Antisense oligonucleotide therapeutic approach for Timothy syndrome.

Timothy syndrome (TS) is a severe, multisystem disorder characterized by autism, epilepsy, long-QT syndrome and other neuropsychiatric conditions. TS type 1 (TS1) is caused by a gain-of-function variant in the alternatively spliced and developmentally enriched CACNA1C exon 8A, as opposed to its counterpart exon 8. We previously uncovered several phenotypes in neurons derived from patients with TS1, including delayed channel inactivation, prolonged depolarization-induced calcium rise, impaired interneuron migration, activity-dependent dendrite retraction and an unanticipated persistent expression of exon 8A.

Maturation and circuit integration of transplanted human cortical organoids.

Self-organizing neural organoids represent a promising in vitro platform with which to model human development and disease. However, organoids lack the connectivity that exists in vivo, which limits maturation and makes integration with other circuits that control behaviour impossible. Here we show that human stem cell-derived cortical organoids transplanted into the somatosensory cortex of newborn athymic rats develop mature cell types that integrate into sensory and motivation-related circuits.

Engineering brain assembloids to interrogate human neural circuits.

The development of neural circuits involves wiring of neurons locally following their generation and migration, as well as establishing long-distance connections between brain regions. Studying these developmental processes in the human nervous system remains difficult because of limited access to tissue that can be maintained as functional over time in vitro. We have previously developed a method to convert human pluripotent stem cells into brain region-specific organoids that can be fused and integrated to form assembloids and study neuronal migration.

Dissecting the molecular basis of human interneuron migration in forebrain assembloids from Timothy syndrome.

Defects in interneuron migration can disrupt the assembly of cortical circuits and lead to neuropsychiatric disease. Using forebrain assembloids derived by integration of cortical and ventral forebrain organoids, we have previously discovered a cortical interneuron migration defect in Timothy syndrome (TS), a severe neurodevelopmental disease caused by a mutation in the L-type calcium channel (LTCC) Ca1.2.

Generation of human striatal organoids and cortico-striatal assembloids from human pluripotent stem cells.

Cortico-striatal projections are critical components of forebrain circuitry that regulate motivated behaviors. To enable the study of the human cortico-striatal pathway and how its dysfunction leads to neuropsychiatric disease, we developed a method to convert human pluripotent stem cells into region-specific brain organoids that resemble the developing human striatum and include electrically active medium spiny neurons. We then assembled these organoids with cerebral cortical organoids in three-dimensional cultures to form cortico-striatal assembloids.

Neuronal defects in a human cellular model of 22q11.2 deletion syndrome.

22q11.2 deletion syndrome (22q11DS) is a highly penetrant and common genetic cause of neuropsychiatric disease. Here we generated induced pluripotent stem cells from 15 individuals with 22q11DS and 15 control individuals and differentiated them into three-dimensional (3D) cerebral cortical organoids.

Retinoid X Receptor α Regulates DHA-Dependent Spinogenesis and Functional Synapse Formation In Vivo.

Coordinated intracellular and extracellular signaling is critical to synapse development and functional neural circuit wiring. Here, we report that unesterified docosahexaenoic acid (DHA) regulates functional synapse formation in vivo via retinoid X receptor α (Rxra) signaling. Using Rxra conditional knockout (cKO) mice and virus-mediated transient gene expression, we show that endogenous Rxra plays important roles in regulating spinogenesis and excitatory synaptic transmission in cortical pyramidal neurons.

A Critical Role of Presynaptic Cadherin/Catenin/p140Cap Complexes in Stabilizing Spines and Functional Synapses in the Neocortex.

The formation of functional synapses requires coordinated assembly of presynaptic transmitter release machinery and postsynaptic trafficking of functional receptors and scaffolds. Here, we demonstrate a critical role of presynaptic cadherin/catenin cell adhesion complexes in stabilizing functional synapses and spines in the developing neocortex. Importantly, presynaptic expression of stabilized β-catenin in either layer (L) 4 excitatory neurons or L2/3 pyramidal neurons significantly upregulated excitatory synaptic transmission and dendritic spine density in L2/3 pyramidal neurons, while its sparse postsynaptic expression in L2/3 neurons had no such effects.

CRISPR-Cas9-mediated genome editing in one blastomere of two-cell embryos reveals a novel Tet3 function in regulating neocortical development.

Studying the early function of essential genes is an important and challenging problem in developmental biology. Here, we established a method for rapidly inducing CRISPR-Cas9-mediated mutations in one blastomere of two-cell stage embryos, termed 2-cell embryo-CRISPR-Cas9 injection (2CC), to study the in vivo function of essential (or unknown) genes in founder chimeric mice. By injecting both Cre mRNA and CRISPR-Cas9 targeting the gene of interest into fluorescent reporter mice, the 2CC method can trace both wild-type and mutant cells at different developmental stages, offering internal control for phenotypic analyses of mutant cells.

Postsynaptic spiking homeostatically induces cell-autonomous regulation of inhibitory inputs via retrograde signaling.

Developing neural circuits face the dual challenge of growing in an activity-induced fashion and maintaining stability through homeostatic mechanisms. Compared to our understanding of homeostatic regulation of excitatory synapses, relatively little is known about the mechanism mediating homeostatic plasticity of inhibitory synapses, especially that following activity elevation. Here, we found that elevating neuronal activity in cultured hippocampal neurons for 4 h significantly increased the frequency and amplitude of mIPSCs, before detectable change at excitatory synapses.