Comparison of the Effects of Forskolin and Dibutyryl Cyclic AMP in Neuroblastoma Cells: Evidence that Some of the Actions of Dibutyryl Cyclic AMP Are Mediated by Butyrate

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Abstract
We have compared the effects of forskolin, N6,2′‐O‐dibutyryladenosine 3′:5′‐cyclic monophosphate (dibutyryl cyclic AMP, Bt2‐cAMP), and butyrate on several aspects of neuroblastoma cell physiology. The morphology of Neuro 2A cells was similar after incubation with forskolin and Bt2‐cAMP, which caused extensive neurite outgrowth, whereas in the presence of butyrate some rudimentary neurites were formed but they were not nearly as extensive. All compounds produced a dose‐dependent inhibition of cell proliferation, but the effect of Bt2‐cAMP was more marked than that caused by forskolin, thus showing that the effect of Bt2‐cAMP is due partially to the butyrate released. Acetylcholinesterase activity was lower in the cells incubated with butyrate or Bt2‐cAMP than in untreated cells or in forskolin‐treated cells. This suggests that cyclic AMP does not play a role in the regulation of this enzyme. Bt2‐cAMP produced histone acetylation, a well‐known effect of butyrate in cultured cells, whereas forskolin did not affect this modification. Consequently, the levels of thyroid hormone receptor, a nuclear protein whose concentration is regulated by butyrate through changes in acetylation of chromatin proteins, were decreased in cells incubated with Bt2‐cAMP or butyrate, but were unaffected by forskolin. Butyrate elevated the concentration of histone H1°, a protein that increases in neuroblastoma cells as a result of different treatments that block cell division. The concentration of H1° in the cells treated with Bt2‐cAMP was at a level intermediate between that found after treatment with butyrate and with forskolin. The present results clearly indicate that some of the effects of Bt2‐cAMP on neuroblastoma cells can be attributed to the butyryl moiety of this compound rather than to the cyclic nucleotide itself.