Combining ketamine with astrocytic inhibitor as a potential analgesic strategy for neuropathic pain ketamine, astrocytic inhibitor and pain

Abstract

Background: Neuropathic pain is an intractable clinical problem. Intrathecal ketamine, a noncompetitive N-methyl-D-aspartate receptor (NMDAR) antagonist, is reported to be useful for treating neuropathic pain in clinic by inhibiting the activity of spinal neurons. Nevertheless, emerging studies have disclosed that spinal astrocytes played a critical role in the initiation and maintenance of neuropathic pain. However, the present clinical therapeutics is still just concerning about neuronal participation. Therefore, the present study is to validate the coadministration effects of a neuronal noncompetitive N-methyl-D-aspartate receptor (NMDAR) antagonist ketamine and astrocytic cytotoxin L-α-aminoadipate (LAA) on spinal nerve ligation (SNL)-induced neuropathic pain.

Results: Intrathecal ketamine (10, 100, 1000 μg/kg) or LAA (10, 50, 100 nmol) alleviated SNL-induced mechanical allodynia in a dose-dependent manner respectively. Phosphorylated NR1 (pNR1) or glial fibrillary acidic protein (GFAP) expression was down-regulated by intrathecal ketamine (100, 1000 μg/kg) or LAA (50, 100 nmol) respectively. The combination of ketamine (100 μg/kg) with LAA (50 nmol) showed superadditive effects on neuropathic pain compared with that of intrathecal administration of either ketamine or LAA alone. Combined administration obviously relieved mechanical allodynia in a quick and stable manner. Moreover, down-regulation of pNR1 and GFAP expression were also enhanced by drugs coadministration.

Conclusions: These results suggest that combining NMDAR antagonist ketamine with an astrocytic inhibitor or cytotoxin, which is suitable for clinical use once synthesized, might be a potential strategy for clinical management of neuropathic pain.

Figure 1

Effects of drugs administration on spinal nerve ligation (SNL) -induced astrocytic activation. A representative spinal section showed that SNL induced a marked astrocytic activation in ipsilateral than that of contralateral spinal dorsal horn (A). B and C show the magnified images of the ipsilateral and contralateral spinal dorsal horn of SNL rats respectively. Compared with Sham-Saline rats (D), SNL induced a marked astrocytic activation (F, J). Intrathecal ketamine (100 μg/kg) didn't affect influence astrocytic activation obviously at both 1 d and 3 d after application (G, K). However, intrathecal LAA (50 nmol) attenuated SNL-induced astrocytic activation at 1 d after administration compared with that of SNL-Saline group (H). Astrocytic activation was further suppressed by coadministration (100 μg/kg of ketamine and 50 nmol of LAA) than that of LAA (50 nmol) injected alone (I). Astrocytic activation was returned obvious 3 d after LAA injection (L). However, drugs coadministration significantly inhibited astrocytic activation compared to that of LAA alone (M). E, Scheme presenting an overview over the detected region. N, Statistics of above results. Scalebar = 800 μm in A, 20 μm in B-C and 200 μm in D, F-M. * or # indicates statistically significant difference with P < 0.05 between groups. KTM: ketamine, LAA: L-α-aminoadipate.

Figure 2

Effects of drugs administration on SNL-induced NR1 phosphorylation. Phosphorylated NR1 (pNR1) level was much higher in SNL-Saline group than that of Sham-Saline group (* P < 0.05). Intrathecal ketamine (100 μg/kg) down-regulated pNR1 level compared with that of SNL-Saline group. Coadministration further inhibited pNR1 expression than that of individual ketamine injection. Furthermore, pNR1 returned to high level at 1 d after individual ketamine injection. However, pNR1 remained low at 1 d in the coadministration group compared with that of individual ketamine injection. Additionally, LAA application alone has no obvious effects on SNL-induced NR1 phosphorylation. *, # and $ each indicates statistically significant difference with P < 0.05 between groups.

Figure 3

Effects of ketamine and/or LAA on rats' motor performance in rotarod test. Immediately after a baseline response was obtained, saline or drugs were administered intrathecally to normal rats and rotarod test was performed at different time points thereafter (6 rats in each group). The score of each group was normalized as a percentage of the baseline value. Compared with baseline, there was no statistical difference obtained from rotarod test after drugs injection, but intrathecally administered 1000 μg/kg of ketamine caused an obvious defect on rotarod test at 10 min after injection. * P < 0.05 compared with baseline response.

Figure 4

Effects of intrathecal ketamine or LAA on SNL-induced mechanical allodynia, respectively. Intrathecal ketamine elevated paw withdrawal thresholds (PWTs) in a dose-dependent manner (A). *, # P < 0.05 compared with that of Saline or 100 μg/kg of ketamine, respectively. Intrathecal LAA attenuated SNL-induced mechanical allodynia individually in a dose-dependent manner (B). *, # P < 0.05 compared with that of Saline or 50 nmol of LAA, respectively.

Figure 5

Effects of combining ketamine with LAA on SNL-induced mechanical allodynia. PWTs of coadministration group were much higher than that of individual ketamine (100 μg/kg) or LAA (50 nmol) injection at any time points after administration (A). #, $ P < 0.05 compared with that of ketamine 100 μg/kg or LAA 50 nmol, respectively. A', Schematic curves according to A implied that compared with that of individual administration, the anti-allodynia effect of coadministration was more quick, stable and persistent. The analgesic effects of coadministration (100 μg/kg of ketamine and 50 nmol of LAA) compared with that of higher doses of individual ketamine (1000 μg/kg) or LAA (100 nmol), respectively (B). * P < 0.05 compared with that of Ketamine-LAA group. #, $ P < 0.05 compared with that of 1000 μg/kg of ketamine or 100 nmol of LAA, respectively. B', Schematic curves according to B implied that even compared with that of higher doses of individual administration, the anti-allodynia effect of coadministration was still more stable and long-lasting.