The in vitro rat spinal cord

Abstract

In this study, in vitro spinal cord preparations have been used to investigate the possible role of excitatory amino acids in nociceptive mechanisms. The in vitro spinal cord dorsal horns showed good viability which was demonstrated by the development of spontaneous dorsal root activity a nd an evoked dorsal root reflex (DRR) 2-3 hours following dissection. Both these activities were blocked by the addition of manganese to the bathing medium, demonstrating them to be of synaptic origin. The DRR evoked by stimulation of an adjacent dorsal root showed evidence of both excitatory and inhibitory phases, similar to that described for hamster spinal cords (Bagust et al. 1982; Bagust et al. 1985a; Bagust et al. 1989). Stimulation of a lumbar dorsal root gave rise to dorsal horn field potentials. Investigations were made into the effects of the N-methyl-D-aspartate (NMDA) antagonists, 801 and the non-NMDA antagonist CNQX on fast and slow dorsal horn field potentials elictied by low and high intensity dorsal root stimulation respectively. None of the NMDA antagonists had any significant effect on fast wave dorsal root field potentials at concentrations up to 100 [jM, FC/NA a nd M K - 8 01 significantly inhibited the slow wave field potential. The non-NMDA antagonist CNQX significantly inhibited the synaptic components of both the fast and slow wave dorsal horn field potentials. It was concluded that low-threshold, non-noxious sensory transmission is mediated by non-NMDA glutamate receptors and that NMDA glutamate receptors have a role in the transmission of high-threshold, noxious sensory information in the dorsal horn of the spinal cord. arcaine sulphate, MK- but D-AP57 - CID - A P5 a nd 7 - CIKYNA, agonist (mGluR) group I The metabotropic glutamate receptor the synaptic components of the fast wave field potential. However, neither the group I antagonist L-AP3 nor the group II antagonist EGLU had any significant effect. The group III mGluR agonist L-AP4 significantly reduced the fast wave synaptic components. These results suggested a possible involvement of group I, II and III mGluRs in the processing of non-noxious information in the dorsal horn. The slow wave field potential elicited by high threshold dorsal root stimulation was inhibited by trans-ACPD, L-AP3 and EGLU. The group I mGluR agonist DHPG facilitated the slow wave at low concentrations. L-AP4 had no significant effect. It was therefore concluded that group I and II mGluRs but not group III mGluRs have a role in the processing of noxious sensory information in the spinal cord. trans-ACPD inhibited Expression of c-fos has previously been demonstrated in the dorsal horn of in vitro spinal cord preparations taken from 19-23 day old rats following high intensity dorsal root stimulation (Zhang et al. 1998). The expression of c-fos occurred only when dorsal root stimulation was sufficient to excite C fibres. Addition of 0 . %M capsaicin to the bathing medium induced intense c-fos staining in the absence of dorsal root stimulation. This provided a positive control suggesting that C fibre activation induces c-fos expression. Cords obtained from animals treated at 1 day old with capsaicin (50mg/kg) to destroy afferent C fibres showed a significant decrease in the number of Fos-positive cells induced by high intensity dorsal root stimulation when compared to vehicle treated animals. The effect of D-AP5, 7-CI KYNA and CNQX on pain-related expression of the immediate early gene c-fos was also investigated. Both D-AP5 and 7-CI KYNA significantly reduced the number of Fos-positive cells suggesting that NMDA receptors have a role in mediating expression. CNQX had no significant effect. This study has confirmed that the in vitro spinal cord is a valuable tool for the investigation of nociceptive processing and modulation in the dorsal horn of the spinal cord. The results support the suggestion that NMDA and group I and II metabotropic glutamate receptors acting at glutamatergic synapses in the dorsal horn of the spinal cord have a role in processing or modulating nociceptive input from C-fibre afferents.

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