Institute for Neurophysiology, University of Cologne, Cologne, Germany.
Department of Ophthalmology, Laboratory for Experimental Immunology of the Eye, University of Cologne, Cologne, Germany.
J Neurochem. 2018 Nov;147(3):310-322. doi: 10.1111/jnc.14546. Epub 2018 Aug 30.
Kainic acid (KA) is a potent agonist at non-N-methyl-D-aspartate (non-NMDA) ionotropic glutamate receptors and commonly used to induce seizures and excitotoxicity in animal models of human temporal lobe epilepsy. Among other factors, Ca 2.3 voltage-gated calcium channels have been implicated in the pathogenesis of KA-induced seizures. At physiologically relevant concentrations, endogenous trace metal ions (Cu , Zn ) occupy an allosteric binding site on the domain I gating module of these channels and interfere with voltage-dependent gating. Using whole-cell patch-clamp recordings in human embryonic kidney (HEK-293) cells stably transfected with human Ca 2.3d and β -subunits, we identified a novel, glutamate receptor-independent mechanism by which KA can potently sensitize these channels. Our findings demonstrate that KA releases these channels from the tonic inhibition exerted by low nanomolar concentrations of Cu and produces a hyperpolarizing shift in channel voltage-dependence by about 10 mV, thereby reconciling the effects of Cu chelation with tricine. When tricine was used as a surrogate to study the receptor-independent action of KA in electroretinographic recordings from the isolated bovine retina, it selectively suppressed a late b-wave component, which we have previously shown to be enhanced by genetic or pharmacological ablation of Ca 2.3 channels. Although the pathophysiological relevance remains to be firmly established, we speculate that reversal of Cu -induced allosteric suppression, presumably via formation of stable kainate-Cu complexes, could contribute to the receptor-mediated excitatory effects of KA. In addition, we discuss experimental implications for the use of KA in vitro, with particular emphasis on the seemingly high incidence of trace metal contamination in common physiological solutions.
海人酸（KA）是一种非 NMDA 型离子型谷氨酸受体的有效激动剂，常用于诱导动物颞叶癫痫模型中的癫痫发作和兴奋性毒性。除其他因素外，Ca 2.3 电压门控钙通道已被牵连到 KA 诱导的癫痫发作的发病机制中。在生理相关浓度下，内源性痕量金属离子（Cu 、Zn ）占据这些通道域 I 门控模块的变构结合位点，并干扰电压依赖性门控。我们使用在稳定转染人 Ca 2.3d 和 β 亚基的人胚肾（HEK-293）细胞中进行全细胞膜片钳记录，确定了一种新的、与谷氨酸受体无关的机制，通过该机制，KA 可以强烈敏化这些通道。我们的发现表明，KA 将这些通道从低纳摩尔浓度的 Cu 施加的紧张抑制中释放出来，并使通道电压依赖性产生约 10 mV 的超极化移位，从而使 Cu 螯合与曲利嗪的作用相协调。当曲利嗪被用作研究 KA 在离体牛视网膜视网膜电图记录中与受体无关的作用的替代物时，它选择性地抑制了晚期 b 波成分，我们之前已经表明，该成分通过 Ca 2.3 通道的遗传或药理学消融而增强。尽管病理生理学相关性仍有待确定，但我们推测，Cu 诱导的变构抑制的逆转，可能通过形成稳定的 KA-Cu 复合物，有助于 KA 的受体介导的兴奋性作用。此外，我们讨论了 KA 在体外使用的实验意义，特别强调了常见生理溶液中痕量金属污染的发生率似乎很高。
