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Nitric Oxide Regulation of Na, K‐ATPase Activity in Ocular Ciliary Epithelium Involves Src Family Kinase
Author(s) -
Shahidullah Mohammad,
Mandal Amritlal,
Wei Guojun,
Delamere Nicholas A.
Publication year - 2014
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.24454
Subject(s) - ouabain , nitric oxide , sodium nitroprusside , protein kinase a , chemistry , src family kinase , phosphorylation , cgmp dependent protein kinase , microbiology and biotechnology , kinase , proto oncogene tyrosine protein kinase src , endocrinology , medicine , biology , biochemistry , sodium , mitogen activated protein kinase kinase , organic chemistry
The nitric oxide (NO) donor sodium nitroprusside (SNP) is known to reduce aqueous humor (AH) secretion in the isolated porcine eye. Previously, SNP was found to inhibit Na,K‐ATPase activity in nonpigmented ciliary epithelium (NPE), AH‐secreting cells, through a cGMP/protein kinase G (PKG)‐mediated pathway. Here we show Src family kinase (SFK) activation in the Na,K‐ATPase activity response to SNP. Ouabain‐sensitive 86 Rb uptake was reduced by >35% in cultured NPE cells exposed to SNP (100 µM) or exogenously added cGMP (8‐Br‐cGMP) (100 µM) and the SFK inhibitor PP2 (10 µM) prevented the response. Ouabain‐sensitive ATP hydrolysis was reduced by ∼40% in samples detected in material obtained from SNP‐ and 8‐Br‐cGMP‐treated cells following homogenization, pointing to an intrinsic change of Na,K‐ATPase activity. Tyrosine‐10 phosphorylation of Na,K‐ATPase α1 subunit was detected in SNP and L ‐arginine‐treated cells and the response prevented by PP2. SNP elicited an increase in cell cGMP. Cells exposed to 8‐Br‐cGMP displayed SFK activation (phosphorylation) and inhibition of both ouabain‐sensitive 86 Rb uptake and Na,K‐ATPase activity that was prevented by PP2. SFK activation, which also occurred in SNP‐treated cells, was suppressed by inhibitors of soluble guanylate cyclase (ODQ; 10 µM) and PKG (KT5823; 1 µM). SNP and 8‐Br‐cGMP also increased phosphorylation of ERK1/2 and p38 MAPK and the response prevented by PP2. However, U0126 did not prevent SNP or 8‐Br‐cGMP‐induced inhibition of Na,K‐ATPase activity. Taken together, the results suggest that NO activates guanylate cyclase to cause a rise in cGMP and subsequent PKG‐dependent SFK activation. Inhibition of Na,K‐ATPase activity depends on SFK activation. J. Cell. Physiol. 229: 343–352, 2014. © 2013 Wiley Periodicals, Inc.