Role of CaMK II α in the Injury of Dorsal Root Ganglion Neurons Induced by Ropivacaine Hydrochloride in the Dorsal Root Ganglion of Rats

Abstract

Objective: To investigate whether CaMKⅡα participates in the dorsal root ganglion neurotoxicity induced by ropivacaine hydrochloride.
 Methods: DRG neurons were isolated from 1-day-old SD rats and cultured in vitro. pAd-shRNA-CaMKⅡα-DRG cells were constructed by RNA interference technique to inhibit the expression of CaMKⅡα. The experiment was divided into six groups: DRG group (DRG group), vector DRG group (vector group), pAd-shRNA- CaMKIIα-DRG group (pAd-shRNA group), DRG + ropivacaine group (DRG + R group), vector DRG + ropivacaine group (vector + R group), pAd-shRNA-CaMKII α - DRG + ropivacaine group (pAd-shRNA + R group), and the last three groups were treated with 3 mM ropivacaine hydrochloride for 4 hours. MTT assay was used to detect cell viability, flow cytometry was used to detect cell apoptosis rate, laser confocal microscopy was used to detect intracellular calcium level, and real-time PCR was used to detect the mRNA expression of CaMKⅡα, Cav3.2 and Cav3.3.
 Results: The cell viability of DRG+R group, vector+R group and pAd-shRNA+R group decreased significantly after 3 mM ropivacaine hydrochloride treatment for 4 h. Compared with DRG+R group, the cell viability of pAd-shRNA+R group was significantly higher. After 3 mM ropivacaine hydrochloride treatment for 4 h, the apoptosis rate of DRG + R group, vector + R group and pAd-shRNA + R group increased significantly. Compared with DRG+R group, the apoptosis rate in pAd shRNA+R group was significantly lower. After 3 mM ropivacaine hydrochloride treatment for 4 h, the intracellular calcium levels in DRG + R group, vector + R group and pAd-shRNA + R group were significantly increased, and the intracellular calcium levels in pAd-shRNA + R group were significantly lower than those in DRG + R group. The mRNA expressions of CaMKⅡα, Cav3.2 and Cav3.3 were significantly decreased in pAd- shRNA group. The mRNA expressions of CaMK Ⅱ α, Cav3.2 and Cav3.3 were up-regulated in DRG + R group, vector + R group and pAd-shRNA + R group after 3 mm ropivacaine treatment for 4 h. The mRNA expressions of CaMKⅡα, Cav3.2 and Cav3.3 in pAd-shRNA + R group were significantly lower than those in DRG + R group.
 Conclusion: Inhibition of CaMKⅡα expression can down regulate the expression of Cav3.2 and Cav3.3 mRNA, increase cell viability of DRG neurons, reduce the apoptosis rate, and improve the dorsal root ganglion neurotoxicity induced by ropivacaine hydrochloride.