神经病理性疼痛的背根神经节炎性机制和交感机制研究
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摘要
目的：背根神经节(dorsal root ganglia, DRG)的炎症反应在病理性疼痛的发生和维持中发挥重要作用。在大鼠脊神经结扎模型(spinal nerve tight ligation, SNL)全身应用糖皮质激素氟羟强的松龙(triamcinolone acetonide, TA)能够抑制机械性疼痛行为,减少DRG内交感神经芽生,降低细胞因子和神经生长因子的生成,降低自发放电的频率。而DRG局部炎症模型却和糖皮质激素的作用恰恰相反。因此我们假设并试图验证全身应用糖皮质激素的镇痛机制主要来源于其在DRG水平的作用。
交感神经活动能加剧多种慢性疼痛例如复杂区域性疼痛综合症。在动物模型中,交感神经纤维在外周神经损伤后芽生入DRG内,与感觉神经元建立起异常联系。本研究旨在研究切断发入腰DRG的交感神经灰交通支对神经病理性疼痛模型机械性痛觉过敏的镇痛作用。
方法：成年雄性Sprague-Dawley大鼠根据术前平均基础痛阈值随机分为4组：SNL组、SNL+TA组、Normal+TA组和Normal对照组。在SNL+TA组和Normal+TA组,在SNL手术或假手术同时局部注射20μ1TA悬浮注射液于L5DRG表面和周围区域。于术后第1,3,5和7天测量机械性痛阈。运用免疫组化手段检测DRG酪氨酸羟化酶(TH),胶质细胞原纤维酸性蛋白(GFAP),神经生长因子(NGF),脊髓CD11B抗体(OX-42)水平。
Sprague-Dawley大鼠予以L5脊神经腹支结扎术。部分实验大鼠在脊神经结扎术前三周或同时,离断交感神经灰交通支。运用免疫组化手段检测DRG酪氨酸羟化酶(TH)水平。测量大鼠机械性痛阈。
结果：在脊神经损伤的同时在腰DRG附近注射糖皮质激素TA能改善机械疼痛过敏,减少DRG交感神经篮状结构形成,降低DRG卫星胶质细胞激活水平和脊髓小胶质细胞激活水平,降低DRG内NGF表达水平。
在脊神经腹支结扎术前三周离断发入L5DRG的灰支,显著降低了手术后3天损伤DRG的交感神经芽生。在脊神经切断术同时离断灰支也能降低机械性疼痛。
结论：本试验证实单剂量局部注射糖皮质激素TA于DRG表面及附近组织的临床应用能够模拟全身应用TA在SNL模型中的镇痛效果。这进一步为临床局部应用糖皮质激素治疗某些临床疾病提供了合理的基础。进一步证明DRG水平的局部炎症是脊神经结扎疼痛模型的重要机制,尽管其归类为神经病理性疼痛。本研究也为疼痛行为与DRG和脊髓胶质细胞激活之间的相关性提供了证据。本试验证实单剂量局部注射糖皮质激素TA能降低SNL模型术后第3天DRG内NGF表达水平,可能是糖皮质激素镇痛的机制之一。这与糖皮质激素减少交感神经芽生和胶质细胞激活的结果相吻合,为糖皮质激素临床疼痛治疗提供了进一步理论基础。
在普遍使用的脊神经结扎模型早期,脊神经背支是DRG交感神经芽生的主要来源。提前离断发入L5DRG的灰支手术,比外科或化学性交感神经切断术更少损伤,能显著降低脊神经结扎术导致的机械性疼痛。脊神经结扎同时离断发入L5DRG的灰支手术也能降低脊神经结扎术导致的机械性疼痛,效果有延迟,但对临床更有意义。
BACKGROUND:Inflammatory responses in the lumbar dorsal root ganglion (DRG) play a key role in the development of a variety of pathologic pain states. Systemic administration of a commonly used anti-inflammatory corticosteroid, triamcinolone acetonide (TA), reduces sympathetic sprouting, mechanical pain behavior, cytokine and nerve growth factor production in the DRG, and incidence of spontaneous bursting activity. We hypothesize that the observed effects of systemic TA are primarily due to local effects at the level of the DRG.
Many chronic pain conditions including complex regional pain syndrome are exacerbated by sympathetic activity. In animal models, sympathetic fibers sprout into the dorsal root ganglia (DRG) after peripheral nerve injury, forming abnormal connections with sensory neurons. We hypothesize that cutting the grey ramus to the L5 DRG reduces mechanical pain behaviors in the SNL model.
