Our current understanding of brain function is still too limited to take advantage of the computational power of even the simplest biological nervous systems. To fill this gap, the Si elegans project (www.si-elegans.eu) aims at developing a computational framework that will replicate the nervous system and rich behavior of the nematode Caenorhabditis elegans, a tiny worm with just 302 neurons. One key element of this emulation testbed is an electro-optical, micromirrorbased connectome. Unlike any other current ICT communication protocol, we expect it to accurately mimic the parallel information transfer between neurons. This strategy promises to give new insights into the nature of two hypothesized key mechanisms - the parallel and precisely timed information flow - that make brains excel von-Neumann-type machines. In this contribution, we briefly introduce the overall Si elegans concept to then describe the requirements for designing a light-based connectome within the given boundary conditions imposed by the hardware infrastructure it will be integrated into.
During development, the axons of neurons in the mammalian central nervous system lose their ability to regenerate after
injury. In order to study the regeneration process, we developed a system integrating an optical tweezers and a laser
dissector to manipulate the sample. A sub-nanosecond pulsed UVA laser was used to inflict a partial damage to the axon
of mouse hippocampal neurons at early days in vitro. Partial axonal transections were performed in a highly controlled
and reproducible way without affecting the regeneration process. Force spectroscopy measurements, during and after the
ablation of the axon, were performed by optical tweezers with a bead attached to the neuronal membrane. Thus, the
release of tension in the neurite could be analyzed in order to quantify the inflicted damage. After dissection, we
monitored the viscoelastic properties of the axonal membrane, the cytoskeleton reorganization, and the dynamics of the
newly formed growth cones during regeneration. In order to follow cytoskeleton dynamics in a long time window by
tracking a bead attached to the neuron, we developed a real-time control of the microscope stage position with sub-millisecond
and nanometer resolution. Axonal regeneration was documented by long-term (24-48 hours) bright-field live
imaging using an optical microscope equipped with a custom-built cell culture incubator.
Francesco Difato, Marco Dal Maschio, Emanuele Marconi, Giuseppe Ronzitti, Alessandro Maccione, Tommasso Fellin, Luca Berdondini, Evelina Chieregatti, Fabio Benfenati, Axel Blau
Regeneration of functional connectivity within a neural network after different degrees of lesion is of utmost clinical importance. To test pharmacological approaches aimed at recovering from a total or partial damage of neuronal connections within a circuit, it is necessary to develop a precise method for controlled ablation of neuronal processes. We combined a UV laser microdissector to ablate neural processes in vitro at single neuron and neural network level with infrared holographic optical tweezers to carry out force spectroscopy measurements. Simultaneous force spectroscopy, down to the sub-pico-Newton range, was performed during laser dissection to quantify the tension release in a partially ablated neurite. Therefore, we could control and measure the damage inflicted to an individual neuronal process. To characterize the effect of the inflicted injury on network level, changes in activity of neural subpopulations were monitored with subcellular resolution and overall network activity with high temporal resolution by concurrent calcium imaging and microelectrode array recording. Neuronal connections have been sequentially ablated and the correlated changes in network activity traced and mapped. With this unique combination of electrophysiological and optical tools, neural activity can be studied and quantified in response to controlled injury at the subcellular, cellular, and network level.
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