Biomedical subjects
Lawrence Fourgeaud
Publications and source records attributed to Lawrence Fourgeaud.
A single in vivo exposure to cocaine abolishes endocannabinoid-mediated long-term depression in the nucleus accumbens.
In the nucleus accumbens (NAc), a key structure to the effects of all addictive drugs, presynaptic cannabinoid CB1 receptors (CB1Rs) and postsynaptic metabotropic glutamate 5 receptors (mGluR5s) are the principal effectors of endocannabinoid (eCB)-mediated retrograde long-term depression (LTD) (eCB-LTD) at the prefrontal cortex-NAc synapses. Both CB1R and mGluR5 are involved in cocaine-related behaviors; however, the impact of in vivo cocaine exposure on eCB-mediated retrograde synaptic plasticity remains unknown. Electrophysiological and biochemical approaches were used, and we report that a single in vivo cocaine administration abolishes eCB-LTD. This effect of cocaine was not present in D1 dopamine receptor (D1R) -/- mice and was prevented when cocaine was coadministered with the selective D1R antagonist 8-chloro-2,3,4,5-tetrahydro-3-5-1h-3-benzazepin-7-ol (0.5 mg/kg) or with the NMDA receptor (NMDAR) blocker (+)-5-methyl-10,11-dihydro-5H-dibenzo [a,d] cyclohepten-5,10-imine maleate (1 mg/kg), suggesting the involvement of D1R and NMDAR. We found that the cocaine-induced blockade of retrograde signaling was correlated with enhanced expression levels of Homer scaffolding proteins containing the coiled-coil domain and accompanied by a strong reduction of mGluR5 surface expression. The results suggest that cocaine-induced loss of eCB retrograde signaling is caused by a reduction in the ability of mGluR5 to translate anterograde glutamate transmission into retrograde eCB signaling.
Active surface transport of metabotropic glutamate receptors through binding to microtubules and actin flow.
Receptors for neurotransmitters are concentrated and stabilized at given sites such as synapses through interactions with scaffolding proteins and cytoskeletal elements. The transport of receptors first involves directed vesicular trafficking of intracellularly stored receptors followed by their targeting to the plasma membrane. Once expressed at the cell surface, receptors are thought to reach their final location by random Brownian diffusion in the plasma membrane plane. Here, we investigate whether the metabotropic glutamate receptor mGluR5 can also be transported actively on the cell surface. We used single particle tracking to follow mGluR5 movement in real time at the surface of neuronal growth cones or fibroblast lamellipodia, both of which bear a particularly active cytoskeleton. We found that after a certain lag time mGluR5 undergoes directed rearward transport, which depends on actin flow. On actin depolymerization, directed movement was suppressed, but receptors still bound to a rigid structure. By contrast, receptor transport and immobilization was fully suppressed by microtubule depolymerization but favored by microtubule stabilization. Furthermore, mGluR5 could be immunoprecipitated with tubulin from rat brains, confirming the ability of mGluR5 to bind to microtubules. We propose that mGluR5 can be transported on the cell surface through actin-mediated retrograde transport of microtubules. This process may play a role in receptor targeting and organization during synapse formation or during glutamate-mediated growth cone chemotaxis.
Dynamin 3 is a component of the postsynapse, where it interacts with mGluR5 and Homer.
The dynamins comprise a large family of mechanoenzymes known to participate in membrane modeling events. All three conventional dynamin genes (Dyn1, Dyn2, Dyn3) are expressed in mammalian brain and produce more than 27 different dynamin proteins as a result of alternative splicing. Past studies have suggested that Dyn1 participates in specialized neuronal functions such as rapid synaptic vesicle recycling, while Dyn2 may mediate the conventional clathrin-mediated uptake of surface receptors. Currently, the distribution, expression, and function of Dyn3 in neurons, or in any other cell type, are completely undefined. Here, we demonstrate that Dyn1 and Dyn3 localize differentially in the synapse. Dyn1 concentrates within the presynaptic compartment, while Dyn3 localizes to dendritic spine tips. Within the postsynaptic density (PSD), we found Dyn3, but not Dyn1, to be part of a biochemically isolated complex comprised of Homer and metabotropic glutamate receptors. Finally, although dominant-negative Dyn3 did not seem to inhibit receptor endocytosis, overexpression of a specific Dyn3 spliced variant in mature neurons caused a marked remodeling of dendritic spines. These data suggest that Dyn3 is a postsynaptic dynamin and, like its binding partner Homer, plays a significant role in dendritic spine morphogenesis and remodeling.
The metabotropic glutamate receptor mGluR5 is endocytosed by a clathrin-independent pathway.
Metabotropic glutamate receptors 5 (mGluR5) are members of the growing group C G protein-coupled receptor family. Widely expressed in mammalian brain, they are involved in modulation of the glutamate transmission. By means of transfection of mGluR5 receptors in COS-7 cells and primary hippocampal neurons in culture followed by immunocytochemistry and quantitative image analysis and by a biochemical assay, we have studied the internalization of mGluR5 splice variants. mGluR5a and -5b were endocytosed in COS-7 cells as well as in axons and dendrites of cultured neurons. Endocytosis occurred even in the absence of receptor activity, because receptors mutated in the glutamate binding site were still internalized as well as receptors in which endogenous activity had been inhibited by an inverse agonist. We have measured a constitutive rate of endocytosis of 11.7%/min for mGluR5a. We report for the first time the endocytosis pathway of mGluR5. Internalization of mGluR5 is not mediated by clathrin-coated pits. Indeed, inhibition of this pathway by Eps15 dominant negative mutants did not disturb their endocytosis. However, the large GTPase dynamin 2 is implicated in the endocytosis of mGluR5 in COS-7. mGluR5 is the first shown member of the group C G-protein coupled receptor family internalized by a nonconventional pathway.
Receptor activation and homer differentially control the lateral mobility of metabotropic glutamate receptor 5 in the neuronal membrane.
Glutamate receptors are clustered at the membrane through interactions with intracellular scaffolding proteins and cytoskeletal elements but can also be found in intracellular compartments or dispersed in the membrane. This distribution results from an equilibrium between the different pools of receptors whose dynamic is poorly known. The group I metabotropic glutamate receptor 5 (mGluR5) is concentrated in an annulus around the postsynaptic density but also found in large amounts in the extrasynaptic membrane. To analyze the dynamic of stabilization of mGluR5, we used single-particle tracking, force measurements, and fluorescence recovery to measure the mobility of mGluR5. We found that receptor activation increases receptor diffusion, whereas the scaffolding protein Homer favors confinement of receptor movements within clusters of Homer-mGluR5. However, this stabilization is reversible, because even in the presence of Homer, receptors still enter and exit from clusters at fast rates. Furthermore, clusters themselves are highly dynamic both in their movements and in their composition, which can vary within tens of seconds. Thus, exchange of receptors between dispersed and clustered states is fast and regulated during physiological processes. These properties may explain certain fast changes in receptor composition observed at postsynaptic densities.