The Motor End‐plate Specific Form of Acetylcholinesterase: Appearance During Embryogenesis and Re‐innervation of Rat Muscle

Abstract

Abstract– We have solubilized three active molecular forms of AChE from rat muscle and have confirmed the presence of one of these forms (EP form, apparent sedimentation coefficient: 16 s) uniquely at the motor end‐plate regions of several skeletal muscles. This form was never detected in smooth muscle extracts. In sternocleidomastoïdian muscle it disappeared after denervation and reappeared after re‐innervation in the region where nerve and muscle had come in contact. During the embryonic development of hind leg muscles the EP form appeared on the 14th or 15th day of gestation. The EP form of muscle AchE appears to be an excellent biochemical marker of the neuromuscular junction.

Presence of tailed, asymmetric forms of acetylcholinesterase in the central nervous system of vertebrates

“…The tailed forms can be found in muscle cells cultured alone, or in cultured sympatheticlike transformed cells (PC12), provided they generate spontaneous action potentials. They are not restricted to (or dependent only upon) choline@ synaptic contacts, as earlier found for the rat adult neuromuscular junction [7]. Thus, they potentially could be found in any active neuron.…”

“…For example, synapse formation in vivo (rat muscle) is accompanied by the induction of '16 S' AChE [7]; the '10 S' AChE becomes predominant in rat brain during post-natal maturation [6], whereas the rise of 10 S in cerebellum correlates well with the differentiation of Purkinje cells and the external granular layer [ 181; chick '11 S' AChE increases in sympathetic ganglia after hatching, when cytologicai maturity has been reached but junctional synaptic activity is still increasing [ 1 I]. The tailed forms (mainly rat or mouse 16 S and chick 20 S), in vivo, in adult vertebrates (rat and chicks) muscles, disappear upon denervation [S,6,8,30], but remain in the denervated rat superior cervical ganglion [IO] and are present in rat muscle cells cultured alone [24] or cultured NGF-treated pheochromocytoma cells (PC 12) [23], when action potentials can be generated.…”

Multiple forms of acetylcholinesterase in clonal muscle cells

“…The muscle cells studied in the present work have all three forms of AChE that have been identified in the rat muscle in vivo [1,3]. Of these three forms, the 16 S form has been found exclusively in the endplate regions where muscles are innervated by motor nerves [1,3].…”

“…AChE was extracted by homogenizing the cells in 0.5-1.0 ml solution A/plate (1.0 M NaC1, 50 mM MgC12, 1% Triton X-100, 10 mM Tris-HC1, pH 7.4) and centrifuging at 20 000 × g for 30 min [3]. Most of the AChE activity in the crude homogenates was recovered in the supernatant fractions (110%).…”

“…Three major forms have been identified with sedimentation coefficients of 4 S, 10 S and 16 S [1][2][3][4]. The 16 S form has been found only at the site of the synapse between neuron and skeletal muscles [1,3].…”