Table 1.

Developmental characteristics of functional innervation in co-cultures of rat spinal cord and human skeletal muscle cells. A contracting unit is a distinct group of cultured myotubes (myofibers) that contract simultaneously at a frequency that is different from the frequencies of other contracting units. Contraction-positive explant is an explant that established functional neuromuscular contacts (i.e., has at least one contracting unit). Presented data are based on: [27,42,46,122,171]. Staging of co-cultures is based on the appearance of basal lamina [24,171].

Co-Culture Stage	Basal Lamina	Formation of Functional Neuromuscular Junctions	Acetylcholinesterase and nAChR
Stage I (Day 1–9)	Not formed	First neurite-myotube contact (Day 3)	Diffuse AChE staining in myotubes and neurites extending from the spinal cord explant
		First α-bungarotoxin-sensitive contractions occur (Day 7)	AChE expressed in all myonuclei and present along the whole myotube length
		No visible cross-striations in myotubes	Immature nAChR clusters at neuromuscular contacts
Stage II (Day 10–21)	Formed	Number of contraction-positive explants attains plateau (Day 10)	AChE expressed predominantely at junctional myonuclei
		Number of contracting units attains plateau (Day 17)	Discrete patches of AChE originating in motor neurons and myotubes
		Cross-striations visible in contracting myotubes	Mature nAChR cluster at neuromuscular contacts AChE and nAChR co-localize in discrete patches