Phospholipase A 2 activation by melittin enhances spontaneous glutamatergic excitatory transmission in rat substantia gelatinosa neurons

Abstract

In order to know a role of phospholipase A 2 in modulating nociceptive transmission, the effect of a secreted phospholipase A 2 activator melittin on spontaneous glutamatergic excitatory transmission was investigated in substantia gelatinosa neurons of an adult rat spinal cord slice by using the whole-cell patch-clamp technique. Bath-applied melittin at concentrations higher than 0.5μM increased both the amplitude and the frequency of spontaneous excitatory postsynaptic current in a manner independent of tetrodotoxin; the latter effect of which was examined in detail. In 80% of the neurons examined ( n=64), melittin superfused for 3 min gradually increased spontaneous excitatory postsynaptic current frequency (by 65±6% at 1μM; n=51) in a dose-dependent manner (effective concentration for half-maximal effect=1.1μM). This effect subsided within 3 min after washout. The spontaneous excitatory postsynaptic current frequency increase produced by melittin was reduced by the phospholipase A 2 inhibitor 4-bromophenacryl bromide (10μM) while being unaffected by the cyclooxygenase inhibitor indomethacin (100μM) and the lipoxygenase inhibitor nordihydroguaiaretic acid (100μM). A similar increase in spontaneous excitatory postsynaptic current frequency was produced by exogenous arachidonic acid (50μM); this effect was also unaffected by the cyclooxygenase or lipoxygenase inhibitor. Melittin failed to increase spontaneous excitatory postsynaptic current frequency in a nominally Ca 2+-free or La 3+-containing Krebs solution. We conclude that melittin increases the spontaneous release of l-glutamate to substantia gelatinosa neurons by activating secreted phospholipase A 2 and increasing Ca 2+ influx through voltage-gated Ca 2+ channels in nerve terminals, probably with an involvement of arachidonic acid but not its metabolites produced by cyclooxygenase and lipoxygenase. Considering that the substantia gelatinosa plays an important role in regulating nociceptive transmission, it is suggested that this transmission may be positively modulated by secreted phospholipase A 2 activation in the substantia gelatinosa.