A novel approach to screen for cytokine effects on neuronal gene expression.
We describe an assay based on reverse transcription-polymerase chain reaction to detect the expression of mRNAs for a variety of transmitter synthetic enzymes and neuropeptides present at low levels in primary neuronal cultures. The assay is specific for mRNA-derived templates and is not affected by the presence of genomic DNA. Using this method, we demonstrate that cholinergic differentiation factor/leukemia inhibitory factor (CDF/LIF) and ciliary neurotrophic factor (CNTF) induce mRNAs for choline acetyltransferase, somatostatin, substance P, vasoactive intestinal polypeptide, cholecystokinin, and enkephalin. The induction of cholecystokinin and enkephalin by CDF/LIF and CNTF had not been shown previously. These data illustrate that the assay can reproduce findings obtained with other methods, as well as provide the sensitivity necessary to produce new results. These results also extend the overlap of CDF/LIF and CNTF in controlling gene expression in cultured sympathetic neurons, supporting the idea that these cytokines may share receptor subunits and signal transduction pathways.
Fann
M J
MJ
Division of Biology, California Institute of Technology, Pasadena 91125.
Patterson
P H
PH
D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D002766:Cholecystokinin / Q000378:metabolism; D020934:Ciliary Neurotrophic Factor; D016207:Cytokines / Q000494:pharmacology*; D004745:Enkephalins / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects*; D006131:Growth Inhibitors / Q000494:pharmacology; D015850:Interleukin-6; D053605:Leukemia Inhibitory Factor; D008222:Lymphokines / Q000494:pharmacology; D015335:Molecular Probes / Q000235:genetics; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D018377:Neurotransmitter Agents / Q000378:metabolism; D016133:Polymerase Chain Reaction / Q000379:methods*; D013564:Sympathetic Nervous System / Q000166:cytology / Q000378:metabolism; D014158:Transcription, Genetic
D020934:Ciliary Neurotrophic Factor; D016207:Cytokines; D004745:Enkephalins; D006131:Growth Inhibitors; D015850:Interleukin-6; D053605:Leukemia Inhibitory Factor; D008222:Lymphokines; D015335:Molecular Probes; D009419:Nerve Tissue Proteins; D018377:Neurotransmitter Agents; D002766:Cholecystokinin