Dumitriu Bogdan, Cohen Jonathan E, Wan Qin, Negroiu Andreea M, Abrams Thomas W
Department of Pharmacology, University of Maryland School of Medicine, Baltimore, MD 21201-1559, USA.
J Neurophysiol. 2006 Apr;95(4):2713-20. doi: 10.1152/jn.00642.2005. Epub 2005 Oct 19.
Highly selective serotonin (5-hydroxytryptamine, 5-HT) receptor antagonists developed for mammals are ineffective in Aplysia due to the evolutionary divergence of neurotransmitter receptors and because the higher ionic strength of physiological saline for marine invertebrates reduces antagonist affinity. It has therefore been difficult to identify antagonists that specifically block individual signaling cascades initiated by 5-HT. We studied two broad-spectrum 5-HT receptor antagonists that have been characterized biochemically in Aplysia CNS: methiothepin and spiperone. Methiothepin is highly effective in inhibiting adenylyl cyclase (AC)-coupled 5-HT receptors in Aplysia. Spiperone, which blocks phospholipase C (PLC)-coupled 5-HT receptors in mammals, does not block AC-coupled 5-HT receptors in Aplysia. In electrophysiological studies, we explored whether methiothepin and spiperone can be used in parallel to distinguish between the AC-cAMP and PLC-protein kinase C (PKC) modulatory cascades that are initiated by 5-HT. 5-HT-induced broadening of the sensory neuron action potential in the presence of tetraethylammonium/nifedipine, which is mediated by modulation of the S-K+ currents, was used an assay for the AC-cAMP cascade. Spike broadening initiated by 5 microM 5-HT was unaffected by 100 microM spiperone, whereas it was effectively blocked by 100 microM methiothepin. Facilitation of highly depressed sensory neuron-to-motor neuron synapses by 5-HT was used as an assay for the PLC-PKC cascade. Spiperone completely blocked facilitation of highly depressed synapses by 5 microM 5-HT. In contrast, methiothepin produced a modest, nonsignificant, reduction in the facilitation of depressed synapses. Interestingly, these experiments revealed that the PLC-PKC cascade undergoes desensitization during exposure to 5-HT.
为哺乳动物开发的高选择性血清素（5-羟色胺，5-HT）受体拮抗剂在海兔中无效，这是由于神经递质受体的进化差异，以及海洋无脊椎动物生理盐水中较高的离子强度会降低拮抗剂的亲和力。因此，很难鉴定出能特异性阻断由5-HT引发的单个信号级联反应的拮抗剂。我们研究了两种在海兔中枢神经系统中已进行生化特性鉴定的广谱5-HT受体拮抗剂：甲硫哒嗪和螺哌隆。甲硫哒嗪在抑制海兔中与腺苷酸环化酶（AC）偶联的5-HT受体方面非常有效。在哺乳动物中阻断与磷脂酶C（PLC）偶联的5-HT受体的螺哌隆，在海兔中并不阻断与AC偶联的5-HT受体。在电生理研究中，我们探讨了甲硫哒嗪和螺哌隆是否可同时用于区分由5-HT引发的AC-cAMP和PLC-蛋白激酶C（PKC）调节级联反应。在存在四乙铵/硝苯地平的情况下，5-HT诱导的感觉神经元动作电位展宽由S-K+电流的调节介导，被用作AC-cAMP级联反应的检测方法。由5 microM 5-HT引发的动作电位展宽不受100 microM螺哌隆的影响，而100 microM甲硫哒嗪可有效阻断。5-HT对高度抑制的感觉神经元到运动神经元突触的易化作用被用作PLC-PKC级联反应的检测方法。螺哌隆完全阻断了5 microM 5-HT对高度抑制突触的易化作用。相比之下，甲硫哒嗪对抑制性突触的易化作用产生了适度的、无统计学意义的降低。有趣的是，这些实验表明，PLC-PKC级联反应在暴露于5-HT期间会发生脱敏。
