D-APV blocking of MC spike potentiation in this in vitro model suggests b-adrenoceptor activation paired with odor input triggers a NMDAR-dependent potentiation of odor-encoding MCs. One route for the b-adrenoceptor-mediated activation of NMDARs on MCs to promote the LTP of MC responses could be disinhibition. The previous in vitro study suggested that b-adrenoceptor activation by isoproterenol suppressed evoked EPSCs in periglomerular cells in the olfactory bulb slice. If disinhibition is important in the opening of NMDARs on mitral cells, we expected increasing glomerular inhibition would counteract the isoproterenol effect in potentiating MCs when paired with TBS, and local glomerular disinhibition could, by itself, lead to NMDAR-dependent potentiation of MC spikes. To test the role of the NMDAR in our in vivo learning model, we first examined whether the NMDAR is activated following early odor Nutlin-3 preference learning and the localization of its activation in the olfactory bulb. It has been shown that phosphorylation of GluN1 affects the kinetics of the NMDAR, resulting in a larger current and greater calcium influx. We used immunohistochemical staining with an antibody recognizing the PKA phosphorylation site of the obligatory GluN1 subunit of the NMDAR. The location of pGluN1 activation is consistent with previous reports using a 2-DG tracing technique showing that peppermint odor activates glomerular ����hot spots���� in the mid-lateral portion of olfactory bulb glomeruli and that these peppermint ����hot spots���� were enlarged in rat pups who underwent odor learning. In the current study, immunohistochemistry revealed that pGluN1 staining in the glomerular layer was seen in processes and may correspond to dendritic structures in the glomeruli such as MC and tufted cell dendrites. We also observed staining in small glial-like cells. We did not further pursue the identity of those cells but glial cells in the olfactory bulb express GluN1 and glial activity in the Axitinib VEGFR/PDGFR inhibitor glomerulus mirrors that of MCs. We next explored a potential causal role of NMDAR activation in mediating odor preference learning. We directly infused isoproterenol into the olfactory bulbs of rat pups during odor training to induce odor preference learning and tested whether co-application of D-APV, a NMDAR antagonist, would block learning. We established a method that allowed us to infuse the drug mainly into the superficial layer of the olfactory bulb on the lateral surface, where enhanced pGluN1 expression was observed following odor preference learning. NMDAR is one of the excitatory synaptic transmission receptors at ON-MC synapses, as well as mediating synaptic transmission from MCs to granule cells. Depending on the synaptic site, NMDAR blockade would have differential effects on MC excitation. Previous research suggests that the NMDAR augments a long-lasting depolarization of MCs to ON stimulation.