The brain is as beautiful as it is complex. Throughout the centuries scientists have attempted to glean understanding from the brain based on its anatomy. Since Santiago Ramn y Cajals detailed drawings at the turn of the 20th Century, neuroscientists of the modern era have been zooming in on the brain to unlock its secrets. Recent improvements in imaging technologies, greater access to microscopes and specialist software have culminated in the cutting-edge imaging currently being performed on brains and brain tissue across the world.1. Recreating the retina: drawing it by hand

Line drawing of the retina. Ramn y Cajal. Wellcome Images.The neuroanatomist and father of modern neuroscience, Santiago Ramn y Cajal looked at brain tissue stained using the Golgi Stain method under his microscope. He then drew what he saw, producing beautiful reconstructions of neurons such as the line drawing of the retina above.

2. Recreating the retina: from electron micrographs

Connectome of the mouse retina. Max Planck Society

Nowadays, whole sections of the retina have been reconstructed in unprecedented three-dimensional detail. In 2013 Helmstaedter et al. published their paper on the connectomic reconstruction of the inner plexiform layer in the mouse retina. The group used a lot of manpower to manually reconstruct the neurons in a small section of the mouse retina by tracing projections through electron micrograph sections.Related:From Cajal to now

3. Antibody-labelling lights up the brainBy using primary antibodies to target a protein of interest, neuroscientists can then use a fluorescent secondary antibody against the primary to increase the fluorescence signal, producing pretty yet insightful images. Such as the following...

Confocal micrograph image of the adult mouse hippocampus, with immunofluorescent staining for perineuronal nets. Blue = DAPI (cell nuclei), green = parvalbumin interneurons, red = wisteria floribunda agglutinin (WFA; commonly used to label perineuronal nets). Credit Adam Ramsaran, The Hospital for Sick Children (Toronto, ON).

Confocal micrograph of the anterior region of the developing zebrafish brain.Some of the neurons (shown in green) express the green fluorescent protein (GFP) under the control of specific gene expression. Axons, tracts and neuropils have been labeled using antibodies that mark tubulin (in red) and synaptic vesicles (in blue). Credit: Monica Folgueira and Steve Wilson, Wellcome Images.

Confocal micrograph of hippocampal interneurons that express serotonin receptors (green) co-labeledin red for calretinin, and counterstained with DAPI (blue), a marker of cell nuclei. Credit: Margaret I. Davis

4. Speeding up labeling

Antibody labeling and imaging in Drosophila melanogaster brains can take up to a week. In this image, genetically encoded chemical tags have been expressed in neurons of interest. It takes just 15 minutes to stain for these tags with their substrates. Credit: Ben Sutcliffe, Jefferis lab. MRC Laboratory of Molecular Biology.

5. Deeper insights into the brain

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YFP mouse brain section. The 720m z stack is color coded for depth, such that red cells are deepest in the tissue and the blue cells shallowest. Credit Dr. Mark Lessard, The Jackson Laboratory, Bar Harbor, Maine, USA.

Multiphoton imaging systems, such as Leica's SP8multiphoton confocal system (used to capture the video above), use longer and less-damaging wavelength beams of light to excite fluorescent proteins deeper in tissue, enabling neuroscientists to see complete structures in intact tissue.

6. CLARITY: See through brains

Despite improvements in imaging systems, such as multiphoton laser scanning microscopes, the brain tissue itself hinders imaging, by causing light to scatter.In 2013, Chung et al. published their seminal paper on their hydrogel method, CLARITY, which makes brain tissue transparent, reducing the scattering of light, affording structural and molecular interrogation of the whole intact brain.

7. Live imaging in the brain

Multiphoton image of microglia (GFP, green) and cerebral blood vessels (Texas-red dextran, red) in a living, anesthetized transgenic mouse. Harris A. Gelbard, 2009 Olympus BioScapes Digital Imaging Competition.

8. Imaging Activity With Genetically Encoded Calcium Indicators

This video from the Sur labshows In vivo imaging of visual cortex neurons expressing the genetically encoded calcium indicator GCamp6 responding to a moving grating (top right corner).

9. A Look Inside the Head With Magnetic Resonance Imaging

This video from the Human Connectome Projectexplains how Principal Diffusion Direction uses colors to map out the different directions that water molecules diffuse through the brain's white matter. This data is then used to reconstruct 3D representations of the white matter tracts in the brain.

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Imaging the Brain in the Era of Modern Neuroscience - Technology Networks