Adenylate cyclase from Fasciola hepatica. 1. Ligand specificity of adenylate cyclase-coupled serotonin receptors
Northup, J.K.; Mansour, T.E.
Molecular Pharmacology 14(5): 804-819
1978
ISSN/ISBN: 0026-895X PMID: 714024 Document Number: 136967
Cell-free particles from the liver fluke F. hepatica contain a highly active serotonin stimulated adenylate cyclase. Serotonin (5-HT) stimulates this enzyme 25-30 fold over basal activity to activities of 0.80 .+-. 0.10 nmol/min .cntdot. mg protein. Histamine, dopamine, octopamine, epinephrine and carbachol failed to activate this adenylate cyclase. The kinetics of activation by 5-HT showed apparent negative cooperativity with a Hill coefficient of 0.7. The lower apparent affinity half-maximal activation by 5-HT occurred at 2.1 .+-. 0.3 .mu.M. Any substitutions on the 5-HT molecule reduced apparent affinity and degree of activation. Apparent affinity and intrinsic activity of indoleamines decreased with decreasing structural similarity to 5-HT. All indoleamines tested compete for the serotonin receptor. Derivatives of lysergic acid also activate this adenylate cyclase, with very high apparent affinity. D-LSD was the most potent derivative activating maximally about 25% of 5-HT stimulated activity. Half-maximal activation by D-LSD occurred at 40 nM. Activation by LSD was totally stereospecific, with the L-isomer inactive even at 1 mM. Both D- and L-LSD antagonized 5-HT stimulation, but the L-isomer had a 500-fold decreased affinity. 2-Bromo LSD (BOL) competitively antagonized 5-HT activation as shown by Schild analysis. BOL directly inhibited basal (nonactivated) adenylate cyclase. The direct inhibition and antagonism of 5-HT by BOL both involved a single population of receptor sites with the same inhibition constant. BOL, inhibits adenylate cyclase by interacting with the 5-HT site and BOL might have a "negative" efficacy for this receptor. A single class of adenylate cyclase regulated only by serotonin receptors was proposed.