Dataset Information

Visualization of ligand-induced dopamine D2S and D2L receptor internalization by TIRF microscopy.

ABSTRACT: G protein-coupled receptors (GPCRs), including the dopamine receptors, represent a group of important pharmacological targets. Upon agonist binding, GPCRs frequently undergo internalization, a process that is known to attenuate functional responses upon prolonged exposure to agonists. In this study, internalization was visualized by means of total internal reflection fluorescence (TIRF) microscopy at a level of discrete single events near the plasma membrane with high spatial resolution. A novel method has been developed to determine the relative extent of internalized fluorescent receptor-ligand complexes by comparative fluorescence quantification in living CHO cells. The procedure entails treatment with the reducing agent sodium borohydride, which converts cyanine-based fluorescent ligands on the membrane surface to a long-lived reduced form. Because the highly polar reducing agent is not able to pass the cell membrane, the fluorescent receptor-ligand complexes located in internalized compartments remain fluorescent under TIRF illumination. We applied the method to investigate differences of the short (D2S) and the long (D2L) isoforms of dopamine D2 receptors in their ability to undergo agonist-induced internalization.

SUBMITTER: Tabor A

PROVIDER: S-EPMC5589927 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Visualization of ligand-induced dopamine D<sub>2S</sub> and D<sub>2L</sub> receptor internalization by TIRF microscopy.

Tabor Alina A Möller Dorothee D Hübner Harald H Kornhuber Johannes J Gmeiner Peter P

Scientific reports 20170907 1

G protein-coupled receptors (GPCRs), including the dopamine receptors, represent a group of important pharmacological targets. Upon agonist binding, GPCRs frequently undergo internalization, a process that is known to attenuate functional responses upon prolonged exposure to agonists. In this study, internalization was visualized by means of total internal reflection fluorescence (TIRF) microscopy at a level of discrete single events near the plasma membrane with high spatial resolution. A novel ...[more]

PMID: 28883522

Similar Datasets

Project description:Dopamine receptors are integral membrane proteins whose endogenous ligand is dopamine. They play a fundamental role in the central nervous system and dysfunction of dopaminergic neurotransmission is responsible for the generation of a variety of neuropsychiatric disorders. From an evolutionary standpoint, phylogenetic relationships among the DRD1 class of dopamine receptors are still a matter of debate as in the literature different tree topologies have been proposed. In contrast, phylogenetic relationships among the DRD 2 group of receptors are well understood. Understanding the time of origin of the different dopamine receptors is also an issue that needs further study, especially for the genes that have restricted phyletic distributions (e.g., DRD2l and DRD4rs). Thus, the goal of this study was to investigate the evolution of dopamine receptors, with emphasis on shedding light on the phylogenetic relationships among the D1 class of dopamine receptors and the time of origin of the DRD2l and DRD4rs gene lineages. Our results recovered the monophyly of the two groups of dopamine receptors. Within the DRD1 group the monophyly of each paralog was recovered with strong support, and phylogenetic relationships among them were well resolved. Within the DRD1 class of dopamine receptors we recovered the sister group relationship between the DRD1C and DRD1E, and this clade was recovered sister to a cyclostome sequence. The DRD1 clade was recovered sister to the aforementioned clade, and the group containing DRD5 receptors was sister to all other DRD1 paralogs. In agreement with the literature, among the DRD2 class of receptors, DRD2 was recovered sister to DRD3, whereas DRD4 was sister to the DRD2/DRD3 clade. According to our phylogenetic tree, the DRD2l and DRD4rs gene lineages would have originated in the ancestor of gnathostomes between 615 and 473 mya. Conservation of sequences required for dopaminergic neurotransmission and small changes in regulatory regions suggest a functional refinement of the dopaminergic pathways along evolution.

Dataset Information

Visualization of ligand-induced dopamine D2S and D2L receptor internalization by TIRF microscopy.

Publications

Visualization of ligand-induced dopamine D<sub>2S</sub> and D<sub>2L</sub> receptor internalization by TIRF microscopy.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets