14898
2015
eng
29
9
article
1
2017-05-22
--
--
Bruchpilot and Synaptotagmin collaborate to drive rapid glutamate release and active zone differentiation
The active zone (AZ) protein Bruchpilot (Brp) is essential for rapid glutamate release at Drosophila melanogaster neuromuscular junctions (NMJs). Quantal time course and measurements of action potential-waveform suggest that presynaptic fusion mechanisms are altered in brp null mutants (brp\(^{69}\)). This could account for their increased evoked excitatory postsynaptic current (EPSC) delay and rise time (by about 1 ms). To test the mechanism of release protraction at brp\(^{69}\) AZs, we performed knock-down of Synaptotagmin-1 (Syt) via RNAi (syt\(^{KD}\)) in wildtype (wt), brp\(^{69}\) and rab3 null mutants (rab3\(^{rup}\)), where Brp is concentrated at a small number of AZs. At wt and rab3\(^{rup}\) synapses, syt\(^{KD}\) lowered EPSC amplitude while increasing rise time and delay, consistent with the role of Syt as a release sensor. In contrast, syt\(^{KD}\) did not alter EPSC amplitude at brp\(^{69}\) synapses, but shortened delay and rise time. In fact, following syt\(^{KD}\), these kinetic properties were strikingly similar in wt and brp\(^{69}\), which supports the notion that Syt protracts release at brp\(^{69}\) synapses. To gain insight into this surprising role of Syt at brp\(^{69}\) AZs, we analyzed the structural and functional differentiation of synaptic boutons at the NMJ. At tonic type Ib motor neurons, distal boutons contain more AZs, more Brp proteins per AZ and show elevated and accelerated glutamate release compared to proximal boutons. The functional differentiation between proximal and distal boutons is Brp-dependent and reduced after syt\(^{KD}\). Notably, syt\(^{KD}\) boutons are smaller, contain fewer Brp positive AZs and these are of similar number in proximal and distal boutons. In addition, super-resolution imaging via dSTORM revealed that syt\(^{KD}\) increases the number and alters the spatial distribution of Brp molecules at AZs, while the gradient of Brp proteins per AZ is diminished. In summary, these data demonstrate that normal structural and functional differentiation of Drosophila AZs requires concerted action of Brp and Syt.
Frontiers in Cellular Neuroscience
10.3389/fncel.2015.00029
urn:nbn:de:bvb:20-opus-148988
Frontiers in Cellular Neuroscience 9:29 (2015). DOI: 10.3389/fncel.2015.00029
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Mila M. Paul
Martin Pauli
Nadine Ehmann
Stefan Hallermann
Markus Sauer
Robert J. Kittel
Manfred Heckmann
eng
uncontrolled
neuromuscular junction
eng
uncontrolled
Bruchpilot
eng
uncontrolled
synaptic delay
eng
uncontrolled
dSTORM
eng
uncontrolled
synaptotagmin
eng
uncontrolled
presynaptic differentiation
eng
uncontrolled
neurotransmitter release
eng
uncontrolled
active zone
eng
uncontrolled
synaptic transmission
eng
uncontrolled
fluorescent probes
Medizin und Gesundheit
open_access
Physiologisches Institut
Theodor-Boveri-Institut für Biowissenschaften
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/14898/052_Paul_Frontiers_in_Cellular_Neuroscience.pdf
29944
2022
eng
16
article
1
--
2022-12-14
--
Endogenous tagging of Unc-13 reveals nanoscale reorganization at active zones during presynaptic homeostatic potentiation
Introduction
Neurotransmitter release at presynaptic active zones (AZs) requires concerted protein interactions within a dense 3D nano-hemisphere. Among the complex protein meshwork the (M)unc-13 family member Unc-13 of Drosophila melanogaster is essential for docking of synaptic vesicles and transmitter release.
Methods
We employ minos-mediated integration cassette (MiMIC)-based gene editing using GFSTF (EGFP-FlAsH-StrepII-TEV-3xFlag) to endogenously tag all annotated Drosophila Unc-13 isoforms enabling visualization of endogenous Unc-13 expression within the central and peripheral nervous system.
