Summary: Researchers discovered a multi-layer circuit that forms in the cortex at an unexpectedly early stage of brain development. Genetically manipulating this circuit results in changes similar to those found in the brains of people with autism.
Using a new approach for studying live embryonic mouse brains at single-cell resolution, researchers have identified an active multi-layer circuit that forms in the cortex during an unexpectedly early stage of development.
Perturbing the circuit genetically led to changes similar to those seen in the brains of people with autism. The findings are reported today in Cell by a team based at the Institute of Molecular and Clinical Ophthalmology Basel.
“Understanding the detailed development of cell types and circuits in the cortex can provide important insights into autism and other neurodevelopmental diseases,” says Botond Roska, Director at IOB and the paper’s corresponding author. “This is what our findings confirm.”
Autism has long been associated with faulty circuits in the cortex, which is the part of the brain that governs sensory perception, cognition, and other high-order functions. Most of the cortex is composed of excitatory cells called pyramidal neurons.
The IOB team wanted to study when and how these neurons assemble into the first active circuits in the cortex, but that posed a difficult challenge. Pyramidal neurons measure only a tenth of the width of a human hair, and any movement during experimental procedures might lead to inaccurate recordings of activity.
To keep the neurons stable for research, the team devised a surgical solution: Embryos were secured inside of agar-filled 3D holding devices within the mother’s abdominal cavity, so that normal embryonic blood flow and temperature could be maintained.
The prevailing view is that the cortex develops in an “inside out fashion”, with the deepest of its six layers appearing first. Seen this way, pyramidal neurons were thought to slowly become active as they migrate to their final locations in the cortex and form connections with each other.
But during the research, “we actually detected a very different activity pattern,” says Arjun Bharioke, a systems neuroscientist in IOB’s Central Visual Circuits Group, and one of the paper’s two lead authors.
Focusing specifically on pyramidal neurons that develop into layer 5 of the cortex, the team discovered a very early transient circuit that was already highly active and correlated even before the six-layer cortex had formed. This indicates that the neurons were already connected prior to their migration to form layer 5.
The transient circuit initially had 2 layers: a deep layer and a superficial layer. Later, the superficial layer became silent and vanished, while the classical layer-by-layer cortical development resumed, with a third intermediate layer forming layer 5.
“We also wanted to understand how this circuit changes in an autism model,” says Martin Munz, an IOB developmental biologist in the Central Visual Circuits Group and the paper’s other lead author.
Working with knock-out mouse lines missing one or both alleles of two autism-associated genes–Chd8 and Grin2b–the team made a key finding. The absence of these genes is known to cause significant autism in children. And in homozygous and heterozygous knockout mice, the superficial layer remained active as a developmental remnant.
“Throughout embryonic development, it never disappeared,” Munz says. Moreover, the knockout mouse brains contained patchy areas of cortical disorganization similar to those seen in people with autism.
The findings suggest that the spatial organization of pyramidal neurons is regulated by the newly-found circuit, and that “changes to embryonic circuits play a role in dysfunctions associated with neurodevelopmental disorders, including autism spectrum disorder,” Bharioke says.
In future research, IOB researchers will “carefully look at the superficial and deep layers of this early circuitry and independently manipulate them,” Roska says. “This will be instructive for learning about the etiology of neurodevelopmental diseases.”
About this neurodevelopment research news
Author: Clara Vuille-dit-Bille
Contact: Clara Vuille-dit-Bille – IOB
Image: The image is credited to Institute of Molecular and Clinical Ophthalmology Basel (IOB)
Original Research: Closed access.
“Pyramidal neurons form active, transient, multilayered circuits perturbed by autism-associated mutations at the inception of neocortex” by Botond Roska et al. Cell
Pyramidal neurons form active, transient, multilayered circuits perturbed by autism-associated mutations at the inception of neocortex
- Mouse embryonic pyramidal neurons display two phases of circuit assembly in vivo
- Pyramidal neurons first form a multi-layered circuit before cortical lamination begins
- This circuit is transiently active with functional synapses and active conductances
- Perturbing autism-associated genes interferes with the switch between the two phases
Cortical circuits are composed predominantly of pyramidal-to-pyramidal neuron connections, yet their assembly during embryonic development is not well understood.
We show that mouse embryonic Rbp4-Cre cortical neurons, transcriptomically closest to layer 5 pyramidal neurons, display two phases of circuit assembly in vivo. At E14.5, they form a multi-layered circuit motif, composed of only embryonic near-projecting-type neurons. By E17.5, this transitions to a second motif involving all three embryonic types, analogous to the three adult layer 5 types. I
n vivo patch clamp recordings and two-photon calcium imaging of embryonic Rbp4-Cre neurons reveal active somas and neurites, tetrodotoxin-sensitive voltage-gated conductances, and functional glutamatergic synapses, from E14.5 onwards. Embryonic Rbp4-Cre neurons strongly express autism-associated genes and perturbing these genes interferes with the switch between the two motifs.
Hence, pyramidal neurons form active, transient, multi-layered pyramidal-to-pyramidal circuits at the inception of neocortex, and studying these circuits could yield insights into the etiology of autism.