A team of researchers from MIT have successfully described a technology pipeline that enabled them to finely process, richly label and sharply image full hemispheres of the brains of two donors at high resolution and speed, helping further analysis of the human brain.
The study used one brain with Alzheimer’s and one without, and then analysed them using an integrated suite of three technologies that allowed the researchers to explore long-sought neuroscience investigations, such as the creation of a comprehensive map or atlas of the entire brain with full hemispheric imaging. Beneficially, this means every cell, circuit and protein can be identified and analysed.
The research, published in Science, provides a ‘proof of concept’ by showing numerous examples of what the pipeline makes possible, such as sweeping landscapes of thousands of neurons within whole brain regions, diverse forests of cells each in individual detail, and tufts of subcellular structures nestled among extracellular molecules. In the paper, the researchers also present a variety of quantitative analytical comparisons focused on a chosen region within the Alzheimer’s and non-Alzheimer’s hemispheres.
Kwanghun Chung, associate professor in The Picower Institute for Learning and Memory, the Departments of Chemical Engineering and Brain and Cognitive Sciences, and the Institute for Medical Engineering and Science at MIT, said: “We performed holistic imaging of human brain tissues at multiple resolutions from single synapses to whole brain hemispheres and we have made that data available. This technology pipeline really enables us to analyse the human brain at multiple scales. Potentially this pipeline can be used for fully mapping human brains.”
Understanding the benefits of mapping with the new process
Being able to map whole hemispheres of human brains intact and down to the resolution of individual synapses is beneficial for understanding the human brain in health and disease for multiple reasons.
Firstly, it enables scientists to conduct integrated explorations of questions using the same brain, rather than having to observe different phenomena in different brains, which can vary significantly, and then trying to construct a composite picture of the whole system.
At the same time, this approach doesn’t degrade the brain tissue. Instead, it makes the tissues extremely durable and repeatedly re-labelable, allowing for highlighting of different cells or molecules as needed for new studies, potentially for years on end. This is demonstrated by the researchers who use 20 different antibody labels to highlight different cells and proteins, though they are already expanding that to 100 or more.
Kwanghun Chung added: “We need to be able to see all these different functional components – cells, their morphology and their connectivity, subcellular architectures, and their individual synaptic connections – ideally within the same brain, considering the high individual variabilities in the human brain and considering the precious nature of human brain samples. This technology pipeline really enables us to extract all these important features from the same brain in a fully integrated manner.”
The researchers are also optimistic that they may be able to create a brain bank of fully imaged brains that researchers could analyse and re-label as needed for new studies to make further comparisons. This is made possible by the pipeline’s relatively high scalability and throughput, allowing for sampling different sexes, ages, and disease, as well as robust follow-up analysis.
Recognising the approaches used by the researchers
The team’s senior and corresponding author stated that the greatest challenge was building a team at MIT that included three especially talented young scientists, each a co-lead author of the paper because of their key roles in producing the three major innovations in the pipeline.
Ji Wang, a mechanical engineer and former postdoc, developed the ‘Megatome’, a device for slicing intact human brain hemispheres so finely that there is no damage to it. Juhyuk Park, a materials engineer and former postdoc, developed the chemistry that makes each brain slice clear, flexible, durable, expandable, and quickly, evenly and repeatedly labelable – a technology called ‘mELAST’. Webster Guan, a former MIT chemical engineering graduate student with a knack for software development, created a computational system called ‘UNSLICE’ that can seamlessly reunify the slabs to reconstruct each hemisphere in full 3D down to the precise alignment of individual blood vessels and neural axons.
These aspects were all key to the research as no technology allows for imaging whole human brain anatomy at subcellular resolution without first slicing it because it is very thick and opaque. With the Megatome, tissue could remain undamaged as its blade could vibrate side-to-side faster and sweep wider than previous vibratome slicers. This produced slices that didn’t lose anatomical information at their separation or anywhere else, at speeds that were previously achievable.
Though the use of mELAST meant that slabs in the pipeline could be thicker, as the approach used hydrogel that infuses with the brain sample to make it optically clear, virtually indestructible and compressible and expandable. Combined with other chemical engineering technologies developed, the samples can then be evenly and quickly infused with the antibody labels that highlight cells and proteins of interest. According to the researchers, by using a light sheet microscope customised in the lab, a whole hemisphere could then be imaged down to individual synapses in about 100 hours.
Juhyuk Park stated: “This advanced polymeric network, which fine-tunes the physicochemical properties of tissues, enabled multiplexed multiscale imaging of the intact human brains.”
After imaging each slab, the researchers worked to restore an intact picture of the whole hemisphere computationally. The UNSLICE does this at multiple scales. For example, at the middle, or ‘meso/ scale, it algorithmically traces blood vessels coming into one layer from adjacent layers and matches them. It can also purposely label neighboring neural axons in different colours (like the wires in an electrical fixture), so that the researchers could match layers up based on tracing the axons.
The results of using the pipeline show how diverse the labeling can be, revealing long axonal connections and the abundance and shape of different cell types including not only neurons but also astrocytes and microglia.
Exploring Alzheimer’s in human brains
Ultimately, the technology has allowed Kwanghun Chung and fellow researchers to image and understand Alzheimer’s disease in human brains. The new pipeline offered an open-ended exploration, first identifying where within a slab of tissue the greatest loss of neurons can be seen. From there, they were able to produce a series of detailed investigations.
Kwanghun Chung said: “We didn’t lay out all these experiments in advance. We just started by saying, ‘OK, let’s image this slab and see what we see.’ We identified brain regions with substantial neuronal loss so let’s see what’s happening there. ‘Let’s dive deeper.’ So we used many different markers to characterise and see the relationships between pathogenic factors and different cell types. This pipeline allows us to have almost unlimited access to the tissue. We can always go back and look at something new.”
As the research produced just two samples, the team is not offering any conclusions about the nature of Alzheimer’s disease, however they have shown that the capability (which can apply to other bodily tissues) now exists to fully image and deeply analyse whole human brain hemispheres to enable exactly that kind of research.
The researchers concluded: “We envision that this scalable technology platform will advance our understanding of the human organ functions and disease mechanisms to spur development of new therapies.”
Lead image: University of Minnesota
