突触外膜GABA_A受体对大鼠脊髓背角痛觉突触可塑性的调控作用
发布时间：2019-03-17 07:16
【摘要】：目的:在成年动物,γ-氨基丁酸(GABA)作为重要的抑制性神经递质,可通过激动A-型GABA受体(GABA_A受体),影响神经元的兴奋性。突触中的GABA_A受体介导瞬时抑制(phasic inhibition),而突触外的GABA_A受体则介导持久性抑制(tonic inhibition)。研究显示:突触外的GABA_A受体主要包含α_5亚基(即:α_5-GABA_A受体)和δ亚基(即:δ-GABA_A受体)。大量的实验已经探讨了GABA能瞬时抑制对谷氨酸能突触传递效率的影响。然而,持久性抑制对脊髓背角突触传递和突触可塑性的影响还不明确。本文探讨了GABA能持久性抑制对C纤维诱发的场电位(C fiber-evoked field potentials)长时程增强(Long-Term Potential,LTP)的调控作用。方法:本文通过电生理方法,在大鼠脊髓背角记录C纤维诱发的场电位;低频电刺激(Low Frequency Stimulation,LFS)诱发LTP;脊髓局部给予α_5-GABA_A受体抑制剂L-655,708(20 nM)或δ-GABA_A受体激动剂THIP(2μM),观察LTP的诱导及幅值的变化,研究突触外膜GABA_A受体对痛觉突触传递的调控作用及其机制。结果:(1)给予大鼠坐骨神经2 min的低频电刺激(60 V,0.5-ms duration,2 Hz)可稳定诱发LTP,且LTP至少可以维持3 h。滴加生理盐水不影响正常场电位及LTP的幅值。(2)脊髓局部滴加α_5-GABA_A受体的特异性抑制剂L-655,708(20 nM)可明显增大C纤维的基础电位幅值,提示:α_5-GABA_A受体的去抑制可增强C纤维支配的突触传递。(3)低频电刺激前用L-655,708(20 nM)预处理脊髓1 h,能够促进C纤维诱发场电位LTP的形成、显著提高LTP的幅值,提示:抑制α_5-GABA_A受体可易化LTP的诱导。(4)低频电刺激前,用L-655,708(20 nM)先作用30 min,再给脊髓局部滴加“包含GluN2B亚基的NMDA型谷氨酸受体(GluN2B受体)”的特异性抑制剂ifenprodil(10μg),能够阻断L-655,708对LTP的增强作用,提示:α_5-GABA_A受体可能通过GluN2B受体调控LTP的诱导。(5)L-655,708(20 nM)作用30 min后,脊髓背角滴加“包含GluN2A亚基的NMDA受体(GluN2A受体)”的选择性抑制剂TCN-201(10μM),不影响低频电刺激诱导LTP。(6)低频电刺激30 min和3 h后,分别用L-655,708(20 nM)处理脊髓背角,能使早期和晚期LTP的幅值进一步增强;(7)脊髓滴加δ-GABA_A受体的特异性激动剂THIP(2μM),对C纤维基础电位幅值无明显影响,但能够降低低频电刺激引发的LTP幅值。(8)低频电刺激3 h后再用THIP(2μM)处理脊髓,可显著降低晚期LTP幅值。结论:介导持久性抑制的突触外膜α_5-GABA_A受体和δ-GABA_A受体参与脊髓背角LTP的调节。
[Abstract]:Aim: in adult animals, 纬-aminobutyric acid (GABA), as an important inhibitory neurotransmitter, can affect the excitability of neurons by activating the A-type GABA receptor (GABA_A receptor). GABA_A receptor in synapse mediates transient inhibition of (phasic inhibition), while extrasynaptic GABA_A receptor mediates persistent inhibition of (tonic inhibition). The results show that the extrasynaptic GABA_A receptors mainly contain 伪 _ 5 subunit (伪 _ 5-GABA_A receptor) and 未 subunit (未-GABA_A receptor). A large number of experiments have investigated the effect of transient inhibition of GABA energy on synaptic transmission efficiency of glutamic acid. However, the effects of persistent inhibition on synaptic transmission and synaptic plasticity in spinal dorsal horn are unclear. The regulatory effect of persistent inhibition of GABA on C-fiber-evoked field potentials) evoked field potentials (C-fiber-evoked field potentials) long-term potentiation (Long-Term Potential,LTP) was investigated in this paper. Methods: the field potentials evoked by C fibers were recorded in the dorsal horn of the spinal cord of rats by electrophysiological method, and the LTP; was induced by low frequency electrical stimulation (Low Frequency Stimulation,LFS). Local administration of 伪 _ 5-GABA_A receptor inhibitor L _ 5-GABA_A receptor inhibitor (20 nM) or 未-GABA_A