The basic secretagogue compound 48/80 activates G proteins indirectly via stimulation of phospholipase D-lysophosphatidic acid receptor axis and 5-HT 1A receptors in rat brain sections

Abstract

1 The basic secretagogues, such as compound 48/80 (c48/80) and mastoparans, are widely used histamine-releasing agents and their mechanism of action is commonly attributed to a direct, receptor-bypassing property to activate the G i/o class of G proteins. 2 We tested here whether c48/80 could directly stimulate [ 35S]guanosine-5′-[γ-thio]triphosphate ([ 35S]GTPγS) binding to rat brain sections in an attempt to visualize the entire signaling pool of G i/o in its native neuroanatomical context. 3 Instead of direct G i/o activation, c48/80 (100 μg ml -1) from various suppliers stimulated brain phospholipase D (PLD) activity, leading to the generation of endogenous phospholipids capable of activating brain white matter-enriched, G i/o-coupled lysophosphatidic acid (LPA) receptors. This response was sensitive to 1-butanol and was potently reversed by the LPA 1/LPA 3 receptor-selective antagonist Ki16425 (IC 50 59 ± 13 nM, mean ± s.e.m.), and showed age-dependent decline, closely reflecting known developmental regulation of the PLD-LPA 1 receptor axis in the CNS. 4 In addition, c48/80 was found to modestly activate hippocampal 5-HT 1A receptors in a pH-dependent and antagonist-sensitive manner. 5 Consistent with the lack of direct G i/o-activating properties in brain sections, c48/80 showed no activity in classical membrane [ 35S]GTPγS binding assays. Instead, c48/80 from one particular manufacturer elicited non-specific effect in these assays, therefore challenging the previous interpretations regarding the compound's ability to activate G proteins directly. 6 We conclude that c48/80 is not a receptor-bypassing general G protein activator but rather activates PLD, leading to generation of endogenous LPA receptor-activating phospholipids. This property may also contribute to the compound's ability to release histamine from mast cells. © 2006 Nature Publishing Group All rights reserved.