Acetylcholine accumulation and release by hybrid NG108-15, glioma and neuroblastoma cells--role of a 16kDa membrane protein in release.
A procedure is described to fill up cells in culture with ACh and study its calcium dependent release, by-passing the synthesis steps. Whether differentiated or not with dbc-AMP, the NG108-15 cells efficiently released ACh when stimulated with calcium and ionophore A23187. The release was also studied in the parent C6-BU-1 and N18TG2 cells. It was found that C6-BU-1 released ACh much better that N18TG2 in spite of their glial origin. The internalization by NG108-15 cells of an antisense oligonucleotide probe hybridizing the 16 kDa proteolipid messenger common to mediatophore and to the V-ATPase reduced ACh release indicated a role of this proteolipid in ACh translocation. This characteristic protein was found in the membrane extract of NG108-15 cells and also in the C6-BU-1 cells, but its amount was strongly reduced in the N18TG2 cell line and in the NG108-15 cells having internalized the antisense probe.