The neural networks consisting of the axonal tracts that interconnect the thalamus and the neocortex are central to information processing where the thalamus acts as the ?gateway' to the neocortex where almost all sensory information passes. Research on TC projection has been relying on electrical stimulation in brain slices, where it has been a big challenge to contain all the projections in a thin slab of tissue. Channelrhodopsin-2 (ChR2), a light-sensitive cation-channel derived from green algae, chlamydomonas reinhardtii, was used as a potential alternative for electrical stimulation. Lentivirus coding for ChR2 was intracranially injected into mice to render neurons photo-sensitive. With the ability to genetically target the expression of ChR2 to neurons using synapsin-I promoter and using lentiviral construct without retrograde infection, successful expression of ChR2 along the TC axonal tract was achieved. Taking advantage of ChR2's ability to spatiotemporally control the activity of neurons by the illumination of pulses of blue light (wavelength ~440nm), we have successfully activated the neurons both directly and synaptically. Synaptic activation allowed light activation of neurons that did not express ChR2 on the cell membrane. Furthermore, activation of brain-region remote from the injection site (thalamus) and close to the surface of the brain (cortex) was possible, taking advantage of the anatomical features of the projection neurons The results of synaptic activation of the axonal terminus in slice studies were extended to in vivo scenario using engineered single-unit optical electrode (SUOE) as the optical activation-electrical recording modality. Optically-evoked temporary-precise multi-unit activities were observed in the cortex upon light illumination of the TC axonal terminals. Kubelka-Munk model was used to calculate the minimum light intensity that was required to elicit response from the TC axonal terminals. Exploration of potential non-viral gene transfer of ChR2 was also attempted. Using silica nanoparticles, successful gene delivry of ChR2-coding DNA into cortical cells of live mouse brain was achieved. Light-evoked modulation of transfected neural cells was confirmed using standard electrophysiology and semi-quantitative comparison with viral infection was conducted.
Urabe, Hayato,
"Optical Elucidation of Neural Microcircuitry"
(2009).
Biomedical Engineering Theses and Dissertations.
Brown Digital Repository. Brown University Library.
https://doi.org/10.7301/Z0RR1WHB
