Cellular mechanisms of acute decrease of glutamate release induced by raloxifene in rat cerebral cortex

Abstract

Raloxifene, a selective estrogen receptor modulator, has been observed to offer a neuroprotective effect in several in vitro models of neurotoxicity. An excessive release of glutamate is considered to be related to neuropathology of several neurological diseases. In this study, we investigated whether raloxifene could affect endogenous glutamate release in nerve terminals of rat cerebral cortex (synaptosomes) and explored the possible mechanism. Raloxifene exhibited a dose-dependent inhibition of 4-aminopyridine (4-AP)-evoked release of glutamate, and this effect was not blocked by the estrogen receptor antagonists. The effect of raloxifene on the evoked glutamate release was prevented by the chelating extracellular Ca(2+) ions, and by the vesicular transporter inhibitor bafilomycin A1, but was insensitive to the glutamate transporter inhibitor DL-TBOA. Raloxifene decreased the depolarization-induced increase in the cytosolic free Ca(2+) concentration ([Ca(2+)](C)), whereas it did not alter the resting synaptosomal membrane potential or 4-AP-mediated depolarization. The effect of raloxifene on evoked glutamate release was prevented by blocking the Ca(v)2.2 (N-type) and Ca(v)2.1 (P/Q-type) channels, but not by blocking intracellular Ca(2+) release or Na(+)/Ca(2+) exchange. In addition, the inhibitory effect of raloxifene on evoked glutamate release was abolished by the mitogen-activated/extracellular signal-regulated kinase kinase (MEK) inhibitors, PD98059 and U0126. Furthermore, raloxifene significantly decreased the depolarization-induced phosphorylation of mitogen-activated protein kinase/extracellular signal-regulated kinase 1 and 2 (MAPK/ERK1/2) and synapsin I, the main presynaptic target of ERK. Thus, the effect of raloxifene on evoked glutamate release is linked to a decrease in [Ca(2+)](i) contributed by Ca(2+) entry through presynaptic voltage-dependent Ca(2+) channels and to the subsequent suppression of the ERK/synapsin I signaling cascade.