title: Painful stimuli induce in vivo phosphorylation and membrane mobilization of mouse spinal cord NKCC1 co-transporter.
abstract: The Na+ --Cl- --K+ isoform 1 (NKCC1) is a co-transporter that increases the intracellular concentration of chloride. NKCC1 plays a critical role in neuronal excitability and it has been recently suggested that it can contribute to hyperalgesic states by modulating the chloride concentration inside nociceptive neurons. In the spinal cord, trafficking of neurotransmitter receptors from the cytosol to the plasma membrane has been demonstrated to contribute to the development of hyperalgesia. However, it is unknown if trafficking of co-transporters can also occur in the nervous system or if it can be induced by painful stimulation. In this study, we have induced referred mechanical hyperalgesia in vivo by intracolonic instillation of capsaicin in mice. Using subcellular fractionation of proteins and cross-linking of membrane proteins we have observed that intracolonic capsaicin induced a 50% increase in NKCC1 in the plasma membrane of lumbosacral spinal cord 90 and 180 min after instillation, in parallel with a similar decrease in the cytosolic fraction. These effects returned to basal levels 6 h after capsaicin treatment. Intracolonic capsaicin also evoked a rapid (10 min) and transient phosphorylation of NKCC1, however, intracolonic saline did not produce significant changes in either NKCC1 trafficking or phosphorylation and none of the treatments induced any alterations of NKCC1 in the thoracic spinal cord. These results suggest that phosphorylation and recruitment of NKCC1 might play a role in referred mechanical hyperalgesia evoked by a painful visceral stimulus. The time course of the effects observed suggests that phosphorylation could contribute to the initial generation of hyperalgesia whereas trafficking could participate in the maintenance of hyperalgesic states observed at longer time points.
authors.affiliation: Anesthesia Research Unit, McGill University, McIntyre Medical Building, 3655 Promenade Sir William Osler, Montreal, Quebec, Canada H3G 1Y6.
authors.forename: A
authors.initials: A
authors.lastname: Galan
authors.forename: F
authors.initials: F
authors.lastname: Cervero
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002211:Capsaicin / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D006930:Hyperalgesia / Q000503:physiopathology; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009434:Neural Pathways / Q000503:physiopathology; D010146:Pain / Q000378:metabolism*; D010766:Phosphorylation; D020133:Reverse Transcriptase Polymerase Chain Reaction; D028021:Sodium-Potassium-Chloride Symporters / Q000378:metabolism*; D064506:Solute Carrier Family 12, Member 2; D013116:Spinal Cord / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism
chemical_list: D008565:Membrane Proteins; C578878:Slc12a2 protein, mouse; D028021:Sodium-Potassium-Chloride Symporters; D064506:Solute Carrier Family 12, Member 2; D002211:Capsaicin