Seeing neurobiology at every scale

Neurobiology functions at different levels of organization, from molecular scale-protein interactions to a whole organism’s coordination of organs and signals. Each level contributes to the complete picture. However, no single imaging system is optimized to capture that complexity. At St. Jude, the Neuroimaging Laboratory (NIML), part of the Department of Developmental Neurobiology, meets this challenge, building bespoke, advanced imaging solutions that empower researchers to make meaningful discoveries. 

NIML is staffed by experts working at the forefront of the fields of imaging science and engineering, such as Daniel Stabley, PhD, NIML director, Sharon King, PhD, and Becky Petersen, PhD, associate scientist. These investigators build advanced microscopes with their own hands, tailored to the nuances of performing imaging experiments for neuroscience applications. NIML experts provide invaluable collaboration, helping researchers navigate the complexities of what imaging is needed and how to troubleshoot project-specific complications if problems arise.

The proof of these capabilities is best represented by the images they capture and the discoveries they enable.

Heterochromatin in a neuronal nucleus

At the subcellular scale, electron microscopy (EM) captures single-nanometer-level details. However, EM preparations cannot easily label a specific protein. Fluorescent imaging, which is much lower resolution, can. Cryogenic Correlative Light and Electron Microscopy (Cryo-CLEM) combines EM with a fluorescence image of the same sample at a resolution of hundreds of nanometers, enabling scientists to pair those data to reveal more biology than either technique could in isolation.

Represented in black, white and gray is an EM image of a neuron’s nucleus, with a bump map of signal intensity. The translucent blue represents the three-dimensional (3D) positions of HP1α, a heterochromatin protein labeled with a fluorescent marker, computationally aligned with the EM image to show where it exists in the nucleus, a finding that was published in Science. By studying this organization, researchers gained a better understanding of how a cancerous neuron organizes its gene expression to promote disease and alter its interactions with surrounding cells.

Junctions between two potentially cancerous neurons

EM data can also be combined with fluorescence to study cells, with scales at the hundreds-of-nanometers level, to see the behaviors of individual or small numbers of cells. Researchers overlaid fluorescent imaging of two adhesion proteins, Drebrin in pink and JAM-C in green, on gray EM images of their cell bodies to study junctions between cerebellar neurons. Documenting how neurons and neighboring cells form junctions helps researchers learn how these cells influence each other during development, and how that behavior changes when they become cancerous. 

Capturing many cells together

Instead of looking at individual or small groups of cells, scientists often picture many cells at once to observe emergent patterns. By looking at fluorescent imaging at the hundreds of nanometers to micrometer levels, researchers can document cell behaviors at a larger scale.

In this image, fluorescently labeled actin proteins are shown in blue and microtubule proteins are labeled in magenta, with the cell nucleus in red. Both proteins form major portions of the cytoskeleton. The cytoskeleton is a large structure of proteins throughout the cell that anchors adhesion proteins, indirectly contributing to cell-to-cell interactions at junctions. Understanding these cytoskeletal components and how they enable cells to influence each other can provide insights into brain cell development and cell migration, as well as tumor emergence.

Imaging of an organoid

At the scale of micrometers to millimeters, scientists have traditionally studied tissue organization and function using cell lines. However, most cell lines grow in a flat plane, which can cause researchers to miss important organizational details. Tissues are 3D in nature, with many groups of cells working together to coordinate development or be disrupted in disease. To account for this dynamism, researchers sometimes use organoids — tissues derived from patient tumors into 3D spheroids. 

In this set of images, scientists created a spheroid of a patient’s tumor sample. They then labeled the inside of cells with a cytosolically expressed fluorescent protein (gold). Examining the whole spheroid (left) and a significant zoom into one small component (right) enables researchers to look at the cellular and organoid levels to connect how changes in small interactions lead to large changes at the tissue level.

Capturing at the scale of the brain

At the level of a whole organ, image scales are measured in millimeters and centimeters. Instead of focusing on how individual cells influence each other, researchers look at how cells work in aggregate in an organized structure, such as during brain development. 

In two sections through the whole brain, researchers used the gold fluorescence to identify where tumors emerged. They found that during brain development, tumors initiated at specific sites based on which cells were interacting with each other and the presence of a mutation in a chromatin protein. The results were published in Nature Communications.

Creating a clearer picture with NIML

At the intersection of the fast-moving fields of physics, biology and computer science, imaging continues to advance rapidly. These tools are revealing new neurobiology and decoding cancer development, providing opportunities for therapeutic interventions. While researchers focus on biological discoveries, the St. Jude NIML can convey the newest developments in image acquisition and help investigators search the width and depth (and sometimes height) of their imaging, at any scale, with the best state-of-the-art techniques. Enabled by their technical support, scientists at St. Jude are already making discoveries that are pushing the boundaries of neurobiology and cancer research forward.