Rabconnectin3α promotes stable activity of the H+ pump on synaptic vesicles in hair cells

• Published in: The Journal of neuroscience 2012-08, Vol.32 (32), p.11144-11156
• Main Authors: Einhorn, Zev, Trapani, Josef G, Liu, Qianyong, Nicolson, Teresa
• Format: Article
• Language: English
• Subjects: Acoustic Stimulation - adverse effects; Action Potentials - genetics; Adaptor Proteins, Signal Transducing - genetics; Adaptor Proteins, Signal Transducing - metabolism; Analysis of Variance; Animals; Animals, Genetically Modified; Danio rerio; Enzyme Inhibitors - pharmacology; Escape Reaction - drug effects; Escape Reaction - physiology; Freshwater; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; Hair Cells, Auditory - cytology; Larva; Lateral Line System - metabolism; Macrolides - pharmacology; Membrane Transport Proteins - genetics; Membrane Transport Proteins - metabolism; Microscopy, Confocal; Molecular Biology; Mutation - genetics; Physical Stimulation; Proton Pumps - metabolism; RNA, Messenger - metabolism; Sensation Disorders - genetics; Synaptic Vesicles - drug effects; Synaptic Vesicles - metabolism; Vacuolar Proton-Translocating ATPases - metabolism; Vision Disorders - genetics; Zebrafish; Zebrafish Proteins - genetics; Zebrafish Proteins - metabolism
• ISSN: 0270-6474; 1529-2401; 1529-2401
• DOI: 10.1523/JNEUROSCI.1705-12.2012

Acidification of synaptic vesicles relies on the vacuolar-type ATPase (V-ATPase) and provides the electrochemical driving force for neurotransmitter exchange. The regulatory mechanisms that ensure assembly of the V-ATPase holoenzyme on synaptic vesicles are unknown. Rabconnectin3α (Rbc3α) is a potential candidate for regulation of V-ATPase activity because of its association with synaptic vesicles and its requirement for acidification of intracellular compartments. Here, we provide the first evidence for a role of Rbc3α in synaptic vesicle acidification and neurotransmission. In this study, we characterized mutant alleles of rbc3α isolated from a large-scale screen for zebrafish with auditory/vestibular defects. We show that Rbc3α is localized to basal regions of hair cells in which synaptic vesicles are present. To determine whether Rbc3α regulates V-ATPase activity, we examined the acidification of synaptic vesicles and localization of the V-ATPase in hair cells. In contrast to wild-type hair cells, we observed that synaptic vesicles had elevated pH, and a cytosolic subunit of the V-ATPase was no longer enriched in synaptic regions of mutant hair cells. As a consequence of defective acidification of synaptic vesicles, afferent neurons in rbc3α mutants had reduced firing rates and reduced accuracy of phase-locked action potentials in response to mechanical stimulation of hair cells. Collectively, our data suggest that Rbc3α modulates synaptic transmission in hair cells by promoting V-ATPase activity in synaptic vesicles.