Subcellular mechanisms of angiotensin II and arginine vasopressin activation of area postrema neurons

Angiotensin II (ANG II) and arginine vasopressin (AVP) act on area postrema (AP) neurons to modulate the baroreflex. Because activation of AP neurons by either ANG II or AVP increases intracellular free Ca²⁺ concentrations ([Ca²⁺](i)), the goal of this study was to analyze the factors affecting the [Ca²⁺](i) responses to ANG II and AVP. Neurons were recovered from 14- to 16-day old rats and studied after 8-14 days in culture by use of the microscopic digital image analysis for fura 2-loaded cells. The effects of ANG II (100 nM) and AVP (100 nM) on [Ca²⁺](i) were determined in normal (2 mM) and low (<10 nM) extracellular Ca²⁺ concentrations. In 143 of 240 neurons, ANG II increased [Ca²⁺](i) 4.65-fold after 20 s, and a similar response was observed in the absence of extracellular Ca²⁺ (3.65-fold after 20 s). After 60 s of observation, steady-state levels of increased [Ca²⁺](i) were still present under both conditions. Pretreatment with AT₁ antagonist or pertussis toxin abolished the response to ANG II. AVP also increased [Ca²⁺](i) (3.6-fold at peak, 20 s) in normal and low extracellular Ca²⁺. Pretreatment with AVP V₁ antagonist or pertussis toxin abolished the response to AVP. This study indicates that ANG II-induced increases in [Ca²⁺](i) are independent of extracellular Ca²⁺ concentrations and involve the activation of AT₁ receptors and a pertussis toxin-sensitive G protein. Although AVP affects a fewer number of AP neurons, the mechanisms of activation are also independent of extracellular Ca²⁺ concentration and are mediated by a pertussis toxin-sensitive G protein.