Thrombin action decreases acetylcholine receptor aggregate number and stability in cultured mouse myotubes

Neurons develop and make very stable, long-term synaptic connections with o ther nerve cells and with muscle. Synaptic stability at the neuromuscular j unction changes over development in that a proliferation of synaptic input are made to individual myotubes and synapses from all but one neuron are lo st during development. In an established co-culture paradigm in which spina l motoneurons synaptically contact myotubes, thrombin and associated protea se inhibitors have been shown to affect the loss of functional synaptic con tacts [6]. Evidence has not been provided which clearly demonstrate whether protease/protrease inhibitors affect either the pre- or postsynaptic termi nal, or both. In an effort to determine whether these reagents directly aff ect postsynaptic receptors on myotubes, myotubes were cultured in the absen ce of neurons and the spontaneous presence and stability of aggregates of a cetylcholine receptors (AChR) in control and thrombin-containing media were evaluated. In dishes fixed after treatment and in dishes in which individu al aggregates were observed live, thrombin action appeared to increase loss of AChR aggregates over time. Hirudin, a specific inhibitor of the thrombi n protease, diminished this loss. Neither reagent affected the overall inco rporation or degradation of AChR; therefore, it appears these protease/prot ease inhibitors affect the state of AChR aggregation. (C) 2000 Elsevier Sci ence B.V. All rights reserved.