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. 2021 Jun 28;12:4004. doi: 10.1038/s41467-021-24241-2

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — a Raw data examples of cortico-tectal projections targeting distinct zones within SC layers at ARA 90. Anterograde AAV injections into PTLp→SC.m (red), VISam→SC.cm (orange), ACAd→SC.cl (green), and MOp→SC.l (purple) terminate with different laminar and nonoverlapping patterns. Together, these representative cortico-tectal projections reveal layer-specific terminals distributed across four distinct zones along the medial-lateral axis of SC. Scale bar at injection sites is 500 µm, and 200 µm in lower SC panels. b The Connection Lens neuroinformatics workflow: Raw tissue image with anterograde labeling is assigned to the corresponding standard atlas level (this example is ARA 96). The tissue is warped based on template atlas borders and reconstructed using in-house neuroinformatics software. Thresholded images are overlapped onto a custom atlas for zone- and layer-specific registration for pixel quantification. c Left: Polar Coordinate analysis method used to quantify angular distribution of thresholded pixel labeling in SC. Angles represented by theta (θ°) values where midline starts at 90° and ranges toward 0° at lateral angles. Right: Custom SC atlas with overlay of angular range shows coarse alignment of manually delineated borders in SC. d Example of probability distribution graph for average distributions of zone-specific cortico-tectal cases. Peaks are aligned with custom SC borders at ARA 90. SC.m (red) is SW180522-04A. SC.cm (orange) is SW121221-03A. SC.cl (green) is SW171130-02A. SC.l (purple) is SW170410-04A. e Layer- and zone-specific SC nomenclature to facilitate referencing and quantification. Below: zone delineations across representative ARA levels (86, 90, 96, 100) spanning rostral-to-caudal SC. ACAd anterior cingulate cortex dorsal part, ARA Allen reference atlas, AAV Adeno-associated virus, PHAL Phaseolus vulgaris leucoagglutinin, PTLp posterior parietal cortex, MOp primary motor cortex, VISam visual cortex anterior medial part, SC superior colliculus, SC Layers: zo zonal, sg superficial gray, op optic, ig intermediate gray, iw intermediate white, dg deep gray, dw deep white.