Illuminating Phenylazopyridines To Photoswitch Metabotropic Glutamate Receptors: From the Flask to the Animals

Abstract

Phenylazopyridines are photoisomerizable compounds with high potential to control biological functions with light. We have obtained a series of phenylazopyridines with light dependent activity as negative allosteric modulators (NAM) of metabotropic glutamate receptor subtype 5 (mGlu5). Here we describe the factors needed to achieve an operational molecular photoisomerization and its effective translation into in vitro and in vivo receptor photoswitching, which includes zebrafish larva motility and the regulation of the antinociceptive effects in mice. The combination of light and some specific phenylazopyridine ligands displays atypical pharmacological profiles, including light-dependent receptor overactivation, which can be observed both in vitro and in vivo. Remarkably, the localized administration of light and a photoswitchable compound in the peripheral tissues of rodents or in the brain amygdalae results in an illumination-dependent analgesic effect. The results reveal a robust translation of the phenylazopyridine photoisomerization to a precise photoregulation of biological activity.

Reversible silencing of endogenous receptors in intact brain tissue using two-photon pharmacology

“…Interestingly, using 1PE of alloswitch, Ca 2+ responses were recovered in a significantly higher fraction of cells in the field of view than those responding to DHPG in the absence of alloswitch ( Figure 5E). Similar effects are observed in cell lines ( Figure 3D) and have been previously reported 16,28 . In brain slices, these results suggest that pharmacological silencing of mGlu 5 receptors followed by rapid photo-rescue in a physiological setting uncovers neglected features of receptor balancing in response to prolonged arrested signaling.…”

“…Several alloswitch analogs have been recently reported that display a variety of pharmacological and photophysical properties 28 , and we selected a few in order to test their ability to photocontrol mGlu 5 receptors at 2PE. We repeated the previous experiments using analogs bearing substitutions at the distal phenyl ring, and found that all of them were responsive at similar wavelengths ( Figure S3).…”

“…Freely-diffusible photoswitchable allosteric modulators of mGlu 5 receptors were obtained as previously described for alloswitch 16 and compounds 1c-d, 1j and 6 28 3.3 "All-2P" experiments workflow (2P-imaging of Fura-2 in mGlu 5 -eYFP cells)…”

“…in the brain), it is necessary to take a photoswitchable ligand having the highest pharmacological selectivity. Currently, the best available option is alloswitch and its analog compounds, which are freely-diffusible allosteric modulators of metabotropic glutamate 5 (mGlu 5 ) receptors 16,28 . These receptors are important modulators of neurotransmission and synaptic plasticity 29 , and are involved in several neurological conditions 30,31,32,33,34,35 .…”

“…receptors, and 2PE produced rapid and localized rescue of mGlu 5 agonist responses. Illumination power, wavelength and other 2PEconditions can be achieved with commercial pulsed lasers(Figure 2)and were effective in all tested alloswitch analogs (four selected from a library of 18,28 ;Figure S3). 2PE of alloswitch achieved mGlu 5 responses comparable to endogenous activity induced by agonist, yet weaker than 1PE in the same conditions(Figure 3).…”