NMDAR mediated calcium transients elicited by glutamate co-release at developing inhibitory synapses in the auditory brainstem

Before hearing onset, synapses in the inhibitory pathway from the medial nucleus of the trapezoid body (MNTB) to the lateral superior olive (LSO) not only release GABA and glycine, but also glutamate. This transient glutamate release from nominally inhibitory synapses coincides with the period in which the MNTB-LSO pathway is tonotopically refined. I hypothesized that by releasing glutamate, developing MNTB-LSO synapses can elicit NMDA receptor (NMDAR) mediated calcium influx, which has been shown to underlie the refinement of excitatory topographic maps. To test my hypothesis, I designed and built a 2-photon microscopy system to perform two complementary series of calcium (Ca2+) imaging experiments. The objective of the first series of experiments was to investigate the potential of developing MNTB-LSO synapses to elicit NMDAR-mediated Ca2+ responses. Experiments were performed under conditions that maximized the ability for GABA/glycinergic conductances to aid in the relief of magnesium (Mg2+) block at NMDARs. Electrical stimulation of the MNTB-LSO pathway consistently elicited local Ca2+ transients in LSO dendrites that were significantly reduced by NMDAR antagonists. Despite their significant contribution to MNTB-elicited dendritic Ca2+ responses, NMDARs did not contribute to somatically recorded electrical responses. In the second series of experiments, I investigated the ability for MNTB-LSO synapses to activate NMDAR-mediated Ca2+ transients using only imaging techniques. Here we found that MNTB-LSO inputs by themselves could not elicit somatic NMDAR-mediated Ca2+ responses when GABA/glycinergic conductances were present with extracellular Mg+, despite having the potential to elicit these Ca2+ responses. These results suggest that GABA/glycinergic conductances shunt the somatically-recorded Ca2+ response. They raise the question of whether MNTB-LSO inputs can elicit local dendritic NMDAR-mediated Ca2+ responses independently from other LSO inputs.Together, these experiments demonstrate the ability of developing MNTB-LSO inputs to activate NMDAR Ca2+ signaling pathways during the time of topographic map refinement. Although this ability was only observed under conditions that altered or abolished GABA/glycinergic conductances or extracellular Mg2+, this ability could endow developing inhibitory synapses with a novel and potentially powerful refinement mechanism to be activated under physiological conditions.