Triphenylamine Sensitized 8-Dimethylaminoquinoline: An Efficient Two-Photon Caging Group for Intracellular Delivery.

Triphenylamine-sensitized 8-dimethylaminoquinoline (TAQ) probes showed fair two-photon absorption and fragmentation cross sections in releasing kainate and GABA ligands. The water-soluble PEG and TEG-analogs allowed cell internalization and efficient light-gated liberation of the rhodamine reporter under UV and two-photon (NIR) irradiation conditions.

Determination of the absolute configuration of phosphinic analogues of glutamate.

A series of phosphinic glutamate derivatives (e.g.LSP1-2111) have been proven to be potent agonists of metabotropic glutamate (mGlu) receptors and shown promising in vivo activity. However, so far all were synthesized and tested as a mixture of two diastereomers whose absolute and relative configurations are not known. In this study, the stereomers were separated on a Crownpack CR(+) column and their absolute configuration was assessed by means of a diastereoselective synthesis. Both separated L-stereomers activated the mGlu4 receptor with EC50's of 0.72 and 4.4 µM for (1S,1'S)-and (1S,1'R)-LSP1-2111, respectively.

A novel selective metabotropic glutamate receptor 4 agonist reveals new possibilities for developing subtype selective ligands with therapeutic potential.

Metabotropic glutamate (mGlu) receptors are promising targets to treat numerous brain disorders. So far, allosteric modulators are the only subtype selective ligands, but pure agonists still have strong therapeutic potential. Here, we aimed at investigating the possibility of developing subtype-selective agonists by extending the glutamate-like structure to hit a nonconsensus binding area. We report the properties of the first mGlu4-selective orthosteric agonist, derived from a virtual screening hit, LSP4-2022 using cell-based assays with recombinant mGlu receptors [EC(50): 0.11 ± 0.02, 11.6 ± 1.9, 29.2 ± 4.2 µM (n>19) in calcium assays on mGlu4, mGlu7, and mGlu8 receptors, respectively, with no activity at the group I and -II mGlu receptors at 100 µM]. LSP4-2022 inhibits neurotransmission in cerebellar slices from wild-type but not mGlu4 receptor-knockout mice. In vivo, it possesses antiparkinsonian properties after central or systemic administration in a haloperidol-induced catalepsy test, revealing its ability to cross the blood-brain barrier. Site-directed mutagenesis and molecular modeling was used to identify the LSP4-2022 binding site, revealing interaction with both the glutamate binding site and a variable pocket responsible for selectivity. These data reveal new approaches for developing selective, hydrophilic, and brain-penetrant mGlu receptor agonists, offering new possibilities to design original bioactive compounds with therapeutic potential.

Increased Potency and Selectivity for Group III Metabotropic Glutamate Receptor Agonists Binding at Dual sites.

A group III metabotropic glutamate (mGlu) receptor agonist (PCEP) was identified by virtual HTS. This orthosteric ligand is composed by an l-AP4-derived fragment that mimics glutamate and a chain that binds into a neighboring pocket, offering possibilities to improve affinity and selectivity. Herein we describe a series of derivatives where the distal chain is replaced by an aromatic or heteroaromatic group. Potent agonists were identified, including some with a mGlu4 subtype preference, e.g., 17m (LSP1-2111) and 16g (LSP4-2022). Molecular modeling suggests that aromatic functional groups may bind at either one of the two chloride regulatory sites. These agonists may thus be considered as particular bitopic/dualsteric ligands. 17m was shown to reduce GABAergic synaptic transmission at striatopallidal synapses. We now demonstrate its inhibitory effect at glutamatergic parallel fiber-Purkinje cell synapses in the cerebellar cortex. Although these ligands have physicochemical properties that are markedly different from typical CNS drugs, they hold significant therapeutic potential.

Phasic and Tonic mGlu7 Receptor Activity Modulates the Thalamocortical Network.

Mutation of the metabotropic glutamate receptor type 7 (mGlu7) induces absence-like epileptic seizures, but its precise role in the somatosensory thalamocortical network remains unknown. By combining electrophysiological recordings, optogenetics, and pharmacology, we dissected the contribution of the mGlu7 receptor at mouse thalamic synapses. We found that mGlu7 is functionally expressed at both glutamatergic and GABAergic synapses, where it can inhibit neurotransmission and regulate short-term plasticity. These effects depend on the PDZ-ligand of the receptor, as they are lost in mutant mice. Interestingly, the very low affinity of mGlu7 receptors for glutamate raises the question of how it can be activated, namely at GABAergic synapses and in basal conditions. Inactivation of the receptor activity with the mGlu7 negative allosteric modulator (NAM), ADX71743, enhances thalamic synaptic transmission. In vivo administration of the NAM induces a lethargic state with spindle and/or spike-and-wave discharges accompanied by a behavioral arrest typical of absence epileptic seizures. This provides evidence for mGlu7 receptor-mediated tonic modulation of a physiological function in vivo preventing synchronous and potentially pathological oscillations.

Identification of positive allosteric modulators VU0155094 (ML397) and VU0422288 (ML396) reveals new insights into the biology of metabotropic glutamate receptor 7.

Metabotropic glutamate receptor 7 (mGlu7) is a member of the group III mGlu receptors (mGlus), encompassed by mGlu4, mGlu6, mGlu7, and mGlu8. mGlu7 is highly expressed in the presynaptic active zones of both excitatory and inhibitory synapses, and activation of the receptor regulates the release of both glutamate and GABA. mGlu7 is thought to be a relevant therapeutic target for a number of neurological and psychiatric disorders, and polymorphisms in the GRM7 gene have been linked to autism, depression, ADHD, and schizophrenia. Here we report two new pan-group III mGlu positive allosteric modulators, VU0155094 and VU0422288, which show differential activity at the various group III mGlus. Additionally, both compounds show probe dependence when assessed in the presence of distinct orthosteric agonists. By pairing studies of these nonselective compounds with a synapse in the hippocampus that expresses only mGlu7, we have validated activity of these compounds in a native tissue setting. These studies provide proof-of-concept evidence that mGlu7 activity can be modulated by positive allosteric modulation, paving the way for future therapeutics development.

A virtual screening hit reveals new possibilities for developing group III metabotropic glutamate receptor agonists.

(R)-PCEP (3-amino-3-carboxypropyl-2'-carboxyethyl phosphinic acid, 1), a new metabotropic glutamate receptor 4 (mGlu4R) agonist, was discovered in a previously reported virtual screening. The (S)-enantiomer and a series of derivatives were synthesized and tested on recombinant mGlu4 receptors. A large number of derivatives activated this receptor but was not able to discriminate between mGlu4 and mGlu8 receptors. The most potent ones 6 and 12 displayed an EC(50) of 1.0 +/- 0.2 microM at mGlu4R. Interestingly these agonists with longer alkyl chains revealed a new binding pocket adjacent to the glutamate binding site, which is lined with residues that differ among the mGluR subtypes and that will allow the design of more selective compounds. Additionally 6 was able to activate mGlu7 receptor with an EC(50) of 43 +/- 16 microM and is thus significantly more potent than L-AP4 (EC(50) of 249 +/- 106 microM).