Adrenergic modulation of astroglial phospholipase D activity and cell proliferation

As phospholipase D (PLD) activation has been associated with mitogenic signalling in several cell types, we tested an association between adrenergic activation of PLD and cellular proliferation in primary cultures of rat cortical astrocytes. In 2-week old cultures, PLD activation by noradrenaline (EC50: 0.49 microM) was inhibited by prazosin, a specific antagonist at alpha1-adrenergic receptors (IC50: 0.23 microM). Adrenergic PLD activation was not affected by genistein, an inhibitor of tyrosine kinases, or by Ro 31-8220, an inhibitor of protein kinase C (PKC), but was dose-dependently depressed in the presence of brefeldin A (1-100 microg/ml), an inhibitor of ARF activation. In experiments measuring cell proliferation, noradrenaline potently (EC50: 20 nM) reduced [3H]thymidine incorporation to 20-30% of basal values. This action was mimicked by the beta-specific agonist isoprenaline and was inhibited by the beta-antagonist propranolol in a concentration-dependent manner. The alpha1-adrenergic agonists, phenylephrine and methoxamine, also reduced DNA synthesis. The adrenergic inhibition of astroglial DNA synthesis was not reduced, but further potentiated in the presence of brefeldin A, ethanol, and 1- and 2-butanol; 1-butanol, a substrate of PLD, was equally effective as 2-butanol, a non-substrate. We conclude that adrenergic PLD activation in astrocytes is not involved in mitogenic signalling. The involvement of ARF in the activation of PLD via alpha1-adrenoceptors indicates a role in protein trafficking.