Results and discussion
Electrophysiological characterization using two-electrode voltage clamp (TEVC) reveals that evoked and spontaneous synaptic transmission remain unaffected in unc-13\(^{GFSTF}\) 3rd instar larvae and acute presynaptic homeostatic potentiation (PHP) can be induced at control levels. Furthermore, multi-color structured-illumination shows precise co-localization of Unc-13\(^{GFSTF}\), Bruchpilot, and GluRIIA-receptor subunits within the synaptic mesoscale. Localization microscopy in combination with HDBSCAN algorithms detect Unc-13\(^{GFSTF}\) subclusters that move toward the AZ center during PHP with unaltered Unc-13\(^{GFSTF}\) protein levels.
Frontiers in Cellular Neuroscience
1662-5102
10.3389/fncel.2022.1074304
urn:nbn:de:bvb:20-opus-299440
2023-01-18T06:19:17+00:00
sword
swordwue
attachment; filename=deposit.zip
b2865a404dcc65a1e7608966b766fbf7
Frontiers in Cellular Neuroscience (2022) 16:1074304. doi:10.3389/fncel.2022.1074304
false
true
Sven Dannhäuser
Achmed Mrestani
Florian Gundelach
Martin Pauli
Fabian Komma
Philip Kollmannsberger
Markus Sauer
Manfred Heckmann
Mila M. Paul
eng
uncontrolled
active zone
eng
uncontrolled
Unc-13
eng
uncontrolled
MiMIC
eng
uncontrolled
presynaptic homeostasis
eng
uncontrolled
nanoarchitecture
eng
uncontrolled
localization microscopy
eng
uncontrolled
STORM
eng
uncontrolled
HDBSCAN
Medizin und Gesundheit
open_access
Physiologisches Institut
Klinik und Poliklinik für Psychiatrie, Psychosomatik und Psychotherapie
Klinik und Poliklinik für Unfall-, Hand-, Plastische und Wiederherstellungschirurgie (Chirurgische Klinik II)
Theodor-Boveri-Institut für Biowissenschaften
Import
Center for Computational and Theoretical Biology
Förderzeitraum 2022
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/29944/fncel-16-1074304.pdf
26549
2021
eng
109770
1
37
article
1
2022-04-04
--
--
Active zone compaction correlates with presynaptic homeostatic potentiation
Neurotransmitter release is stabilized by homeostatic plasticity. Presynaptic homeostatic potentiation (PHP) operates on timescales ranging from minute- to life-long adaptations and likely involves reorganization of presynaptic active zones (AZs). At Drosophila melanogaster neuromuscular junctions, earlier work ascribed AZ enlargement by incorporating more Bruchpilot (Brp) scaffold protein a role in PHP. We use localization microscopy (direct stochastic optical reconstruction microscopy [dSTORM]) and hierarchical density-based spatial clustering of applications with noise (HDBSCAN) to study AZ plasticity during PHP at the synaptic mesoscale. We find compaction of individual AZs in acute philanthotoxin-induced and chronic genetically induced PHP but unchanged copy numbers of AZ proteins. Compaction even occurs at the level of Brp subclusters, which move toward AZ centers, and in Rab3 interacting molecule (RIM)-binding protein (RBP) subclusters. Furthermore, correlative confocal and dSTORM imaging reveals how AZ compaction in PHP translates into apparent increases in AZ area and Brp protein content, as implied earlier.
