Targeting neurotransmitter receptors with nanoparticles in vivo allows single molecule tracking in acute brain slices

Single-molecule imaging has changed the way we understand many biological mechanisms, particularly in neurobiology, by shedding light on intricate molecular events down to the nanoscale. However, current single-molecule studies in neuroscience have been limited to cultured neurons or organotypic slices, leaving as an open question the existence of fast receptor diffusion in intact brain tissue. Here, for the first time, we targeted dopamine receptors in vivo with functionalized quantum dots and were able to perform single-molecule tracking in acute rat brain slices. We propose a novel delocalized and non-inflammatory way of delivering nanoparticles (NPs) in vivo to the brain, which allowed us to label and track genetically engineered surface dopamine receptors in neocortical neurons, revealing inherent behaviour and receptor activity regulations. We thus propose a NP-based platform for single-molecule studies in the living brain, opening new avenues of research in physiological and pathological animal models

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

International audienceDopamine 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

Unraveling the Functions of Endogenous Receptor Oligomers in the Brain Using Interfering Peptide: The Example of D1R/NMDAR Heteromers

Decoding signaling pathways in different brain structures is crucial to develop pharmacological strategies for neurological diseases. In this perspective, the targeting of receptors by selective ligands is one of the classical therapeutic strategies. Nonetheless, this approach often results in a decrease of efficiency over time and deleterious side effects because physiological functions can be affected. An emerging concept has been to target mechanisms that fine-tune receptor signaling, such as heteromerization, the process by which physical receptor-receptor interaction at the membrane allows the reciprocal modulation of receptors' signaling. Because of the central role of the synergistic transmission mediated by dopamine (DA) and glutamate (Glu) in brain physiology and pathophysiology, heteromerization between DA and Glu receptors has received a lot of attention. However, the study of endogenous heteromers has been challenging because of the lack of appropriate tools. Over the last years, progress has been made in the development of techniques to study their expression in the brain, regulation and function. In this chapter, we provide a methodological framework for the design and use of interfering peptides to study endogenous receptor oligomers through the example of the dopamine type 1 receptor (D1R) and the GluN1 subunit of NMDA receptor heteromers

Dopamine Receptors in the Subthalamic Nucleus: Identification and Localization of D5 Receptors

International audienceHerein we present methodological approaches for the identification and characterization of dopamine receptors in the subthalamic nucleus, a component nucleus of the basal ganglia, at pre-and postsynaptic locations and of their roles with an emphasis given to the dopamine D5 receptor subtype. This chapter focuses on the possible sources of divergence between electrophysiological studies and describes the pharmacological tools available for functional studies of this receptor. The procedures for single-cell reverse transcription PCR (polymerase chain reaction) identification of dopamine D5 receptor mRNA and the immunochemical detection of the receptor at cellular and subcellular levels are presented

Hints on the Lateralization of Dopamine Binding to D1 Receptors in Rat Striatum

Dopamine receptors in striatum are important for healthy brain functioning and are the target of levodopa-based therapy in Parkinson's disease. Lateralization of dopaminergic neurotransmission in striata from different hemispheres occurs in patients, but also in healthy individuals. Our data show that the affinity of dopamine binding to dopamine D1 receptors is significantly higher in left than in right striatum. Analysis of data from radioligand binding to striatal samples from naïve, 6-hydroxydopamine lesioned, levodopa-treated and levodopa-induced dyskinetic rats shows differential receptor structure and gives hints on the causes of right/left lateralization of dopamine binding to striatal D1 receptors. Moreover, binding data showed loss of lateralization in levodopa (L-DOPA)-induced dyskinetic rats