Single-molecule imaging of the functional crosstalk between surface NMDA and dopamine D1 receptors

Dopamine is a powerful modulator of glutamatergic neurotransmission and NMDA receptor-dependent synaptic plasticity. Although several intracellular cascades participating in this functional dialogue have been identified over the last few decades, the molecular crosstalk between surface dopamine and glutamate NMDA receptor (NMDAR) signaling still remains poorly understood. Using a combination of single-molecule detection imaging and electrophysiology in live hippocampal neurons, we demonstrate here that dopamine D1 receptors (D1Rs) and NMDARs form dynamic surface clusters in the vicinity of glutamate synapses. Strikingly, D1R activation or D1R/NMDAR direct interaction disruption decreases the size of these clusters, increases NMDAR synaptic content through a fast lateral redistribution of the receptors, and favors long-term synaptic potentiation. Together, these data demonstrate the presence of dynamic D1R/NMDAR perisynaptic reservoirs favoring a rapid and bidirectional surface crosstalk between receptors and set the plasma membrane as the primary stage of the dopamine–glutamate interplay.Hippocampal dopaminergic neuromodulation participates in several cognitive functions including novelty detection and long-term memory storage (1, 2). As a consequence, impairments in hippocampal neuromodulatory transmission affect synaptic plasticity at glutamatergic synapses, prevent learning and memory formation, and have been proposed to be a cellular substrate for neurodevelopmental psychiatric disorders such as schizophrenia (3). In the hippocampus and cortex, pyramidal neurons express mostly dopamine D1 and D5 receptors along their dendritic tree (4–6). Their recruitment affects the trafficking and surface expression of glutamate NMDA receptors (NMDARs), two processes that are essential for excitatory neurotransmission and synaptic plasticity. Indeed, activating dopamine D1 receptors (D1Rs) promotes the surface expression and function of NMDAR and thereby favors the long-term potentiation of excitatory glutamate synapses (7–10). Reciprocally, the activation of NMDAR modulates D1R surface expression and signaling (11). The bidirectional dialogue between dopamine and glutamate NMDAR-associated signaling thus involves changes in membrane receptor content and trafficking.Although this functional interaction is usually considered as relying on intracellular protein kinase signaling cascades (7, 10, 12), physical interactions between D1R and NMDAR at the plasma membrane were recently reported to stabilize laterally diffusing surface D1R in spines, modulate D1R- and NMDAR-mediated signaling, and influence working memory (13–16). Thus, direct interactions between these receptors could contribute to the regulation of their surface distributions and play a major role in the dopamine–glutamate interplay (15, 17). In particular, because the regulation of NMDAR synaptic content involves surface diffusion processes in and out of synaptic and extrasynaptic compartments (18), the possibility emerges that dopamine might modulate NMDAR-dependent synaptic transmission by tuning NMDAR lateral dynamics through D1R–NMDAR physical interactions. To address this question and investigate the role of the D1R/NMDAR surface crosstalk in synaptic physiology, we here assessed the surface distribution and trafficking of D1 and NMDA receptors in rat hippocampal neurons using a combination of high-resolution single-nanoparticle tracking, bulk imaging, and electrophysiology.