The SNARE protein vti1a functions in dense-core vesicle biogenesis.

The SNARE protein vti1a is proposed to drive fusion of intracellular organelles, but recent data also implicated vti1a in exocytosis. Here we show that vti1a is absent from mature secretory vesicles in adrenal chromaffin cells, but localizes to a compartment near the trans-Golgi network, partially overlapping with syntaxin-6. Exocytosis is impaired in vti1a null cells, partly due to fewer Ca(2+)-channels at the plasma membrane, partly due to fewer vesicles of reduced size and synaptobrevin-2 content. In contrast, release kinetics and Ca(2+)-sensitivity remain unchanged, indicating that the final fusion reaction leading to transmitter release is unperturbed. Additional deletion of the closest related SNARE, vti1b, does not exacerbate the vti1a phenotype, and vti1b null cells show no secretion defects, indicating that vti1b does not participate in exocytosis. Long-term re-expression of vti1a (days) was necessary for restoration of secretory capacity, whereas strong short-term expression (hours) was ineffective, consistent with vti1a involvement in an upstream step related to vesicle generation, rather than in fusion. We conclude that vti1a functions in vesicle generation and Ca(2+)-channel trafficking, but is dispensable for transmitter release.

Medical Team Evaluation: Effect on Emergency Department Waiting Time and Length of Stay.

Emergency Departments (ED) are trying to alleviate crowding using various interventions. We assessed the effect of an alternative model of care, the Medical Team Evaluation (MTE) concept, encompassing team triage, quick registration, redesign of triage rooms and electronic medical records (EMR) on door-to-doctor (waiting) time and ED length of stay (LOS). We conducted an observational, before-and-after study at an urban academic tertiary care centre. On July 17th 2014, MTE was initiated from 9:00 a.m. to 10 p.m., 7 days a week. A registered triage nurse was teamed with an additional senior ED physician. Data of the 5-month pre-MTE and the 5-month MTE period were analysed. A matched comparison of waiting times and ED LOS of discharged and admitted patients pertaining to various Emergency Severity Index (ESI) triage categories was performed based on propensity scores. With MTE, the median waiting times improved from 41.2 (24.8-66.6) to 10.2 (5.7-18.1) minutes (min; P < 0.01). Though being beneficial for all strata, the improvement was somewhat greater for discharged, than for admitted patients. With a reduction from 54.3 (34.2-84.7) to 10.5 (5.9-18.4) min (P < 0.01), in terms of waiting times, MTE was most advantageous for ESI4 patients. The overall median ED LOS increased for about 15 min (P < 0.01), increasing from 3.4 (2.1-5.3) to 3.7 (2.3-5.6) hours. A significant increase was observed for all the strata, except for ESI5 patients. Their median ED LOS dropped by 73% from 1.2 (0.8-1.8) to 0.3 (0.2-0.5) hours (P < 0.01). In the same period the total orders for diagnostic radiology increased by 1,178 (11%) from 10,924 to 12,102 orders, with more imaging tests being ordered for ESI 2, 3 and 4 patients. Despite improved waiting times a decrease of ED LOS was only seen in ESI level 5 patients, whereas in all the other strata ED LOS increased. We speculate that this was brought about by the tendency of triage physicians to order more diagnostic radiology, anticipating that it may be better for the downstream physician to have more information rather than less.

Neurobeachin regulates neurotransmitter receptor trafficking to synapses.

The surface density of neurotransmitter receptors at synapses is a key determinant of synaptic efficacy. Synaptic receptor accumulation is regulated by the transport, postsynaptic anchoring, and turnover of receptors, involving multiple trafficking, sorting, motor, and scaffold proteins. We found that neurons lacking the BEACH (beige-Chediak/Higashi) domain protein Neurobeachin (Nbea) had strongly reduced synaptic responses caused by a reduction in surface levels of glutamate and GABA(A) receptors. In the absence of Nbea, immature AMPA receptors accumulated early in the biosynthetic pathway, and mature N-methyl-d-aspartate, kainate, and GABA(A) receptors did not reach the synapse, whereas maturation and surface expression of other membrane proteins, synapse formation, and presynaptic function were unaffected. These data show that Nbea regulates synaptic transmission under basal conditions by targeting neurotransmitter receptors to synapses.

Synapse associated protein 102 (SAP102) binds the C-terminal part of the scaffolding protein neurobeachin.

Neurobeachin (Nbea) is a multidomain scaffold protein abundant in the brain, where it is highly expressed during development. Nbea-null mice have severe defects in neuromuscular synaptic transmission resulting in lethal paralysis of the newborns. Recently, it became clear that Nbea is important also for the functioning of central synapses, where it is suggested to play a role in trafficking membrane proteins to both, the pre- and post-synaptic sites. So far, only few binding partners of Nbea have been found and the precise mechanism of their trafficking remains unclear. Here, we used mass spectrometry to identify SAP102, a MAGUK protein implicated in trafficking of the ionotropic glutamate AMPA- and NMDA-type receptors during synaptogenesis, as a novel Nbea interacting protein in mouse brain. Experiments in heterologous cells confirmed this interaction and revealed that SAP102 binds to the C-terminal part of Nbea that contains the DUF, PH, BEACH and WD40 domains. Furthermore, we discovered that introducing a mutation in Nbea's PH domain, which disrupts its interaction with the BEACH domain, abolishes this binding, thereby creating an excellent starting point to further investigate Nbea-SAP102 function in the central nervous system.