Cell Reports
10.1016/j.celrep.2021.109770
urn:nbn:de:bvb:20-opus-265497
publish
Cell Reports (2021) 37:1, 109770. https://doi.org/10.1016/j.celrep.2021.109770
false
true
CC BY-NC-ND: Creative-Commons-Lizenz: Namensnennung, Nicht kommerziell, Keine Bearbeitungen 4.0 International
Achmed Mrestani
Martin Pauli
Philip Kollmannsberger
Felix Repp
Robert J. Kittel
Jens Eilers
Sören Doose
Markus Sauer
Anna-Leena Sirén
Manfred Heckmann
Mila M. Paul
eng
uncontrolled
active zone
eng
uncontrolled
Bruchpilot
eng
uncontrolled
RIM-binding protein
eng
uncontrolled
compaction
eng
uncontrolled
homeostasis
eng
uncontrolled
presynaptic plasticity
eng
uncontrolled
super-resolution microscopy
Medizin und Gesundheit
open_access
Neurochirurgische Klinik und Poliklinik
Physiologisches Institut
Klinik und Poliklinik für Unfall-, Hand-, Plastische und Wiederherstellungschirurgie (Chirurgische Klinik II)
Theodor-Boveri-Institut für Biowissenschaften
Center for Computational and Theoretical Biology
Förderzeitraum 2021
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/26549/1-s2.0-S2211124721012249-main.pdf
29978
2022
eng
1
12
article
1
--
--
--
Plate osteosynthesis combined with bone cement provides the highest stability for tibial head depression fractures under high loading conditions
Older patients sustaining tibial head depression fractures often cannot follow the post-operative rehabilitation protocols with partial weight-bearing of the affected limb, leading to osteosynthesis failure, cartilage step-off and arthritis development. Therefore, the aim of this study was to analyse the biomechanical performance of different types of osteosyntheses alone and in combination with bone cement simulating cyclically high loading conditions of tibial head depression fractures. Lateral tibial head depression fractures (AO: 41-B2.2; Schatzker type III) were created in synthetic bones and stabilized using three different osteosyntheses alone and in combination with a commonly used bone cement (chronOS™): 2 screws, 4 screws in the jail technique and a lateral angle-stable buttress plate. After fixation, the lateral tibial plateau was axially loaded in two, from each other independent testing series: In the first test protocol, 5000 cycles with 500 N and in the end load-to-failure tests were performed. In the second test protocol, the cyclic loading was increased to 1000 N. Parameters of interest were the displacement of the articular fracture fragment, the stiffness and the maximum load. The osteosyntheses revealed a higher stiffness in combination with bone cement compared to the same type of osteosynthesis alone (e.g., 500 N level: 2 screws 383 ± 43 N/mm vs. 2 screws + chronOs 520 ± 108 N/mm, increase by 36%, p < 0.01; 4 screws 368 ± 97 N/mm vs. 4 screws + chronOS 516 ± 109 N/mm, increase by 40%, p < 0.01; plate: 509 ± 73 N/mm vs. plate + chronOs 792 ± 150 N/mm, increase by 56%, p < 0.01). Bone cement reduced the displacement of the plate significantly (500 N level: plate: 8.9 ± 2.8 mm vs. plate + chronOs: 3.1 ± 1.4 mm, reduction by 65%, p < 0.01; 1000 N level: 16.9 ± 3.6 mm vs 5.6 ± 1.3 mm, reduction by 67%, p < 0.01). Thus, the highest stiffness and lowest displacement values were found when using the plate with bone cement in both loading conditions (500 N level: 2 screws + chronOs 3.7 ± 1.3 mm, 4 screws + chronOs 6.2 ± 2.4 mm; 1000 N level: 2 screws + chronOs 6.5 ± 1.2 mm, 4 screws + chronOs 5.7 ± 0.8 mm). From a biomechanical perspective, plate osteosynthesis of tibial head depression fractures should always be combined with bone cement, provides higher stability than 2-screw and 4-screw fixation and is a valid treatment option in cases where extraordinary stability is required.
