Abstract
BackgroundGlycine receptors (GlyRs) are involved in the development of spinal pain sensitization. The GlyRα3 subunit has recently emerged as a key factor in inflammatory pain pathways in the spinal cord dorsal horn (DH). Our study is to identify the extent of location and cell types expressing different GlyR subunits in spinal cord and dorsal root ganglion (DRGs). To tease out the possible actions of GlyRs on pain transmission, we investigate the effects produced by GlyRs on acute inflammatory pain by behavioral testing using prostaglandin E2 (PGE2) intrathecal injection models. Furthermore, we investigate the changes of GlyR expression in DRGs and spinal cord in rats after the induction of acute inflammatory pain.ResultsCompared to the vehicle administration, the PGE2 intrathecal injection model produced significantly higher hyperalgesia, which started 3 h after PGE2 injection and lasted more than 5 h. PGE2 intrathecal injection significantly decreased GlyRα1 and GlyRα3 protein expressions in the L5 DH at 1 h and lasted to 5 h, and similar results were observed in the L5 DRG at 5 h. Confocal microscopic images showed the co-existence of punctate gephyrin and GlyRα3 immunoreactivity (IR) throughout the gray matter of the spinal cord, mainly in DH laminae I–III neurons and in ventral horn neurons. It also showed the co-existence of punctate gephyrin and GlyRα3 IR in DRG neurons.ConclusionsIn this study, PGE2 intrathecal injection significantly decreased protein expression of gephyrin, GlyRα1 and GlyRα3 in spinal cord DH and DRG. The gephyrin and GlyRα3 were localized on neuron cells both in the DH and DRG.
Highlights
Glycine receptors (GlyRs) are involved in the development of spinal pain sensitization
The membranes were firstly blocked with 5% milk in phosphate-buffered saline (PBS) with 0.1% Tween-20 for 1 h at room temperature, and probed overnight at 4 °C with Rabbit antiGlyRα1 Polyclonal Antibody (1:1000, AGR-001, Alomone Labs, Israel), Rabbit anti-gephyrin Polyclonal Antibody (1:2000, AIP-005, Alomone Labs, Israel), goat AntiGlyRα3 polyclonal Antibody (1:500, SC-17282, Santa Cruz Biotechnology, Santa Cruz, CA, USA), or mouse anti-actin monoclonal antibody (1:10,000, MAB1501, Indianapolis, IN, USA) primary antibody to detect the expression of GlyRα1, GlyRα3 and gephyrin in the dorsal root ganglion (DRG) and spinal cord
IF revealed the gephyrin and GlyRα3 were localized on neuron cells
Summary
Glycine receptors (GlyRs) are involved in the development of spinal pain sensitization. The GlyRα3 subunit has recently emerged as a key factor in inflammatory pain pathways in the spinal cord dorsal horn (DH). Our study is to identify the extent of location and cell types expressing different GlyR subunits in spinal cord and dorsal root ganglion (DRGs). We investigate the changes of GlyR expression in DRGs and spinal cord in rats after the induc‐ tion of acute inflammatory pain. The GlyRα3 subunit has recently become a key factor in inflammatory pain pathways in the dorsal horn of the spinal cord [5], The completeness of the position and the identification of the cell types expressing different GlyR subunits are not clear. Quantitative methods allow the comparison of GlyRs activity at different times following inflammation
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