title: A New Fiji-Based Algorithm That Systematically Quantifies Nine Synaptic Parameters Provides Insights into Drosophila NMJ Morphometry.
abstract: The morphology of synapses is of central interest in neuroscience because of the intimate relation with synaptic efficacy. Two decades of gene manipulation studies in different animal models have revealed a repertoire of molecules that contribute to synapse development. However, since such studies often assessed only one, or at best a few, morphological features at a given synapse, it remained unaddressed how different structural aspects relate to one another. Furthermore, such focused and sometimes only qualitative approaches likely left many of the more subtle players unnoticed. Here, we present the image analysis algorithm 'Drosophila_NMJ_Morphometrics', available as a Fiji-compatible macro, for quantitative, accurate and objective synapse morphometry of the Drosophila larval neuromuscular junction (NMJ), a well-established glutamatergic model synapse. We developed this methodology for semi-automated multiparametric analyses of NMJ terminals immunolabeled for the commonly used markers Dlg1 and Brp and showed that it also works for Hrp, Csp and Syt. We demonstrate that gender, genetic background and identity of abdominal body segment consistently and significantly contribute to variability in our data, suggesting that controlling for these parameters is important to minimize variability in quantitative analyses. Correlation and principal component analyses (PCA) were performed to investigate which morphometric parameters are inter-dependent and which ones are regulated rather independently. Based on nine acquired parameters, we identified five morphometric groups: NMJ size, geometry, muscle size, number of NMJ islands and number of active zones. Based on our finding that the parameters of the first two principal components hardly correlated with each other, we suggest that different molecular processes underlie these two morphometric groups. Our study sets the stage for systems morphometry approaches at the well-studied Drosophila NMJ.
authors.affiliation: Department of Human Genetics, Donders Institute for Brain, Cognition and Behaviour, Radboud University Medical Center, Nijmegen, the Netherlands.
authors.forename: Bonnie
authors.initials: B
authors.lastname: Nijhof
authors.affiliation: Department of Human Genetics, Donders Institute for Brain, Cognition and Behaviour, Radboud University Medical Center, Nijmegen, the Netherlands.
authors.forename: Anna
authors.initials: A
authors.lastname: Castells-Nobau
authors.affiliation: Microscopical Imaging Centre (MIC), Radboud University Medical Center, Nijmegen, the Netherlands.
authors.forename: Louis
authors.initials: L
authors.lastname: Wolf
authors.affiliation: Department of Human Genetics, Donders Institute for Brain, Cognition and Behaviour, Radboud University Medical Center, Nijmegen, the Netherlands.
authors.forename: Jolanda M
authors.initials: JM
authors.lastname: Scheffer-de Gooyert
authors.affiliation: Department of Biology, Universidad Autónoma de Madrid, Madrid, Spain.|Department of Clinical and Experimental Medicine, Linköping University, Linköping, Sweden.
authors.forename: Ignacio
authors.initials: I
authors.lastname: Monedero
authors.affiliation: Department of Biology, Universidad Autónoma de Madrid, Madrid, Spain.
authors.forename: Laura
authors.initials: L
authors.lastname: Torroja
authors.affiliation: Research Group on Statistics, Econometrics and Health (GRECS) and CIBER of Epidemiology and Public Health (CIBERESP), University of Girona, Girona, Spain.
authors.forename: Lluis
authors.initials: L
authors.lastname: Coromina
authors.affiliation: Department of Pathology, Radboud University Medical Center, Nijmegen, the Netherlands.|Microscopical Imaging Centre (MIC), Radboud University Medical Center, Nijmegen, the Netherlands.
authors.forename: Jeroen A W M
authors.initials: JA
authors.lastname: van der Laak
authors.affiliation: Department of Human Genetics, Donders Institute for Brain, Cognition and Behaviour, Radboud University Medical Center, Nijmegen, the Netherlands.
authors.forename: Annette
authors.initials: A
authors.lastname: Schenck
mesh_terms: D000465:Algorithms; D000818:Animals; D057225:Data Mining; D016208:Databases, Factual; D004330:Drosophila / Q000166:cytology*; D007090:Image Interpretation, Computer-Assisted / Q000379:methods*; D008953:Models, Anatomic; D008959:Models, Neurological; D009469:Neuromuscular Junction / Q000166:cytology*