Odorant receptors regulate the final glomerular coalescence of olfactory sensory neuron axons

Abstract

Odorant receptors (OR) are strongly implicated in coalescence of olfactory sensory neuron (OSN) axons and the formation of olfactory bulb (OB) glomeruli. However, when ORs are first expressed relative to basal cell division and OSN axon extension is unknown. We developed an in vivo fate-mapping strategy that enabled us to follow OSN maturation and axon extension beginning at basal cell division. In parallel, we mapped the molecular development of OSNs beginning at basal cell division, including the onset of OR expression. Our data show that ORs are first expressed around 4 d following basal cell division, 24 h after OSN axons have reached the OB. Over the next 6+ days the OSN axons navigate the OB nerve layer and ultimately coalesce in glomeruli. These data provide a previously unidentified perspective on the role of ORs in homophilic OSN axon adhesion and lead us to propose a new model dividing axon extension into two phases. Phase I is OR-independent and accounts for up to 50% of the time during which axons approach the OB and begin navigating the olfactory nerve layer. Phase II is OR-dependent and concludes as OSN axons coalesce in glomeruli.olfactory epithelium | axon guidance | tamoxifen | olfactory marker protein | Ascl1 I n the mouse olfactory system, olfactory sensory neurons (OSNs) extend their axons from the olfactory epithelium (OE) to the olfactory bulb (OB), where they converge to form glomeruli. Each OSN expresses only 1 of ∼2,400 candidate odorant receptor (OR) alleles. OSNs expressing the same OR can be widely dispersed in the OE, yet their axons converge in only two to three molecularly specific glomeruli of a possible 3,700 (1). It was first recognized almost 20 y ago that substituting an OR-coding region with that of a different OR resulted in the glomerular convergence of axons at an ectopic location relative to that of the native ORs (2). This led to the suggestion that ORs have an instructive role in the extension and glomerular coalescence of OSN axons, most likely mediated by homophilic fasciculation (3-5).Postnatal OSNs are derived from self-renewing precursors located proximal to the deep basal lamina of the OE (6). Following the division of globose basal stem cells, OSN neuroblasts transiently express Achaete-scute homolog 1 (Ascl1) followed by two phases of differentiation (6-8). Initially, they express growthassociated protein-43 (GAP-43), a marker of immature cells. Subsequently, the OSNs down-regulate GAP-43 and express olfactory marker protein (OMP), a universal marker of mature OSNs.Although there is a consensus on the involvement of ORs in OSN axon glomerular convergence, when ORs exert that influence following basal cell division or axon extension is not known. Moreover, the developmental progression of GAP-43 to OMP, as a measure of OSN differentiation and maturation, or of adenylate cyclase 3 (AC3), a downstream signaling molecule also implicated in axon extension, has not been considered in the context of OR expression or OSN dynamics. Here, we determined when ORs exert thei...

Alteration of Nrp1 signaling at different stages of olfactory neuron maturation promotes glomerular shifts along distinct axes in the olfactory bulb

“…3B). OR transcription can be observed prior to OMP expression (Hanchate et al, 2015;Rodriguez-Gil et al, 2015). To control for the stage at which the specific knock-in lines express the Cre recombinase, Gng8 cre ;R26 flRFP , M71 cre ;R26 flRFP and Omp cre ;R26 flRFP mice were analyzed, and the positions along the apicobasal axis of the septal sensory epithelium of the youngest red-fluorescent neurons were located (that is, those closest to the basal lamina with a one-neuronwide apicobasal sliding window; Fig.…”

“…Two levels of guidance have been identified. The first directs axons to their target zones in the bulb and the second guides local axon sorting, which ends in glomerular segregation Imai et al, 2006Imai et al, , 2009Nakashima et al, 2013;Nishizumi and Sakano, 2015;Rodriguez-Gil et al, 2015;Serizawa et al, 2006;Takeuchi et al, 2010;Wang et al, 1998). The current model explaining global targeting (at least on the medial side of the bulb) proposes the existence of guidance cues that determine the position of the target along two axes in the bulb: one dorsoventral and the other anteroposterior.…”

The Odorant Receptor-Dependent Role of Olfactory Marker Protein in Olfactory Receptor Neurons

“…The ciliary cAMP increase opens the olfactory cyclic nucleotide-gated (CNG) channel (Nakamura and Gold, 1987;Firestein et al, 1991) and influx of Na ϩ and Ca 2ϩ (Dzeja et al, 1999) causes ORN depolarization. Ca 2ϩ triggers the activation of the second olfactory transduction channel, an excitatory Ca 2ϩ -activated Cl Ϫ channel TMEM16B/ANO2 (Kleene, 1993;Lowe and Gold, 1993;Stephan et al, 2009;Hengl et al, 2010;Rasche et al, 2010;Sagheddu et al, 2010;Billig et al, 2011). Transduction leads to the generation of APs to convey odor information to the olfactory bulb (OB).…”

“…Transduction leads to the generation of APs to convey odor information to the olfactory bulb (OB). Mammalian ORNs fire APs at increasing frequency as the odor concentration increases, while the number of fired APs first increases up to intermediate odor concentrations and then decreases at higher concentrations (Gesteland and Sigwart, 1977;Reisert and Matthews, 2001;Rospars et al, 2008). For the odorant response to terminate, cAMP and Ca 2ϩ must return to basal levels for CNG and Cl Ϫ channels to close.…”

“…Electro-olfactograms from OMP knock-out (KO) mice have slower onset and decay kinetics compared with wild type (WT; Buiakova et al, 1996). Single-cell recordings from isolated OMP KO ORNs showed that ORNs' ability to fire APs was also altered (Reisert et al, 2007). OMP speeds up signal transduction by acting upstream of cAMP production, probably at the level of AC3.…”