Estrogens are among the most promising candidate substances to reduce the cardiovascular risk,however,their use is still controversial. It is established that E2 has a vasorelaxant effect on vascular smooth muscle, which is direct, do not necessarily needs endothelium implication, is non-genomic, involves both a and b estrogen receptors, is mediated by a calcium dependent mechanism and the opening of potassium channels.

Methods: Smooth cells isolation: thoracic aortas were removed and carefully cleaned. The endothelium was removed by gently rubbing the "intima" surface. We opened the aorta longitudinally, and with a scalpel the muscle layer was dissociated scraping longitudinally the aorta. The adventitia layer was discarded. Later, the dissociated muscle layer was submitted to enzymatic disaggregation. Finally the samples were filtered with a of 50 micron mesh. Membrane potential determination: We used a cytometric adaptation of the Sguilla[14] method. The quantitative measurement was performed in isolated aorta smooth muscle cells with the bis-oxonol sensitive probe DISBAC2(3) (Invitrogen).The cells were excited at 488 nm and the fluorescence emitted read at 525±25 nm with a flow cytometer Epics Elite(Coulter, Hielah,Fl.)equipped with a flow chamber of 70 microns diameter.

The results obtained are shown in the following table:

E2 concentration Control 10 mM 33 mM 100 mM

D Fluorescence % (mean) 0,38 -3,37 -22,10 -43,93

D Fluorescence % (S.D.) 3,25 5,41 10,61 11,45

From our result we are found a direct relation between E2 concentration and fluorescence decrease in the range of 10-100 mM This result explain the relaxing effect of E2 in this concentration but not the relaxing effect at lower concentrations.