The defect in hippocampal synaptic plasticity in Neto1 tlz/tlz mutant mice suggested these mice might be impaired in hippocampal-dependant spatial learning and memory. Neto1tlz/tlz mutant mice were analyzed in the Morris water maze task and were found to have impaired spatial learning. The reduced LTP and the disruption of spatial learning in Neto1tlz/tlz mice are consistent with the decrease of NMDA-mediated currents, which can be explained by fewer receptors at hippocampal excitatory synapses. These findings highlight the important role of Neto1 in the regulation of synaptic NMDA receptors, and suggest it may play an important role in diseases affecting synaptic plasticity.Neto1 and Neto2 are homologous CUB (Clr, Cls/Uegf/Bmp1) domain proteins related to neuropilin and tolloid. Both Neto1 and Neto2 are expressed in the developing mouse embryo and the mature brain. To begin to understand the role of the Neto genes in the nervous system, Neto1 and Neto2 knockout mice were generated. While Neto1tlz/tlz and Neto2-/- mice are viable, fertile, and have no major morphological abnormalities of the brain, they show defects in axon projections and exhibit myoclonic-like seizures. These findings suggest that the Neto proteins are important for neural development and maintenance.Neto1 localizes to the postsynaptic density (PSD) of excitatory synapses and associates bivalently with N-methyl-D-aspartate (NMDA) receptors. One interaction between Neto1 and NMDA receptors is mediated by PSD-95, a major PSD scaffold protein, via the C-terminal PDZ tripeptide ligand of Neto1. Another interaction occurs independently of PSD-95, and is mediated by the first extracellular CUB domain of Neto1. Thus, Neto1 associates with NMDA receptors through intracellular and extracellular contacts.To determine whether NMDA receptor function is disrupted in Neto1tlz/tlz mice, long-term potentiation (LTP) was measured at Schaffer collateral-CA1 synapses in hippocampal slices. LTP and NMDA receptor-mediated currents were depressed at Neto1 tlz/tlz synapses. Furthermore, Neto1tlz/tlz mice have a specific reduction of NR2A-containing NMDA receptors at hippocampal synapses, but not in the overall expression level of these receptors in cells. These findings suggest that Neto1 plays a critical role in NMDA receptor-dependant synaptic plasticity, and in the delivery or stability of NR2A-containing NMDA receptors at these synapses.