{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,21]],"date-time":"2026-03-21T05:03:56Z","timestamp":1774069436408,"version":"3.50.1"},"update-to":[{"DOI":"10.1371\/journal.pcbi.1009051","type":"new_version","label":"New version","source":"publisher","updated":{"date-parts":[[2022,2,8]],"date-time":"2022-02-08T00:00:00Z","timestamp":1644278400000}}],"reference-count":42,"publisher":"Public Library of Science (PLoS)","issue":"12","license":[{"start":{"date-parts":[[2021,12,8]],"date-time":"2021-12-08T00:00:00Z","timestamp":1638921600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"European Union\u2019s Horizon 2020 Research and Innovation Program","award":["945539"],"award-info":[{"award-number":["945539"]}]},{"DOI":"10.13039\/501100004837","name":"Ministerio de Ciencia e Innovaci\u00f3n de Espa\u00f1a","doi-asserted-by":"crossref","award":["PCI2019-111900-2"],"award-info":[{"award-number":["PCI2019-111900-2"]}],"id":[{"id":"10.13039\/501100004837","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/501100004837","name":"Ministerio de Ciencia e Innovaci\u00f3n de Espa\u00f1a","doi-asserted-by":"crossref","award":["BFU2017-88549"],"award-info":[{"award-number":["BFU2017-88549"]}],"id":[{"id":"10.13039\/501100004837","id-type":"DOI","asserted-by":"crossref"}]},{"name":"Fundaci\u00f3n Bancaria \u201cla Caixa\u201d","award":["100010434"],"award-info":[{"award-number":["100010434"]}]}],"content-domain":{"domain":["www.ploscompbiol.org"],"crossmark-restriction":false},"short-container-title":["PLoS Comput Biol"],"abstract":"<jats:p>\n                    Projection neurons are the commonest neuronal type in the mammalian forebrain and their individual characterization is a crucial step to understand how neural circuitry operates. These cells have an axon whose arborizations extend over long distances, branching in complex patterns and\/or in multiple brain regions. Axon length is a principal estimate of the functional impact of the neuron, as it directly correlates with the number of synapses formed by the axon in its target regions; however, its measurement by direct 3D axonal tracing is a slow and labor-intensive method. On the contrary, axon length estimations have been recently proposed as an effective and accessible alternative, allowing a fast approach to the functional significance of the single neuron. Here, we analyze the accuracy and efficiency of the most used length estimation tools\u2014design-based stereology by virtual planes or spheres, and mathematical correction of the 2D projected-axon length\u2014in contrast with direct measurement, to quantify individual axon length. To this end, we computationally simulated each tool, applied them over a dataset of 951 3D-reconstructed axons (from\n                    <jats:ext-link xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" ext-link-type=\"uri\" xlink:href=\"http:\/\/NeuroMorpho.org\" xlink:type=\"simple\">NeuroMorpho.org<\/jats:ext-link>\n                    ), and compared the generated length values with their 3D reconstruction counterparts. The evaluated reliability of each axon length estimation method was then balanced with the required human effort, experience and know-how, and economic affordability. Subsequently, computational results were contrasted with measurements performed on actual brain tissue sections. We show that the plane-based stereological method balances acceptable errors (~5%) with robustness to biases, whereas the projection-based method, despite its accuracy, is prone to inherent biases when implemented in the laboratory. This work, therefore, aims to provide a constructive benchmark to help guide the selection of the most efficient method for measuring specific axonal morphologies according to the particular circumstances of the conducted research.\n                  <\/jats:p>","DOI":"10.1371\/journal.pcbi.1009051","type":"journal-article","created":{"date-parts":[[2021,12,8]],"date-time":"2021-12-08T13:44:12Z","timestamp":1638971052000},"page":"e1009051","update-policy":"https:\/\/doi.org\/10.1371\/journal.pcbi.corrections_policy","source":"Crossref","is-referenced-by-count":6,"title":["Benchmarking of tools for axon length measurement in individually-labeled projection