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Wild-Type Monomeric ?-Synuclein Can Impair Vesicle Endocytosis and Synaptic Fidelity via Tubulin Polymerization at the Calyx of Held.

ABSTRACT: ?-Synuclein is a presynaptic protein the function of which has yet to be identified, but its neuronal content increases in patients of synucleinopathies including Parkinson's disease. Chronic overexpression of ?-synuclein reportedly expresses various phenotypes of synaptic dysfunction, but the primary target of its toxicity has not been determined. To investigate this, we acutely loaded human recombinant ?-synuclein or its pathological mutants in their monomeric forms into the calyces of Held presynaptic terminals in slices from auditorily mature and immature rats of either sex. Membrane capacitance measurements revealed significant and specific inhibitory effects of WT monomeric ?-synuclein on vesicle endocytosis throughout development. However, the ?-synuclein A53T mutant affected vesicle endocytosis only at immature calyces, whereas the A30P mutant had no effect throughout. The endocytic impairment by WT ?-synuclein was rescued by intraterminal coloading of the microtubule (MT) polymerization blocker nocodazole. Furthermore, it was reversibly rescued by presynaptically loaded photostatin-1, a photoswitcheable inhibitor of MT polymerization, in a light-wavelength-dependent manner. In contrast, endocytic inhibition by the A53T mutant at immature calyces was not rescued by nocodazole. Functionally, presynaptically loaded WT ?-synuclein had no effect on basal synaptic transmission evoked at a low frequency, but significantly attenuated exocytosis and impaired the fidelity of neurotransmission during prolonged high-frequency stimulation. We conclude that monomeric WT ?-synuclein primarily inhibits vesicle endocytosis via MT overassembly, thereby impairing high-frequency neurotransmission.SIGNIFICANCE STATEMENT Abnormal ?-synuclein abundance is associated with synucleinopathies including Parkinson's disease, but neither the primary target of ?-synuclein toxicity nor its mechanism is identified. Here, we loaded monomeric ?-synuclein directly into mammalian glutamatergic nerve terminals and found that it primarily inhibits vesicle endocytosis and subsequently impairs exocytosis and neurotransmission fidelity during prolonged high-frequency stimulation. Such ?-synuclein toxicity could be rescued by blocking microtubule polymerization, suggesting that microtubule overassembly underlies the toxicity of acutely elevated ?-synuclein in the nerve terminal.

SUBMITTER: Eguchi K

PROVIDER: S-EPMC6596501 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Wild-Type Monomeric α-Synuclein Can Impair Vesicle Endocytosis and Synaptic Fidelity via Tubulin Polymerization at the Calyx of Held.

Eguchi Kohgaku K Taoufiq Zacharie Z Thorn-Seshold Oliver O Trauner Dirk D Hasegawa Masato M Takahashi Tomoyuki T

The Journal of neuroscience : the official journal of the Society for Neuroscience 20170602 25

α-Synuclein is a presynaptic protein the function of which has yet to be identified, but its neuronal content increases in patients of synucleinopathies including Parkinson's disease. Chronic overexpression of α-synuclein reportedly expresses various phenotypes of synaptic dysfunction, but the primary target of its toxicity has not been determined. To investigate this, we acutely loaded human recombinant α-synuclein or its pathological mutants in their monomeric forms into the calyces of Held pr ...[more]

PMID: 28576942

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Project description:Key pointsDuring development the giant, auditory calyx of Held forms a one-to-one connection with a principal neuron of the medial nucleus of the trapezoid body. While anatomical studies described that most of the target cells are temporarily contacted by multiple calyces, multi-calyceal innervation was only sporadically observed in in vivo recordings, suggesting a structure-function discrepancy. We correlated synaptic strength of inputs, identified in in vivo recordings, with post hoc labelling of the recorded neuron and synaptic terminals containing vesicular glutamate transporters (VGluT). During development only one input increased to the level of the calyx of Held synapse, and its strength correlated with the large VGluT cluster contacting the postsynaptic soma. As neither competing strong inputs nor multiple large VGluT clusters on a single cell were observed, our findings did not indicate a structure-function discrepancy.AbstractIn adult rodents, a principal neuron in the medial nucleus of the trapezoid (MNTB) is generally contacted by a single, giant axosomatic terminal called the calyx of Held. How this one-on-one relation is established is still unknown, but anatomical evidence suggests that during development principal neurons are innervated by multiple calyces, which may indicate calyceal competition. However, in vivo electrophysiological recordings from principal neurons indicated that only a single strong synaptic connection forms per cell. To test whether a mismatch exists between synaptic strength and terminal size, we compared the strength of synaptic inputs with the morphology of the synaptic terminals. In vivo whole-cell recordings of the MNTB neurons from newborn Wistar rats of either sex were made while stimulating their afferent axons, allowing us to identify multiple inputs. The strength of the strongest input increased to calyceal levels in a few days across cells, while the strength of the second strongest input was stable. The recorded cells were subsequently immunolabelled for vesicular glutamate transporters (VGluT) to reveal axosomatic terminals with structured-illumination microscopy. Synaptic strength of the strongest input was correlated with the contact area of the largest VGluT cluster at the soma (r = 0.8), and no indication of a mismatch between structure and strength was observed. Together, our data agree with a developmental scheme in which one input strengthens and becomes the calyx of Held, but not with multi-calyceal competition.

Dataset Information

Wild-Type Monomeric ?-Synuclein Can Impair Vesicle Endocytosis and Synaptic Fidelity via Tubulin Polymerization at the Calyx of Held.

Publications

Wild-Type Monomeric α-Synuclein Can Impair Vesicle Endocytosis and Synaptic Fidelity via Tubulin Polymerization at the Calyx of Held.

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