VanSaun M, Humburg B C, Arnett M G, Pence M, Werle M J
Department of Anatomy and Cell Biology, University of Kansas Medical Center, Kansas City, KS 66160, USA.
Dev Neurobiol. 2007 Sep 15;67(11):1488-97. doi: 10.1002/dneu.20523.
The extracellular matrix surrounding the neuromuscular junction is a highly specialized and dynamic structure. Matrix Metalloproteinases are enzymes that sculpt the extracellular matrix. Since synaptic activity is critical to the structure and function of this synapse, we investigated whether changes in synaptic activity levels could alter the activity of Matrix Metalloproteinases at the neuromuscular junction. In particular, we focused on Matrix Metalloproteinase 3 (MMP3), since antibodies to MMP3 recognize molecules at the frog neuromuscular junction, and MMP3 cleaves a number of synaptic basal lamina molecules, including agrin. Here we show that the fluorogenic compound (M2300) can be used to perform in vivo proteolytic imaging of the frog neuromuscular junction to directly measure the activity state of MMP3. Application of this compound reveals that active MMP3 is concentrated at the normal frog neuromuscular junction, and is tightly associated with the terminal Schwann cell. Blocking presynaptic activity via denervation, or TTX nerve blockade, results in a decreased level of active MMP3 at the neuromuscular junction. The loss of active MMP3 at the neuromuscular junction in denervated muscles can result from decreased activation of pro-MMP3, or it could result from increased inhibition of MMP3. These results support the hypothesis that changes in synaptic activity can alter the level of active MMP3 at the neuromuscular junction.
神经肌肉接头周围的细胞外基质是一种高度特化且动态的结构。基质金属蛋白酶是塑造细胞外基质的酶。由于突触活动对于该突触的结构和功能至关重要，我们研究了突触活动水平的变化是否会改变神经肌肉接头处基质金属蛋白酶的活性。特别地，我们聚焦于基质金属蛋白酶3（MMP3），因为针对MMP3的抗体可识别青蛙神经肌肉接头处的分子，且MMP3可切割多种突触基膜分子，包括聚集蛋白。在此我们表明，荧光化合物（M2300）可用于对青蛙神经肌肉接头进行体内蛋白水解成像，以直接测量MMP3的活性状态。应用该化合物显示，活性MMP3集中于正常青蛙神经肌肉接头处，并与终末施万细胞紧密相关。通过去神经支配或TTX神经阻滞阻断突触前活动，会导致神经肌肉接头处活性MMP3水平降低。去神经支配肌肉的神经肌肉接头处活性MMP3的丧失可能是由于前体MMP3的激活减少，或者可能是由于对MMP3的抑制增加。这些结果支持了突触活动的变化可改变神经肌肉接头处活性MMP3水平这一假说。
