Since no modification in protein expression was observed, this result indicates that a post-translational modification is responsible for the inhibition of the activity of both ATPases. Modulation of different protein kinases in Na+ pumps has been studied extensively (Cabral et al., 2010; Ramia and Kreydiyyeh, 2010). The question arises whether there is modulation of Na+ pump activity by kinase-mediated regulatory phosphorylation. To investigate this question, the identification Regorafenib mw of

PKA and PKC, known modulators of renal Na+ ATPases, was performed as well the kinase activity. Unexpectedly both kinase activity and expression of PKA and PKC proteins were unaltered in rats exposed to MCYST-LR compared with the CTRL group Natural Product Library (data not shown). This result could be explained by the known role of MCYST as a protein phosphatase inhibitor (MacKintosh et al., 1990; Runnegar et al., 1995). These results suggest that the ATPases from MCYST-LR exposed rats have a higher regulatory phosphorylation status, due to a sustained kinase-mediated phosphorylation caused by inhibition of phosphatases. Since Na+ handling is a key factor in blood pressure control, the decrease in Na+ reabsorption and the increase in Na+ clearance (Table 1) could be related to the decreased arterial pressure observed

by Theiss et al. (1988). The equilibrium of phosphatases and kinases plays an essential role in cellular metabolism and any modification to maintain Na+ cellular homeostasis could be responsible for Sitaxentan renal cellular injury and malfunction of the kidney. In the present work we have demonstrated that a sublethal dose of MCYST-LR is capable of altering the structure and function of the kidney in Wistar rats within 24 h of exposure to MCYST-LR. These alterations involve different parameters, such as increased formation of ROS, expansion of the interstitial space probably filled

with collagen deposition in both the cortex and medulla of the kidney. These modifications have a relevant role in the function of the organ, which was observed by the increased GFR and imbalance of Na+ handling, caused by the inhibition of both Na+ pumps. This inhibition is a result of a sustained kinase-mediated regulatory phosphorylation status of the ATPases, caused by the well-described role of MCYST as a phosphatase inhibitor. These findings confirm the importance of understanding the mechanisms of MCYST at lower doses, which could be more severe with chronic exposure. This study received financial support from: The Brazilian Council for Scientific and Technological Development (CNPq), The Foundation for the Coordination of Higher Education and Graduate Training (CAPES), The Carlos Chagas Filho Rio de Janeiro State Research Supporting Foundation (FAPERJ, Pensa Rio), The Macaé Educational Foundation (FUNEMAC), The National Institute for Science and Technology in Structural Biology and Bioimage (INCT-INBEB).