Wen Hua, Hubbard Jeffrey Michael, Wang Wei-Chun, Brehm Paul
Oregon Health and Sciences University, Portland, Oregon 97239.
Oregon Health and Sciences University, Portland, Oregon 97239
J Neurosci. 2016 Oct 19;36(42):10870-10882. doi: 10.1523/JNEUROSCI.0505-16.2016.
Rapsyn-deficient myasthenic syndrome is characterized by a weakness in voluntary muscle contraction, a direct consequence of greatly reduced synaptic responses that result from poorly clustered acetylcholine receptors. As with other myasthenic syndromes, the general muscle weakness is also accompanied by use-dependent fatigue. Here, we used paired motor neuron target muscle patch-clamp recordings from a rapsyn-deficient mutant line of zebrafish to explore for the first time the mechanisms causal to fatigue. We find that synaptic responses in mutant fish can follow faithfully low-frequency stimuli despite the reduced amplitude. This is in part helped by a compensatory increase in the number of presynaptic release sites in the mutant fish. In response to high-frequency stimulation, both wild-type and mutant neuromuscular junctions depress to steady-state response levels, but the latter shows exaggerated depression. Analysis of the steady-state transmission revealed that vesicle reloading and release at individual release sites is significantly slower in mutant fish during high-frequency activities. Therefore, reductions in postsynaptic receptor density and compromised presynaptic release collectively serve to reduce synaptic strength to levels that fall below the threshold for muscle action potential generation, thus accounting for use-dependent fatigue. Our findings raise the possibility that defects in motor neuron function may also be at play in other myasthenic syndromes that have been mapped to mutations in muscle-specific proteins.
Use-dependent fatigue accompanies many neuromuscular myasthenic syndromes, including muscle rapsyn deficiency. Here, using a rapsyn-deficient line of zebrafish, we performed paired motor neuron target muscle patch-clamp recordings to investigate the mechanisms causal to this phenomenon. Our findings indicate that the reduced postsynaptic receptor density resulting from defective rapsyn contributes to weakness, but is not solely responsible for use-dependent fatigue. Instead, we find unexpected involvement of altered transmitter release from the motor neuron. Specifically, slowed reloading of vesicle release sites leads to augmented synaptic depression during repeated action potentials. Even at moderate stimulus frequencies, the depression levels for evoked synaptic responses fall below the threshold for the generation of muscle action potentials. The associated contraction failures are manifest as use-dependent fatigue.
无rapsyn的肌无力综合征的特征是随意肌收缩无力，这是由于乙酰胆碱受体聚集不良导致突触反应大幅降低的直接后果。与其他肌无力综合征一样，全身肌肉无力也伴有使用依赖性疲劳。在此，我们首次使用来自无rapsyn的斑马鱼突变系的配对运动神经元-靶肌肉膜片钳记录来探究导致疲劳的机制。我们发现，尽管突变鱼的突触反应幅度降低，但仍能忠实地跟随低频刺激。这部分得益于突变鱼突触前释放位点数量的代偿性增加。在高频刺激下，野生型和突变型神经肌肉接头均会抑制至稳态反应水平，但后者的抑制更为明显。对稳态传递的分析表明，在高频活动期间，突变鱼单个释放位点的囊泡重新装载和释放明显较慢。因此，突触后受体密度降低和突触前释放受损共同作用，将突触强度降低到低于肌肉动作电位产生阈值的水平，从而导致使用依赖性疲劳。我们的研究结果提出了一种可能性，即运动神经元功能缺陷可能在其他已定位到肌肉特异性蛋白突变的肌无力综合征中也起作用。
使用依赖性疲劳伴随许多神经肌肉性肌无力综合征，包括肌肉rapsyn缺乏症。在此，我们使用无rapsyn的斑马鱼品系，进行了配对运动神经元-靶肌肉膜片钳记录，以研究导致这种现象的机制。我们的研究结果表明，rapsyn缺陷导致的突触后受体密度降低是肌无力的原因之一，但并非使用依赖性疲劳的唯一原因。相反，我们发现运动神经元递质释放改变意外地参与其中。具体而言，囊泡释放位点重新装载减慢导致重复动作电位期间突触抑制增强。即使在中等刺激频率下，诱发突触反应的抑制水平也会降至肌肉动作电位产生阈值以下。相关的收缩失败表现为使用依赖性疲劳。
