TY  - JOUR
A1  - Kaltdorf, Kristin Verena
A1  - Schulze, Katja
A1  - Helmprobst, Frederik
A1  - Kollmannsberger, Philip
A1  - Dandekar, Thomas
A1  - Stigloher, Christian
T1  - Fiji macro 3D ART VeSElecT: 3D automated reconstruction tool for vesicle structures of electron tomograms
JF  - PLoS Computational Biology
N2  - Automatic image reconstruction is critical to cope with steadily increasing data from advanced microscopy. We describe here the Fiji macro 3D ART VeSElecT which we developed to study synaptic vesicles in electron tomograms. We apply this tool to quantify vesicle properties (i) in embryonic Danio rerio 4 and 8 days past fertilization (dpf) and (ii) to compare Caenorhabditis elegans N2 neuromuscular junctions (NMJ) wild-type and its septin mutant (unc-59(e261)). We demonstrate development-specific and mutant-specific changes in synaptic vesicle pools in both models. We confirm the functionality of our macro by applying our 3D ART VeSElecT on zebrafish NMJ showing smaller vesicles in 8 dpf embryos then 4 dpf, which was validated by manual reconstruction of the vesicle pool. Furthermore, we analyze the impact of C. elegans septin mutant unc-59(e261) on vesicle pool formation and vesicle size. Automated vesicle registration and characterization was implemented in Fiji as two macros (registration and measurement). This flexible arrangement allows in particular reducing false positives by an optional manual revision step. Preprocessing and contrast enhancement work on image-stacks of 1nm/pixel in x and y direction. Semi-automated cell selection was integrated. 3D ART VeSElecT removes interfering components, detects vesicles by 3D segmentation and calculates vesicle volume and diameter (spherical approximation, inner/outer diameter). Results are collected in color using the RoiManager plugin including the possibility of manual removal of non-matching confounder vesicles. Detailed evaluation considered performance (detected vesicles) and specificity (true vesicles) as well as precision and recall. We furthermore show gain in segmentation and morphological filtering compared to learning based methods and a large time gain compared to manual segmentation. 3D ART VeSElecT shows small error rates and its speed gain can be up to 68 times faster in comparison to manual annotation. Both automatic and semi-automatic modes are explained including a tutorial.
KW  - Biology
KW  - Vesicles
KW  - Caenorhabditis elegans
KW  - Zebrafish
KW  - Septins
KW  - Synaptic vesicles
KW  - Neuromuscular junctions
KW  - Computer software
KW  - Synapses
Y1  - 2017
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-172112
VL  - 13
IS  - 1
ER  - 
TY  - JOUR
A1  - Helmprobst, Frederik
A1  - Lillesaar, Christina
A1  - Stigloher, Christian
T1  - Expression of sept3, sept5a and sept5b in the Developing and Adult Nervous System of the Zebrafish (Danio rerio)
JF  - Frontiers in Neuroanatomy
N2  - Septins are a highly conserved family of small GTPases that form cytoskeletal filaments. Their cellular functions, especially in the nervous system, still remain largely enigmatic, but there are accumulating lines of evidence that septins play important roles in neuronal physiology and pathology. In order to further dissect septin function in the nervous system a detailed temporal resolved analysis in the genetically well tractable model vertebrate zebrafish (Danio rerio) is crucially necessary. To close this knowledge gap we here provide a reference dataset describing the expression of selected septins (sept3, sept5a and sept5b) in the zebrafish central nervous system. Strikingly, proliferation zones are devoid of expression of all three septins investigated, suggesting that they have a role in post-mitotic neural cells. Our finding that three septins are mainly expressed in non-proliferative regions was further confirmed by double-stainings with a proliferative marker. Our RNA in situ hybridization (ISH) study, detecting sept3, sept5a and sept5b mRNAs, shows that all three septins are expressed in largely overlapping regions of the developing brain. However, the expression of sept5a is much more confined compared to sept3 and sept5b. In contrast, the expression of all the three analyzed septins is largely similar in the adult brain.
KW  - retinal development
KW  - sept5b
KW  - septin
KW  - RNA in situ hybridization
KW  - neuronal development
KW  - sept3
KW  - sept5a
Y1  - 2017
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-157625
VL  - 11
IS  - 6
ER  -