title: Rapid constitutive and ligand-activated endocytic trafficking of P2X receptor.
abstract: P2X receptors mediate a variety of physiological actions, including smooth muscle contraction, neuro-endocrine secretion and synaptic transmission. Among P2X receptors, the P2X(3) subtype is expressed in sensory neurons of dorsal root- and trigeminal-ganglia, where it performs a well-recognized role in sensory and pain transmission. Recent evidence indicates that the strength of P2X(3)-mediated responses is modulated in vivo by altering the number of receptors at the plasma membrane. In the present study, we investigate the trafficking properties of P2X(3) receptor in transfected HEK293 cells and in primary cultures of dorsal root ganglion neurons, finding that P2X(3) receptor undergoes rapid constitutive and cholesterol-dependent endocytosis. We also show that endocytosis is accompanied by preferential targeting of the receptor to late endosomes/lysosomes, with subsequent degradation. Furthermore, we observe that at steady state the receptor localizes predominantly in lamp1-positive intracellular structures, with a minor fraction present at the plasma membrane. Finally, the level of functional receptor expressed on the cell surface is rapidly up-regulated in response to agonist stimulation, which also augments receptor endocytosis. The findings presented in this work underscore a very dynamic trafficking behavior of P2X(3) receptor and disclose a possible mechanism for the rapid modulation of ATP-mediated responses potentially relevant during physiological and pathological conditions.
authors.affiliation: Laboratory of Cellular Neurobiology, Santa Lucia Foundation, Rome, Italy. fabrizio.vacca@unige.ch
authors.forename: Fabrizio
authors.initials: F
authors.lastname: Vacca
authors.forename: Michela
authors.initials: M
authors.lastname: Giustizieri
authors.forename: Maria Teresa
authors.initials: MT
authors.lastname: Ciotti
authors.forename: Nicola Biagio
authors.initials: NB
authors.lastname: Mercuri
authors.forename: Cinzia
authors.initials: C
authors.lastname: Volonté
mesh_terms: D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000502:physiology; D000595:Amino Acid Sequence; D000818:Animals; D019921:Biotinylation; D015153:Blotting, Western; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D004594:Electrophysiology; D004705:Endocytosis / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D005455:Fluorescent Antibody Technique; D005727:Ganglia, Spinal / Q000166:cytology / Q000187:drug effects; D006801:Humans; D008024:Ligands; D008247:Lysosomes / Q000502:physiology; D008969:Molecular Sequence Data; D018408:Patch-Clamp Techniques; D058910:Purinergic P2 Receptor Agonists; D051381:Rats; D017208:Rats, Wistar; D018048:Receptors, Purinergic P2 / Q000235:genetics / Q000502:physiology*; D058477:Receptors, Purinergic P2X3; D015398:Signal Transduction / Q000502:physiology; D014162:Transfection
chemical_list: D008024:Ligands; C546407:P2RX3 protein, human; D058910:Purinergic P2 Receptor Agonists; D018048:Receptors, Purinergic P2; D058477:Receptors, Purinergic P2X3; D000255:Adenosine Triphosphate; C002630:alpha,beta-methyleneadenosine 5'-triphosphate