Abstract
Metabotropic glutamate receptor 1a (mGlu1a) is a G protein coupled receptor linked with phosphoinositides (PI) hydrolysis and with β-arrestin-1-mediated sustained ERK phosphorylation and cytoprotective signaling. Previously, we reported the existence of ligand bias at this receptor, since glutamate induced both effects, while quisqualate induced only PI hydrolysis. In the current study, we show that such mGlu1 agonists as glutamate, aspartate and L-cysteate were unbiased and activated both signaling pathways, while quisqualate and DHPG stimulated only PI hydrolysis. Moreover, competitive antagonists inhibited only PI hydrolysis, but not the β-arrestin-dependent pathway, while a noncompetitive mGlu1 antagonist blocked both pathways. A mutational analysis of the ligand binding domain of the mGlu1a receptor revealed that Thr188 residues were essential for PI hydrolysis, but not for protective signaling, while Arg323 and Lys409 residues were required for β-arrestin-1-mediated sustained ERK phosphorylation and cytoprotective signaling, but not for PI hydrolysis. Therefore, the mechanism of ligand bias appears to involve different modes of agonist interaction with the receptor ligand binding domain. While some mGlu1a agonists are biased toward PI hydrolysis, we identified two endogenous compounds, glutaric and succinic acids, as new mGlu1 agonists fully biased toward β-arrestin-mediated protective signaling. Pharmacological studies indicate that in producing the two effects, glutamate interacts in two distinct ways with mGlu1 receptors, since cytoprotective signaling was not inhibited by competitive mGlu1 antagonists which blocked PI hydrolysis. Moreover, quisqualate, a ligand biased toward PI hydrolysis failed to inhibit glutamate-induced protection; and glutaric acid, which is biased toward protection did not interfere with glutamate-induced PI hydrolysis. Taken together, these data indicate that ligand bias at mGlu1 receptors is due to different modes of receptor-glutamate interactions which are differentially coupled to PI hydrolysis and β-arrestin-mediated cytoprotective signaling, and reveal the existence of new endogenous agonists acting at mGlu1 receptors.
- Metabotropic glutamate
- Gq/11 family
- Phospholipase C's
- GRKs, barrestins
- Mutagenesis/Chimeric approaches
- Excitotoxicity, neurodegeneration
- Received February 22, 2012.
- Revision received May 9, 2012.
- Accepted May 10, 2012.
- The American Society for Pharmacology and Experimental Therapeutics