Visualization and analysis of whole depot adipose tissue neural innervation
Jake W. Willows,
Magdalena Blaszkiewicz,
Amy Lamore,
Samuel Borer,
Amanda L. Dubois,
Emma Garner,
William P. Breeding,
Karissa B. Tilbury,
Andre Khalil,
Kristy L. Townsend
Affiliations
Jake W. Willows
School of Biology and Ecology, University of Maine, Orono, ME, USA; Department of Neurological Surgery, The Ohio State University, 1014 Biomedical Research Tower, 460 W. 12th Avenue, Columbus, OH, USA
Magdalena Blaszkiewicz
School of Biology and Ecology, University of Maine, Orono, ME, USA; Graduate School of Biomedical Science and Engineering, University of Maine, Orono, ME, USA; Department of Neurological Surgery, The Ohio State University, 1014 Biomedical Research Tower, 460 W. 12th Avenue, Columbus, OH, USA
Amy Lamore
School of Molecular and Biomedical Sciences, University of Maine, Orono, ME, USA
Samuel Borer
School of Biology and Ecology, University of Maine, Orono, ME, USA
Amanda L. Dubois
School of Molecular and Biomedical Sciences, University of Maine, Orono, ME, USA
Emma Garner
School of Biology and Ecology, University of Maine, Orono, ME, USA
William P. Breeding
Department of Chemical and Biomedical Engineering, University of Maine, Orono, ME, USA
Karissa B. Tilbury
Graduate School of Biomedical Science and Engineering, University of Maine, Orono, ME, USA; Department of Chemical and Biomedical Engineering, University of Maine, Orono, ME, USA
Andre Khalil
Graduate School of Biomedical Science and Engineering, University of Maine, Orono, ME, USA; Department of Chemical and Biomedical Engineering, University of Maine, Orono, ME, USA; CompuMAINE Laboratory, University of Maine, Orono, ME, USA
Kristy L. Townsend
School of Biology and Ecology, University of Maine, Orono, ME, USA; Graduate School of Biomedical Science and Engineering, University of Maine, Orono, ME, USA; School of Molecular and Biomedical Sciences, University of Maine, Orono, ME, USA; Department of Chemical and Biomedical Engineering, University of Maine, Orono, ME, USA; Department of Neurological Surgery, The Ohio State University, 1014 Biomedical Research Tower, 460 W. 12th Avenue, Columbus, OH, USA; Corresponding author
Summary: Little is known about the diversity and function of adipose tissue nerves, due in part to the inability to effectively visualize the tissue’s diverse nerve subtypes and the patterns of innervation across an intact depot. The tools to image and quantify adipose tissue innervation are currently limited. Here, we present a method of tissue processing that decreases tissue thickness in the z-axis while leaving cells intact for subsequent immunostaining. This was combined with autofluorescence quenching techniques to permit intact whole tissues to be mounted on slides and imaged by confocal microscopy, with a complementary means to perform whole tissue neurite density quantification after capture of tiled z-stack images. Additionally, we demonstrate how to visualize nerve terminals (the neuro-adipose nexus) in intact blocks of adipose tissue without z-depth reduction. We have included examples of data demonstrating nerve subtypes, neurovascular interactions, label-free imaging of collagen, and nerve bundle digital cross-sections.
