Yuko Muroyama

In the mouse olfactory bulb, sensory information detected by ∼1,000 types of olfactory sensory neurons (OSNs) is represented by the glomerular map. The second-order neurons, mitral and tufted cells, connect a single primary dendrite to one glomerulus. This forms discrete connectivity between the ∼1,000 types of input and output neurons. It has remained unknown how this discrete dendrite wiring is established during development. We found that genetically silencing neuronal activity in mitral cells, but not from OSNs, perturbs the dendrite pruning of mitral cells. In vivo calcium imaging of awake neonatal animals revealed two types of spontaneous neuronal activity in mitral/tufted cells, but not in OSNs. Pharmacological and knockout experiments revealed a role for glutamate and NMDARs. The genetic blockade of neurotransmission among mitral/tufted cells reduced spontaneous activity and perturbed dendrite wiring. Thus, spontaneous network activity generated within the olfactory bulb self-organizes the parallel discrete connections in the mouse olfactory system.

Mitral cells (MCs) of the mammalian olfactory bulb have a single primary dendrite extending into a single glomerulus, where they receive odor information from olfactory sensory neurons (OSNs). Molecular mechanisms for controlling dendritic arbors of MCs, which dynamically change during development, are largely unknown. Here we found that MCs displayed more complex dendritic morphologies in mouse mutants of Maml1, a crucial gene in Notch signaling. Similar phenotypes were observed by conditionally misexpressing a dominant negative form of MAML1 (dnMAML1) in MCs after their migration. Conversely, conditional misexpression of a constitutively active form of Notch reduced their dendritic complexity. Furthermore, the intracellular domain of Notch1 (NICD1) was localized to nuclei of MCs. These findings suggest that Notch signaling at embryonic stages is involved in the dendritic complexity of MCs. After the embryonic misexpression of dnMAML1, many MCs aberrantly extended dendrites to more than one glomerulus at postnatal stages, suggesting that Notch signaling is essential for proper formation of olfactory circuits. Moreover, dendrites in cultured MCs were shortened by Jag1-expressing cells. Finally, blocking the activity of Notch ligands in OSNs led to an increase in dendritic complexity as well as a decrease in NICD1 signals in MCs. These results demonstrate that the dendritic complexity of MCs is controlled by their presynaptic partners, OSNs. Olfactory circuits are critical for the survival of many animals. Odor information is transmitted from olfactory sensory neurons (OSNs) to relay neurons, the morphology of which is crucial for processing of the information and similar among species. The major relay neurons, mitral cells (MCs) in mammals and projection neurons in flies, have a single primary dendrite at the mature stage. Molecular mechanisms to control the formation of the dendrite are largely unknown. MCs dynamically change their dendrites during development. In this study, we show that the dendritic morphologies of MCs are controlled by Notch signaling, many factors of which are well conserved among species. Moreover, we have found that Notch signaling in MCs is activated by OSNs, and that Notch operates in the relay neurons in the mouse olfactory system, in contrast to the fly system, where Notch functions in OSNs. Therefore, our study has revealed a novel step for shaping the dendritic morphologies of MCs.

The glomerular map in the olfactory bulb (OB) is the basis for odor recognition. Once established during development, the glomerular map is stably maintained throughout the life of an animal despite the continuous turnover of olfactory sensory neurons (OSNs). However, traumatic damage to OSN axons in the adult often leads to dysosmia, a qualitative and quantitative change in olfaction in humans. A mouse model of dysosmia has previously indicated that there is an altered glomerular map in the OB after the OSN axon injury; however, the underlying mechanisms that cause the map distortion remain unknown. In this study, we examined how the glomerular map is disturbed and how the odor information processing in the OB is affected in the dysosmia model mice. We found that the anterior–posterior coarse targeting of OSN axons is disrupted after OSN axon injury, while the local axon sorting mechanisms remained. We also found that the connectivity of mitral/tufted cell dendrites is reduced after injury, leading to attenuated odor responses in mitral/tufted cells. These results suggest that existing OSN axons are an essential scaffold for maintaining the integrity of the olfactory circuit, both OSN axons and mitral/tufted cell dendrites, in the adult.

In the developing neocortex, neural progenitor cells (NPCs) produce projection neurons of the six cortical layers in a temporal order. Over the course of cortical neurogenesis, maintenance of NPCs is essential for the generation of distinct types of neurons at the required time. Notch signaling plays a pivotal role in the maintenance of NPCs by inhibiting neuronal differentiation. Although Hairy and Enhancer-of-split (Hes)-type proteins are central to Notch signaling, it remains unclear whether other essential effectors take part in the pathway. In this study, we identify Nepro, a gene expressed in the developing mouse neocortex at early stages that encodes a 63 kDa protein that has no known structural motif except a nuclear localization signal. Misexpression of Nepro inhibits neuronal differentiation only in the early neocortex. Furthermore, knockdown of Nepro by siRNA causes precocious differentiation of neurons. Expression of Nepro is activated by the constitutively active form of Notch but not by Hes genes. Nepro represses expression of proneural genes without affecting the expression of Hes genes. Finally, we show that the combination of Nepro and Hes maintains NPCs even when Notch signaling is blocked. These results indicate that Nepro is involved in the maintenance of NPCs in the early neocortex downstream of Notch.

In the developing central nervous system, cellular diversity depends in part on organising signals that establish regionally restricted progenitor domains, each of which produces distinct types of differentiated neurons. However, the mechanisms of neuronal subtype specification within each progenitor domain remain poorly understood. The p2 progenitor domain in the ventral spinal cord gives rise to two interneuron (IN) subtypes, V2a and V2b,which integrate into local neuronal networks that control motor activity and locomotion. Foxn4, a forkhead transcription factor, is expressed in the common progenitors of V2a and V2b INs and is required directly for V2b but not for V2a development. We show here in experiments conducted using mouse and chick that Foxn4 induces expression of delta-like 4 (Dll4) and Mash1 (Ascl1). Dll4 then signals through Notch1 to subdivide the p2 progenitor pool. Foxn4, Mash1 and activated Notch1 trigger the genetic cascade leading to V2b INs, whereas the complementary set of progenitors, without active Notch1, generates V2a INs. Thus, Foxn4 plays a dual role in V2 IN development: (1) by initiating Notch-Delta signalling, it introduces the asymmetry required for development of V2a and V2b INs from their common progenitors; (2) it simultaneously activates the V2b genetic programme.

Astrocytes are the most abundant and functionally diverse glial population in the vertebrate central nervous system (CNS)¹. However, the mechanisms underlying astrocyte specification are poorly understood. It is well established that cellular diversification of neurons in the embryo is generated by position-dependent extrinsic signals and combinatorial interactions of transcription factors that direct specific cell fates by suppressing alternative fates². It is unknown whether a comparable process determines embryonic astrocyte identity. Indeed, astrocyte development is generally thought to take place in a position-independent manner^3,4. Here we show multiple functions of Stem cell leukaemia (Scl, also known as Tal1), which encodes a basic helix–loop–helix (bHLH) transcription factor, in the regulation of both astrocyte versus oligodendrocyte cell fate acquisition and V2b versus V2a interneuron cell fate acquisition in the p2 domain of the developing vertebrate spinal cord. Our findings demonstrate a regionally restricted transcriptional programme necessary for astrocyte and V2b interneuron development, with striking parallels to the involvement of SCL in haematopoiesis. They further indicate that acquisition of embryonic glial subtype identity might be regulated by genetic interactions between SCL and the transcription factor Olig2 in the ventral neural tube.