receptor agonist THIP (2 渭 M) was given to the spinal cord to observe the induction and amplitude change of LTP. To study the regulation and mechanism of GABA_A receptor on pain synaptic transmission. Results: (1) low-frequency electrical stimulation (60 V, 0.5 Ms duration,2 Hz) of sciatic nerve (60 V, 0.5 Ms duration,2 Hz) could induce LTP, stably and LTP could be maintained for at least 3 h. The normal field potential and the amplitude of LTP were not affected by the drip of saline. (2) the basal potential amplitude of C fiber was significantly increased by local drip of 伪 _ 5-GABA_A receptor inhibitor L, 655708 (20 nM) in spinal cord. These results suggest that de-inhibition of 伪 _ 5-GABA_A receptor can enhance synaptic transmission innervated by C fibers. (3) pretreatment of spinal cord with low frequency electrical stimulation for 1 hour can promote the formation of evoked field potential (LTP) of C fibers. It was suggested that the inhibition of 伪 _ 5-GABA_A receptor could facilitate the induction of LTP. (4) before low-frequency electrical stimulation, the inhibition of 伪 _ 5-GABA_A receptor could induce the induction of LTP. (4) before low-frequency electrical stimulation, the inhibitory effect of L _ 5-GABA_A receptor on the induction of LTP was 30 min, first. Ifenprodil (10 渭 g), a specific inhibitor of "NMDA glutamic acid receptor (GluN2B receptor) containing GluN2B subunit" was added to the spinal cord to block the enhancement of LTP induced by LA 655708. These results suggest that 伪 _ 5-GABA_A receptor may regulate the induction of LTP through GluN2B receptor. (5) after 30 min treatment with Lac 655708 (20 nM), it is suggested that 伪 _ TNF receptor may regulate the induction of LTP. The addition of selective inhibitor TCN-201 (10 渭 M) to "NMDA receptor (GluN2A receptor) containing GluN2A subunit" in spinal dorsal horn did not affect the low frequency electrical stimulation of LTP. (6) induced by low frequency electrical stimulation for 30 min and 3 h. The amplitude of LTP in the early and late stage of spinal cord could be further enhanced by the treatment of spinal dorsal horn with L, 655708 (20 nM), respectively. (7) THIP (2 渭 M), a specific agonist of 未-GABA_A receptor, did not affect the amplitude of basic potential of C fiber. (8) after 3 hours of low frequency electrical stimulation, the spinal cord was treated with THIP (2 渭 M), and the amplitude of late LTP was decreased significantly. (3) the amplitude of LTP induced by low frequency electrical stimulation was decreased significantly after 3 hours of low frequency electrical stimulation. Conclusion: 伪 _ 5-GABA_A receptor and 未-GABA_A receptor, which mediate persistent inhibition, are involved in the regulation of LTP in spinal dorsal horn.
