Immunoaffinity purification and dose-response of cholinergic neuronal differentiation factor.
A glycoprotein from heart cell-conditioned medium, cholinergic neuronal differentiation factor (CDF), causes a transition from noradrenergic to cholinergic phenotype in cultured rat sympathetic neurons. Although the transition has been known to occur in a dose-dependent manner and CDF has been purified, the examination of a complete dose-response of neurons to CDF has not been possible because sufficient quantities of pure CDF have not been available. A complete dose-response curve is essential for evaluating the biological response of the neurons, for assessing the physiological role of CDF and for understanding the mechanism of action of CDF. We report here an immunoaffinity-purification procedure for CDF with a 73.1% recovery using antibodies raised against a synthetic peptide homologous with the N-terminal region of CDF. This method produced pure CDF in quantities sufficient for examination of the full dose-response range of the neurons. Our main findings are the following. The dose-responses of acetylcholine and catecholamine metabolisms to CDF are different, although the same molecule affects both transmitters. While the half-maximal concentrations for acetylcholine induction (0.20 nM) and for catecholamine suppression (0.28 nM) are similar, the response of catecholamine metabolism begins slowly and saturates at a CDF concentration (5-20 nM) considerably higher than that of acetylcholine (0.6 nM). This may indicate that CDF affects multiple processes in catecholamine metabolism.
Fukada
K
K
Department of Anatomy and Cell Biology, State University of New York, Brooklyn 11203.
Rushbrook
J I
JI
Towle
M F
MF
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D004305:Dose-Response Relationship, Drug; D006023:Glycoproteins / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D007158:Immunologic Techniques; D008969:Molecular Sequence Data; D020880:Neuregulins; D009474:Neurons / Q000187:drug effects*; D013564:Sympathetic Nervous System / Q000166:cytology / Q000187:drug effects
D000596:Amino Acids; D006023:Glycoproteins; D020880:Neuregulins