A Signaling Pathway for Stimulation of Na⁺ Uptake Induced by Angiotensin II in Primary Cultured Rabbit Renal Proximal Tubule Cells

Abstract

The aim of the present study was to examine the signaling pathways for a low dose of angiotensin II (ANG II) on Na⁺ uptake of primary cultured rabbit renal proximal tubule cells (PTCs) in hormonally defined serum-free medium. The results were as follows; ANG II (10^-11 M) stimulated the proliferation of PTCs. 10^-11 M ANG II stimulated Na⁺ uptake by 20%, whereas 10^-9 M ANG II inhibited it by 20% (p<0.05). The stimulatory effect of 10^-11 M ANG II on Na⁺ uptake was inhibited by amiloride (10^-3 M) and by losartan (ANG II receptor subtype 1 antagonist, 10^-8 M) but not by PD123319 (ANG II receptor subtype 2 antagonist, 10^-8 M). Pertussis toxin (PTX, 50 ng/ml) prevented the ANG II-induced stimulation of Na⁺ uptake (p<0.01). 8-Bromoadenosine 3', 5'-cyclic monophosphate (8-Br-cAMP, 10^-6 M) did not affect Na⁺ uptake. SQ 22536 (adenylate cyclase inhibitor, 10^-6 M) also did not change the ANG II-induced stimulation of Na⁺ uptake. ANG II did not stimulate cAMP production. In contrast, 12-O-tetradecanoylphorbol-13-acetate (TPA, 0.01 ng/ml) produced significant increase in Na⁺ uptake. When ANG II and TPA were added together to the PTCs, there was no additive effect on Na⁺ uptake. Staurosporine (calcium-dependant protein kinase C inhibitor, 10^-6 M) led to a complete inhibition of ANG II-induced stimulation of Na⁺ uptake. ANG II-treatment resulted in a 26% increase in total protein kinase C (PKC) activity. However, 10^-11 M ANG II did not change [Ca²⁺]_i mobilization and [³H]-AA release while 10^-9 M ANG II increased both of them. In conclusion, the PTX-sensitive PKC pathway may be the main signaling cascade in the stimulatory effects of low dose of ANG II (10^-11 M) on Na⁺ uptake in the primary cultured rabbit renal proximal tubule cells in hormonally defined serum-free medium.