (+) content. Shrinkage of the rat erythrocytes resulted in 2-fold activation of Na (+) influx. All tested PPs inhibitors additionally activated the Na (+) influx by 70-100% above basal shrinkage-induced level. Amiloride and bumetanide have diminished both the shrinkage-induced and PPs-inhibitors-induced Na (+) influxes. Thus, our observations clearly indicate that activities of Na (+)/H (+) exchanger and Na-K-2Cl cotransporter in rat erythrocytes are regulated by protein phosphatases and stimulated when protein dephosphorylation is inhibited. |
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