The neuromuscular junction (NMJ) is the peripheral synapse that controls the coordinated movement of many organisms. The NMJ is also an archetypical model to study synaptic morphology and function. As the NMJ is the primary target of neuromuscular diseases and traumatic injuries, the establishment of suitable models to study the contribution of specific postsynaptic muscle-derived proteins on NMJ maintenance and regeneration is a permanent need. Considering the unique experimental advantages of the levator auris longus (LAL) muscle, here we present a method allowing for efficient electroporation-mediated gene transfer and subsequent detailed studies of the morphology and function of the NMJ and muscle fibers. Also, we have standardized efficient facial nerve injury protocols to analyze LAL muscle NMJ degeneration and regeneration. Our results show that the expression of a control fluorescent protein does not alter either the muscle structural organization, the apposition of the pre- and post-synaptic domains, or the functional neurotransmission parameters of the LAL muscle NMJs; in turn, the overexpression of MuSK, a major regulator of postsynaptic assembly, induces the formation of ectopic acetylcholine receptor clusters. Our NMJ denervation experiments showed complete reinnervation of LAL muscle NMJs four weeks after facial nerve injury. Together, these experimental strategies in the LAL muscle constitute effective methods to combine protein expression with accurate analyses at the levels of structure, function, and regeneration of the NMJ.

神经肌肉接头（NMJ）是控制许多生物体协调运动的外周突触。NMJ也是研究突触形态和功能的原型模型。由于NMJ是神经肌肉疾病和外伤的主要目标，因此永久性地需要建立合适的模型来研究特定的突触后肌肉衍生蛋白对NMJ维持和再生的贡献。考虑到实验的独特优势长耳提肌（LAL）肌肉，这里我们介绍一种允许有效电穿孔介导的基因转移以及随后对NMJ和肌肉纤维的形态和功能进行详细研究的方法。此外，我们已经标准化了有效的面神经损伤方案，以分析LAL肌肉NMJ的退化和再生。我们的结果表明，对照荧光蛋白的表达不会改变肌肉的结构组织，突触前和突触后结构域的位置或LAL肌肉NMJ的功能性神经传递参数。反过来，MuSK（突触后组装的主要调节剂）的过表达诱导异位乙酰胆碱受体簇的形成。我们的NMJ去神经实验显示，面神经损伤后四周，LAL肌肉NMJ完全被神经支配。一起，