During corticogenesis, distinct subtypes of neurons are sequentially born from ventricular zone progenitors. How these cells are molecularly temporally patterned is poorly understood. We used single-cell RNA sequencing at high temporal resolution to trace the lineage of the molecular identities of successive generations of apical progenitors (APs) and their daughter neurons in mouse embryos. We identified a core set of evolutionarily conserved, temporally patterned genes that drive APs from internally driven to more exteroceptive states. We found that the Polycomb repressor complex 2 (PRC2) epigenetically regulates AP temporal progression. Embryonic age–dependent AP molecular states are transmitted to their progeny as successive ground states, onto which essentially conserved early postmitotic differentiation programs are applied, and are complemented by later-occurring environment-dependent signals. Thus, epigenetically regulated temporal molecular birthmarks present in progenitors act in their postmitotic progeny to seed adult neuronal diversity.

The molecular mechanisms that control how progenitors generate distinct subtypes of neurons, and how undifferentiated neurons acquire their specific identity during corticogenesis, are increasingly understood. However, whether postmitotic neurons can change their identity at late stages of differentiation remains unknown. To study this question, we developed an electrochemical in vivo gene delivery method to rapidly manipulate gene expression specifically in postmitotic neurons. Using this approach, we found that the molecular identity, morphology, physiology and functional input-output connectivity of layer 4 mouse spiny neurons could be specifically reprogrammed during the first postnatal week by ectopic expression of the layer 5B output neuron-specific transcription factor Fezf2. These findings reveal a high degree of plasticity in the identity of postmitotic neocortical neurons and provide a proof of principle for postnatal re-engineering of specific neural microcircuits in vivo.

During neurogenesis, mitotic progenitor cells lining the ventricles of the embryonic mouse brain undergo their final rounds of cell division, giving rise to a wide spectrum of postmitotic neurons and glia1,2. The link between developmental lineage and cell-type diversity remains an open question. Here we used massively parallel tagging of progenitors to track clonal relationships and transcriptomic signatures during mouse forebrain development. We quantified clonal divergence and convergence across all major cell classes postnatally, and found diverse types of GABAergic neuron that share a common lineage. Divergence of GABAergic clones occurred during embryogenesis upon cell-cycle exit, suggesting that differentiation into subtypes is initiated as a lineage-dependent process at the progenitor cell level.

During development, thalamocortical (TC) input has a critical role in the spatial delineation and patterning of cortical areas, yet the underlying cellular and molecular mechanisms that drive cortical neuron differentiation are poorly understood. In the primary (S1) and secondary (S2) somatosensory cortex, layer 4 (L4) neurons receive mutually exclusive input originating from two thalamic nuclei: the ventrobasalis (VB), which conveys tactile input, and the posterior nucleus (Po), which conveys modulatory and nociceptive input. Recently, we have shown that L4 neuron identity is not fully committed postnatally, implying a capacity for TC input to influence differentiation during cortical circuit assembly. Here we investigate whether the cell-type-specific molecular and functional identity of L4 neurons is instructed by the origin of their TC input. Genetic ablation of the VB at birth resulted in an anatomical and functional rewiring of Po projections onto L4 neurons in S1. This induced acquisition of Po input led to a respecification of postsynaptic L4 neurons, which developed functional molecular features of Po-target neurons while repressing VB-target traits. Respecified L4 neurons were able to respond both to touch and to noxious stimuli, in sharp contrast to the normal segregation of these sensory modalities in distinct cortical circuits. These findings reveal a behaviourally relevant TC-input-type-specific control over the molecular and functional differentiation of postsynaptic L4 neurons and cognate intracortical circuits, which instructs the development of modality-specific neuronal and circuit properties during corticogenesis.

During corticogenesis, ventricular zone progenitors sequentially generate distinct subtypes of neurons, accounting for the diversity of neocortical cells and the circuits they form. While activity-dependent processes are critical for the differentiation and circuit assembly of postmitotic neurons, how bioelectrical processes affect nonexcitable cells, such as progenitors, remains largely unknown. Here, we reveal that, in the developing mouse neocortex, ventricular zone progenitors become more hyperpolarized as they generate successive subtypes of neurons. Experimental in vivo hyperpolarization shifted the transcriptional programs and division modes of these progenitors to a later developmental status, with precocious generation of intermediate progenitors and a forward shift in the laminar, molecular, morphological, and circuit features of their neuronal progeny. These effects occurred through inhibition of the Wnt-beta-catenin signaling pathway by hyperpolarization. Thus, during corticogenesis, bioelectric membrane properties are permissive for specific molecular pathways to coordinate the temporal progression of progenitor developmental programs and thus neocortical neuron diversity.