Endogenous NMDA-Receptor Activation Regulates Glutamate Release in Cultured Spinal Neurons

Robert, Antoine, Joel A. Black, and Stephen G. Waxman. En- AMPARs and NMDARs are localized at postsynaptic memdogenous NMDA-receptor activation regulates glutamate release branes in the mature CNS (Kullmann and Siegelbaum 1995;in cultured spinal neurons. J. Neurophysiol. 80: 196–208, 1998. Kullmann et al. 1996; Petralia et al. 1994) .N-methyl-D-aspartate (NMDA) receptor activation plays a funda- Such a view of postsynaptic NMDARs and AMPARs fomental role in the genesis of electrical activity of immature neurons and may participate in activity-dependent aspects of CNS develop- cused at excitatory synapses is contradicted in developing ment. A recent study has suggested that NMDA-receptor–mediated neurons by observations showing that the NMDAR subunits glutamatergic neurotransmission might occur in the developing are expressed diffusely, rather than focused at synapses, in spinal cord via activation of nonsynaptic receptors, but the details cultured spinal cord neurons (O’Brien et al. 1997) or are of NMDA-receptor activation in the developing CNS are not yet disseminated in scattered extrasynaptic clusters in cultured well understood. We describe here a model of cultured spinal neu- hippocampal neurons (Rao and Craig 1997) . In contrast,rons that display ongoing a-amino-3-hydroxy-5-methyl-4-isoxazo- AMPAR subunits are clustered at postsynaptic membranes lepropionic acid (AMPA) receptor activity characterized by spon- (Baude et al. 1995; Craig et al. 1993) . These observations taneous excitatory postsynaptic currents (EPSCs) , with NMDA- raise the question of whether, at early stages of development receptor activity detectable only as single channel events. DL-2-amino-5-phosphonovaleric acid (100 mM) and tetrodotoxin (TTX) of the spinal cord, NMDARs might not participate in excit100 nM each reduced the occurrence of spontaneous AMPA atory neurotransmission. An active role for NMDARs seems,EPSCs; quantal analysis showed a decrease in the number of re- however, likely for several reasons. First, during developleased quanta but no changes in quantal size, indicating that ment, NMDAR activation plays a crucial role in activity-NMDA-receptor activation and Na/ channel activity affect the dependent developmental processes, e.g., in the visual cortex generation of spontaneous AMPA EPSCs, at least in part, via mech- of mammals (Bear et al. 1990; Carmignoto and Vicini 1992;anisms that impinge on the presynaptic terminal. Once the Mg2/- Constantine-Paton et al. 1990; Fox et al. 1989) and in motor block was released, activity of NMDA receptors dramatically in- neuron dendritic outgrowth in the spinal cord (Kalb 1994; creased the release of quantal and multiquantal amounts of gluta-mate, indicating that the NMDA receptors are physiologically cou- Kalb and Hockfield 1994) , where the NMDA receptors are pled to glutamate release. In Mg2/-free solution, TTX application expressed transiently at a high level during the first weeks elicited an increase in the number of quantal AMPA EPSCs and after birth (Kalb et al. 1992) . Second, electrophysiological a reduction in the number of multiquantal EPSCs, consistent with studies have shown that NMDARs contribute to both spontaan effect of NMDA-receptor activation on presynaptic terminals. neous and induced activity, as evidenced by giant synaptic Our results suggest that endogenous activity at a small number of potentials (Ben-Ari et al. 1989) , Ca2/oscillations (Leineku-NMDA receptors can regulate the release of neurotransmitters at gel et al. 1997) , and polysynaptic responses in postnatal developing AMPA synapses..............................