BACKGROUND:Injection of glutamate (Glu) in normal animals can cause neuronal c-Fos expression; however,whether Glu can induce spinal neuronal c-Fos expression in pain models is unclear. OBJECTIVE:To examine the effects of intraplantar and intrathecal injection of Glu on c-Fos expression in the L5 spinal cord dorsal horn Ⅰ/Ⅱand Ⅲ/Ⅳ layers after spinal nerve ligation,and to study the effects of the N-methyl-D-aspartic acid (NMDA) receptor antagonist,D-2-amino-5-phosphonopentanoate (D-AP5),and a selective group I mGluR antagonist,7-hydroyiminocyclo propan[a]chromen-1a-carboxylic acid ethyl ester (cpccoEt). DESIGN,TIME AND SETTING:A randomized,controlled animal study was performed at the Department of Pharmacology,Oral Anatomy,and Neurobiology,Osaka University Graduate School of Dentistry,from December 2005 to December 2006. MATERIALS:Glu (5 μmol),D-AP5 (50 nmol) and cpccoEt (250 nmol) were provided by Wako Pure Chemical Industries,Osaka,Japan,and diluted in saline (50 μL). The pH of all solutions was adjusted to 7.4. METHODS:Twelve rats were randomly divided into sham operation (n = 6) and spinal nerve ligation (SNL; n = 6) groups for behavioral assessments of neuropathic pain after ligation surgery of the left L5-6 nerve segment. Another 60 rats were randomly divided into sham operation,SNL,saline-intraplantar,saline-intrathecal,Glu-intraplantar,Glu-intrathecal,D-AP5-intrathecal,Glu-D-AP5-intrathecal,cpccoEt-intrathecal,and Glu-cpccoEt-intrathecal groups,with 6 rats in each group. All groups except sham operation group received a similar SNL. On day 14,rats received a 50-μL injection of saline,Glu,D-AP5,and/or cpccoEt into the left intraplantar or intrathecal L5-6 segments. MAIN OUTCOME MEASURES:The number of c-Fos positive neurons in both Ⅰ/Ⅱ and Ⅲ/Ⅳ spinal layers at L6 was observed using immunohistochemistry 2 hours after administration. RESULTS:(1) SNL increased the level of c-Fos expression in two sides of the spinal cord,particularly on Ⅲ/Ⅳ spinal layers of the ligated side. (2) Intraplantar or intrathecal administration of saline significantly increased the c-Fos labeled neurons in Ⅰ/Ⅱ spinal layers of the ligated side,compared with SNL alone (P < 0.01). (3) Intraplantar Glu (5 μmol) increased the number of c-Fos positive neurons in Ⅰ/Ⅱ spinal layers compared with intraplantar saline (P < 0.01). (4) The number of c-Fos neurons in Ⅰ/Ⅱspinal layers on both the ipsilateral and contralateral side after intraplantar Glu was lower than intrathecal Glu (P < 0.01),with a 3-fold higher induction by intrathecal Glu. (5) Co-administration of D-AP5 or cpccoEt reduced the effects of intrathecal Glu (P < 0.01). CONCLUSION:Intrathecal Glu increases c-Fos expression more than intraplantar Glu. Antagonists of NMDA and group I mGluRs block this effect. BACKGROUND: Injection of glutamate (Glu) in normal animals can cause neuronal c-Fos expression; however, whether Glu can induce spinal neuronal c-Fos expression in pain models is unclear. OBJECTIVE: To examine the effects of intraplantar and intrathecal injection of Glu on c-Fos expression in the L5 spinal cord dorsal horn Ⅰ / Ⅱ and Ⅲ / Ⅳ layers after spinal nerve ligation, and to study the effects of the N-methyl-D-aspartic acid (NMDA) receptor antagonist, D-2- -5-phosphonopentanoate (D-AP5), and a selective group I mGluR antagonist, 7-hydroyiminocyclo propan [a] chromen-1a- carboxylic acid ethyl ester (cpccoEt) performed at the Department of Pharmacology, Oral Anatomy, and Neurobiology, Osaka University Graduate School of Dentistry, from December 2005 to December 2006. MATERIALS: Glu (5 μmol), D-AP5 (50 nmol) and cpccoEt by Wako Pure Chemical Industries, Osaka, Japan, and diluted in saline (50 μL). The p H of all solutions was adjusted to 7.4. METHODS: Twelve rats were randomly divided into sham operation (n = 6) and spinal nerve ligation (SNL; n = 6) groups for behavioral assessments of neuropathic pain after ligation surgery of the left L5- Another 60 rats were randomly divided into sham operation, SNL, saline-intraplantar, saline-intrathecal, Glu-intraplantar, Glu- intrathecal, D-AP5- intrathecal, Glu- D-AP5- intrathecal, cpccoEt-intrathecal, and Glu-cpccoEt-intrathecal groups, with 6 rats in each group. On day 14, rats received a 50-μL injection of saline, Glu, D-AP5, and / or cpccoEt MAIN OUTCOME MEASURES: The number of c-Fos positive neurons in both I / II and III / IV spinal layers at L6 was observed using immunohistochemistry for 2 hours after administration. RESULTS: (1) SNL increased the level of c-Fos expression in two sides of the spinal cord, particularly on III / IV spinal layers of the l(2) Intraplantar or intrathecal administration of corticobasal or intrathecal administration of c-Fos labeled neurons in I / II spinal layers of the ligated side, compared with SNL alone (P <0.01) (4) The number of c-Fos neurons in I / IIspinal layers on both the ipsilateral and contralateral side after intraplantar Glu was lower than intrathecal Glu (P <0.01), with a 3-fold higher induction by intrathecal Glu. (5) Co-administration of D-AP5 or cpccoEt reduced the effects of intrathecal Glu increases c-Fos expression more than intraplantar Glu. Antagonists of NMDA and group I mGluRs block this effect.