RT Journal Article
SR Electronic
T1 Allosteric Interactions of Staurosporine and Other Indolocarbazoles with<em>N</em>-[<em>methyl</em>-<sup>3</sup>H]Scopolamine and Acetylcholine at Muscarinic Receptor Subtypes: Identification of a Second Allosteric Site
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP 194
OP 207
DO 10.1124/mol.58.1.194
VO 58
IS 1
A1 Sebastian Lazareno
A1 Angela Popham
A1 Nigel J. M. Birdsall
YR 2000
UL http://molpharm.aspetjournals.org/content/58/1/194.abstract
AB We have studied the interactions of five indolocarbazoles withN-[methyl-3H]scopolamine (NMS) and unlabeled acetylcholine at M1-M4 muscarinic receptors, using equilibrium and nonequilibrium radioligand binding studies. The results are consistent with an allosteric model in which the primary and allosteric ligands bind simultaneously to the receptor and modify each other's affinities. The compounds were generally most active at M1 receptors. [3H]NMS binding was enhanced by staurosporine, KT5720, and KT5823 at M1 and M2 receptors, and by K-252a at M1 receptors. Gö 7874 reduced [3H]NMS affinity by up to threefold for all subtypes. A range of cooperative effects with acetylcholine was seen, and, at the M1 receptor, KT5720 had a log affinity of 6.4 and enhanced acetylcholine affinity by 40%. The compounds inhibited the dissociation of [3H]NMS to different extents across the receptor subtypes, with the largest effects at M1 receptors. In equilibrium binding studies the inhibitory potency of gallamine at M1 receptors was not affected by KT5720, indicating that these agents bind to two distinct allosteric sites and have neutral cooperativity with each other. In contrast, gallamine and staurosporine had a negatively cooperative or competitive interaction at M1 receptors. Similarly, the potency and relative effectiveness of KT5720 for inhibiting [3H]NMS dissociation from M1 receptors were not affected by gallamine or brucine, but were affected in a complex manner by staurosporine. These results demonstrate that there are at least two distinct allosteric sites on the M1 receptor, both of which can support positive cooperativity with acetylcholine.