Nociceptin receptor coupling to a potassium conductance in rat locus coeruleus neurones in vitro
Nociceptin receptor coupling to a potassium conductance in rat locus coeruleus neurones in vitro
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DOI：
10.1111/j.1476-5381.1996.tb16080.x
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发表时间：
1996-12-01
影响因子：
7.3
通讯作者：
Christie, MJ
中科院分区：
文献类型：
--
作者：
Connor, M;Vaughan, CW;Christie, MJ
1在这项研究中，我们使用细胞内和全细胞膜片钳记录，检测了孤啡肽（阿片样受体ORL的内源性配体）对体外大鼠蓝斑（LC）神经元膜特性的影响。2当蓝斑神经元电压钳位至-60 mV时，孤啡肽的应用在所有检测的细胞中引起外向电流（n=49），EC（50）为90 nM。在存在肽酶抑制剂bestatin（20 μ M）或thiorphan（2 μ M）的情况下，伤害感受素的效力和最大效应均未改变。3在2.5 mM细胞外K+中由伤害感受素引起的外向电流在-123 mV时极性反转，比预测的-105 mV K+反转电位更负。将细胞外K+增加至6.5 mM导致+25 mV的逆转电位偏移，该偏移与K+电导一致。由伤害感受素激活的电导显示出轻微的内向整流。4应用高浓度的伤害感受素（3 μ M）阻断了由同时应用高浓度的甲硫氨酸-脑啡肽（10 μ M）、（3 μ M）生长抑素和UK 14304（3 μ M）产生的电流，表明痛敏素激活与μ-阿片样物质和生长抑素受体以及α（2）-肾上腺素受体相同的传导性。痛敏素的作用被阿片样物质拮抗剂微弱地拮抗，纳洛酮，pK（B）由2个细胞估算，分别为-4.23和-4.33。μ阿片拮抗剂CTAP（D-Phe-Cys-Tyr-D-Trp-Arg-Pen-Thr-NH 2，1 μ M）、阿片样物质拮抗剂纳洛啡（30 μ M）或生长抑素拮抗剂CPP（环（7-氨基庚酰基-Phe-D-Trp-Lys-Thr[Bz 1]）3 μ M）不影响痛敏肽诱导的电流。6强啡肽A（3 μ M），另一种假定的ORL内源性配体（1），在蓝斑神经元中引起了强烈的外向电流，然而，可被中等浓度纳洛酮完全拮抗（300 nM-1 μ M）。7连续应用痛敏肽（3 μ M）导致外向电流降低至最大响应的70%的稳定水平，t（1/2）120 s的。脱敏在很大程度上是同源的，因为同时应用甲硫氨酸脑啡肽（30 μ M）在脱敏期间的伤害感受素反应导致的外向电流是92%的控制反应，甲硫氨酸脑啡肽在相同的细胞。相反，持续应用甲硫氨酸脑啡肽（30 μ M）导致甲硫氨酸脑啡肽电流下降到稳定水平，为初始电流的54%。在此脱敏期间，应用伤害感受素（3 μ M）导致的电流是相同细胞中对伤害感受素的对照反应的78%。8因此，伤害感受素有效地激活蓝斑神经元中的内向整流K+电导，其药理学特征与ORL（1）受体的激活一致。强啡肽A似乎不是大鼠蓝斑神经元中ORL（1）的配体。
1 In this study we have examined the effects of nociceptin, an endogenous ligand for the opioid-like receptor ORL(1), on the membrane properties of rat locus coeruleus (LC) neurones in vitro, using intracellular and whole cell patch clamp recording.2 When locus coeruleus neurones were voltage clamped to -60 mV, application of nociceptin caused an outward current in all cells examined (n=49), with an EC(50) of 90 nM. Neither the potency nor the maximal effect of nociceptin was altered in the presence of the peptidase inhibitors, bestatin (20 mu M) or thiorphan (2 mu M).3 The outward currents caused by nociceptin in 2.5 mM extracellular K+ reversed polarity at -123 mV, more negative than the predicted K+ reversal potential of -105 mV. Increasing extracellular K+ to 6.5 mM resulted in a shift of the reversal potential of +25 mV, a shift consistent with a K+ conductance. The conductance activated by nociceptin showed mild inward rectification.4 Application of a high concentration of nociceptin (3 mu M) occluded the current produced by simultaneous application of high concentrations of Met-enkephalin (10 mu M), (3 mu M) somatostatin and UK 14304 (3 mu M), indicating that nociceptin activated the same conductance as mu-opioid and somatostatin receptors and alpha(2)-adrenoceptors.5 The actions of nociceptin were weakly antagonized by the opioid antagonist, naloxone, with pK(b)'s estimated from 2 cells of -4.23 and -4.33. The mu-opioid antagonist, CTAP (D-Phe-Cys-Tyr-D-Trp-Arg-Pen-Thr-NH2, 1 mu M), the opioid antagonist, nalorphine (30 mu M), or the somatostatin antagonist, CPP (cyclo(7-aminoheptanoyl-Phe-D-Trp-Lys-Thr[Bz1]) 3 mu M) did not affect the nociceptin-induced current.6 Dynorphin A (3 mu M), another putative endogenous ligand for ORL(1), caused a robust outward current in locus coeruleus neurones that was, however, completely antagonized by moderate concentrations of naloxone (300 nM-1 mu M).7 Continuous application of nociceptin (3 mu M) resulted in a decrease of the outward current to a steady level of 70% of the maximum response with a t(1/2) of 120s. Desensitization was largely homologous because simultaneous application of Met-enkephalin (30 mu M) during the desensitized period of the nociceptin response resulted in an outward current that was 92% of control responses to Met-enkephalin in the same cells. Conversely, continuous application of Met-enkephalin (30 mu M) resulted in a decrease of Met-enkephalin current to a steady level that was 54% of the initial current. During this desensitized period application of nociceptin (3 mu M) resulted in a current that was 78% of the control responses to nociceptin in the same cells.8 Thus nociceptin potently activates an inwardly rectifying K+ conductance in locus coeruleus neurones, with a pharmacological profile consistent with activation of the ORL(1) receptor. Dynorphin A does not appear to be a ligand for ORL(1) in rat locus coeruleus neurones.