picrotoxinin and (1-2-5-6-tetrahydropyridin-4-yl)methylphosphinic-acid

Retinal degenerative diseases can have many possible causes and are currently difficult to treat. As an alternative to therapies that require genetic manipulation or the implantation of electronic devices, photopharmacology has emerged as a viable approach to restore visual responses. Here, we present a new photopharmacological strategy that relies on a photoswitchable excitatory amino acid, ATA. This freely diffusible molecule selectively activates AMPA receptors in a light-dependent fashion. It primarily acts on amacrine and retinal ganglion cells, although a minor effect on bipolar cells has been observed. As such, it complements previous pharmacological approaches based on photochromic channel blockers and increases the potential of photopharmacology in vision restoration.

Topics: Action Potentials; Animals; Animals, Newborn; Blindness; Cyclic Nucleotide-Gated Cation Channels; Disease Models, Animal; GABA Agents; GluK2 Kainate Receptor; HEK293 Cells; Hippocampus; Humans; Light; Mice; Mice, Inbred C57BL; Mice, Knockout; Neurons; Phosphinic Acids; Picrotoxin; Pyridines; Receptors, AMPA; Receptors, Kainic Acid; Retinal Ganglion Cells; rho GTP-Binding Proteins; Rod Opsins; Sesterterpenes

The relative impact on picrotoxinin activity of residues at the intracellular (2' and 6' residues) and extracellular (15' and 17' residues) ends of the second transmembrane (M2) domain of the human gamma-aminobutyric acid-C (GABA(C)) rho1 receptor was investigated. A series of GABA(C) rho1 subunits were produced containing either single or multiple mutations at the positions of interest. Wild-type and mutant subunits (containing one or more of the following mutations: P2'S, T6'M, I15'N, G17'H) were expressed in Xenopus oocytes and characterized using agonists, partial agonists and antagonists. Changes in agonist activity were observed for mutant receptors. Most notably, mutation at the 2' position resulted in decreased agonist potency, while mutation at the 15' and 17' residues increased agonist potency. The affinity of the competitive antagonist (1,2,5,6-tetrahydropyridine-4-yl)methylphosphinic acid (TPMPA) was unchanged compared to wild-type at all mutant receptors. Of the four residues studied, mutation of residues at the 2' and 6' positions had the greatest impact on picrotoxinin activity. Inclusion of the P2'S mutation typically produced receptors with increased picrotoxinin potency, while the T6'M mutation reduced picrotoxinin potency. Picrotoxinin is a mixed antagonist at wild-type and all mutant receptors, with the exception of the double mutant rho1P2'S/T6'M receptors at which the non-competitive component was isolated. It is proposed that the contribution of M2 domain residues to picrotoxinin activity is potentially two-fold: (1) their role as a potential picrotoxinin binding site within the pore; and (2) they are critical for receptor activation properties of the receptor, thus may alter the allosteric mechanism of picrotoxinin.

Topics: Allosteric Regulation; Animals; Binding Sites; Binding, Competitive; GABA Antagonists; Humans; Mutation; Oocytes; Phosphinic Acids; Picrotoxin; Protein Subunits; Pyridines; Receptors, GABA; Receptors, GABA-B; Sesterterpenes; Xenopus laevis