METHODS:Male Sprague-Dawley rats were used for all experiments. Rats were randomly divided into 4 groups:SNL (tight ligation of spinal nerves), SNL+TA, Normal+TA, and Normal. In normal rats (Normal+TA) and in SNL rats (SNL+TA), a single dose of 20μ1 TA injectable suspension (10mg/ml) was slowly injected onto the surface of DRG and and surrounding region at the time of SNL or sham surgery. Cutaneous sensitivity to mechanical stimulation was tested on postoperative days 1,3,5, and 7. Immunohistochemical staining was performed to examine tyrosine hydroxylase (TH), glial fibrillary acidic protein (GFAP) and nerve growth factor (NGF) in DRG, and CD11B antibody (OX-42) in spinal cord.
Male Sprague-Dawley rats received a unilateral ligation of the ventral ramus of the L5 spinal nerve following the original description by Kim and Chung. In some experiments, the grey ramus located in close proximity to the DRG was isolated and cut, either at the time of, or three weeks before, the spinal nerve ligation surgery. Immunohistochemical staining was performed to examine tyrosine hydroxylase (TH) in DRG. Cutaneous sensitivity to mechanical stimulation was tested.
RESULTS:Local TA treatment attenuated mechanical sensitivity, reduced sympathetic sprouting in the DRG, and decreased satellite glia activation, NGF expression level in the DRG and microglia activation in the spinal cord after SNL. Cutting the grey ramus to the L5 DRG reduces mechanical sensitivity in the SNL model.
CONCLUSION:This study demonstrates that a single injection of in the vicinity of the axotomized DRG can mimic many of the effects of systemic TA in mitigating the behavioral and cellular abnormalities induced by spinal nerve ligation. This provides a further rational basis for the clinical use of localized steroid injections in some clinical conditions, and provides further support for the idea that localized inflammation at the level of the DRG is an important component of the spinal nerve ligation model even though this is commonly classified as neuropathic pain model. And the study provides further evidence of a correlation between pain behaviors and activation of glia in the DRG and spinal cord. This study demonstrates that a single injection of in the vicinity of the axotomized DRG can decrease NGF expression level in the DRG and likely one of mechanisms of local TA treatment attenuate mechanical sensitivity. This is consistent with the result that TA can reduce sympathetic sprouting and decrease satellite glia activation. And provides further theory basis of corticosteroid clinical application for pain treatment.
This study presented evidence that the dorsal ramus of the spinal nerve is a source of sympathetic fibers that sprout into the DRG during early phases of the commonly used spinal nerve ligation model. Pre-cutting the grey ramus to the L5 DRG, a much less invasive procedure than surgical or chemical sympathectomy, markedly reduces the mechanical pain induced by spinal nerve ligation. More relevant to clinical situations, cutting the grey ramus also reduced mechanical pain (albeit with a delay) when done at the time of spinal nerve ligation.
交感神经活动能加剧多种慢性疼痛例如复杂区域性疼痛综合症。在动物模型中,交感神经纤维在外周神经损伤后芽生入DRG内,与感觉神经元建立起异常联系。本研究旨在研究切断发入腰DRG的交感神经灰交通支对神经病理性疼痛模型机械性痛觉过敏的镇痛作用。
方法：成年雄性Sprague-Dawley大鼠根据术前平均基础痛阈值随机分为4组：SNL组、SNL+TA组、Normal+TA组和Normal对照组。在SNL+TA组和Normal+TA组,在SNL手术或假手术同时局部注射20μ1TA悬浮注射液于L5DRG表面和周围区域。于术后第1,3,5和7天测量机械性痛阈。运用免疫组化手段检测DRG酪氨酸羟化酶(TH),胶质细胞原纤维酸性蛋白(GFAP),神经生长因子(NGF),脊髓CD11B抗体(OX-42)水平。
Sprague-Dawley大鼠予以L5脊神经腹支结扎术。部分实验大鼠在脊神经结扎术前三周或同时,离断交感神经灰交通支。运用免疫组化手段检测DRG酪氨酸羟化酶(TH)水平。测量大鼠机械性痛阈。
结果：在脊神经损伤的同时在腰DRG附近注射糖皮质激素TA能改善机械疼痛过敏,减少DRG交感神经篮状结构形成,降低DRG卫星胶质细胞激活水平和脊髓小胶质细胞激活水平,降低DRG内NGF表达水平。
在脊神经腹支结扎术前三周离断发入L5DRG的灰支,显著降低了手术后3天损伤DRG的交感神经芽生。在脊神经切断术同时离断灰支也能降低机械性疼痛。
结论：本试验证实单剂量局部注射糖皮质激素TA于DRG表面及附近组织的临床应用能够模拟全身应用TA在SNL模型中的镇痛效果。这进一步为临床局部应用糖皮质激素治疗某些临床疾病提供了合理的基础。进一步证明DRG水平的局部炎症是脊神经结扎疼痛模型的重要机制,尽管其归类为神经病理性疼痛。本研究也为疼痛行为与DRG和脊髓胶质细胞激活之间的相关性提供了证据。本试验证实单剂量局部注射糖皮质激素TA能降低SNL模型术后第3天DRG内NGF表达水平,可能是糖皮质激素镇痛的机制之一。这与糖皮质激素减少交感神经芽生和胶质细胞激活的结果相吻合,为糖皮质激素临床疼痛治疗提供了进一步理论基础。
在普遍使用的脊神经结扎模型早期,脊神经背支是DRG交感神经芽生的主要来源。提前离断发入L5DRG的灰支手术,比外科或化学性交感神经切断术更少损伤,能显著降低脊神经结扎术导致的机械性疼痛。脊神经结扎同时离断发入L5DRG的灰支手术也能降低脊神经结扎术导致的机械性疼痛,效果有延迟,但对临床更有意义。
