In the olfactory epithelium (OE), generation of new neurons by neuronal progenitors is inhibited by a signal from neurons themselves. Here we provide evidence that this feedback inhibitory signal is growth and differentiation factor 11 (GDF11). Both GDF11 and its receptors are expressed by OE neurons and progenitors, and GDF11 inhibits OE neurogenesis in vitro by inducing p27(Kip1) and reversible cell cycle arrest in progenitors. Mice lacking functional GDF11 have more progenitors and neurons in the OE, whereas mice lacking follistatin, a GDF11 antagonist, show dramatically decreased neurogenesis. This negative autoregulatory action of GDF11 is strikingly like that of its homolog, GDF8/myostatin, in skeletal muscle, suggesting that similar strategies establish and maintain proper cell number during neural and muscular development.

MASH1, a basic helix-loop-helix transcription factor, is widely expressed by neuronal progenitors in the CNS and PNS, suggesting that it plays a role in the development of many neural regions. However, in mice lacking a functional Mash1 gene, major alterations have been reported in only a few neuronal populations; among these is a generalized loss of olfactory receptor neurons of the olfactory epithelium. Here, we use a transgenic reporter mouse line, in which the cell bodies and growing axons of subsets of central and peripheral neurons are marked by expression of a tau-lacZ reporter gene (the Tattler-4 allele), to look both more broadly and deeply at defects in the nervous system of Mash1-/- mice. In addition to the expected lack of olfactory receptor neurons in the main olfactory epithelium, developing Mash1-/-;Tattler-4+/- mice exhibited reductions in neuronal cell number in the vomeronasal organ and in the olfactory bulb; the morphology of the rostral migratory stream, which gives rise to olfactory bulb interneurons, was also abnormal. Further examination of cell proliferation, cell death, and cell type-specific markers in Mash1-/- animals uncovered parallels between the main olfactory epithelium and the vomeronasal organ in the regulation of sensory neuron development. Interestingly, this analysis also revealed that, in the olfactory epithelium of Mash1-/- animals, there is an overproduction of proliferating cells that co-express markers of both neuronal progenitors and supporting cells. This finding suggests that olfactory receptor neurons and olfactory epithelium supporting cells may share a common progenitor, and that expression of Mash1 may be an important factor in determining whether these progenitors ultimately generate neurons or glia.

The olfactory epithelium of the mouse has many properties that make it an ideal system for studying the molecular regulation of neurogenesis. We have used a combination of in vitro and in vivo approaches to identify three distinct stages of neuronal progenitors in the olfactory receptor neuron lineage. The neuronal stem cell, which is ultimately responsible for continual neuron renewal in this system, gives rise to a transit amplifying progenitor identified by its expression of a transcription factor, MASH1. The MASH1-expressing progenitor gives rise to a second transit amplifying progenitor, the Immediate Neuronal Precursor, which is distinct from the stem cell and MASH1-expressing progenitor, and gives rise quantitatively to olfactory receptor neurons. Regulation of progenitor cell proliferation and differentiation occurs at each of these three cell stages, and growth factors of the fibroblast growth factor (FGF) and bone morphogenetic protein (BMP) families appear to play particularly important roles in these processes. Analyses of the actions of FGFs and BMPs reveal that negative signaling plays at least as important a role as positive signaling in the regulation of olfactory neurogenesis.

To understand how signaling molecules regulate the generation of neurons from proliferating stem cells and neuronal progenitors in the developing and regenerating nervous system, we have studied neurogenesis in a model neurogenic epithelium, the olfactory epithelium (OE) of the mouse. Our studies have employed a candidate approach to test signaling molecules of potential importance in regulating neurogenesis and have utilized methods that include tissue culture, in situ hybridization and mouse genetics. Using these approaches, we have identified three distinct stages of stem and transit amplifying progenitor cells in the differentiation pathway of olfactory receptor neurons (ORNs) and have identified mechanisms by which the development of each of these progenitor cell types is regulated by signals produced both within the OE itself and by its underlying stroma. Our results indicate that regulation of olfactory neurogenesis is critically dependent on multiple signaling molecules from two different polypeptide growth factor superfamilies, the fibroblast growth factors and the transforming growth factor β (TGF-β) group. In addition, they indicate that these signaling molecules interact in at least two important ways: first, opposing signals converge on cells at specific developmental stages in the ORN pathway to regulate proliferation and differentiation; and second, these signaling molecules – particularly the TGF-βs and their antagonists – play key roles in feedback loops that regulate the size of progenitor cell pools and thereby neuron number, during development and regeneration.