Regulation Of Glycine Receptor α1^ins Subunit By Adenosine

Objective:Glycine receptors（GlyRs）are pentameric proteins composed ofα（α1-α4）andβsubunits.In spinal cord dorsal horn of adult animals,α1 subunit-containing glycine receptors mediate inhibitory synaptic transmission,which plays an important role in the pain regulation.Theα1^ins subunit is an isoform ofα1 subunit.Accumulating evidence has indicated that adenosine can alleviate the painful responses by enhancing the function of GlyRs.However,the mechanisms by which adenosine regulates GlyRs have remained elusive.This study was designed to explore the regulatory effects of adenosine on glycine receptorα1^ins subunit.Methods:The model of inflammatory pain was established by intraplantar injection of Complete Freund’s adjuvant（CFA）.Behavioral tests were performed to examine the effects of adenosine on the mechanical allodynia and thermal hyperalgesia.The distribution ofα1^ins subunits in the spinal cord dorsal horn neurons was investigated by immunohistochemistry and immunocytochemistry.The patch-clamp electrophysiological recordings were used to examine the changes ofα1ⁱnsns currents.The effects of adenosine on the phosphorylation ofα1^ins and extracellular signal-regulated kinase 1/2（ERK1/2）were studied by immunoblotting analysis.Results:（1）Adenosine increased the values of Paw Withdrawal Thresholds（PWTs）evoked by mechanical stimuli and of Paw Withdrawal Latencies（PWLs）evoked by thermal stimuli in inflamed mice.（2）Adenosine increased postsynaptic GlyRs currents.（3）Theα1^ins subunit was a specific target for adenosine A1 receptors（A1Rs）regulation.Activation of A1Rs was able to enhance theα1^ins currents,without any significant effects onα1 currents.（4）Theα1^ins subunit was abundant in the superficial dorsal horn neurons of spinal cord and was localized at the inhibitory synapses.（5）The G proteinβγsubunits（Gβγ）were critical for A1Rs to regulateα1^ins.Inhibition of Gβγ,but not cAMP-dependent protein kinase（PKA）,eliminated the potentiating effects of adenosine onα1^ins currents.（6）Adenosine reversed the increase ofα1^ins phosphorylation at Ser380 induced by peripheral inflammation through inhibition of ERK1/2.Conclusion:The analgesic action of adenosine against inflammatory pain involved the potentiation ofα1^ins-mediated glycinergic transmission.