The 2014 Nobel Prize for Medicine has awarded the discovery of neuronal mechanisms that transform spatial movements into cerebral reference maps. The Prize for Chemistry has been assigned to the three scientists that developed super-resolved fluorescence microscopy up to a nanoscale resolution
Giuliana Miglierini
Spatial orienteering is a primary capacity of humans as well as all other animals. The sense of sight, one might think, should represent a relevant part of the complex mechanism at the base of the recognition of space inside which we move around and of the building of the consequent mental reference maps. May-Britt and Edvard Moser together with John O’Keefe demonstrated this is not the case, the process occurs entirely at the neurological level without the need of visible point marks, and the results they obtained have been awarded this year with the Nobel Prize for Physiology and Medicine.
Neuronal punctual activation
May-Britt and Edvard Moser are Norwegians working in their lab in Trondheim, which they built in 2007 with a grant of the Kavli Foundation. Far before that, they moved their first steps in neurophysiological research in the lab of John O’Keefe at the University College in London. O’Keefe, now 72, is an American who moved to the UK being fascinated by the British culture and particularly by the BBC, the NHS and Ordnance Survey map, as he told BBC Radio. In his free time, O’Keefe likes to walk in the country side and find his way around and maybe the passion for maps and orienteering suggested him to address its scientific attention towards the study of how the brain build the neuronal map of the space inside which we move. O’Keefe has been awarded with the Nobel Prize for his discovery, in 1971, of the so called place cells, a special kind of neuronal cells located into the hippocampus. Place cell activate and emit neurological signals as rats (but the same applies to all animal as well as to humans) move into the space, a squared box: a different set of activated place cells corresponds to each spatial point the rat pass on moving around, thus indicating that place cells should be somehow involved in the construction of the spatial map for movement. John O’Keefe is now director of the Sainsbury Wellcome Centre for Neural Circuits & Behaviour and professor of Cognitive neurosciences at the UCL University.
Grid cells mapping the space
In 2005 May-Britt and Edvard Moser published in Nature their first results about the role of the socalled grid cells, another type of specialised cells located into the entorhinal cortex of the hippocampus. In the paper, the Mosers described the how grid cells are directionally oriented and how they are activated upon passage of the animal on specific points in space, building a spatial map made up of an ensemble of equilteral triangles. Rats are implanted with cerebral electrodes able to detect a single neuron activation. In such a way, the Mosers recognised a one-to-one correlation between spatial points and grid cell’s activation, ending up in a global map of the spatial movement made of regular hexagons as rats move around in the box. Hexagonal geometry is often used by nature every time there is need for an highly efficient spatial order or energetic stability: examples are the cells of the beehive or the hexagonal geometry of benzene and other aromatic compounds, even at the nanolevel as graphene. The Mosers’ hypothesis is that the brain uses an hexagonal code to represent and transfer spatial information into neurological networks. Hexagonal reference schemes are persistent also in the dark and are apparently somehow linked and oriented by ‘connections’ with the box’s borders. Grid cells are stratified within the entorhinal cortex, from up to bottom, according to the size of the hexagonal grid they generate.
As rats move around, each grid cell is specifically activated and generates an electric signal in correspondence to a single point in space. The resulting code, summing all different activations and thus mapping the space, is then transmitted to the hippocampus, which is responsible for the associations between space, time and action that are at the base of thoughts and memory. Grid cells functionality and behaviour might be involved in early stage neuronal pathologies such as Alzheimer, where loss of the capability of spatial orienteering is often an early symptom, suggest Mosers’ results. The Nobel winning couple is now planning to further investigate how spatial recognition mechanisms develop from birth on in order to adapt to the external environment.
Nanoscale optical microscopy
A wall has been torned apart by the three winners of the 2014 Nobel Prize for Chemistry: thanks to the innovations the introduced, optical microscopy is no longer limited by the Abbe’s limit (Box “Limits for Optical Microscopy”), corresponding approximately to half the wavelength of the light used by the microscope. Traditional optical microscopy has a resolution up to approx. 200 nm, thus it is useful to detect bacteria but not viruses, proteins or small molecules. The new techniques introduced by Stefan Hell, director of the Max Planck Institute for Biophysical Chemistry in Gottingen, William E. Moerner, physical chemist from Stanford University and Eric Betzig, physics at the Howard Hughes Medical Institute, allow now to reach resolutions up to nanometers, and ‘nanoscopy’ might become a more convenient term to identify a technique that today may detect single molecules without the disadvantages typical of electronic microscopy (i.e. the need of high vacuum which prevent the use of living cells as samples).
Stefan Hell introduced in 2000 the Stimulated Emission Depletion (Sted) method of analysis: a first laser beam enlights the fluorescently marked sample; a second laser is used to ‘remove’ fluorescence, like an eraser, except from a nanometer-wide portion of the sample which is thus magnified. The global image of the sample is reconstructed upon acquisition and addition of many single images at the nano-scale level, thus allowing to overcome Abbe’s limit.
Blue light for single molecule detection
Betzig and Moerner further developed the method and separately introduced Single fluorphore microscopy (SFP), a technique where a small quantity of a fluorescent protein (i.e. GFP, green fluorescent protein) is added to the analytical sample. Using a very weak blue light the protein become temporarily fluorescent: the repeated enlighting of the sample allows for the detection, at each passage, of a different subset of fluorescent proteins. The global image of the sample is obtained summing the single frames. Using the technique of GFP’s dispersion into gel, Moerner has been the first scientist able to detect a single molecule using optical microscopy. Eric Betzig has applied the technique to the study of specific components within the cells.
Antoni van Leeuwenhoek invented optical microscopy in the second half of 1600 as a method to better test carpet’s quality; he was the first to observe red blood cells, somatic muscle cells and protozoans. After 400 years, a new era is opening for optical microscopy thanks to Nobel Prize’s winning innovation.
LIMITS FOR OPTICAL MICROSCOPY
Ernst Abbe in 1873 established the optical limit for the resolution of two different points in a sample in half of the wavelength of the radiating light, corresponding to approximately 200 nm.
A traditional optical microscope can be used to detect animal cells (50 m) and, with some more effort, bacteria (500 nm). Viruses (100 nm), proteins (10 nm) and small molecules (few nm) are not detectable this way.
Very small microorganisms or molecules are detectable with electronic microscopy with resolutions up to 100 pm. Yet, this technique too has a limit: it requires high vacuum conditions, thus it is not compatible with in vivo cell analysis.