Brain K L, Cunnane T C
Department of Pharmacology, University of Oxford, Oxford, UK.
Br J Pharmacol. 2008 Feb;153(4):831-9. doi: 10.1038/sj.bjp.0707623. Epub 2007 Dec 10.
The antidysrhythmic bretylium is useful experimentally because it selectively abolishes neurotransmitter release from sympathetic peripheral nerve terminals. Its mechanism of action seemed settled, but recent results from optical monitoring of single terminals now suggests a new interpretation.
Orthograde transport of a dextran-conjugated Ca(2+) indicator to monitor Ca(2+) in nerve terminals of mouse isolated vas deferens with a confocal microscope. In some experiments, local neurotransmitter release was detected by monitoring neuroeffector Ca(2+) transients (NCTs) in adjacent smooth muscles, a local measure of purinergic transmission. Sympathetic terminals were identified with catecholamine fluorescence (UV excitation) or post-experiment immunohistochemistry.
Bretylium (10 microM) abolished NCTs at 60/61 junctions over the course of 2 h, indicating effective abolition of neurotransmitter release. However, bretylium did not abolish the field stimulus-induced Ca(2+) transient in most nerve terminals, but did increase both action potential delay (by 2+/-0.4 ms) and absolute refractory period (by 4+/-2 ms). Immunohistochemistry demonstrated that 85-96% of terminals orthogradely filled with a dextran-conjugated fluorescent probe contained Neuropeptide Y (NPY). A formaldehyde-glutaraldehyde-induced catecholamine fluorescence (FAGLU) technique was modified to allow sympathetic terminals to be identified with a Ca(2+) indicator present. Most terminals contained catecholamines (based on FAGLU) or secrete ATP (as NCTs in adjacent smooth muscle cells are abolished).
Bretylium can inhibit neurotransmitter release downstream of Ca(2+) influx without abolishing the nerve terminal action potential. Bretylium-induced increases in the absolute refractory period permit living sympathetic terminals to be identified.
抗心律失常药溴苄铵在实验中很有用，因为它能选择性地消除交感神经外周神经末梢的神经递质释放。其作用机制似乎已确定，但最近对单个末梢进行光学监测的结果提示了一种新的解释。
将葡聚糖偶联的Ca(2+)指示剂进行顺行运输，用共聚焦显微镜监测小鼠离体输精管神经末梢中的Ca(2+)。在一些实验中，通过监测相邻平滑肌中的神经效应Ca(2+)瞬变（NCTs）来检测局部神经递质释放，这是嘌呤能传递的一种局部测量方法。通过儿茶酚胺荧光（紫外线激发）或实验后免疫组织化学鉴定交感神经末梢。
在2小时内，溴苄铵（10 microM）使60/61个连接处的NCTs消失，表明神经递质释放被有效消除。然而，溴苄铵并没有消除大多数神经末梢中电场刺激诱导的Ca(2+)瞬变，但确实增加了动作电位延迟（增加2±0.4毫秒）和绝对不应期（增加4±2毫秒）。免疫组织化学显示，85 - 96%顺行填充有葡聚糖偶联荧光探针的末梢含有神经肽Y（NPY）。对甲醛 - 戊二醛诱导的儿茶酚胺荧光（FAGLU）技术进行了改进，以便用存在的Ca(2+)指示剂鉴定交感神经末梢。大多数末梢含有儿茶酚胺（基于FAGLU）或分泌ATP（因为相邻平滑肌细胞中的NCTs消失）。
溴苄铵可在Ca(2+)内流下游抑制神经递质释放，而不消除神经末梢动作电位。溴苄铵诱导的绝对不应期增加使得能够识别存活的交感神经末梢。
