Abstract
Vincristine (VCR), an alkaloid isolated from vinca, is a commonly used chemotherapeutic drug. However, VCR therapy can lead to dose-dependent peripheral neurotoxicity, mainly manifesting as neuropathic pain, which is one of the dominant reasons for limiting its utility. Experimentally, we discovered that VCR-induced neuropathic pain (VINP) was accompanied by astrocyte activation; the upregulation of phospho-CaMKII (p-CaMKII), CaV3.2, and Connexin-43 (Cx43) expression; and the production and release of inflammatory cytokines and chemokines in the spinal cord. Similar situations were also observed in astrocyte cultures. Interestingly, these alterations were all reversed by intrathecal injection of KN-93 (a CaMKII inhibitor) or l-Ascorbic acid (a CaV3.2 inhibitor). In addition, KN-93 and l-Ascorbic acid inhibited the increase in [Ca2+]i associated with astrocyte activation. We also verified that knocking down or inhibiting Cx43 level via intrathecal injection of Cx43 siRNA or Gap27 (a Cx43 mimetic peptide) relieved pain hypersensitivity and reduced the release of inflammatory factors; however, they did not affect astrocyte activation or p-CaMKII and CaV3.2 expression. Besides, the overexpression of Cx43 through the transfection of the Cx43 plasmid did not affect p-CaMKII and CaV3.2 expressions in vitro. Therefore, CaMKII and CaV3.2 may activate astrocytes by increasing [Ca2+]i, thereby mediating Cx43-dependent inflammation in VINP. Moreover, we demonstrated that the CaMKII signalling pathway was involved in VCR-induced inflammation, apoptosis, and mitochondrial damage. Collectively, our findings show a novel mechanism by which CaMKII and CaV3.2 mediate Cx43-dependent inflammation by activating astrocytes in neuropathic pain induced by VCR.
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Acknowledgements
The authors acknowledge technical and equipment support for this work provided by the Key Laboratory of Pediatrics, Children’s Hospital of Nanjing Medical University.
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This work was supported by Grants from the National Natural Science Foundation of China (No. 81803642 and 82073845), the China Postdoctoral Science Foundation (No. 2019M651903), the National Major Scientific and Technological Special Project for “Significant New Drugs Development” during the Thirteenth Five-year Plan Period (No. 2018ZX09301043-001), and the Jiangsu Research Hospital Association for Precision Medication (No. JY202010).
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Yunman Li, Yahui Hu, and Feng Chen developed the idea of the study, participated in its design and coordination, and revised the manuscript. Guizhou Li, Yahui Hu, Qingyan Yang, and Di Fu performed the experiments and analysed data. Guizhou Li, Yini Lu, and Yahui Hu drafted the manuscript. All authors read and approved the final manuscript.
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Highlights
1. CaMKII, CaV3.2, and Cx43 play vital roles in neuropathic pain.
2. CaMKII and CaV3.2 activate astrocytes by mediating calcium ions.
3. CaMKII and CaV3.2 facilitate Cx43-mediated inflammatory factor release in spinal dorsal horn astrocytes.
4. CaMKII mediates VCR-induced inflammation and apoptosis.
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Supplementary S1.
(A, B) The PWT and PWL of mice in sham and sham treated with Gap27 (144 nM) groups. (C, D) The PWT and PWL of mice in sham and sham treated with KN-93 (70 nM) groups. (E, F) The PWT and PWL of mice in sham and sham treated with l-Ascorbic acid (300 μM) groups. ANOVA followed by Tukey’s post hoc test. n = 8 mice/group. All data are represented as means ± SEM. (PNG 429 kb)
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Li, Gz., Hu, Yh., Lu, Yn. et al. CaMKII and CaV3.2 T-type calcium channel mediate Connexin-43-dependent inflammation by activating astrocytes in vincristine-induced neuropathic pain. Cell Biol Toxicol 39, 679–702 (2023). https://doi.org/10.1007/s10565-021-09631-y
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DOI: https://doi.org/10.1007/s10565-021-09631-y