摘要
Objective The rostral anterior cingulate cortex （rACC） is implicated in processing the emotional component of pain. N-methyl-D-aspartate receptors （NMDARs） are highly expressed in the rACC and mediate painrelated affect by activating a signaling pathway that involves cyclic adenosine monophosphate （cAMP）/protein ki- nase A （PKA） and/or extracellular regulated kinase （ERK）/cAMP-response element-binding protein （CREB）. The present study investigated the contributions of the NMDAR glycine site and GluN2B subunit to the activation of ERK and CREB both in vitro and in vivo in rat rACC. Methods Immunohistochemistry and Western blot analy- sis were used to separately assess the expression of phospho-ERK （pERK） and phospho-CREB （pCREB） in vitro and in vivo. Double immunostaining was also used to determine the colocalization of pERK and pCREB. Results Both bath application of NMDA in brain slices in vitro and intraplantar injection of formalin into the rat hindpaw in vivo induced significant up-regulation of pERK and pCREB in the rACC, which was inhibited by the NMDAR antago- nist DL-2-amino-5-phospho-novaleric acid. Selective blockade of the NMDAR GluN2B subunit and the glycine- binding site, or degradation of endogenous D-serine, a co-agonist for the glycine site, significantly decreased the up- regulation of pERK and pCREB expression in the rACC. Further, the activated ERK predominantly colocalized with CREB. Conclusion Either the glycine site or the GluN2B subunit of NMDARs participates in the phosphorylation of ERK and CREB induced by bath application of NMDA in brain slices or hindpaw injection of 5% formalin in rats, and these might be fundamental molecular mechanisms underlying pain affect.
Objective The rostral anterior cingulate cortex （rACC） is implicated in processing the emotional component of pain. N-methyl-D-aspartate receptors （NMDARs） are highly expressed in the rACC and mediate painrelated affect by activating a signaling pathway that involves cyclic adenosine monophosphate （cAMP）/protein ki- nase A （PKA） and/or extracellular regulated kinase （ERK）/cAMP-response element-binding protein （CREB）. The present study investigated the contributions of the NMDAR glycine site and GluN2B subunit to the activation of ERK and CREB both in vitro and in vivo in rat rACC. Methods Immunohistochemistry and Western blot analy- sis were used to separately assess the expression of phospho-ERK （pERK） and phospho-CREB （pCREB） in vitro and in vivo. Double immunostaining was also used to determine the colocalization of pERK and pCREB. Results Both bath application of NMDA in brain slices in vitro and intraplantar injection of formalin into the rat hindpaw in vivo induced significant up-regulation of pERK and pCREB in the rACC, which was inhibited by the NMDAR antago- nist DL-2-amino-5-phospho-novaleric acid. Selective blockade of the NMDAR GluN2B subunit and the glycine- binding site, or degradation of endogenous D-serine, a co-agonist for the glycine site, significantly decreased the up- regulation of pERK and pCREB expression in the rACC. Further, the activated ERK predominantly colocalized with CREB. Conclusion Either the glycine site or the GluN2B subunit of NMDARs participates in the phosphorylation of ERK and CREB induced by bath application of NMDA in brain slices or hindpaw injection of 5% formalin in rats, and these might be fundamental molecular mechanisms underlying pain affect.
基金
supported by the National Natural Science Foundation of China (30900444,31070973,30870835,31121061 and 30830044)
