TY - JOUR
T1 - Upregulated acetylcholine synthesis during early differentiation in the embryonic stem cell line CGR8.
AU - Wessler, Ignaz
AU - Michel-Schmidt, Rosmarie
AU - Schmidt, Harald
AU - Kaltwasser, Susanne
AU - Unger, Ronald
AU - Kirkpatrick, Charles James
PY - 2013/6/28
Y1 - 2013/6/28
N2 - Stem cells are used to generate differentiated somatic cells including neuronal cells. Synthesis and release of acetylcholine, a neurotransmitter and widely expressed signaling molecule, were investigated in the murine embryonic stem cell line CGR8 during early differentiation, i.e. in the presence of leukemia inhibitory factor (LIF) to maintain pluripotency and in the absence of LIF to induce early differentiation. CGR8 cells express choline acetyltransferase (ChAT) as demonstrated by measurement of enzyme activity and substantial inhibition by bromoacetylcholine. Pluripotent CGR8 cells showed a ChAT activity of 250 pmol acetylcholine/mg/h, contained 1.1 pmol acetylcholine/106 cells and released about 12.00 pmol acetylcholine/1 x 106 cells/6 h. Removal of LIF induced early differentiation as evidenced by reduced transcription factors Oct-4 and Nanog and a substantial slowing of the proliferation rate. Under this condition acetylcholine synthesis increased to 1640 pmol/mg/h; related to the pluripotent state the content of acetylcholine increased 10-fold and the release to about 32 pmol acetylcholine/1 x 106 cells/6 h. Enzyme kinetic analysis showed a significant increase of the K(m) for the precursor acetyl-CoA and of V(max) without a change of the K(m) for the precursor choline. In conclusion, early differentiation of the stem cell line CGR8 is associated with a substantial increase in ChAT activity and acetylcholine release.
AB - Stem cells are used to generate differentiated somatic cells including neuronal cells. Synthesis and release of acetylcholine, a neurotransmitter and widely expressed signaling molecule, were investigated in the murine embryonic stem cell line CGR8 during early differentiation, i.e. in the presence of leukemia inhibitory factor (LIF) to maintain pluripotency and in the absence of LIF to induce early differentiation. CGR8 cells express choline acetyltransferase (ChAT) as demonstrated by measurement of enzyme activity and substantial inhibition by bromoacetylcholine. Pluripotent CGR8 cells showed a ChAT activity of 250 pmol acetylcholine/mg/h, contained 1.1 pmol acetylcholine/106 cells and released about 12.00 pmol acetylcholine/1 x 106 cells/6 h. Removal of LIF induced early differentiation as evidenced by reduced transcription factors Oct-4 and Nanog and a substantial slowing of the proliferation rate. Under this condition acetylcholine synthesis increased to 1640 pmol/mg/h; related to the pluripotent state the content of acetylcholine increased 10-fold and the release to about 32 pmol acetylcholine/1 x 106 cells/6 h. Enzyme kinetic analysis showed a significant increase of the K(m) for the precursor acetyl-CoA and of V(max) without a change of the K(m) for the precursor choline. In conclusion, early differentiation of the stem cell line CGR8 is associated with a substantial increase in ChAT activity and acetylcholine release.
UR - http://www.scopus.com/inward/record.url?scp=84892405826&partnerID=8YFLogxK
U2 - 10.1016/j.neulet.2013.04.052
DO - 10.1016/j.neulet.2013.04.052
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C2 - 23669640
AN - SCOPUS:84892405826
SN - 0304-3940
VL - 547
SP - 32
EP - 36
JO - Neuroscience Letters
JF - Neuroscience Letters
ER -