B-50/GAP43 Gene Expression in the Rat Olfactory System During Postnatal Development and Aging

The olfactory neuroepithelium exhibits neurogenesis throughout adult life, and in response to lesions, a phenomenon that distinguishes this neural tissue from the rest of the mammalian brain. The newly formed primary olfactory neurons elaborate axons into the olfactory bulb. Thus, denervation and subsequent re-innervation of olfactory bulb neurons may occur throughout life. In this study the authors demonstrate the distribution of the growth-associated phosphoprotein B-50/GAP43 and its mRNA in the olfactory neuroepithelium and olfactory bulb during development and aging. In neonatal rats B-50/GAP43 mRNA was expressed in primary olfactory neurons throughout the olfactory epithelium and in their target neurons in the olfactory bulb, the mitral, juxtaglomerular and tufted cells. In contrast, in adult (7.5 weeks) and aging animals (6 - 18 months of age) B-50/GAP43 mRNA expression was progressively restricted to neurons in the basal region of the neuroepithelium and to some of their target mitral and juxtaglomerular cells in the olfactory bulb. The continuing expression of B-50/GAP43 mRNA in mitral- and juxtaglomerular cells in mature animals is thought to be related to their capacity to respond to continuously changing input from the primary olfactory neurons present in the olfactory neuroepithelium.     

Expression of calmodulin mRNA in rat olfactory neuroepithelium.

A calmodulin (CaM) cDNA was isolated by differential hybridization screening of a lambda gt10 library prepared from rat olfactory mucosa. This cDNA fragment, containing most of the open reading frame of the rat CaMI gene, was subcloned and used to characterize steady-state expression of CaM mRNA in rat olfactory neuroepithelium and bulb. Within the bulb mitral cells are the primary neuronal population expressing CaM mRNA. The major CaM mRNA expressed in the olfactory mucosa is 1.7 kb with smaller contributions from mRNAs of 4.0 and 1.4 kb. CaM mRNA was primarily associated with the olfactory neurons and, despite the cellular complexity of the tissue and the known involvement of CaM in diverse cellular processes, was only minimally evident in sustentacular cells, gland cells or respiratory epithelium. Following bulbectomy CaM mRNA declines in the olfactory neuroepithelium as does olfactory marker protein (OMP) mRNA. In contrast to the latter, CaM mRNA makes a partial recovery by one month after surgery. These results, coupled with those from in situ hybridization, indicate that CaM mRNA is expressed in both mature and immature olfactory neurons. The program regulating CaM gene expression in olfactory neurons is distinct from those controlling expression of B50/GAP43 in immature, or OMP in mature, neurons respectively.     

Carnosine in primary afferents of the olfactory system: an autoradiographic and biochemical study

Previous in vivo studies have shown that beta-alanine is incorporated specifically into the dipeptide L-carnosine (beta-alanyl-L-histidine). In the present study, we administered beta-[3H]alanine to the nasal cavity of hamsters and used biochemical analyses to identify the radioactively labeled compounds in the olfactory epithelium and olfactory bulb and autoradiography to demonstrate the localization and transport of the label in the primary afferents of the olfactory system. The olfactory epithelium and lamina propria were labeled intensely 6 hr after intranasal beta-[3H]alanine administration. At this survival time, 61% of the radioactivity in the olfactory epithelium was present in the carnosine fraction, while 37% of the label remained in the beta-alanine fraction. After 24-hr and 4-day survival periods, greater than 82% of the radioactivity was present in the carnosine fraction, and the olfactory receptors and bundles of axons were labeled preferentially. The olfactory nerve and glomerular layers of the main olfactory bulb were labeled intensely at 6 and 24 hr after beta-[3H]alanine administration; much less label was present in these layers at 4 days survival. At all three of these survival times, greater than 84% of the radioactivity in the olfactory bulb was present in the carnosine fraction. No label was present in the olfactory epithelium or bulb 18 days after beta-[3H]alanine administration. While the autoradiographic labeling over the structures of the accessory olfactory system was consistently less intense than that over the main olfactory system structures, the patterns of labeling were similar over the four survival times. Intranasal alpha-[3H]alanine administration resulted in some labeling in the primary afferent fibers, but the labeling did not have the specificity nor the same time course over the four survival times that was observed after beta-[3H]alanine administration. The results are consistent with the hypothesis that carnosine is a neurotransmitter or neuromodulator in the olfactory neurons. The results also suggest that carnosine may play a similar role in the vomeronasal neurons.     

