鞘内注射PSD-93反义寡核苷酸对神经病理性痛大鼠脊髓nNOS的影响

Effects of intrathecal PSD-93 antisense oligonucleotide on meuronal nitric oxide synthase in spinal cord in a rat model of neuropathic pain

目的探讨鞘内注射PSD-93反义寡核苷酸对神经病理性痛大鼠脊髓神经型一氧化氮合酶（nNOS）的影响。方法雄性SD大鼠32只，体重250～350g，随机分为4组（n=8）：假手术＋生理盐水组（C组）；C，脊神经压迫＋生理盐水组（N组）；C7脊神经压迫＋PSD-93误义寡核苷酸10μg组（M组）；C7脊神经压迫＋PSD-93反义寡核苷酸10μg组（A组）。N组、M组和A组采用60g微血管夹压迫大鼠右侧C7脊神经15min制备神经病理性痛模型，经枕骨大孔鞘内置管至颈膨大处。术毕当日开始给药，每日1次，连续4d。于术前2d（T0）、术后1、3、5和7d（T1-4）时测定机械痛阈和热痛阈，术后7d处死大鼠取C7段脊髓，免疫组化法检测神经压迫侧脊髓PSD-93蛋白和nNOS的表达。结果与Tn时比较，N组和M组各时点机械痛阈及热痛阈降低（P〈0．05）；与C组比较，N组和M组各时点机械痛阈及热痛阈降低，N组、M组和A组脊髓背角PSD-93蛋白和nNOS表达上调（P〈0．05）；与N组和M组比较，A组各时点机械痛阈及热痛闽升高，脊髓背角PSD-93蛋白和nNOS表达下调（P〈0．05）。结论鞘内注射PSD-93反义寡核苷酸可抑制神经病理性痛大鼠脊髓nNOS表达，nNOS在神经病理性痛中的作用可能受PSD-93蛋白调节。

Objective To evaluate the effects of intrathecal （IT） PSD-93 antisense oligonucleotide （ASODN） on neuronal nitric oxide synthase （nNOS） in the spinal cord in a rat model of neuropathic pain. Methods Thirty-two male SD rats weighing 250-350 g were randomly divided into 4 groups （ n = 8 each） ： group Ⅰ sham operation; group Ⅱ C7 spinal nerve compression （C7 SNC ）; group Ⅲ C7 SNC ＋ mismatch oligonucleotide （MSODN） 10 μg and group Ⅳ C7 SNC ＋ ASODN 10 μg. The animals were anesthetized with intraperitoneal 10% chloral hydrate 350 mg/kg. Neuropathic pain was produced by clamping the right C7 spinal nerve for 15 min with a 60 g mini-clamp （C7 SNC）. A catheter was inserted into the subarachnoid space via foramen magnum. The tip of the catheter roached cervical enlargement. MSODN and ASODN were administered IT once a day for 4 consecutive days after operation in group Ⅲ and Ⅳ respectively. In group Ⅰ and Ⅱ normal saline （NS） was injected IT instead. Paw withdrawal threshold （PWT） to noxious thermal and mechanical stimuli was measured 2 days before surgery （To baseline） and on 1st, 3rd, 5th and 7th day after operation （T1-4 ）. All rats were killed on the 7th day after operation and the C7 segment of the spinal cord was removed for determination of the expression of PSD-93 protein and nNOS in the spinal cord on the SNC side by immuno-histochemistry. Results The mechanical and thermal pain threshold at T1-4 were significantly decreased as compared with the baseline values at To in group Ⅱ and Ⅲ and were significantly lower than those in group Ⅰ. The expression of PSD-93 protein and nNOS in the dorsal horn of the spinal cord on the SNC side was significantly higher in group Ⅱ ,Ⅲ and Ⅳ than in group Ⅰ but was significantly decreased by IT ASODN in group Ⅳ as compared with group Ⅱ and Ⅲ. Conclusion Intrathecal PSD-93 ASODN can inhibit the expression of nNOS in the spinal cord in rats with neuropathic pain. The role of nNOS in reducing neuropathic pain may be modulated by PSD-93 protein.