The influence of cellular bioelectric properties, such as the resting membrane potential (Vm), on the behaviour of non-excitable cells, such as neural progenitors (APs), is not well understood. Here, Vitali et al. show that as cortical development progresses, the Vm of APs in the ventricular zone becomes progressively more hyperpolarized and drives the generation of successive subclasses of neurons.

During corticogenesis, subclasses of excitatory neurons are generated sequentially and migrate to populate cortical layers from the inside out, with the peak of layer 4 (L4) neurons being born at embryonic day 14.5 (E14.5). To investigate whether activity manipulation affected neuron differentiation, the authors used in utero electroporation (IUE) on E14.5 in mice to overexpress the inward-rectifying K+ channel Kir2.1 in ventricular zone APs, and then looked at how the resulting hyperpolarization affected neuronal identity in primary sensory cortex postnatally. In contrast to controls, E14.5-born L4 neurons in Kir2.1 IUE mice were found in L4 as expected, but also L2/3, which would normally occur only for neurons born at E15.5. These precociously located neurons showed numerous characteristics of L2/3 neurons, including expressing L2/3 transcriptional programmes, displaying apical dendrites and symmetrical dendritic arbours, as well as receiving input from L2 neurons and sending callosal projections. Together, these findings suggest that Kir2.1 IUE results in a forward temporal shift in laminar, morphological, molecular and circuit-level characteristics. The authors then investigated whether this identity reassignment was the result of a pre- or post-mitotic effect of hyperpolarization. The temporally selective hyperpolarization of APs replicated the identity shift previously reported. Moreover, whole-cell patch clamp of apical progenitors in E12.5–15.5 cortical slices revealed that there was a sharp drop in the Vm over this period, similar in magnitude to that induced by Kir2.1 IUE. As corticogenesis progresses, a greater proportion of APs are reported to differentiate into neurons via intermediate progenitors (indirect neurogenesis). Differential labelling of APs, intermediate progenitors and new-born neurons showed that Kir2.1 IUE induced a precocious shift from direct to indirect neurogenesis.

The authors hypothesized that changes in the Vm might constitute an endogenous mechanism for regulating neuronal diversity in developing cortex. Using single-cell RNA sequencing, they found that Kir2.1 overexpression in APs at E14.5 repressed genes that are involved in WNT signalling. Temporally selective repression of WNT in APs replicated the effects of Kir2.1 upregulation on neuronal fate. Overall, these findings suggest that membrane hyperpolarization is permissive for specific molecular pathways, including WNT signalling, which in turn coordinate AP developmental programmes and neocortical neuron diversity.