Characterization of Acetylcholinesterase Secretion in Neuronal Cultures and Regulation by High K⁺ and Soluble Factors from Target Cells

Abstract

We have observed that cultured neurons from chick spinal cord and the neuroblastoma hybrid line 108CC15 released lower amounts of acetylcholinesterase (AChE) when compared with the parental line, N18TG2. AChE activity extracted by hypotonic buffer, which can be regarded as the source of the released enzyme, was considerably higher in the parental than in the hybrid 108CC15 (respectively, ∼80% and ∼40% of cellular activity). On the other hand, evaluation of ectocellular, with respect to total, AChE activity showed that in N18TG2 cells only 7% of AChE was localized on the plasmalemma, whereas in the hybrid line the percentage of ectocellular activity was 3.7 times higher than in the parental line. We have also examined the effect of cytochalasin B and nocodazole. In the N18TG2 line, the former did not affect AChE release, which was significantly reduced by the latter. High K+ level in the culture medium, of both N18TG2 and hybrid 108CC15 cultures, induced an increase in AChE secretion; Ca2+ presence was required for high K+‐induced release. Muscle extracts increased AChE secretion in both the hybrid 108CC15 and the spinal cord neurons. The present data suggest that AChE secretion during neuronal development is modulated by depolarizing stimuli and by soluble factors produced by target cells and may be involved in the control of neuronal differentiation.

The readthrough variant of acetylcholinesterase remains very minor after heat shock, organophosphate inhibition and stress, in cell culture and in vivo

“…The basal level of AChE mRNA was higher in the neuroblastoma‐glioma NG108CC15 cells than in N18TG2, in agreement with their relative AChE activities (Melone et al . 1987; Biagioni et al . 1995) (Fig.…”

“…As expected, HSP70 transcripts were highly increased after heat shock in the two cell lines; PRiMA transcripts were unaffected. In NG108CC15 cells, which express a higher level of AChE activity than N18TG2 cells (Biagioni et al . 1995), we found no change in total AChE transcripts or in the AChE R variant; in contrast, we observed a modest increase of AChE transcripts in N18TG2, and a sixfold increase of AChE R , which reached approximately the same level as in NG108CC15 cells.…”

Modulation of Acetylcholinesterase and Voltage‐Gated Na+ Channels in Choline Acetyltransferase‐ Transfected Neuroblastoma Clones

“…A further consideration pointing to AChE as a possible modulatory molecule for cell behavior is its ability to be associated to different cell compartments; it is known that in different cell types and possibly in relation to developmental stage the enzyme is either secreted in the extracellular space or remains associated to the plasma membrane. In neuroblastoma cultures its cellular localization has been related to different stages of neuronal differentiation (Melone et al, 1987), and the comparative analysis of AChE release in different neuronal populations has suggested that acquisition of a cholinergic phenotype is characterized by a low level of AChE release (Biagioni et al, 1995; Coleman and Taylor, 1996). The data reported in Fig.…”

Alternative acetylcholinesterase molecular forms exhibit similar ability to induce neurite outgrowth

“…This allows the investigation of AChE morphogenetic action, avoiding the possible interference of self‐produced neurotransmitter. This neuroblastoma clone exhibits some neuronal traits, such as the presence of tetrodotoxin‐sensitive voltage‐gated Na + channels and cholinergic receptors (De Jaco et al, 2000); it expresses moderate amounts of the tetrameric AChE form that is mostly secreted in the extracellular medium (Biagioni et al, 1995).…”