A Cellular Positioning System to Probe Morphological Heterogeneity Among Mouse CA3 Hippocampal Pyramidal Neurons

Handwerk, C. J.; Bland, K. M.; Brett, C. A.; Denzler, C. J.; Kalinowski, A. R.; Swinehart, B. D.; Rodriguez, H. V.; Cook, H. N.; Vidal, G. S.

doi:10.1101/2022.08.12.503761

bioRxiv · 10.1101/2022.08.12.503761

A Cellular Positioning System to Probe Morphological Heterogeneity Among Mouse CA3 Hippocampal Pyramidal Neurons

Handwerk, C. J.·Bland, K. M.·Brett, C. A.·Denzler, C. J.·Kalinowski, A. R.·Swinehart, B. D.·Rodriguez, H. V.·Cook, H. N.·Vidal, G. S.

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Abstract

BackgroundPyramidal neurons throughout hippocampal CA3 are diverse in their dendritic morphology, and CA3 is not homogenous in its structure or function. Nonetheless, few structural studies have captured the precise 3D somatic position and the 3D dendritic morphology of CA3 pyramidal neurons simultaneously. New methodHere, we present a simple approach to reconstruct the apical dendritic morphology of CA3 pyramidal neurons using the transgenic fluorescent Thy1-GFP-M line. The approach simultaneously tracks the dorsoventral, tangential, and radial positions of reconstructed neurons within the hippocampus. It is especially designed for use with transgenic fluorescent mouse lines, which are commonly used in genetic studies of neuronal morphology and development. ResultsWe demonstrate how topographic and morphological data are captured from transgenic fluorescent mouse CA3 pyramidal neurons. Comparison with existing methodsThere is no need to select and label CA3 pyramidal neurons with the transgenic fluorescent Thy1-GFP-M line. By taking transverse (not coronal) serial sections, we preserve fine dorsoventral, tangential, and radial somatic positioning of 3D-reconstructed neurons. Because CA2 is well defined by PCP4 immunohistochemistry, we use that technique here to to increase precision in defining tangential position along CA3. ConclusionsWe developed a method for simultaneously collecting precise somatic positioning as well as 3D morphological data among transgenic fluorescent mouse hippocampal pyramidal neurons. This fluorescent method should be compatible with many other transgenic fluorescent reporter lines and immunohistochemical methods, facilitating the capture of topographic and morphological data from a wide variety of genetic experiments in mouse hippocampus. HighlightsO_LISimultaneous capture of 3D location and pyramidal dendritic morphology in CA3 C_LIO_LIMethod utilizes replicable techniques and reagents available to most laboratories C_LIO_LIMethod is adaptable to other transgenic mouse lines and immunohistochemical studies C_LI

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Handwerk, C. J., Bland, K. M., Brett, C. A., Denzler, C. J., Kalinowski, A. R., Swinehart, B. D., Rodriguez, H. V., Cook, H. N., Vidal, G. S.. 2022-08-15. A Cellular Positioning System to Probe Morphological Heterogeneity Among Mouse CA3 Hippocampal Pyramidal Neurons. https://doi.org/10.1101/2022.08.12.503761

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