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What is the role of the axonal connections between the hemispheres in sensory processing?

Periodic Reporting for period 1 - CALLAX (What is the role of the axonal connections between the hemispheres in sensory processing?)

Reporting period: 2018-03-01 to 2020-02-29

Why does our brain have two hemispheres and what is the role of connections between them? While it is well known that in humans, each hemisphere has functional specializations, we have little mechanistic understanding of how circuits communicate across the corpus callosum that connects the hemispheres. Moreover, the role of these callosal axons, and the information they convey is highly debated. There is a century-old debate among cognitive psychologists and neuroscientists: Do callosal axons have mainly an inhibitory action in the other hemisphere, producing a dominant and a repressed hemisphere; or do these projections faithfully report the activity of one hemisphere to the other, facilitating the cooperation between the two hemispheres?

In my proposal I aimed to elucidate if the callosal projections within cortical circuits are either inhibitory or excitatory; and what kind of information is conveyed by these callosal axons. Furthermore, I wanted to test if the lack of callosal information modifies the activity of neurons in the cortical circuit. I used two-photon microscopy and calcium indicators expressed in neurons to study the effects of callosal inputs arriving to a cortical area from the contralateral homotopic brain region. During the calcium imaging sessions mice were performing a behavioral task head-fixed, in which they had to estimate position based on tactile stimuli. Using this high spatial resolution technique, I recorded the activity of cortical pyramidal cells as well as callosal axons. Finally, I used optogenetics to inhibit callosal input specifically and measured how this influenced cortical circuits and behavior.
Conclusions of the action: Our results show that callosal axons are derived from excitatory neurons, and not just convey reliably information from one hemisphere to the other, but they also shape the activity of the neuron population in the homotopic brain area. Our study strengthens the hypothesis that homotopic brain regions collaborate with each other, instead of having one dominant and one repressed hemisphere.
The combination of state-of-the-art microscopy, novel viral methods and mouse behavior made this project very timely.

Recently, more and more human studies uncover evidence that altered corpus callosum morphology associated with autism spectrum disorder and schizophrenia. As both medical conditions develop during childhood or young adulthood, these become a lifelong burden for families, healthcare systems, and societies. Therefore, it is essential to explore the normal functioning of the involved brain structures, as well as to understand the pathogenesis and find new therapies. My findings provide for the first time a mechanistic understanding of the role of the connections between our hemispheres and it may provide a framework for understanding diseases that affect the corpus callosum.
Do callosal axons derive from inhibitory or excitatory neurons?
Our first goal was to examine if either excitatory or inhibitory neurons (or both types) send their axons to the other hemisphere. I labelled either excitatory or inhibitory neurons with a green fluorescent protein in one hemisphere by utilizing transgenic mouse lines and specific viruses. After anatomical processing, I searched for labelled axons in the other hemisphere in the homotopic cortical area. The results show that several excitatory cells project to the other hemisphere’s homotopic region, whereas inhibitory neurons do not send callosal axons.

What kind of information is being transferred between the hemispheres?
First, I imaged the activity of cell bodies in cortical L2/3 region during the behavioral task to characterize the regular cell activity during the task. On average, 20-30% of the cells had reliable task-specific responses: many cells responded to tactile stimuli or to the reward, and some cells got activated sequentially during a trial. Next, I compared the activity of callosal axons by recording calcium transients (GCaMP-expressing virus was injected to the contralateral hemisphere, where cell bodies were labelled, whereas I recorded in the other hemisphere, where only callosal axons expressed GCaMP). The data shows that callosal axons represent the activity of the cell bodies of the other hemisphere. Our results suggest that the hemispheres inform faithfully each other about what kind of activity is going on in their cortical circuits.

How does silencing callosal input affect circuit activity?
We applied pharmacological methods and optogenetics to investigate this question. First, we used pharmacological tools to silence the cortical region in one hemisphere, while we were recording the activity of pyramidal cells in the other hemisphere before and during the silencing. Second, we applied a red-shifted inhibitory opsin to selectively silence the callosal axons, meanwhile we recorded the activity of pyramidal cells within the same hemisphere. This way we could compare the activity of pyramidal cells in control condition and during silencing either the other hemisphere or the callosal axons coming from the other hemisphere’s homotopic region. The results show that ~20% of the pyramidal cells change their activity when the callosal axons are inhibited.

Exploitation and dissemination of results: Currently we are writing the manuscript, which planned to be submitted by this autumn in a high-impact, open access journal. The results have been presented and discussed several times at meetings at the University of Oslo, and on the Society for Neuroscience Conference (October, 2018), and on the Spring Hippocampal Research Conference (June 2019, Taormina, Italy). Additionally, the results will be presented this year at the Society for Neuroscience Conference (October, 2020).
I have been using state of the art techniques during my fellowship: high-resolution two-photon calcium imaging, novel transgenic mouse lines and viral constructs, recently developed behavioral paradigms, and optogenetics. We went beyond the state of the art by optimizing our two-photon system that we were able to simultaneously perform calcium imaging and optogenetic inhibition in the same brain area. This technical development might be an essential tool for other researchers as well to explore what kind of information is conveyed, either among neurons within a network or between brain areas, in unprecedented fine details. These future studies might lead to better understanding of how our brain works and may provide hints that can open cutting-edge therapeutic avenues to treat brain diseases.
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