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Diffusion barriers constrain receptors at synapses.

ABSTRACT: The flux of neurotransmitter receptors in and out of synapses depends on receptor interaction with scaffolding molecules. However, the crowd of transmembrane proteins and the rich cytoskeletal environment may constitute obstacles to the diffusion of receptors within the synapse. To address this question, we studied the membrane diffusion of the ?-aminobutyric acid type A receptor (GABA(A)R) subunits clustered (?2) or not (?5) at inhibitory synapses in rat hippocampal dissociated neurons. Relative to the extrasynaptic region, ?2 and ?5 showed reduced diffusion and increased confinement at both inhibitory and excitatory synapses but they dwelled for a short time at excitatory synapses. In contrast, ?2 was ~3-fold more confined and dwelled ~3-fold longer in inhibitory synapses than ?5, indicating faster synaptic escape of ?5. Furthermore, using a gephyrin dominant-negative approach, we showed that the increased residency time of ?2 at inhibitory synapses was due to receptor-scaffold interactions. As shown for GABA(A)R, the excitatory glutamate receptor 2 subunit (GluA2) of the ?-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) had lower mobility in both excitatory and inhibitory synapses but a higher residency time at excitatory synapses. Therefore barriers impose significant diffusion constraints onto receptors at synapses where they accumulate or not. Our data further reveal that the confinement and the dwell time but not the diffusion coefficient report on the synapse specific sorting, trapping and accumulation of receptors.

SUBMITTER: Renner M

PROVIDER: S-EPMC3418229 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Diffusion barriers constrain receptors at synapses.

Renner Marianne M Schweizer Claude C Bannai Hiroko H Triller Antoine A Lévi Sabine S

PloS one 20120813 8

The flux of neurotransmitter receptors in and out of synapses depends on receptor interaction with scaffolding molecules. However, the crowd of transmembrane proteins and the rich cytoskeletal environment may constitute obstacles to the diffusion of receptors within the synapse. To address this question, we studied the membrane diffusion of the γ-aminobutyric acid type A receptor (GABA(A)R) subunits clustered (γ2) or not (α5) at inhibitory synapses in rat hippocampal dissociated neurons. Relativ ...[more]

PMID: 22912780

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Project description:Using both optical and electrical methods, we document that solute diffusion in the cytoplasm of BL6 murine cardiac myocytes becomes restricted >30-fold as molecular weight increases from 30 to 2000, roughly as expected for pores with dimensions of cardiac porin channels. The Bodipy-FL ATP analogue diffuses ∼50-fold slower in BL6 cardiac cytoplasm than in free water. From several fluorophores analyzed, our estimates of bound fluorophore fractions range from 0.1 for a 2 kD FITC-labeled polyethylene glycol to 0.93 for sulforhodamine. We estimate that diffusion coefficients of unbound fluorophores range from 0.5 to 8 x 10 -7 cm 2 /s. Analysis of Na/K pump and veratridine-modified Na channel currents confirms that Na diffusion is nearly unrestricted (time constant for equilibration with the pipette tip, ∼20 s). Using three different approaches, we estimate that ATP diffuses 8 to 10-times slower in the cytoplasm of BL6 myocytes than in free water. To address whether restrictions are caused more by cytoplasmic protein or membrane networks, we verified first that a protein gel, 10 gram% gelatin, restricts solute diffusion with strong dependence on molecular weight. Solute diffusion in membrane-extracted cardiac myofilaments, confined laterally by suction into large-diameter pipette tips, is however less restricted than in intact myocytes. Notably, myofilaments from equivalently extracted skeletal (diaphragm) myocytes restrict diffusion less than cardiac myofilaments. Solute diffusion in myocytes with sarcolemma permeabilized by β-escin (80 µM) is similarly restricted as in intact myocytes. Diffusion restriction in cardiac myocytes is strain-dependent, being about two-fold greater in BL6 myocytes than in myocytes with a CD1/J6/129svJ background. Furthermore, diffusion is 2.5-fold more restricted in CD1/J6/129svJ myocytes lacking the mitochondrial porin, Vdac1, than in WT CD1/J6/129svJ myocytes. We conclude that both myofilaments and mitochondria networks restrict diffusion in cardiac myocytes. As a result, long-range solute diffusion may preferentially occur via passage through porin channels and intramembrane mitochondrial spaces, where diffusion is less restricted than in myofilament spaces.

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Diffusion barriers constrain receptors at synapses.

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Diffusion barriers constrain receptors at synapses.

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