Modulation of synaptic transmission as the therapeutic strategies for chronic pain

1.为阐明加巴喷丁的脊髓上作用机制，采用全细胞膜片钳技术，对周围神经损伤小鼠和假手术小鼠脑干脑片蓝斑(LC)神经元进行了全细胞膜片钳记录，研究了加巴喷丁对突触传递的影响。加巴喷丁(10-100μM)可浓度依赖性地减少部分神经结扎小鼠脑片中GABA受体介导的IPSCs(Seltzer模式)，而谷氨酸介导的EPSCs几乎不受影响。相比之下，加巴喷丁没有减少从假手术小鼠身上提取的切片中的IPSCs。由于加巴喷丁减少了IPSCs，增加了成对脉冲比率，它作用于LC内的突触前GABA能神经末梢。此外，蛋白激酶A抑制剂11-89而不是蛋白激酶C抑制剂白屈菜红碱可取消加巴喷丁对…的抑制作用，提示PICA介导的磷酸化似乎影响了IPSCs加巴喷丁对脊髓上加巴喷丁的反应性在神经病理性条件下更为重要。加巴喷丁共同产生PICA依赖的突触前抑制GABA能突触传递，从而消除仅在神经病理性疼痛状态下对LC神经元的抑制影响(去甲肾上腺素下行系统的去抑制)。我们探讨了外源性或通过阻断甘氨酸转运蛋白1(GlyT1)选择性抑制剂肌氨酸和NFPS增加脊髓甘氨酸对小鼠神经病理性和炎症性疼痛的治疗效果。甘氨酸、肌氨酸和NFPS可改善Seltzer模型小鼠的热和机械超敏反应，降低STZ注射糖尿病小鼠的机械超敏反应。此外，它们还选择性地抑制福尔马林引起的舔咬行为的第二阶段。在Seltzer模型小鼠的海马片中，LTP维持在显著低于假手术组的水平。当在NFPS存在的情况下诱导LTP时，从未观察到这种损伤。总之，通过GlyT1阻滞剂增加内源性甘氨酸不仅在脊髓水平对疼痛传递产生净抑制影响，而且还在棘上缓解突触效能下降，这可能与慢性疼痛住院患者经常描述的认知障碍有关。在初级传入纤维与背角浅层神经元之间有关痛传递的药理学和生理学研究中，我们使用了大鼠脊髓切片。然而，使用小鼠脊髓切片连接背根进行突触传递很难得到评估。我们将这种大鼠脊髓切片制备方法应用于成年小鼠，并探讨了周围神经损伤后，HCN通道是否参与了A-δ和C-纤维介导的EPSCs。与我们的行为学研究表明鞘内注射HCN通道阻滞剂ZD7288降低Seltzer模型小鼠的热和机械超敏反应一致，7288减少了Aδ和C纤维介导的EPSCs。较少

1. To clarify the supraspinal mechanism of action of gabapentin, whole-cell patch-clamp recordings were performed on locus coeruleus (LC) neurons in brainstem slices prepared from mice after peripheral nerve injury or mice subjected to a sham operation, and the effects of gabapentin on synaptic transmission were studied. Gabapentin (10-100 μM) concentration-dependently reduced the GABAAreceptor-mediated IPSCs in slices prepared from partial nerve-ligated mice (the Seltzer mode)), whereas glutamate-mediated EPSCs were hardly affected. By contrast, gabapentin did not reduce IPSCs in slices taken from mice given a sham operation. Since gabapentin reducedIPSCs together with an increase in the paired-pulse ratio, it acts on the presynaptic GABAergic nerve terminals in the LC. Moreover, the protein kinase A inhibitor 11-89 but not the protein kinase C inhibitor chelerythrine abolished the inhibitory action of gabapentin on IPSCs, suggesting that PICA-mediated phosphorylation seems to be impo … More rtant for supraspinal gabapentin responsiveness in neuropathic conditions.Together, gabapentin generates PICA-dependent presynaptic inhibition of GABAergic synaptic transmission, and thereby removes the inhibitory influence on LC neurons (disinhibition of the descending noradrenergic system) only under neuropathic pain states.2. We addressed the therapeutic impact of increasing spinal glycine applied exogenously or via blockade of glycine transporter 1 (GlyT1) using its selective inhibitors sarcosine and NFPS on neuropathic and inflammatory pain in mice. Glycine, sarcosine and NFPS ameliorated thermal and mechanical hypersensitivity in Seltzer model mice, and reduced mechanical hypersensitivity in STZ injected diabetic mice. Moreover, they selectively inhibited the second phase of formalin evoked licking/biting behavior. In hippocampal slices prepared from Seltzer model mice, LTP was maintained at a significantly lower level than that in sham-treated mice. Such impairment of LTP was never observed when it was induced in the presence of NFPS. Together, an increase in endogenous glycine via GlyT1 blockade not only results in a net inhibitory influence on pain transmission at the spinal level but also supraspinally relieves decreased synaptic efficacy presumably related to cognitive disturbance often described inpatients with chronic pain.3. In the pharmacological and physiological studies related with pain transmission between primary afferent fibers and superficial dorsal horn neurons, spinal cord slices with attached dorsal roots prepared from rats have been used. However, synaptic transmission using mice spinal cord slices attached dorsal roots is hardly evaluated. We adapted this rat spinal slice preparation to adult mice, and addressed whether HCN channels contribute to Aδ-and C-fiber-mediated EPSCs after peripheral nerve injury. Consistent with our behavioral study demonstrating that intrathecal injection of the HCN channel blocker ZD7288 reduced thermal and machanical hypersensitivity in Seltzer model mice, 7288 reduced Aδ-and C-fiber-mediated EPSCs. Less