PI-PLCβ is involved in the modulation of the proximal tubule Na+-ATPase by angiotensin II

Abstract In previous papers we showed that Ang II increases the proximal tubule Na + -ATPase activity through AT 1 /PKC pathway [L.B. Rangel, C. Caruso-Neves, L.S. Lara, A.G. Lopes, Angiotensin II stimulates renal proximal tubule Na + -ATPase activity through the activation of protein kinase C. Biochim. Biophys. Acta 1564 (2002) 310–316, L.B.A. Rangel, A.G. Lopes, L.S. Lara, C. Caruso-Neves, Angiotensin II stimulates renal proximal tubule Na + -ATPase activity through the activation of protein kinase C. Biochim. Biophys. Acta 1564 (2002) 310–316]. In the present paper, we study the involvement of PI-PLCβ on the stimulatory effect of angiotensin II (Ang II) on the proximal tubule Na + -ATPase activity. Western blotting assays, using a polyclonal antibody for PI-PLCβ, show a single band of about 150 KDa, which correspond to PI-PLCβ isoforms. Ang II induces a rapid decrease in PIP 2 levels, a PI-PLCβ substrate, being the maximal effect observed after 30 s incubation. This effect of Ang II is completely abolished by 5×10 −8 M U73122, a specific inhibitor of PI-PLCβ. In this way, the effect of 10 −8 M Ang II on the proximal tubule basolateral membrane (BLM) Na + -ATPase activity is completely abolished by 5×10 −8 M U73122. The increase in diacylglycerol (DAG) concentration, an product of PI-PLCβ, from 0.1 to 10 nM raises the Na + -ATPase activity from 6.1±0.2 to 13.1±1.8 nmol Pi mg −1 min −1 . This effect is similar and non-additive to that observed with Ang II. Furthermore, the stimulatory effect of 10 nM DAG is completely reversed by 10 −8 M calphostin C (Calph C), an inhibitor of PKC. Taken together these data indicate that Ang II stimulates the Na + -ATPase activity of proximal tubule BLM through a PI-PLCβ/PKC pathway.