Thrombin reduces MuSK and acetylcholine receptor expression along with neuromuscular contact size in vitro.
In the course of studies on thrombin and its inhibitor(s) in synaptic plasticity, we addressed the question of their roles in the formation of neuromuscular junctions (NMJ) and used a model of rat neuron-myotube cocultures. We report that the size of acetylcholinesterase (AChE) patches used as a marker of neuromuscular contacts was decreased in the presence of either thrombin or SFLLRN, the agonist peptide of the thrombin receptor PAR-1, whereas it was increased with hirudin, a specific thrombin inhibitor. In an attempt to relate these neuromuscular contact size variations to molecular changes, we studied muscle-specific tyrosine kinase receptor (MuSK), acetylcholine receptor (AChR) and rapsyn expression in the presence of thrombin. We showed that thrombin did not change rapsyn gene and protein expression. However, the expression of MuSK and surface AChR proteins was diminished in both myotube cultures and neuron-myotube cocultures. These reductions in protein expression were associated with a decrease in MuSK and AChR alpha-subunit gene expression in myotube cultures but not in neuron-myotube cocultures. Moreover, the expression of the AChR epsilon-subunit gene, specifically enhanced by neuron-released factors, was not modified by thrombin in neuron-myotube cocultures. This suggests that thrombin did not affect the expression of synaptic AChRs enhanced by neuron-released factors but rather reduced the level of extrasynaptic AChRs. Taken together, these results indicate that thrombin in balance with its inhibitor(s) could modulate the formation of neuromuscular contacts in vitro by affecting the expression of two essential molecules in NMJ postsynaptic differentiation, MuSK and AChR.
Faraut
Brice
B
INSERM U582, Institut de Myologie, Groupe Hospitalier Pitié-Salpêtrière, 47, Boulevard de l'Hôpital, 75651 Paris Cedex 13, France.
Ravel-Chapuis
Aymeric
A
Bonavaud
Sylvie
S
Jandrot-Perrus
Martine
M
Verdière-Sahuqué
Martine
M
Schaeffer
Laurent
L
Koenig
Jeanine
J
Hantaï
Daniel
D
D000818:Animals; D002478:Cells, Cultured; D005260:Female; D009469:Neuromuscular Junction / Q000187:drug effects* / Q000378:metabolism*; D011247:Pregnancy; D051381:Rats; D017207:Rats, Sprague-Dawley; D020794:Receptor Protein-Tyrosine Kinases / Q000096:biosynthesis*; D011950:Receptors, Cholinergic / Q000096:biosynthesis*; D013917:Thrombin / Q000494:pharmacology*
D011950:Receptors, Cholinergic; C095714:MUSK protein, human; D020794:Receptor Protein-Tyrosine Kinases; D013917:Thrombin