Elsevier

NeuroImage

Volume 58, Issue 3, 1 October 2011, Pages 732-740
NeuroImage

Segmentation of the mouse hippocampal formation in magnetic resonance images

https://doi.org/10.1016/j.neuroimage.2011.06.025Get rights and content

Abstract

The hippocampal formation plays an important role in cognition, spatial navigation, learning, and memory. High resolution magnetic resonance (MR) imaging makes it possible to study in vivo changes in the hippocampus over time and is useful for comparing hippocampal volume and structure in wild type and mutant mice. Such comparisons demand a reliable way to segment the hippocampal formation. We have developed a method for the systematic segmentation of the hippocampal formation using the perfusion-fixed C57BL/6 mouse brain for application in longitudinal and comparative studies. Our aim was to develop a guide for segmenting over 40 structures in an adult mouse brain using 30 μm isotropic resolution images acquired with a 16.4 T MR imaging system and combined using super-resolution reconstruction.

Research highlights

► Development of an MR-based mouse hippocampal atlas ► Delineation of the anatomical boundaries of 40 hippocampal sub-regions ► Volumetric analysis of hippocampal structures in the adult mouse brain

Introduction

The hippocampal formation is made up of a number of distinct regions. It includes the hippocampus proper (which consists of the cornu ammonis (CA) fields 1–3 and the dentate gyrus (DG)), the subiculum and adjacent presubiculum, postsubiculum, and parasubiculum, the entorhinal cortex, the perirhinal cortex, and rudimentary components of the hippocampus (indusium griseum, dorsal tenia tecta, and ventral tenia tecta) (Amaral et al., 2007, Andersen, 2007). Structural changes in the hippocampus have recently been studied using MR in mouse models of human disease, including Alzheimer's disease (Borthakur et al., 2006, Lau et al., 2008) and Huntington's disease (Sawiak et al., 2009). Until recently, the value of digital atlases of the hippocampus has been limited by inadequate resolution in MR images, resulting in difficulty distinguishing regional boundaries (Ma et al., 2005, Chen et al., 2006). The use of high resolution MR has made possible the visualization of anatomical structures at a level of detail approaching that of histological preparations (Mueller et al., 2007, Boretius et al., 2009). This has resulted in the emergence of MR imaging atlases with increasing levels of anatomical detail (Benveniste et al., 2000, MacKenzie-Graham et al., 2004, Kovacevic et al., 2005, Badea et al., 2007, Dorr et al., 2008, Johnson et al., 2010).

Harnessing the resolution available with high-field MR imaging requires reproducible methods that allow consistent segmentation of hippocampal structures across mouse brains. One of the difficulties in separating the many sub-regions within the hippocampal formation is the lack of agreed guidelines for defining the boundaries of each region. Our aim was to generate a set of operational criteria for the segmentation of over 40 structures in the hippocampal formation in MR images. These operational criteria are supplemented with a digital atlas and brief description of each region to assist others who may wish to utilize our segmentation protocol. We developed a hippocampal atlas that is consistent with the segmentation of images of histological sections.

Section snippets

C57BL/6J mouse brain preparation and magnetic resonance imaging

All procedures involving animals were approved by the University of Queensland Animal Ethics Committee (AEC: CMR/907/08/NHMRC). A twelve-week old male inbred C57BL/6J mouse was initially anesthetized with isoflurane. Prior to perfusion, the mouse was injected with 0.02 ml/g of pentobarbitone sodium (Lethabarb). Using a Perfusion One® system (MyNeuroLab, St Louis, MO, USA), the mouse circulation was flushed with 0.1 M phosphate buffer (PB; pH 7.4) and then fixed with a solution containing 4%

Results

The hippocampal formation was segmented into major regions and sub-regions as summarized in Table 1. The level of contrast in the CA and DG sub-regions and their layers enabled ready delineation (Fig. 1). On the other hand, subdivisions of the entorhinal cortex (Fig. 2) were difficult to segment because of the lack of definitive contrast. We supplemented our interpretation of the MR images with data from Nissl stained sections as well as in situ hybridization data for gene expression from

Discussion

We have developed a method for the systematic segmentation of the hippocampal formation in MR images in the mouse. In developing the operational criteria for segmentation we have supplemented MR findings with data from cytoarchitecture and gene expression studies. This is the first MR image segmentation that outlines the hippocampal structures in such detail. The method we have developed is applicable to studies that attempt to analyze changes in the hippocampus in mouse models of neurological

Conclusion

This study presents a new method for the systematic and detailed segmentation of MR images of the hippocampal formation in the mouse brain. This method provides the criteria for future development of a probabilistic atlas of the hippocampal formation, where data acquired from a large number of brains will be used to produce a canonical reference to which mutant strains can be compared.

Acknowledgments

We thank the Queensland NMR Network (QNN) and the National Imaging Facility (NIF) for instrument access and technical support. This work was funded by the National Health and Medical Research Council (NHMRC) of Australia.

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