Immuno-pharmacological characterization of group II metabotropic glutamate receptors controlling glutamate exocytosis in mouse cortex and spinal cord.

Background And Purpose: We recently proposed the existence of mGlu -preferring autoreceptors in spinal cord terminals and of mGlu -preferring autoreceptors in cortical terminals. This study aims to verify our previous conclusions and to extend their pharmacological characterization.

Experimental Approach: We studied the effect of LY566332, an mGlu receptor positive allosteric modulator (PAM), and of LY2389575, a selective mGlu receptor negative allosteric (NAM) modulator, on the mGlu agonist LY379268-mediated inhibition of glutamate exocytosis [measured as KCl-evoked release of preloaded [ H]-D-aspartate]. The mGlu PAM BINA and the mGlu NAM ML337, as well as selective antibodies recognizing the N-terminal of the receptor proteins, were used to confirm the pharmacological characterization of the native receptors.

Key Results: Cortical synaptosomes possess LY566332-sensitive autoreceptors that are slightly, although significantly, susceptible to LY2389575. In contrast, LY566332-insensitive and LY2389575-sensitive autoreceptors are present in spinal cord terminals. BINA and ML337 mimicked LY566332 and LY2389575, respectively, in controlling LY379268-mediated inhibition of glutamate exocytosis from both cortical and spinal cord synaptosomes. Incubation of cortical synaptosomes with anti-mGlu antibody prevented the LY379268-induced inhibition of glutamate exocytosis, and this response was partially reduced by the anti-mGlu antibody. Incubation of spinal cord synaptosomes with the anti-mGlu antibody abolished LY379268-mediated reduction of glutamate exocytosis from these terminals, while the anti-mGlu antibody was inactive. Western blot analysis and confocal microscopy data were largely consistent with these functional observations.

Conclusions And Implications: We confirmed that mGlu -preferring autoreceptors exist in spinal cord terminals. Differently, cortical glutamatergic terminals possess mGlu /mGlu heterodimers, whose inhibitory effect is largely mediated by mGlu receptors.