Gylys K H, Mellin C, Amstutz R, Jenden D J
Department of Pharmacology, University of California School of Medicine, Los Angeles 90024.
J Neurochem. 1992 Oct;59(4):1302-8. doi: 10.1111/j.1471-4159.1992.tb08441.x.
The inhibition of high-affinity choline transport by hemicholinium mustard (HCM), an alkylating analogue of hemicholinium-3, was examined in rat brain synaptosomes and guinea pig myenteric plexus. In synaptosomes, 50% high-affinity choline transport inhibition occurs with an HCM concentration of 104 nM (4-min incubation). A 10-min preincubation with 10 microM HCM results in essentially complete (greater than 95%) inactivation that persists after washing. Low-affinity choline transport in synaptosomes is unaffected by HCM inhibition at all concentrations examined (1-50 microM). Time course experiments indicate that the maximum irreversible inhibition (58%) seen after a 1-min preincubation with 500 nM HCM decreases to 46% inhibition after a 15-min preincubation; however, analysis of variance reveals that this difference is not significant. HCM inhibition of acetylcholine release from myenteric plexus-longitudinal muscle preparations persists for at least 2 h after removal of drug from the incubation bath; this inactivation can be prevented by coincubation with a high choline concentration during treatment with the mustard. In contrast, inhibition produced by the parent compound hemicholinium-3 is largely reversed by washing in both preparations examined. The observed potency and selectivity of HCM suggest its usefulness as a covalent probe for high-affinity choline transport.
在大鼠脑突触体和豚鼠肠肌丛中研究了半胱氨酸芥（HCM）（半胱氨酸-3的烷基化类似物）对高亲和力胆碱转运的抑制作用。在突触体中，HCM浓度为104 nM（孵育4分钟）时，高亲和力胆碱转运受到50%的抑制。用10 microM HCM预孵育10分钟会导致基本上完全（大于95%）失活，且在洗涤后仍持续存在。在所检测的所有浓度（1 - 50 microM）下，突触体中的低亲和力胆碱转运不受HCM抑制的影响。时间进程实验表明，用500 nM HCM预孵育1分钟后观察到的最大不可逆抑制（58%）在预孵育15分钟后降至46%抑制；然而，方差分析显示这种差异不显著。从孵育浴中去除药物后，HCM对肠肌丛 - 纵肌制剂中乙酰胆碱释放的抑制作用至少持续2小时；在用芥子处理期间与高浓度胆碱共同孵育可防止这种失活。相比之下，在所检测的两种制剂中，母体化合物半胱氨酸-3产生的抑制作用在洗涤后大部分可逆转。观察到的HCM的效力和选择性表明其作为高亲和力胆碱转运的共价探针是有用的。
