Adrenocorticotropic hormone activation of adenylate cyclase in raphe neurons: multiple regulatory pathways control serotonergic neuronal differentiation.
The RN46A cell line was derived from embryonic day 13 rat medullary raphe cells by infection with a retrovirus encoding the temperature-sensitive mutant of SV40 large T antigen (tsT-ag). The RN46A cell line is neuronally restricted and constitutively differentiates following a shift to nonpermissive temperature. Differentiated RN46A cells express low levels of tryptophan hydroxylase (TPH) but no detectable levels of serotonin (5-HT). Treatment of cultures with the adrenocorticotropic hormone peptide ACTH4-10 up-regulates the expression of TPH immunoreactivity in differentiated RN46A cells, but 5-HT synthesis requires initial treatment with ACTH4-10, followed by partial membrane depolarizing conditions. Up-regulation of TPH by ACTH4-10 is apparently due to activation of adenylate cyclase, whereas the increased 5-HT synthesis with membrane depolarization can be blocked with the voltage-sensitive Ca(2+)-channel blockers nifedipine and omega-conotoxin. ACTH4-10 treatment also markedly up-regulates the expression of the 5-HT reuptake transporter, as do dibutyryl cyclic AMP and forskolin; chronic membrane depolarization has no effect on 5-HT reuptake. The expression of the high-affinity 5-HT1A receptor is increased threefold by ACTH4-10 treatment during differentiation and fivefold by differentiation under partial membrane depolarizing conditions. Combining ACTH4-10 treatment and membrane depolarization does not increase expression of the 5-HT1A receptor further. 5-HT release is constitutive in ACTH-treated RN46A cells and linked to spontaneous synaptic vesicle fusion in RN46A cells. Considered with previous results, these data indicate that multiple effectors, ACTH, brain-derived neurotrophic factor, and membrane depolarization, have both distinct and overlapping effects that regulate specific elements of the serotonergic neuronal phenotype during differentiation and maturation.
Eaton
M J
MJ
Miami Project, Department of Neurological Surgery, University of Miami School of Medicine, Florida 33136, USA.
Whittemore
S R
SR
D000262:Adenylyl Cyclases / Q000819:agonists*; D000324:Adrenocorticotropic Hormone / Q000494:pharmacology*; D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004789:Enzyme Activation; D008564:Membrane Potentials / Q000187:drug effects; D009474:Neurons / Q000166:cytology / Q000187:drug effects*; D010446:Peptide Fragments / Q000494:pharmacology; D011903:Raphe Nuclei / Q000166:cytology / Q000187:drug effects*; D051381:Rats; D012701:Serotonin / Q000096:biosynthesis / Q000378:metabolism; D014365:Tryptophan Hydroxylase / Q000096:biosynthesis; D015854:Up-Regulation / Q000187:drug effects
D010446:Peptide Fragments; D012701:Serotonin; D000324:Adrenocorticotropic Hormone; D014365:Tryptophan Hydroxylase; D000262:Adenylyl Cyclases; C011394:ACTH (4-10)