NORADRENALINE RELEASE FROM RAT SYMPATHETIC NEURONS TRIGGERED BY ACTIVATION OF B-2 BRADYKININ RECEPTORS

1. The role of bradykinin receptors in the regulation of sympathetic t ransmitter release was investigated in primary cultures of neurones di ssociated from superior cervical ganglia of neonatal rats. These cultu res were loaded with [H-3]-noradrenaline and the outflow of radioactiv ity was determined under continuous superfusion. 2. Bradykinin (100 nm ol l(-1) applied for 10 min) caused a transient increase in tritium ou tflow that reached a peak within four minutes after the beginning of t he application and then declined towards the baseline, despite the con tinuing presence of the peptide. ATP (100 mu mol l(-1)) and nicotine ( 10 mu mol l(-1)) caused elevations in H-3 outflow with similar kinetic s, whereas outflow remained elevated during a 10 min period of electri cal held stimulation (0.5 ms, 50 mA, 50 V cm(-1), 1.0 Hz). 3. When bra dykinin was applied for periods of 2 min, the evoked H-3 overflow was half-maximal at 12 nmol l(-1) and reached a maximum of 2.3% of cellula r radioactivity. The preferential B-1 receptor agonist des-Arg(9)-brad ykinin failed to alter H-3 outflow. The B-2 receptor antagonists, [D-P he(7)]-bradykinin (1 mu mol l(-1)) and Hoe 140 (10 nmol l(-1)), per se did not alter H-3 outflow, but shifted the concentration-response cur ve for bradykinin-evoked H-3 overflow to the right by a factor of 7.9 and 4.3, respectively. 4. Bradykinin-induced overflow was abolished in the absence of extracellular Ca2+ and in the presence of either 1 mu mol l(-1) tetrodotoxin or 300 mu mol l(-1) Cd2+, as was electrically-i nduced overflow. Activation of alpha(2)-adrenoceptors by 1 mu mol l(-1 ) UK 14,304 reduced both bradykinin- and electrically-triggered overfl ow. The Ca2+-ATPase inhibitor thapsigargin (0.3 mu mol l(-1)) failed t o alter either type of stimulated overflow. Caffeine (10 mmol l(-1)) e nhanced bradykinin-induced overflow, but reduced overflow triggered by electrical field stimulation. 5. Inclusion of Ba2+ (0.1 to 1 mmol l(- 1)) in the superfusion medium enhanced electrically induced overflow b y approximately 100% and potentiated bradykinin-triggered overflow by almost 400%. Application of 1 mmol l(-1) Ba2+ for periods of 2 min tri ggered H-3 overflow, and this overflow was abolished by 1 mu mol l(-1) tetrodotoxin and enhanced by 10 mmol l(-1) caffeine. In contrast, inc lusion of tetraethylammonium (0.1 to 1 mmol l(-1)) in the superfusion buffer caused similar increases of bradykinin-and electrically evoked H-3 overflow (by about 100%), and tetraethylammonium, when applied for 2 min, failed to alter H-3 outflow. 6. Treatment of cultures with 100 ng ml(-1) pertussis toxin caused a significant increase in bradykinin -, but not in electrically-, evoked tritium overflow. Treatment with 1 00 ng ml(-1) cholera toxin reduced both types of stimulated H-3 overfl ow. 7. These data reveal bradykinin as a potent stimulant of action po tential-mediated and Ca2+-dependent transmitter release from rat sympa thetic neurones in primary cell culture. This neurosecretory effect of bradykinin involves activation of B-2-receptors, presumably linked to pertussis- and cholera toxin-insensitive G proteins, most likely memb ers of the Gq family. Results obtained with inhibitors of muscarinic K + (K-M) channels, like caffeine and Ba2+, indicate that the secretagog ue action of bradykinin probably involves inhibition of these K+ chann els.