The luminal membrane of rat thick limb expresses AT₁ receptor and aminopeptidase activities.

Endogenous intratubular angiotensin II (Ang II) supports an autocrine tonic stimulation of NaCl absorption in the proximal tubule, and its production may be regulated independently of circulating Ang II. In addition, endogenous Ang II activity may be regulated at the brush border membrane (BBM), by the rate of aminopeptidase A and N (APA and APN) activities and the rate of Ca²⁺-independent phospholipase A₂ (PLA₂-dependent endocytosis and recycling of the complex Ang II subtype 1 (AT₁) receptor (AT₁-R). The aim of the present study was to look for subcellular localization of AT₁-R, and APA and APN activities in the medullary thick ascending limb of Henle (mTAL), as well as search for an asymmetric coupling of AT₁-R to signal transduction pathways.

Preparations of isolated basolateral membrane (BLMV) and luminal (LMV) membrane vesicles from rat mTAL were used to localize first, AT₁-R by ¹²⁵I-[Sar¹, Ile⁸] Ang II binding studies and immunoblot experiments with a specific AT₁-R antibody, and second, APA and APN activities. Microfluorometric monitoring of cytosolic Ca²⁺ with a Fura-2 probe was performed in mTAL microperfused in vitro, after apical or basolateral application of Ang II.

AT₁-R were present in both LMV and BLMV, with a similar K_d (nmol/L range) and B_max. Accordingly, BLMV and LMV preparations similarly stained specific AT₁-R antibody. APA and APN activities were selectively localized in LMV, although to a lesser extent than those measured in BBM. In the in vitro microperfused mTAL, basolateral but not apical Ang II induced a transient increase in cytosolic [Ca²⁺].

Besides the presence of basolateral AT₁-R in mTAL coupled to the classical Ca²⁺-dependent transduction pathways, AT₁-R are present in LMV, not coupled with Ca²⁺ signaling, and co-localized with APA and APN activities. Thus, apical APA and APN may play an important role in modulating endogenous Ang II activity on NaCl reabsorption in mTAL.