Corticotropin‐Peptide Regulation of Intracellular Cyclic AMP Production in Cortical Neurons in Primary Culture

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Abstract
Previous studies have provided evidence for adrenocorticotropic hormone (ACTH) effects on a wide variety of behaviors. However, the precise sites of action and the mechanisms by which these effects may be mediated have yet to be clearly elucidated. Although ACTH was shown to augment cyclic AMP levels in glial cells isolated from whole brain, other studies found little or no effect of ACTH peptides on cyclic nucleotide metabolism in slices of cerebral cortex or homogenates of whole brain. In the present study, our objective was to determine whether ACTH peptides regulate intracellular cyclic AMP levels in neurons of the cerebral cortex in primary culture. ACTH peptides stimulated cyclic AMP synthesis up to threefold in a dose-dependent manner; stimulation was complete within 5–10 min of exposure to agonists. Neurohormone efficacy was augmented by 0.1 μM forskolin (which was virtually ineffective alone); potency was unaffected. The order of potency (EC50) for increasing intracellular cyclic AMP levels was as follows: ACTH (1–24), ACTH (1–17) (10 nM) > α-melanocyte stimulating hormone, β-melanocyte stimulating hormone (α-MSH, β-MSH) (100 nM) > ACTH (1–10) (1 μM) > ACTH (4–10) (5 μM). The hexapeptide ACTH (4–9) as well as ACTH (11–24) were inactive at concentrations as high as 10 μM. Other neuropeptides derived from proopiocortin, such as β-endorphin and Met- and Leu-enkephalin were without effect on basal or hormonally stimulated cyclic AMP synthesis. In order to determine whether distinct receptors for ACTH are present on cortical neurons, saturating concentrations of the peptide were coincubated with either vasoactive intestinal polypeptide (VIP) or the β-adrenergic agonist, isoproterenol (INE). The response to combinations of ACTH and INE were clearly additive. However, neither ACTH nor INE could further augment cyclic AMP formation at saturating concentrations of VIP. Comparison of structure-activity relationships suggest that ACTH receptors mediating the elevation of cyclic AMP formation in cortical neurons may be similar to those associated with the peptide actions on arousal rather than conditioned behavior.