Apoptosis in co-cultures of human muscle and rat spinal cord during the formation of the neuromuscular junction.

Abstract

The motor neurons and skeletal muscle fibers they innervate are strongly dependent on each other. Important communication between both tissues is mediated through the neuromuscular junction. However, release and reception of various factors at other parts of both tissues must also be considered as means of mutual influences. In vitro innervated human muscle is the only experimental model to investigate nerve - muscle interactions during synaptogenesis of human neuromuscular junction. Here we studied the occurrence of programmed cell death in this model. In order to find out if mutual interactions between neurons and myotubes control the extent of apoptosis in both tissues, we compared the number of apoptotic cells in spinal cord explants and myotubes in co-cultures and mono-cultures of these tissues. Apoptotic cells were detected by TUNEL at selected time intervals (10, 21 and 30 days after co-culture) coresponding to various stages of synaptogenesis of neuromuscular junction. The number of apoptotic cells in spinal cord explants was higher at earlier (day 10) in comparison to later stages (days 21 and 30). Co-cultures and mono-cultures did not differ in this respect. TUNEL positive cells were not found in mono-cultured human myotubes under our experimental conditions. These results suggest that the process of apoptosis seems to be rather independent on neuron - muscle interactions at least at the time points examine.