【学位授予单位】：兰州大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R402
[Abstract]:Aim: in adult animals, 纬-aminobutyric acid (GABA), as an important inhibitory neurotransmitter, can affect the excitability of neurons by activating the A-type GABA receptor (GABA_A receptor). GABA_A receptor in synapse mediates transient inhibition of (phasic inhibition), while extrasynaptic GABA_A receptor mediates persistent inhibition of (tonic inhibition). The results show that the extrasynaptic GABA_A receptors mainly contain 伪 _ 5 subunit (伪 _ 5-GABA_A receptor) and 未 subunit (未-GABA_A receptor). A large number of experiments have investigated the effect of transient inhibition of GABA energy on synaptic transmission efficiency of glutamic acid. However, the effects of persistent inhibition on synaptic transmission and synaptic plasticity in spinal dorsal horn are unclear. The regulatory effect of persistent inhibition of GABA on C-fiber-evoked field potentials) evoked field potentials (C-fiber-evoked field potentials) long-term potentiation (Long-Term Potential,LTP) was investigated in this paper. Methods: the field potentials evoked by C fibers were recorded in the dorsal horn of the spinal cord of rats by electrophysiological method, and the LTP; was induced by low frequency electrical stimulation (Low Frequency Stimulation,LFS). Local administration of 伪 _ 5-GABA_A receptor inhibitor L _ 5-GABA_A receptor inhibitor (20 nM) or 未-GABA_A receptor agonist THIP (2 渭 M) was given to the spinal cord to observe the induction and amplitude change of LTP. To study the regulation and mechanism of GABA_A receptor on pain synaptic transmission. Results: (1) low-frequency electrical stimulation (60 V, 0.5 Ms duration,2 Hz) of sciatic nerve (60 V, 0.5 Ms duration,2 Hz) could induce LTP, stably and LTP could be maintained for at least 3 h. The normal field potential and the amplitude of LTP were not affected by the drip of saline. (2) the basal potential amplitude of C fiber was significantly increased by local drip of 伪 _ 5-GABA_A receptor inhibitor L, 655708 (20 nM) in spinal cord. These results suggest that de-inhibition of 伪 _ 5-GABA_A receptor can enhance synaptic transmission innervated by C fibers. (3) pretreatment of spinal cord with low frequency electrical stimulation for 1 hour can promote the formation of evoked field potential (LTP) of C fibers. It was suggested that the inhibition of 伪 _ 5-GABA_A receptor could facilitate the induction of LTP. (4) before low-frequency electrical stimulation, the inhibition of 伪 _ 5-GABA_A receptor could induce the induction of LTP. (4) before low-frequency electrical stimulation, the inhibitory effect of L _ 5-GABA_A receptor on the induction of LTP was 30 min, first. Ifenprodil (10 渭 g), a specific inhibitor of "NMDA glutamic acid receptor (GluN2B receptor) containing GluN2B subunit" was added to the spinal cord to block the enhancement of LTP induced by LA 655708. These results suggest that 伪 _ 5-GABA_A receptor may regulate the induction of LTP through GluN2B receptor. (5) after 30 min treatment with Lac 655708 (20 nM), it is suggested that 伪 _ TNF receptor may regulate the induction of LTP. The addition of selective inhibitor TCN-201 (10 渭 M) to "NMDA receptor (GluN2A receptor) containing GluN2A subunit" in spinal dorsal horn did not affect the low frequency electrical stimulation of LTP. (6) induced by low frequency electrical stimulation for 30 min and 3 h. The amplitude of LTP in the early and late stage of spinal cord could be further enhanced by the treatment of spinal dorsal horn with L, 655708 (20 nM), respectively. (7) THIP (2 渭 M), a specific agonist of 未-GABA_A receptor, did not affect the amplitude of basic potential of C fiber. (8) after 3 hours of low frequency electrical stimulation, the spinal cord was treated with THIP (2 渭 M), and the amplitude of late LTP was decreased significantly. (3) the amplitude of LTP induced by low frequency electrical stimulation was decreased significantly after 3 hours of low frequency electrical stimulation. Conclusion: 伪 _ 5-GABA_A receptor and 未-GABA_A receptor, which mediate persistent inhibition, are involved in the regulation of LTP in spinal dorsal horn.
【学位授予单位】：兰州大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R402
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