Abstract
Following prolonged activity blockade, amplitudes of miniature excitatory postsynaptic currents (mEPSCs) increase, a form of plasticity termed “homeostatic synaptic plasticity.” We previously showed that a presynaptic protein, the small GTPase Rab3A, is required for full expression of the increase in miniature endplate current amplitudes following prolonged blockade of action potential activity at the mouse neuromuscular junction in vivo (Wang et al., 2011), but it is unknown whether this form of Rab3A-dependent homeostatic plasticity shares any characteristics with central synapses. We show here that homeostatic synaptic plasticity of mEPSCs is impaired in mouse cortical neuron cultures prepared from Rab3A−/− and mutant mice expressing a single point mutation of Rab3A, Rab3A Earlybird mice. To determine if Rab3A is involved in the well-established homeostatic increase in postsynaptic AMPA-type receptors (AMPARs), we performed a series of experiments in which electrophysiological recordings of mEPSCs and confocal imaging of synaptic AMPAR immunofluorescence were assessed within the same cultures. We found that Rab3A was required for the increase in synaptic AMPARs following prolonged activity blockade, but the increase in mEPSC amplitudes was not always accompanied by an increase in postsynaptic AMPAR levels, suggesting other factors may contribute. Finally, we demonstrate that Rab3A is acting in neurons because only selective loss of Rab3A in neurons, not glia, disrupted the homeostatic increase in mEPSC amplitudes. This is the first demonstration that neuronal Rab3A is required for homeostatic synaptic plasticity and that it does so partially through regulation of the surface expression of AMPA receptors.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
The authors have no competing interests.
Major Changes: 1. We revised our model (old Fig. 10; new Fig. 9) to focus on the data shown in the current study. The mechanisms depicted are confined to pre– or post-synaptic Rab3A participating in either controlling release of a trophic factor that regulates surface GluA2 receptors (pre– or postsynaptic) or directly affecting fusion of GluA2-receptor containing vesicles (postsynaptic). 2. We replaced all cumulative density function plots and ratio plots, based on multiple quantile samples per cell, with box plots of cell means. This affects new Figures 1, 2, 3, 5, 6, 7 and 8. All references to scaling, divergent scaling, or uniform scaling, have been removed. The number of cultures is provided in the figure legends. 3. We have added frequency data to Figures 1, 2 and 8 and included text to describe these data. 4. We added analysis of VGLUT1 levels for the same synaptic sites that we previously analyzed GluA2 levels, and these data are described in Results, lines 344 to 371, and appear in new Table 2. In contrast to previous studies, we did not find any evidence for an increase in VGLUT1 levels after activity blockade. We reviewed those studies to determine whether there might be differences in the experimental details that could explain the lack of effect we observed.
