REGULATION OF Na+-K+-2Cl--COTRANSPORTER ACTIVITY IN RAT SKELETAL MUSCLE AND INTESTINAL
EPITHELIAL CELLS
A. R. Gosmanov, D. B. Thomason
Department of Physiology, College of Medicine University of Tennessee Health Science Center, Memphis, USA;
e-mail: gosmanov@physiol.utmem.edu
In mammalian cells, Na+-K+-2Cl--cotransporter activity participates in regulation of ion and volume homeostasis.
This makes NKCC indispensable for normal cell growth and proliferation. We recently reported the existence of two mechanisms that can regulate NKCC activity in mature
skeletal muscle. In isosmotic conditions, signaling through ERK ÌÀÐÊ pathway is necessary, while inhibition of the cAMP-dependent protein kinase A (PKA) pathway stimulates
NKCC activity during hyperosmotic challenge. Both pathways are involved in regulating cell proliferation in wide variety of cells of epithelial and non-epithelial origin, so we tested
which pathway regulated NKCC activity in cultured cells. In cultured rat skeletal muscle (L6) and intestinal epithelial (IEC-6) cells, NKCC activity in the basal state comprised
30 to 50 % of total potassium influx, assessed as bumetanide-sensitive 86Rb-uptake. This NKCC activity could not be abolished by inhibi-tors of ERK ÌÀÐÊ
(PD98059 and U0126), PKC (GF109203X), or PI 3-K (wortmannin, LY294002). In L6 myoblasts and in IEC-6 cells, elevation of cAMP levels with isoproterenol or forskolin led to a
60 % inhibi-tion on NKCC activity. In contrast, incubation of IEC-6 cells with the ÐÊÀ-inhibitor H-89 resulted in 50 % increase of NKCC activity compared with the basal level. In
conclusion, it appears that in cultured cells the cAMP-PKA pathway regulates NKCC activity. This resembles hyperosmotic regulation of NKCC activity.
Key words: Na+-K+-2Cl--cotransporter, bumetanide, L6 cells, Extracelluar Signal-regulated Kinase Mitogen Activated
Protein Kinase, isoproterenol, forskolin, cAMP, Protein Kinase A
