Glycine Transporter 1 Modulates Gaba Release from Amacrine Cells 1 by Controlling Occupancy of Coagonist Binding Site of Nmda 2 Receptors

26 The occupancy of coagonist binding sites of NMDA receptors (NMDARs) by glycine or D-27 serine has been thought to mediate NMDAR-dependent excitatory signaling, as simultaneous 28 binding of glutamate and a coagonist is obligatory for NMDAR activation. Amacrine cells (ACs) 29 mediating GABAergic feedback inhibition of mixed bipolar cells (Mbs) in the goldfish retina 30 have been shown to express NMDARs. Here we studied whether NMDAR mediated 31 GABAergic inhibitory currents (IGABA) recorded from the axon terminals of Mbs are influenced 32 by experimental manipulations altering retinal glycine and D-serine levels. 33 Feedback IGABA in Mb axon terminals was triggered by focal NMDA application or by 34 synaptically released glutamate from depolarized Mb terminals. In both cases, blocking the 35 coagonist binding sites of NMDARs eliminated the NMDAR-dependent IGABA demonstrating 36 that coagonist binding is critical in mediating NMDAR activity-triggered GABA release. Glycine 37 transporter 1 (GLYT1) inhibition increased IGABA indicating that coagonist binding sites of 38 NMDARs on ACs providing GABAergic feedback inhibition to Mbs were not saturated. Focal 39 glycine application, in the presence of ionotropic glycine receptor blocker strychnine, triggered 40 a GLYT1-dependent current in ACs, suggesting that GLYT1 expressed by putative glycinergic 41 ACs control the saturation level of NMDARs' coagonist sites. External D-serine also increased 42 NMDAR activation triggered IGABA in Mbs, further substantiating that the coagonist sites were 43 unsaturated. Together, our findings demonstrate that coagonist modulation of glutamatergic 44 input to GABAergic ACs via NMDARs is strongly reflected in the AC neuronal output (i.e. 45 transmitter release) thus it is critical in GABAergic signal transfer function in the inner retina.