The protein brain-derived neurotrophic factor (BDNF) has been postulated to be a retrograde or paracrine synaptic messenger in long-term potentiation and other forms of activity-dependent synaptic plasticity. Although crucial for this concept, direct evidence for the activity-dependent synaptic release of BDNF is lacking. Here we investigate secretion of BDNF labelled with green¯uorescent protein (BDNF±GFP) by monitoring the changes in¯uores-cence intensity of dendritic BDNF±GFP vesicles at glutamatergic synaptic junctions of living hippocampal neurons. We show that high-frequency activation of glutamatergic synapses triggers the release of BDNF±GFP from synaptically localized secretory granules. This release depends on activation of postsynaptic ionotropic glutamate receptors and on postsynaptic Ca 2+ in¯ux. Release of BDNF±GFP is also observed from extrasynaptic dendritic vesicle clusters, suggesting that a possible spatial restriction of BDNF release to speci®c synaptic sites can only occur if the postsynaptic depolarization remains local. These results support the concept of BDNF being a synaptic messenger of activity-dependent synaptic plasticity, which is released from postsynaptic neurons.

Neurotrophins are a family of secreted neuronal survival and plasticity factors comprising NGF, BDNF, neurotrophin-3 (NT-3), and NT-4. Whereas synaptic secretion of BDNF has been described, the routes of intracellular targeting and secretion of NGF, NT-3, and NT-4 in neurons are poorly understood.To allow for a direct comparison of intracellular targeting and release properties, all four mammalian neurotrophins were expressed as green fluorescent protein fusion proteins in cultured rat hippocampal neurons. We show that BDNF and NT-3 are targeted more efficiently to dendritic secretory granules of the regulated pathway of secretion (BDNF, in 98% of cells; NT-3, 85%) than NGF (46%) and NT-4 (23%). For all NTs, the remaining cells showed targeting to the constitutive secretory pathway. Fusing the BDNF pre-pro sequence to NT-4 directed NT-4 more efficiently to the regulated pathway of secretion.

BACKGROUND: Allogeneic blood products transfusion during liver transplantation (LT) can be associated with increased morbidity and mortality. Data on thromboelastometry (ROTEM)-guided coagulation management with coagulation factor concentrates (CFCs)-fibrinogen concentrate and/or prothrombin complex concentrate (PCC)-are sparse. We aimed to retrospectively evaluate the safety events observed with this approach in our clinic. STUDY DESIGN AND METHODS: LT patients from January 2009 to December 2010 (n = 266) were identified by chart review. A ROTEM-based algorithm with CFC guided the hemostatic therapy. Doppler ultrasound was used to evaluate thrombosis in the hepatic artery, portal vein, and hepatic veins. Stroke, myocardial isch-emia, pulmonary embolism, and transfusion variables were recorded. Patients receiving CFC were included in the CFC group (n = 156); those not receiving CFC were included in the non-CFC group (n = 110). Safety events were compared between these two groups. RESULTS: Allogeneic transfusion(s) in the 266 patients was low, with medians of 2 (interquartile range [IQR], 0-5), 0 (IQR 0-0), and 0 (IQR 0-1) units for red blood cells (RBCs), fresh-frozen plasma (FFP), and platelets (PLTs), respectively. Ninety-seven of 266 LTs (36.5%) were performed without RBCs transfusion, 227 (85.3%) without FFP, and 190 (71.4%) without PLTs. There were no significant differences in thrombotic, thrombo-embolic, and ischemic adverse events occurrence between the CFC group and the non-CFC group (11/ 156 patients vs. 5/110; p = 0.31). CONCLUSION: In LT, ROTEM-guided treatment with fibrinogen concentrate and/or PCC did not appear to increase the occurrence of thrombosis and ischemic events compared to patients who did not receive these concentrates. T he median model of end-stage liver disease (MELD) score in EUROTRANSPLANT has increased from 25 to 35 (match-MELD) in the past 6 years, attributable to the adoption of MELD score as the basis for organ allocation in liver trans-plantation (LT). 1 This increase is associated with an increased risk of bleeding. 2 However, chronic liver disease is associated with multiple changes in coagulation status. On the one hand, the activity and levels of vitamin K-dependent coagulation factors (II, VII, IX, and X) and coagulation inhibitors (protein C and S) are decreased, as well as platelet (PLT) count. 3 Levels of tissue factor-expressing cells, von Willebrand factor, and coagulation Factor (F)VIII are often increased. 4,5 The concomitant reduction of pro-and anticoagulants typically leads to rebalanced hemostasis, but the low levels of pro-and anti-coagulants mean that the balance can be easily disturbed, ABBREVIATIONS: ALI = acute lung injury; aPTT = activated partial thromboplastin time; CFC(s) = coagulation factor concentrate(s); HAT = hepatic artery thrombosis; ICU = intensive care unit; INR = international normalized ratio; IQR = interquartile range; LT(s) = liver transplantation(s); MELD = median model of end-stage liver disease; PCC(s) = prothrombin complex concentrate(...

IntroductionHemolysis can be induced in sepsis via various mechanisms, its pathophysiological importance has been demonstrated in experimental sepsis. However, no data on free hemoglobin concentrations in human sepsis are available. In the present study we measured free hemoglobin in patients with severe sepsis as well as in postoperative patients using four methods. It was our aim to determine the potential value of free hemoglobin as a biomarker for diagnosis and outcome of severe sepsis in critical illness.MethodsPlasma concentration of free hemoglobin was determined in patients with severe sepsis (n = 161) and postoperative patients (n = 136) on day 1 of diagnosis and surgery. For the measurement of free hemoglobin, an enzyme linked immunosorbent assay and three spectrophotometric algorithms were used. Moreover, SAPS II- and SOFA scores as well as procalcitonin concentration and outcome were determined. Kaplan-Meier analysis was performed and odds ratios were determined after classification of free hemoglobin concentrations in a high and low concentration group according to the median. For statistical evaluation the Mann-Whitney test and logistic regression analysis were used.ResultsIn non-survivors of severe sepsis, free hemoglobin concentration was twice the concentration compared to survivors. Thirty-day survival of patients, as evidenced by Kaplan-Meier analysis, was markedly lower in patients with high free hemoglobin concentration than in patients with low free hemoglobin concentration. Best discrimination of outcome was achieved with the spectrophotometric method of Harboe (51.3% vs. 86.4% survival, p < 0.001; odds ratio 6.1). Multivariate analysis including free hemoglobin, age, SAPS II- and SOFA-score and procalcitonin demonstrated that free hemoglobin, as determined by all 4 methods, was the best and an independent predictor for death in severe sepsis (p = 0.022 to p < 0.001). Free hemoglobin concentrations were not significantly different in postoperative and septic patients in three of four assays. Thus, free hemoglobin can not be used to diagnose severe sepsis in critical illness.ConclusionsFree hemoglobin is an important new predictor of survival in severe sepsis.