ROS mediates regulation of intracellular Ca2 +induced by angiotensin II in primarily cultured medullary neurons / 中国病理生理杂志

ABSTRACT

AIM:To investigate the role of reactive oxygen species (ROS) in the regulation of intracellular Ca2+induced by angiotensin II ( Ang II) in the primarily cultured medullary neurons .METHODS:Primarily cultured me-dullary neurons were prepared from 14-day-old embryos of Sprague-Dawley rats in the study .The identification of medullary neurons was assessed by double-labeling immunofluorescence .To explore the role of ROS , mainly the superoxide ( O2 ·-) , the O2 ·-generation was measured using the fluorogenic probe dihydroethidium ( DHE) .To determine intracellular free cal-cium concentration ( [ Ca2+] i ) , the neurons were loaded with the Ca 2+-specific dye Fura-2/AM.The cell viability after adding Ang II was also examined using CCK-8 assay.RESULTS:Most of the cultured cells were medullary neurons , more than 80%of which were glutamate positive neurons .Ang II (5 μmol/L) increased the level of ROS within 10 min in the medullary neurons .Ang II at 5μmol/L induced a significant [ Ca2+] i increase in the medullary neurons , and the effect of Ang II occurred rapidly and reached a peak within 20 min after administration.The level of [Ca2+]i started to decline after washout .The Ca2+elevation induced by Ang II was significantly decreased by apocynin or TEMPOL .No significant differ-ence in the cell viability between control group and 5μmol/L Ang II treatment group was observed .CONCLUSION:ROS is involved in the regulation of [Ca2+]i induced by Ang II in the primarily cultured medullary neurons , suggesting a poten-tial intracellular signaling mechanism involved in the Ang II-mediated oxidant regulation of central neural control of blood pressure.