Ubiquitination Acutely Regulates Presynaptic Neurotransmitter Release in Mammalian Neurons

Abstract

The ubiquitin proteasome system (UPS) plays a crucial role in modulating synaptic physiology both presynaptically and postsynaptically, but the regulatory mechanisms remain obscure. To determine acute effects of proteasome inhibition on neurotransmission, we performed whole-cell voltage-clamp recordings from cultured rodent hippocampal neurons. We find that proteasome inhibitors induce a strikingly fast, severalfold increase in the frequency of both miniature (mini) and spontaneous synaptic currents at excitatory and inhibitory synapses. The lack of change in mini amplitude and kinetics indicates a presynaptic site of action. This effect does not depend on increased levels of presynaptic proteins, previously suggested as proteasomal targets. Furthermore, blockade of the UPS using E1-activating enzyme inhibitors also increases mini frequency, demonstrating that accumulation of ubiquitinated proteins is not required. Overall, these data suggest that the UPS not only orchestrates protein turnover, but also dynamically regulates the activity state of presynaptic proteins, thus crucially shaping synaptic transmission.

Extensive Remodeling of the Presynaptic Cytomatrix upon Homeostatic Adaptation to Network Activity Silencing

“…MG132 induced, however, a significant increase in mEPSC frequencies in control cultures. This fits well to a recent report by Rinetti and Schweizer (2010), who observed a severalfold increase in frequencies of miniature and spontaneous EPSCs and IPSCs without any changes in amplitudes within minutes after MG132 application. Also, in our experiments, proteasome blockade did not influence the mEPSC amplitudes in spontaneously active control neurons, but led to significant elevation of amplitudes in chronically silenced cultures.…”

The ubiquitin-proteasome pathway and synaptic plasticity

“…In cultured mammalian hippocampal neurons, applications of proteasome inhibitors increase mEPSC frequency without any effect on the amplitude, indicating a presynaptic role for the UPP. Contrary to expectations, stabilization of the presynaptic proteins (RIM1 or Munc13) was not observed (Rinetti and Schweizer 2010).…”

“…The results from these two studies seem to be at odds with each other even though both used postnatal rat hippocampal neurons in culture and antibodies against Rim 1 and Munc 13 from the same commercial sources. Perhaps the discrepancy was due to the fact that the study by Jiang et al (2010) measured Rim 1 and Munc 13 after K+-induced depolarization, whereas the study by Rinetti and Schweizer (2010) tested Rim 1 and Munc 13 levels in relation to changes in mEPSCs and spontaneous EPSCs. Therefore, it is likely that the degradation of Rim 1 and Munc 13 is triggered by neuronal depolarization rather than baseline activity.…”

Increased Neurotransmitter Release at the Neuromuscular Junction in a Mouse Model of Polyglutamine Disease

“…Therefore, the general transient impairment of the UPS could become a rather long-lasting impairment at the synaptic terminals, perhaps because the microtubule-dependent sequestration of expanded htt (Kopito, 2000;Muchowski et al, 2002) is less efficient at the synaptic compartments. In addition, by unknown molecular mechanisms, pharmacological inhibition of the UPS increases neurotransmitter release in hippocampal cultures (Willeumier et al, 2006;Rinetti and Schweizer, 2010). Although we do not have yet enough elements to build up a mechanistic molecular model to explain the enhanced release in R6/1 synapses, it is likely that the upregulation of two SNARE proteins (synaptobrevins and SNAP-25) might be part of the molecular changes contributing to the gain of function phenotype in neurotransmitter release.…”