Submitted on October 30, 2007
Accepted on April 22, 2008
The effect of Angiotensin II on intracellular pH is mediated by AT1 receptor translocation
Karina Thieme1, Debora Mai Nakamura Eguti2, Margarida Mello-Aires3, and Maria Oliveira-Souza2*
1 Physiolgy and Biophysics, Instituto de Ciencias Biomedicas, University of Sao Paulo, Sao Paulo, Brazil
2 Physiology and Biophysics, Instituto de Ciencias Biomedicas, University of Sao Paulo, Sao Paulo, Brazil
3 Physiology and Biophysics, Instituto de Ciencias Biomedicas, Brazil
* To whom correspondence should be addressed. E-mail: souza{at}icb.usp.br.
The effect of ANG II on pHi recovery rate and AT1 receptor translocation were investigated in transfected MDCK cells. The pHi recovery rate was evaluated by fluorescence microscopy using the fluorescent probe BCECF/AM. The hAT1 receptor translocation was analyzed by immunofluorescence and confocal microscope. Our data show that, transfected cells, in control situation, have a pHi recovery rate of [0.219 ± 0.017 pH units/min (n = 11)]. This value was similar to non transfected cells [0.211 ± 0.009 pH units/min (n = 12)]. Both values were significantly increased with ANG II (10-9 M) but not with ANG II (10-6 M). Losartan (10-7 M) and Dimethyl-Bapta-AM (10-7 M) decreased significantly the stimulatory effect of ANG II (10-9 M) and induced an increase on NHE1 activity with ANG II (10-6 M). Immunofluorescence studies indicated that in control situation, the hAT1 receptor was predominantly expressed in cytosol. However, it was translocated to plasma membrane with ANG II (10-9 M) and internalized with ANG II (10-6 M). Losartan (10-7 M) induced hAT1 translocation to plasma membrane in all studied groups. Dimethyl-Bapta-AM (10-7 M) did not change the effect of ANG II (10-9 M) on the hAT1 receptor distribution, but induced its accumulation at plasma membrane in cells treated with ANG II (10-6 M). With Ionomycin (10-6 M), the receptor was accumulated in cytosol. The results indicate that, in MDCK cells, the effect of ANG II on NHE1 activity is associated to ligand binding to AT1 receptor and intracellular signaling events related to AT1 translocation.
