Activators of cyclic adenosine 3':5'-monophosphate accumulation in rat hippocampal slices: Action of vasoactive intestinal peptide (VIP)

Abstract

The present experiments tested the ability of putative neurotransmitters and neuromodulators to regulate cyclic adenosine 3':5'-monophosphate (cAMP) levels in rat hippocampal slices. Slices from ovariectomized adult female rats were equilibrated for 1 hr and incubated for 20 min with various test compounds, and cAMP was extracted and quantified using a competitive protein-binding assay. Norepinephrine, adenosine, histamine, and prostaglandins E1 and E(2α), induced moderate (1.5- to 5-fold) increase in cellular cAMP, whereas dopamine, serotonin, prostaglandin F(2α), and glutamate were relatively ineffective. Most striking was the observation that vasoactive intestinal peptide (VIP) produced marked elevation (approximately 80-fold at 6 μM) of hippocampal slice cAMP content. In contrast, other peptides produced only 2-fold increases (glucagon, somatostatin) or no change in cellular cAMP levels (enkephalins, LHRH, ACTH analogue, arginine vasopressin). Significant elevations in cAMP were seen with VIP concentrations as low as 20 nM; the cAMP response was half-maximal at 1 μM VIP and maximized between 10 and 20 μM. At maximally effective concentrations, VIP was 86% as effective in increasing cAMP as maximal concentrations of forskolin, a compound which activates adenylate cyclase in most cell types. The cAMP response to 10 μM VIP was pronounced after a 1-min incubation (16-fold elevations) and was maximal at 30 min (140-fold elevation). When slices from other brain areas were compared, it was found that regions known to contain high levels of VIP (cerebral cortex) also responded to VIP treatment with 30- to 50-fold elevations in cAMP. Hypothalamus-preoptic area, midbrain, and cerebellum, whose endogenous VIP levels decrease in that order, showed reduced cAMP responses to VIP treatment (9-, 7-, and 1.2-fold elevations, respectively). Further experiments demonstrated that removal of calcium from the bathing medium severely attenuated the ability of VIP to stimulate cAMP accumulation in hippocampal slices. Moreover, VIP-induced cAMP elevations were not dependent on neural firing since co-incubation of slices with VIP and 2 μM tetrodotoxin did not alter the cAMP response to VIP. These data suggest that VIP may be an important physiological regulator of hippocampal cAMP, probably via direct interactions of the peptide with VIP-specific receptors.