A new DNA-sequencing based idea for mapping the connectome is presented in Plos biology.
The defining purpose of neurons is information transmission and processing within a network. Hence, to appreciate neural function we must look at the interactions between neurons; understand their connections; know which other neurons they synapse with.
The idea of documenting all the connections within brains – mapping the ‘connectome’ – is receiving a lot of attention (and money) at the moment. However, the only comprehensive technique so far available is the painful reconstruction of the synaptic map from electron micrographs of serially sectioned brains. Seeing as the human brain is estimated to contain 85 billion neurons making 1014/1015 synaptic connections, one does have to wonder whether even if we could accurately document the connectome and afford it, would we really appreciate it?
The only organism to have had it’s connectome documented in this way is the nematode worm, C. elegans. Mapping its’ 302 neurons and their 7000 connections required over 50 person-years of labour. Despite the utility of this information, and the intrinsic glory of the knowledge, it should be noted that it most certainly can’t be said that ‘we understand the C. elegans nervous system’.
Another, very elegant technique being developed, with the capacity to map connectomes at the mesoscopic level, is the use of rabies-type viruses. These viruses can be transferred across synapses, but engineered to only do so once. Carrying fluorescent protein encoding reporter genes, they can be used to track the connecting neurons within a network. As one can engineer these viruses to carry allsorts of genetic trickery, the mapping of networks can go hand in hand with functional experimentation.
The trans-synaptic transfer of rabies viruses, along with the use of randomising recombination to create hundreds of different combinations of fluorophores in ‘brainbow’ neuronal imaging, are major inspirations underlying a new theoretical connectome mapping technique laid out by Zador et al.
The technique, termed BOINC (barcoding of individual neuronal connections) converts the mapping of the connectome from an anatomical problem into one that can be tackled by DNA sequencing. As the costs of sequencing are currently dropping through the floor, BOINC would make mapping the connectome a repeatable assay rather than a one-off mega-mission.
The method can be divided into three phases. In the first stage, each neuron is labelled with a unique DNA sequence – a barcode. The authors calculate that a random sequence of 20 nucleotides would be sufficient to individually label the entire neuronal complement of the mouse brain (1012 possible sequences to <108 neurons). Zador et al. are sketchy about the specifics of how this could be achieved; suffice to say that it’s conceptually similar to the generation of antibody diversity by recombination.
The second stage is the association of barcodes from synaptically connected neurons. This would be achieved by the single transynaptic spread of rabies-like viruses. The barcode would therefore be carried within the virus genome. In the third phase, the barcodes must be joined together. Each neuron will therefore contain its’ own barcode combined with barcodes from every cell that it synapses with. These tags would then be sequenced – yielding a connectivity matrix.
This term perhaps clarifies the biggest shortcoming of the technique; it creates a matrix, devoid of anatomical or functional detail. However, these dimensions would surely become gradually coupled with repeated experimentation.
In short, BOINC is a very clever idea. If they pull it off, it will be a great advance, allowing cheap, repeated screening of the brain’s circuitry. Obviously, it’s unlikely to be much help for studying the human brain, but providing the technical hurdles are surmountable it could revolutionize the neurobiology of model systems such as the mouse.
Zador, A.M., Dubnau, J., Oyibo, H.K., Zhan, H., Cao, G.,Peikon, I.D. (2012). Sequencing the Connectome Plos Biology, 10 (10) : 10.1371/journal.pbio.1001411
The original paper is available here
Check out some excellent blogposts by Mo Costandi at Neurophilosophy; A book review discussing the connectome concept (with special bilious email from Andrew Lumsden), and descriptions of work using Rabies viruses for mapping connectivity and functional neurobiology.