Scientific Reports
10.1038/s41598-022-19107-6
urn:nbn:de:bvb:20-opus-299782
@articleHeilig.2022, author = Heilig, Philipp and Faerber, Lars-Christopher and Paul, Mila M. and Kupczyk, Eva and Meffert, Rainer H. and Jordan, Martin C. and Hoelscher-Doht, Stefanie, year = 2022, title = Plate osteosynthesis combined with bone cement provides the highest stability for tibial head depression fractures under high loading conditions, pages = 15481, volume = 12, number = 1, journal = Scientific reports, doi = 10.1038/s41598-022-19107-6,
md5:141c1dc808a5ea532571205a54440c66
2023-01-18T07:00:17+00:00
/tmp/phpWSWY4m
bibtex
63c79901842c02.74872320
Scientific Reports 2022, 12(1):15481. DOI: 10.1038/s41598-022-19107-6
false
true
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Philipp Heilig
Lars-Christopher Faerber
Mila M. Paul
Eva Kupczyk
Rainer H. Meffert
Martin C. Jordan
Stefanie Hoelscher-Doht
eng
uncontrolled
head depression fractures
eng
uncontrolled
osteosynthesis
eng
uncontrolled
arthritis
Medizin und Gesundheit
open_access
Klinik und Poliklinik für Unfall-, Hand-, Plastische und Wiederherstellungschirurgie (Chirurgische Klinik II)
Förderzeitraum 2022
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/29978/Scientific_Reports_Heilig.pdf
28279
2022
eng
10
article
1
--
2022-08-04
--
Surgical fixation of calcaneal beak fractures — biomechanical analysis of different osteosynthesis techniques
The calcaneal beak fracture is a rare avulsion fracture of the tuber calcanei characterized by a solid bony fragment at the Achilles tendon insertion. Treatment usually requires osteosynthesis. However, lack of biomechanical understanding of the ideal fixation technique persists. A beak fracture was simulated in synthetic bones and assigned to five different groups of fixation: A) 6.5-mm partial threaded cannulated screws, B) 4.0-mm partial threaded cannulated screws, C) 5.0-mm headless cannulated compression screws, D) 2.3-mm locking plate, and E) 2.8-mm locking plate. Different traction force levels were applied through an Achilles tendon surrogate in a material-testing machine on all stabilized synthetic bones. Outcome measures were peak-to-peak displacement, total displacement, plastic deformation, stiffness, visual-fracture-line displacement, and mode of implant failure. The 2.3- and 2.8-mm plating groups showed a high drop-out rate at 100 N tension force and failed under higher tension levels of 200 N. The fracture fixation using 4.0-mm partial threaded screws showed a significantly higher repair strength and was able to withhold cyclic loading up to 300 N. The lowest peak-to-peak displacement and the highest load-to-failure and stiffness were provided by fracture fixation using 6.5-mm partial threaded cannulated screws or 5.0-mm headless cannulated compression screws. As anticipated, large 6.5-mm screw diameters provide the best biomechanical fixation. Surprisingly, the 5.0-mm headless cannulated compression screws yield reliable stability despite the absent screw head and washer. When such large screws cannot be applied, 4.0-mm screws also allow reasonable fixation strength. Plate fixation should be implemented with precaution and in combination with a restrictive postoperative motion protocol. Finally, clinical cases about the surgical application and recovery are included.