neurons"],"prefix":"10.1371","volume":"17","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-1405-9885","authenticated-orcid":true,"given":"Mario","family":"Rubio-Teves","sequence":"first","affiliation":[]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3830-7297","authenticated-orcid":true,"given":"Sergio","family":"D\u00edez-Hermano","sequence":"additional","affiliation":[]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2170-2453","authenticated-orcid":true,"given":"C\u00e9sar","family":"Porrero","sequence":"additional","affiliation":[]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7978-3436","authenticated-orcid":true,"given":"Abel","family":"S\u00e1nchez-Jim\u00e9nez","sequence":"additional","affiliation":[]},{"given":"Luc\u00eda","family":"Prensa","sequence":"additional","affiliation":[]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0718-1337","authenticated-orcid":true,"given":"Francisco","family":"Clasc\u00e1","sequence":"additional","affiliation":[]},{"given":"Mar\u00eda","family":"Garc\u00eda-Amado","sequence":"additional","affiliation":[]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9220-1226","authenticated-orcid":true,"given":"Jos\u00e9 Antonio","family":"Villacorta-Atienza","sequence":"additional","affiliation":[]}],"member":"340","published-online":{"date-parts":[[2021,12,8]]},"reference":[{"issue":"12","key":"pcbi.1009051.ref001","doi-asserted-by":"crossref","first-page":"1497","DOI":"10.1177\/002215540104901203","article-title":"In vivo transduction of central neurons using recombinant Sindbis virus: Golgi-like labeling of dendrites and axons with membrane-targeted fluorescent proteins","volume":"49","author":"T Furuta","year":"2001","journal-title":"Journal of Histochemistry & Cytochemistry"},{"issue":"9","key":"pcbi.1009051.ref002","doi-asserted-by":"crossref","first-page":"2065","DOI":"10.1093\/cercor\/bhn231","article-title":"Kaneko, T. Two types of thalamocortical projections from the motor thalamic nuclei of the rat: a single neuron-tracing study using viral vectors","volume":"19","author":"E Kuramoto","year":"2009","journal-title":"Cerebral cortex"},{"key":"pcbi.1009051.ref003","doi-asserted-by":"crossref","first-page":"110","DOI":"10.3389\/fnana.2016.00110","article-title":"Connectomic analysis of brain networks: Novel techniques and future directions","volume":"10","author":"JL Cazemier","year":"2016","journal-title":"Frontiers in neuroanatomy"},{"key":"pcbi.1009051.ref004","doi-asserted-by":"crossref","first-page":"e10566","DOI":"10.7554\/eLife.10566","article-title":"A platform for brain-wide imaging and reconstruction of individual neurons","volume":"5","author":"MN Economo","year":"2016","journal-title":"Elife"},{"key":"pcbi.1009051.ref005","doi-asserted-by":"crossref","first-page":"27","DOI":"10.3389\/fnana.2016.00027","article-title":"A simple and efficient in vivo non-viral RNA transfection method for labeling the whole axonal tree of individual adult long-range projection neurons","volume":"10","author":"C Porrero","year":"2016","journal-title":"Frontiers in neuroanatomy"},{"issue":"1","key":"pcbi.1009051.ref006","doi-asserted-by":"crossref","first-page":"268","DOI":"10.1016\/j.cell.2019.07.042","article-title":"Reconstruction of 1,000 projection neurons reveals new cell types and organization of long-range connectivity in the mouse brain","volume":"179","author":"J Winnubst","year":"2019","journal-title":"Cell"},{"issue":"1","key":"pcbi.1009051.ref007","doi-asserted-by":"crossref","first-page":"166","DOI":"10.1002\/cne.24054","article-title":"Individual mediodorsal thalamic neurons project to multiple areas of the rat prefrontal cortex: A single neuron-tracing study using virus vectors","volume":"525","author":"E Kuramoto","year":"2017","journal-title":"Journal of Comparative Neurology"},{"issue":"12","key":"pcbi.1009051.ref008","doi-asserted-by":"crossref","first-page":"2840","DOI":"10.1093\/cercor\/bhr356","article-title":"A morphological analysis of thalamocortical axon fibers of rat posterior thalamic nuclei: a single neuron tracing study with viral vectors","volume":"22","author":"S Ohno","year":"2012","journal-title":"Cerebral cortex"},{"issue":"13","key":"pcbi.1009051.ref009","doi-asserted-by":"crossref","first-page":"2663","DOI":"10.1523\/JNEUROSCI.2886-19.2020","article-title":"Area-specific synapse structure in branched posterior nucleus axons reveals a new level of complexity in thalamocortical networks","volume":"40","author":"J Rodriguez-Moreno","year":"2020","journal-title":"Journal of Neuroscience"},{"issue":"12","key":"pcbi.1009051.ref010","doi-asserted-by":"crossref","first-page":"1713","DOI":"10.1038\/nn.4157","article-title":"Dendritic integration: 60 years of progress","volume":"18","author":"GJ Stuart","year":"2015","journal-title":"Nature neuroscience"},{"issue":"5\u20136","key":"pcbi.1009051.ref011","doi-asserted-by":"crossref","first-page":"817","DOI":"10.1007\/s00285-007-0113-7","article-title":"Mathematical foundations of the dendritic growth models","volume":"55","author":"JA Villacorta-Atienza","year":"2007","journal-title":"Journal of Mathematical Biology"},{"issue":"2","key":"pcbi.1009051.ref012","doi-asserted-by":"crossref","first-page":"275","DOI":"10.1016\/S0896-6273(02)00652-9","article-title":"Geometry and structural plasticity of synaptic connectivity","volume":"34","author":"A. 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