Neuronal activity regulates the synaptic strength of neuronal networks. However, it is still unclear how diminished activity changes connection patterns in neuronal circuits. To address this issue, we analyzed neuronal connectivity and relevant mechanisms using hippocampal cultures in which developmental synaptogenesis had occurred. We show that diminution of network activity in mature neuronal circuit promotes reorganization of neuronal circuits via NR2B subunit-containing NMDA-type glutamate receptors (NR2B-NMDARs), which mediate silent synapse formation. Simultaneous double whole-cell recordings revealed that diminishing neuronal circuit activity for 48 h increased the number of synaptically connected neuron pairs with both silent and functional synapses. This increase was accompanied by the specific expression of NR2B-NMDARs at synaptic sites. Analysis of miniature EPSCs (mEPSCs) showed that the frequency of NMDAR-mediated, but not AMPAR-mediated, mEPSCs increased, indicating that diminished neuronal activity promotes silent synapse formation via the surface delivering NR2B-NMDARs in mature neurons. After activation of neuronal circuit by releasing from TTX blockade (referred as circuit reactivation), the frequency of AMPAR-mediated mEPSCs increased instead, and this increase was prevented by ifenprodil. The circuit reactivation also caused an increased colocalization of glutamate receptor 1-specfic and synaptic NR2B-specific puncta. These results indicate that the circuit reactivation converts rapidly silent synapses formed during activity suppression to functional synapses. These data may provide a new example of homeostatic circuit plasticity that entails the modulation of neuron-neuron connectivity by synaptic activity.

Multiple system atrophy (MSA) is a neurodegenerative disease caused by an accumulation of ␣-synuclein (␣-syn) in oligodendrocytes. Little is known about the cellular mechanisms by which ␣-syn accumulation causes neuronal degeneration in MSA. Our previous research, however, revealed that in a mouse model of MSA, oligodendrocytic inclusions of ␣-syn induced neuronal accumulation of ␣-syn, as well as progressive neuronal degeneration. Here we identify the mechanisms that underlie neuronal accumulation of ␣-syn in a mouse MSA model. We found that the ␣-syn protein binds to ␤-III tubulin in microtubules to form an insoluble complex. The insoluble ␣-syn complex progressively accumulates in neurons and leads to neuronal dysfunction. Furthermore, we demonstrated that the neuronal accumulation of insoluble ␣-syn is suppressed by treatment with a microtubule depolymerizing agent. The underlying pathological process appeared to also be inhibited by this treatment , providing promise for future therapeutic approaches.

Dentatorubral‐pallidoluysian atrophy is caused by polyglutamine (polyQ) expansion in atrophin‐1 (ATN1). Recent studies have shown that nuclear accumulation of ATN1 and cleaved fragments with expanded polyQ is the pathological process underlying neurodegeneration in dentatorubral‐pallidoluysian atrophy. However, the mechanism underlying the proteolytic processing of ATN1 remains unclear. In the present study, we examined the proteolytic processing of ATN1 aiming to understand the mechanisms of ATN1 accumulation with polyQ expansion. Using COS‐7 and Neuro2a cells that express the ATN1 gene, in which ATN1 was accumulated by increasing the number of polyQs, we identified a novel C‐terminal fragment containing a polyQ tract. The mutant C‐terminal fragment with expanded polyQ selectively accumulated in the cells, and this was also demonstrated in the brain tissues of patients with dentatorubral‐pallidoluysian atrophy. Immunocytochemical and biochemical studies revealed that full‐length ATN1 and C‐terminal fragments displayed individual localization. The mutant C‐terminal fragment was preferentially found in the cytoplasmic membrane/organelle and insoluble fractions. Accordingly, it is assumed that the proteolytic processing of ATN1 regulates the localization of C‐terminal fragments. Accumulation of the C‐terminal fragment was enhanced by inhibition of caspases in the cytoplasm of COS‐7 cells. Collectively, these results suggest that the C‐terminal fragment plays a principal role in the pathological accumulation of ATN1 in dentatorubral‐pallidoluysian atrophy.

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