Frontiers in Bioengineering and Biotechnology
2296-4185
10.3389/fbioe.2022.896790
urn:nbn:de:bvb:20-opus-282792
2022-08-18T10:08:16+00:00
sword
swordwue
attachment; filename=deposit.zip
957ed3573224139f6ae7ccb0d2cf6ee8
Frontiers in Bioengineering and Biotechnology (2022) 10:896790. DOI:10.3389/fbioe.2022.896790
false
true
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Martin C. Jordan
Lukas Hufnagel
Miriam McDonogh
Mila M. Paul
Jonas Schmalzl
Eva Kupczyk
Hendrik Jansen
Philipp Heilig
Rainer H. Meffert
Stefanie Hoelscher-Doht
eng
uncontrolled
foot
eng
uncontrolled
ankle
eng
uncontrolled
Achilles
eng
uncontrolled
tendon
eng
uncontrolled
fracture
Chirurgie und verwandte medizinische Fachrichtungen
open_access
Klinik und Poliklinik für Unfall-, Hand-, Plastische und Wiederherstellungschirurgie (Chirurgische Klinik II)
Import
Förderzeitraum 2022
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/28279/fbioe-10-896790.pdf
25983
2021
eng
407
4
article
1
2022-03-10
--
--
Targeted volumetric single-molecule localization microscopy of defined presynaptic structures in brain sections
Revealing the molecular organization of anatomically precisely defined brain regions is necessary for refined understanding of synaptic plasticity. Although three-dimensional (3D) single-molecule localization microscopy can provide the required resolution, imaging more than a few micrometers deep into tissue remains challenging. To quantify presynaptic active zones (AZ) of entire, large, conditional detonator hippocampal mossy fiber (MF) boutons with diameters as large as 10 mu m, we developed a method for targeted volumetric direct stochastic optical reconstruction microscopy (dSTORM). An optimized protocol for fast repeated axial scanning and efficient sequential labeling of the AZ scaffold Bassoon and membrane bound GFP with Alexa Fluor 647 enabled 3D-dSTORM imaging of 25 mu m thick mouse brain sections and assignment of AZs to specific neuronal substructures. Quantitative data analysis revealed large differences in Bassoon cluster size and density for distinct hippocampal regions with largest clusters in MF boutons. Pauli et al. develop targeted volumetric dSTORM in order to image large hippocampal mossy fiber boutons (MFBs) in brain slices. They can identify synaptic targets of individual MFBs and measured size and density of Bassoon clusters within individual untruncated MFBs at nanoscopic resolution.
Communications Biology
10.1038/s42003-021-01939-z
urn:nbn:de:bvb:20-opus-259830
publish
Communications Biology (2021) 4:407. DOI:10.1038/s42003-021-01939-z
false
true
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Martin Pauli
Mila M. Paul
Sven Proppert
Achmed Mrestani
Marzieh Sharifi
Felix Repp
Lydia Kürzinger
Philip Kollmannsberger
Markus Sauer
Manfred Heckmann
Anna-Leena Sirén
eng
uncontrolled
mossy fiber synapses
eng
uncontrolled
CA3 pyrimidal cells
eng
uncontrolled
CA2+ channels
eng
uncontrolled
active zone
eng
uncontrolled
hippocampal
eng
uncontrolled
release
eng
uncontrolled
plasticity
eng
uncontrolled
proteins
eng
uncontrolled
platform
eng
uncontrolled
reveals
Medizin und Gesundheit
open_access
Neurochirurgische Klinik und Poliklinik
Physiologisches Institut
Theodor-Boveri-Institut für Biowissenschaften
Center for Computational and Theoretical Biology
Förderzeitraum 2021
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/25983/s42003-021-01939-z.pdf
30490
2023
eng
3
24
article
1
--
2023-01-21
--
Single-molecule localization microscopy of presynaptic active zones in Drosophila melanogaster after rapid cryofixation
Single-molecule localization microscopy (SMLM) greatly advances structural studies of diverse biological tissues. For example, presynaptic active zone (AZ) nanotopology is resolved in increasing detail. Immunofluorescence imaging of AZ proteins usually relies on epitope preservation using aldehyde-based immunocompetent fixation. Cryofixation techniques, such as high-pressure freezing (HPF) and freeze substitution (FS), are widely used for ultrastructural studies of presynaptic architecture in electron microscopy (EM). HPF/FS demonstrated nearer-to-native preservation of AZ ultrastructure, e.g., by facilitating single filamentous structures. Here, we present a protocol combining the advantages of HPF/FS and direct stochastic optical reconstruction microscopy (dSTORM) to quantify nanotopology of the AZ scaffold protein Bruchpilot (Brp) at neuromuscular junctions (NMJs) of Drosophila melanogaster. Using this standardized model, we tested for preservation of Brp clusters in different FS protocols compared to classical aldehyde fixation. In HPF/FS samples, presynaptic boutons were structurally well preserved with ~22% smaller Brp clusters that allowed quantification of subcluster topology. In summary, we established a standardized near-to-native preparation and immunohistochemistry protocol for SMLM analyses of AZ protein clusters in a defined model synapse. Our protocol could be adapted to study protein arrangements at single-molecule resolution in other intact tissue preparations.
