Protein sorting represents a potential point of regulation in neurotransmission because it dictates the protein composition of synaptic vesicles, the organelle that mediates transmitter release. Although the average number of most vesicle proteins has been estimated using bulk biochemical approaches (Takamori et al., 2006), no information exists on the intervesicle variability of protein number, and thus on the precision with which proteins are sorted to vesicles. To address this, we adapted a single molecule quantification approach (Mutch et al., 2007) and used it to quantify both the average number and variance of seven integral membrane proteins in brain synaptic vesicles. We report that four vesicle proteins, SV2, the proton ATPase, Vglut1, and synaptotagmin 1, showed little intervesicle variation in number, indicating they are sorted to vesicles with high precision. In contrast, the apparent number of VAMP2/synaptobrevin 2, synaptophysin, and synaptogyrin demonstrated significant intervesicle variability. These findings place constraints on models of protein function at the synapse and raise the possibility that changes in vesicle protein expression affect vesicle composition and functioning.

The size of a synaptic vesicle (SV) is generally thought to be determined by the amount of lipid and membrane protein it contains. Once formed, it is thought to remain constant in size. Using fluorescence correlation spectroscopy and cryogenic electron microscopy, we show that glutamatergic vesicles reversibly increase their size upon filling with glutamate. The increase ( approximately 25% in diameter) corresponds to an increase in surface area of approximately 50% and in volume of approximately 100%. This large size increase implies a large structural change in the SV upon loading with neurotransmitters. Vesicles lacking SV protein 2A (SV2A) did not manifest a change in size after loading with glutamate, indicating that SV2A is required for this phenomenon.

Community transmission of influenza A pandemic (H1N1) 2009 was followed by high rates of hospital admissions in the Wellington region of New Zealand, particularly among Maori and Pacific Islanders. These findings may help health authorities anticipate the effects of pandemic (H1N1) 2009 in other communities.

This article describes two complementary techniques, single-particle tracking and correlation spectroscopy, for accurately sizing nanoparticles confined within picoliter volume aqueous droplets. Single-particle tracking works well with bright particles that can be continuously illuminated and imaged, and we demonstrated this approach for sizing single fluorescent beads. Fluorescence correlation spectroscopy detects small intensity bursts from particles or molecules diffusing through the confocal probe volume, which works well with dim and rapidly diffusing particles or molecules; we demonstrated FCS for sizing synaptic vesicles confined in aqueous droplets. In combination with recent advances in droplet manipulations and analysis, we anticipate this capability to size single nanoparticles and molecules in free solution will complement existing tools for probing cellular systems, subcellular organelles, and nanoparticles.

SummaryWe report a case of subcutaneous emphysema following multiple attempts to cannulate both subclavian veins for pacemaker insertion. Initial bilateral pneumothoraces were complicated by subcutaneous emphysema which became so severe that respiration and cardiac output became seriously impaired. The insertion of a single large bore subcutaneous drain produced a dramatic improvement. The aetiology , complications and management of tension subcutaneous emphysema are discussed. Key wordsComplication; pneumothorax, subcutaneous emphysema. Equipment; central venous catheter, chest drain.Subcutaneous emphysema is relatively common in anaesthetic and surgical practice and is rarely of more than temporary cosmetic significance. We report an unusual case of tension subcutaneous emphysema that resulted in acute thoracic constriction causing respiratory and cardiovascular compromise. This was promptly treated in an innovative manner by the insertion of a large bore subcutaneous drain with excellent results. Case historyA 77-year-old man with a history of dizziness and fainting due to P wave asystole attended a cardiology outpatient clinic as a day case for insertion of a permanent pacemaker. A number of unsuccessful attempts were made to cannulate the left subclavian vein, after which attention was switched to the contralateral side. The right subclavian vein was eventually cannulated, the pacing wire passed and the subcutaneous pacemaker attached. A check chest X ray taken 1 h later showed a small right-sided and a much larger left-sided pneumothorax. The patient was complaining of central chest pain and dyspnoea.Under local anaesthesia a 28FG chest drain was inserted through the fourth left intercostal space and, after attachment to an underwater seal, was shown to be functioning satisfactorily. Oxygen was administered by facemask at a rate of 5 1.min-I and the patient was admitted to a medical ward for observation. Next morning his clinical condition had improved and a repeat chest X ray showed resolution of the right-sided pneumothorax and a reduction in the size of the left-sided pneumothorax. Mild subcutaneous emphysema had, however, developed and was apparent both clinically and radiologically (Fig. 1).The patient remained stable for a further 24 h until the chest drain was clamped. Over the next 2-3 h his condition rapidly deteriorated as he became increasingly tachypnoeic and distressed. As well as dyspnoea, he had also developed marked dysphonia and dysphagia. An emergency chest X ray showed that the left-sided pneumothorax had recurred and occupied about 50% of the left hemithorax. The drain was unclamped and a large volume of air bubbled out rapidly. During this period, however, the subcutaneous emphysema was seen to be increasing dramatically, and extended from knees to eyebrows, with the peri-orbital emphysema actually causing the eyelids to close over. The patient's appearance resembled the classic 'Michelin Man'. There was a decrease in respiratory rate to 6 breath.min-'. Hypotension and tachycardia with...