BACKGROUND:Inflammatory responses in the lumbar dorsal root ganglion (DRG) play a key role in the development of a variety of pathologic pain states. Systemic administration of a commonly used anti-inflammatory corticosteroid, triamcinolone acetonide (TA), reduces sympathetic sprouting, mechanical pain behavior, cytokine and nerve growth factor production in the DRG, and incidence of spontaneous bursting activity. We hypothesize that the observed effects of systemic TA are primarily due to local effects at the level of the DRG.
Many chronic pain conditions including complex regional pain syndrome are exacerbated by sympathetic activity. In animal models, sympathetic fibers sprout into the dorsal root ganglia (DRG) after peripheral nerve injury, forming abnormal connections with sensory neurons. We hypothesize that cutting the grey ramus to the L5 DRG reduces mechanical pain behaviors in the SNL model.
METHODS:Male Sprague-Dawley rats were used for all experiments. Rats were randomly divided into 4 groups:SNL (tight ligation of spinal nerves), SNL+TA, Normal+TA, and Normal. In normal rats (Normal+TA) and in SNL rats (SNL+TA), a single dose of 20μ1 TA injectable suspension (10mg/ml) was slowly injected onto the surface of DRG and and surrounding region at the time of SNL or sham surgery. Cutaneous sensitivity to mechanical stimulation was tested on postoperative days 1,3,5, and 7. Immunohistochemical staining was performed to examine tyrosine hydroxylase (TH), glial fibrillary acidic protein (GFAP) and nerve growth factor (NGF) in DRG, and CD11B antibody (OX-42) in spinal cord.
Male Sprague-Dawley rats received a unilateral ligation of the ventral ramus of the L5 spinal nerve following the original description by Kim and Chung. In some experiments, the grey ramus located in close proximity to the DRG was isolated and cut, either at the time of, or three weeks before, the spinal nerve ligation surgery. Immunohistochemical staining was performed to examine tyrosine hydroxylase (TH) in DRG. Cutaneous sensitivity to mechanical stimulation was tested.
RESULTS:Local TA treatment attenuated mechanical sensitivity, reduced sympathetic sprouting in the DRG, and decreased satellite glia activation, NGF expression level in the DRG and microglia activation in the spinal cord after SNL. Cutting the grey ramus to the L5 DRG reduces mechanical sensitivity in the SNL model.
CONCLUSION:This study demonstrates that a single injection of in the vicinity of the axotomized DRG can mimic many of the effects of systemic TA in mitigating the behavioral and cellular abnormalities induced by spinal nerve ligation. This provides a further rational basis for the clinical use of localized steroid injections in some clinical conditions, and provides further support for the idea that localized inflammation at the level of the DRG is an important component of the spinal nerve ligation model even though this is commonly classified as neuropathic pain model. And the study provides further evidence of a correlation between pain behaviors and activation of glia in the DRG and spinal cord. This study demonstrates that a single injection of in the vicinity of the axotomized DRG can decrease NGF expression level in the DRG and likely one of mechanisms of local TA treatment attenuate mechanical sensitivity. This is consistent with the result that TA can reduce sympathetic sprouting and decrease satellite glia activation. And provides further theory basis of corticosteroid clinical application for pain treatment.
This study presented evidence that the dorsal ramus of the spinal nerve is a source of sympathetic fibers that sprout into the DRG during early phases of the commonly used spinal nerve ligation model. Pre-cutting the grey ramus to the L5 DRG, a much less invasive procedure than surgical or chemical sympathectomy, markedly reduces the mechanical pain induced by spinal nerve ligation. More relevant to clinical situations, cutting the grey ramus also reduced mechanical pain (albeit with a delay) when done at the time of spinal nerve ligation.
引文
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