Three cholinergic neuroblastoma hybrid cell lines that form few synapses on myotubes are deficient in acetylcholine receptor aggregation molecules and large dense core vesicles

Neuroblastoma × spinal cord (NSC) hybrid cell lines resemble developing motor neurons

“…Whether AChR clusters represent redistribution of previously expressed AChR (Magill-Solc and McMahan, 1988;Reist et al, 1987), or AChR upregulation and membrane insertion (Jessell et al, 1979;Falls et al, 1991) is currently unknown. No distinct AChR aggregates were observed on myotubes cultured alone or with N18TG2 cells (confirming Busis et al, 1984) or NSC-19 cells.…”

“…We investigated whether NSC 19, NSC 34, or parent neuroblastoma N18TG2 cells were capable of inducing AChR clusters on cultured myotubes, which is a n early event in myoneural synaptogenesis (Anderson and Cohen, 1977). Only NSC-34 cells were capable of inducing AChR clusters, similar to those reported for NG108-15 cells (Busis et al, 1984). NSC-34 cells were added to neonatal mouse forelimb cultures at the time of fusion of myocytes into multinucleated myotubes.…”

“…Under the culture conditions, spontaneous myotube twitching (without neural elements) was observed in -10% of high power fields examined, which was not increased by the addition of N18TG2 cells, even after 3 days prior "induction" with 1 mM dbcAMP (Table 3). Four NSC cell lines, all of which expressed ChAT, increased myotube twitching 2.5-to 7-fold compared to aneural cultures (Table 3), to a similar degree as dbcAMP-induced NG108-15 cells, a previously described neuroblastoma-glioma hybrid cell line which can form myoneural synapses in vitro (Hamprecht, 1977;Nirenberg et al, 1983;Busis et al, 1984;Amano et al, 1974). Twitching was not significantly enhanced by prior culture of NSC cells with dbcAMP.…”

“…Four reported hybrid cell lines model one property of motor neurons by forming transient synapses with myotubes when induced with dibutyryl cyclic AMP (dbcAMP), theophylline, or prostaglandin E-all maneuvers to increase cellular CAMP (Hamprecht, 1977;Nirenberg et al, 1983;Busis et al, 1984): NG108-15 (N18TG2 x C6 rat glioma), NBr-10 and NBr-20 (N18TG2 x BRL-30E rat liver cells), and NCB-20 (N18TG2 x Chinese hamster embryonic brain cells). However, formation of myoneural synapses cannot be regarded as the sine qua non of motor neuron phenotype, since many neuronal types, including sympa- thetic ganglion cells and retinal cells, are capable of forming transient synapses with myotubes in vitro (Nurse and O'Lague, 1975;Puro and Nirenberg, 1976).…”

Muscle Induces Neuronal Expression of Acetylcholinesterase in Neuron-Muscle Co-culture

“…Among these possible AChE regulatory elements, a cAMP-responsive element (CRE) site located on the human AChE promoter has been shown to regulate the level of AChE transcripts in muscle (25,26) and neuron (27). Here, we would like to determine the regulation mechanism of neuronal AChE mediated by the interacting muscle in a co-culture system of neuroblastoma ϫ glioma hybrid cells NG108-15 with chick myotubes; this coculture has been used to resemble the formation of nmjs in vitro previously (14,28,29). By using a luciferase-tagged human AChE promoter construct as a tool, the transcriptional element that mediated the muscle-induced neuronal AChE expression in the neuron-muscle co-culture was elucidated.…”