Integration of thalamocortical and callosal inputs by optogenetic activation of the rat corpus callosum (CC) with MRI-guided robotic arm (MgRA) (#2)
Yi Chen1, Filip Sobczak1, Patricia Pais1, Cornelius Schwarz2, Alan P. Koretsky3, Xin Yu1, 4
1 Max Planck Institute for Biological Cybernetics, Tübingen, Baden-Württemberg, Germany
The hypothesis that CC inhibits contralateral cortex can explain dampened neural responses in cortex in human and rodents(1-4), e.g., the first stimulus suppressed neural responses to the subsequent stimulus on the other eye within a certain time. Here, we optogenetically activated CC(8) and provided direct evidence for CC-mediated interhemispheric inhibition(II), showing that the direct callosal inputs suppressed evoked calcium and BOLD signals in barrel cortex(BC) by whisker stimulation. Our work links callosal circuit-specific regulation to the global brain dynamic changes based on II(5-7).
AAV.CaMKII.ChR2.mCherry was injected into the BC of rats, expressed in callosal projection neurons (CPN) and along their axonal fiber bundles projecting to the opposite BC (Fig.1a), where the GCaMP6f was expressed (Fig.1f). Optogenetic stimulation will be delivered on corpus callosum, followed by a whisker stimulus to the whisker pad with different intervals (0-200 ms) and the paired conditions of each trail were randomized (Fig. 2c). Whole brain BOLD signals were acquired with simultaneous calcium signal in the BC while an MRI-guided robotic arm was used to precisely target the callosal fiber bundle to deliver blue light pulses (473nm) at 2Hz, 10ms width for the fMRI block design (8s on/52s off,13 epochs,Fig.2b,c). Whole brain 3D EPI: TR,1.5s, 400×400×400 μm3 spatial resolution.
Upon the optogenetic stim on CC, salient BOLD signal was detected due to the antidromic activity from the axonal fibers backward to the soma of callosal projection neurons in the ipsilateral BC (Fig.1c,d), further confirmed by LFP (Fig.1e). For the orthodromic activity, there was clear spike for each stimulus at 2Hz, while with higher frequencies, light flashes 2-16 induced responses were consistently weaker than the first response (Fig.1g), Moreover, there was a baseline drift during the whole 40Hz stimulation period (Fig.1g), therefore, confirming the CC-mediated interhemispheric inhibition. With two stimuli paradigm, the anti-dromic activity in the right cortex kept similar for 6 conditions, while the BOLD and calcium signals in the left cortex induced by paired whisker stimuli was the strongest for OW condition, kept suppressed for the O50W and O100W conditions (Fig.2d-h), almost recovered for the O200W condition.
By taking advantage of fMRI, the optogenetic stimuli on CC and cell-specific calcium signal recordings for layer 5 pyramidal excitatory neurons in the BC, we confirmed the CC-mediated interhemispheric inhibition, further provided direct evidence for the dampened neural responses to subsequent contralateral stimulus after an ipsilateral stimulus for a period of several hundred milliseconds in human and rodents at function level.
We thank Mr. Shanyi Yu for building up the first prototype of the robotic arm, Fine Mechanic and Electronic Workshop at MPI for Biological Cybernetics for MgRA system automation. The financial support of the Max-Planck-Society and the China Scholarship Council (Ph.D. fellowship to Y. Chen) are gratefully acknowledged.
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Anti-dromic and orthodromic activation by corpus callosum optogenetic stimulation.
a Schematic of experimental design and CaMKII.mCherry expression.
b Overview of the MgRA for optical fiber insertion inside 14.1T scanner.
c Averaged fMRI map of brain-wide activity upon optogenetic stimulation on CC from 8 rats.
D Average time courses of BOLD in right BC(n = 8) upon light stimulation. Error bars represent mean±SD.
e The representative local field potential for antidromic activation.
f Schematic of experimental design and CaMKII.mCherry expressed in the right BC while GCaMP6f in the left BC.
g Representative calcium signal changes for 8 s of the orthodromic activation responses.
Simultaneous BOLD and calcium signals upon CC opto stim and whisker stim with varying intervals
a Stimulation scheme. 6 conditions: W, O, O50W, O100W, O200W.
b Experimental setup.
c Typical calcium signals for condition W(blue dotted box) and O100W(red dotted box).
d fMRI map of brain-wide activity for 6 conditions(n=6).
e Averaged normalized calcium signal in left BC.
f Normalized calcium signal for individual rat.
g Averaged BOLD in the left BC (left) evoked by whisker stimulation and right BC (right) evoked by CC stimulation.
h Averaged normalized calcium signal changes across 6 rats for different conditions.
i Averaged normalized calcium signal changes across 4 rats for different conditions.
Keywords: Corpus Callosum, Interhemispheric Inhibition, Thalamocortical Pathway