Elsevier

Brain Research

Volume 1015, Issues 1–2, 23 July 2004, Pages 145-153
Brain Research

Research report
Intrathecal interleukin-1β administration induces thermal hyperalgesia by activating inducible nitric oxide synthase expression in the rat spinal cord

https://doi.org/10.1016/j.brainres.2004.04.068Get rights and content

Abstract

The effect of the pro-inflammatory cytokine interleukin-1β (IL-1β) on the inducible nitric oxide synthase-nitric oxide (iNOS-NO) cascade in nociceptive signal transduction was examined in the intact rat spinal cord. All rats were implanted with an intrathecal (i.t.) catheter; some were also implanted with an i.t. microdialysis probe. The paw withdrawal latency to radiant heat was used to assess thermal hyperalgesia. The iNOS protein expression in the spinal cord dorsal horn was examined by western blot analysis and NOS activity assay. NO production in the CSF dialysate was also measured. IL-1β i.t. (100 ng) produced thermal hyperalgesia from 4 to 24 h after i.t. injection. The iNOS protein expression was induced at 4 h after i.t. IL-1β injection, peaked at the 6th hour, and disappeared at 24 h. The iNOS activity showed a similar time-dependent change as the iNOS protein expression. NO release increased by 1.1- to 1.9-fold between 4 and 12 h, also with a peak at the 6th hour, after i.t. IL-1β administration. Pretreatment with the iNOS inhibitor 1400W (10 μg, i.t.) 1 h before i.t. IL-1β injection prevented all the responses of IL-1β. Neither 1400W nor artificial CSF (aCSF) affected the thermal nociceptive threshold and NO production. These results demonstrate that i.t. administration of IL-1β induced thermal hyperalgesia by activating the iNOS-NO cascade in the rat spinal cord. On the basis of the present findings, we suggest that i.t. administration of iNOS inhibitors may have potential in the treatment of inflammatory and neuropathic pain syndromes.

Introduction

Interleukin-1β (IL-1β), a pro-inflammatory cytokine, is involved in the immune response and signal transduction both in the periphery and the central nervous system (CNS) [13], [14], [17], [18], [28], [36]. It is released during inflammatory and neuropathic pain conditions [8], [34], [39], [43]. Samad et al. [34] found that injection of complete Freund's adjuvant (CFA) into the rat's hindpaw induced peripheral inflammation and IL-1β upregulation (>10,000-fold) in the inflamed hindpaw soon after CFA administration; this effect lasted for several days and was correlated with the peripheral inflammation. Moreover, in their study, the IL-1β concentration in the CSF was increased 50- and 20-fold, at 2 and 4 h, respectively, after CFA injection, and this was followed by an increased cyclooxygenase-2 (COX-2) level in the spinal cord. They also found that intrathecal (i.t.) injection of IL-1β (50 ng) induced a 30-fold increase in COX-2 mRNA levels in the rat spinal cord. Administration of IL-1β produces hyperalgesia; in contrast, blockade of IL-1β reduces pain [10], [30], [32], [34], [35], [37]. It is apparent that IL-1β is an important mediator in the cross-talk between the nervous and immune systems, and also participates in modulation of the nociceptive threshold.

Nitric oxide (NO), a short-lived diffusible molecule of great biological importance, plays a key role in host defense, signal transduction, and neurotransmission. There is increasing evidence that NO is also involved in nociceptive processing after inflammation and neuropathy [26]. Three isoforms of nitric oxide synthase (NOS) are known; two are constitutively expressed (nNOS and eNOS), and the third is inducible NOS (iNOS). Both nNOS (neuronal) and eNOS (endothelial) are activated in response to physical stimuli, and rapidly produce low levels of NO. In contrast, iNOS is activated by stimuli such as IL-1β, and produces a high level of NO over a long period of time [6], [23]. Upregulation of NOS expression and subsequent NO production in the spinal cord contributes to pain following nociceptive stimulation in animals [20], [26], [33], [41], [46]. i.t. administration of NOS inhibitors significantly reduces nociceptive behavior in response to peripheral inflammation [31], [33], [46]. Moreover, IL-1β was found to increase both iNOS expression and NO production in cultured cells [12], [15]. Although both IL-1β and the NOS-NO cascade have been suggested to modulate nociception, there is little information concerning the interaction between IL-1β and the iNOS-NO cascade in the spinal cord. We hypothesized that, i.t. IL-1β causes a markedly increased NO production via induction of iNOS expression in the rat spinal cord. If an IL-1β induced elevation in NO can be confirmed, it might reveal a significant pathophysiological pathway of intrathecal IL-1β-induced thermal hyperalgesia. The present study was designed to examine the role of the NOS-NO cascade, and its correspondence to nociceptive behaviors, in the spinal cord of i.t. IL-1β-treated rats.

Section snippets

Construction and implantation of the intrathecal catheter and microdialysis probe

The i.t. catheter was constructed using a 9-cm long polyethylene (PE5) tube and a 3.5-cm long silastic tube. The silastic tube was inserted into the PE5 tube and sealed with epoxy resin and silicon rubber. In addition, the spinal microdialysis probe construction technique was modified and adapted in previous studies [21], [45]. The microdialysis probe was constructed using two 7 cm PE5 tubes (0.008 inch inner diameter, 0.014 inch outer diameter) and a 4-cm cuprophan hollow fiber (300 μm outer

Effect of iNOS inhibitor 1400W on IL-1β-induced thermal hyperalgesia

After i.t. injection of IL-1β (100 ng), thermal hyperalgesia was observed with the PWL reduced to 50–60% of baseline level between 4 and 24 h (Fig. 1). This hypersensitivity to the heat was significantly increased at 4 h, peaked at 6 h, and lasted for 24 h after IL-1β i.t. administration. Either aCSF or 1400W (10 μg) alone did not affect the PWL to the heat, whereas i.t. pretreatment with 1400W (10 μg) 1 h prior to IL-1β (100 ng) injection significantly attenuated the IL-1β-induced thermal

Discussion

In the present study, thermal hyperalgesia was observed 4 h after IL-1β injection and lasted for 24 h. The iNOS selective inhibitor 1400W prevented this thermal hyperalgesia and the corresponding biochemical changes. I.t. administration of IL-1β induced iNOS protein expression and the subsequent NO release, in a time-dependent manner, in the rat spinal cord. iNOS protein induction and activation were observed at 4 h after IL-1β injection, peaked at 6 h, then declined gradually, and disappeared

Acknowledgements

The authors wish to thank Dr. Yoshito Takano for his helpful communications on the algesic effect of spinal IL-1β and Mr. Yao-Chang Chen for the statistical analysis. This work was supported by grants from the National Science Council (NSC 91-2314-B-075-070) and Ministry of Defense (DOD-93-2-08) of the Republic of China.

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