International Journal of Molecular Sciences
1422-0067
10.3390/ijms24032128
urn:nbn:de:bvb:20-opus-304904
2023-03-14T06:02:46+00:00
sword
swordwue
attachment; filename=deposit.zip
9a3b69c8e3fd636316b0ba75befe71f1
International Journal of Molecular Sciences (2023) 24:3, 2128. https://doi.org/10.3390/ijms24032128
false
true
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Achmed Mrestani
Katharina Lichter
Anna-Leena Sirén
Manfred Heckmann
Mila M. Paul
Martin Pauli
eng
uncontrolled
active zone
eng
uncontrolled
nanotopology
eng
uncontrolled
neuromuscular junction
eng
uncontrolled
high-pressure freezing/freeze substitution
eng
uncontrolled
PFA in ethanol
eng
uncontrolled
dSTORM
eng
uncontrolled
Drosophila melanogaster
Medizin und Gesundheit
open_access
Neurochirurgische Klinik und Poliklinik
Physiologisches Institut
Klinik und Poliklinik für Unfall-, Hand-, Plastische und Wiederherstellungschirurgie (Chirurgische Klinik II)
Import
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/30490/ijms-24-02128.pdf
31936
2023
eng
11
12
article
1
--
2023-05-31
--
Impact of a femoral fracture on outcome after traumatic brain injury — a matched-pair analysis of the TraumaRegister DGU\(^®\)
Traumatic brain injury (TBI) is the leading cause of death and disability in polytrauma and is often accompanied by concomitant injuries. We conducted a retrospective matched-pair analysis of data from a 10-year period from the multicenter database TraumaRegister DGU\(^®\) to analyze the impact of a concomitant femoral fracture on the outcome of TBI patients. A total of 4508 patients with moderate to critical TBI were included and matched by severity of TBI, American Society of Anesthesiologists (ASA) risk classification, initial Glasgow Coma Scale (GCS), age, and sex. Patients who suffered combined TBI and femoral fracture showed increased mortality and worse outcome at the time of discharge, a higher chance of multi-organ failure, and a rate of neurosurgical intervention. Especially those with moderate TBI showed enhanced in-hospital mortality when presenting with a concomitant femoral fracture (p = 0.037). The choice of fracture treatment (damage control orthopedics vs. early total care) did not impact mortality. In summary, patients with combined TBI and femoral fracture have higher mortality, more in-hospital complications, an increased need for neurosurgical intervention, and inferior outcome compared to patients with TBI solely. More investigations are needed to decipher the pathophysiological consequences of a long-bone fracture on the outcome after TBI.
Journal of Clinical Medicine
2077-0383
10.3390/jcm12113802
urn:nbn:de:bvb:20-opus-319363
2023-06-07T11:41:39+00:00
sword
swordwue
attachment; filename=deposit.zip
5b83db31cde282f9e7d4e661c6493487
Journal of Clinical Medicine (2023) 12:11, 3802. https://doi.org/10.3390/jcm12113802
false
true
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Mila M. Paul
Hannah J. Mieden
Rolf Lefering
Eva K. Kupczyk
Martin C. Jordan
Fabian Gilbert
Rainer H. Meffert
Anna-Leena Sirén
Stefanie Hoelscher-Doht
eng
uncontrolled
traumatic brain injury
eng
uncontrolled
femoral fracture
eng
uncontrolled
damage control orthopedics
eng
uncontrolled
mortality
Medizin und Gesundheit
open_access
Neurochirurgische Klinik und Poliklinik
Physiologisches Institut
Klinik und Poliklinik für Unfall-, Hand-, Plastische und Wiederherstellungschirurgie (Chirurgische Klinik II)
Import
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/31936/jcm-12-03802-v2.pdf