Alteration of carnosine expression in olfactory system of mouse after unilateral naris closure and partial bulbectomy

To investigate the function of the dipeptide carnosine in the olfactory system, alterations of carnosine in the primary olfactory nerve after unilateral naris closure or olfactory bulb semilesion were studied by means of immunocytochemistry. After unilateral naris closure, carnosine staining showed no alteration, but immunoreactivity of tyrosine hydroxylase (TH), the rate-limiting enzyme for biosynthesis of dopamine in dopaminergic neurons, decreased dramatically in periglomerular neurons. This behavior of carnosine is consistent with that reported for the olfactory marker protein (OMP). On the other hand, following partial lesion of the olfactory bulb, ordinary to strong carnosine expression was seen in newly innervated glomeruli that exceeded that seen in the conventional targets. Ectopic, TH-positive, periglomerular cell-like neurons appeared around the newly formed glomeruli. These data suggest that carnosine may have some role in regulating TH expression in post-synaptic neurons. © 1993 Wiley-Liss, Inc.     

Neuroplasticity in the olfactory system: differential effects of central and peripheral lesions of the primary olfactory pathway on the expression of B-50/GAP43 and the olfactory marker protein

The regeneration of the olfactory neuroepithelium following olfactory bulbectomy or peripheral deafferentation was studied with mRNA probes and antibodies for B-50/GAP43 and for olfactory marker protein (OMP). Two stages in the regeneration of the olfactory epithelium could be discerned with these reagents. The first stage occurs following either peripheral deafferentation of the olfactory epithelium with Triton X-100 (TX-100) or after bulbectomy and is characterized by the formation of a large population of immature olfactory receptor neurons. These newly formed neurons express B-50/GAP43, a phosphoprotein related to neuronal growth and plasticity. During the second stage of the regeneration process the newly formed olfactory neurons mature, as evidenced by a decrease in their expression of B-50/GAP43 and an increase in the expression of OMP. This stage is only manifested if the developing neurons have access to the target olfactory bulb. Formation of a full complement of OMP-expressing neurons occurs only after peripheral lesion with TX-100. In contrast, following bulbectomy the reconstituted olfactory epithelium lacks its normal target and is compromised in its ability to recover from nerve damage, as evidenced by the presence of a large number of B-50/GAP43-expressing neurons up to 3 months after the lesion and its failure to establish a full complement of OMP-expressing neurons. These results demonstrate that the olfactory epithelium is capable of replacing its sensory neurons independently of the presence of its target, the olfactory bulb. However, the differential patterns of expression of B-50/GAP43 and OMP at long times after peripheral lesion with TX-100 or bulbectomy illustrate the profound effect the olfactory bulb has on neuronal maturation in reconstituted olfactory neuroepithelium.     

Neurogenesis of sensory neurons in the primate olfactory system after section of the fila olfactoria

Axotomy of the olfactory sensory neurons in the adult primate squirrel monkey induces retrograde degeneration of the perikarya in the nasal neuroepithelium. The process of neuronal degeneration is rapid and by the 10th day the olfactory neuroepithelium is deprived of all mature neurons. Basal cells, supporting cells and the Bowman's glands are unaffected by the surgical procedure. The degeneration of the neurons is followed by intense mitotic activity of the basal cells of the neuroepithelium. At 30 days survival several young, mature neurons are present again in the neuroepithelium. At 60–90 survival days the neuroepithelium reacquires a population of neurons similar to controls. The persistence of neurogenesis and the replacement of experimentally degenerated neurons in an adult, non-human primate is briefly discussed.     

Experimental studies on the olfactory marker protein. III. The olfactory marker protein in the olfactory neuroepithelium lacking connections with the forebrain

Total unilateral bulbectomy induces degeneration of the mature olfactory neurons and disappearance of the olfactory marker protein from the primary sensory pathway. Owing to the presence of a neurogenetic matrix in the neuroepithelium, reconstitution of a new population of neuronal elements occurs. In this experiment, connections of the regrown olfactory axons with the spared forebrain are barred by the formation of scar tissue. In spite of the absence of a target, new neurons differentiate and produce olfactory marker protein.     

Sensory reinnervation after partial removal of the olfactory bulb.

In this study, in order to provide the anatomical basis for future behavioral and electrophysiological experiments, we describe the effects of unilateral bulbar lesion on the peripheral sensory neurons and the parameters of reinnervation of the damaged olfactory bulb. Neonatal mice and rats were subjected to removal of portions of the olfactory bulb. At survival times from 2 to 6 months, the animals were killed by transcardial perfusion and processed for light (histological, immunohistochemical, autoradiographic) and electron microscopic observations. As a result of this surgery, in the basal layer of the olfactory neuroepithelium the rate of mitotic activity increased while the number of mature olfactory neurons was greatly reduced. The regrowing olfactory axons, by forming ectopic glomerular structures in the damaged target, profoundly influenced its reorganization. The typical layered morphology of the olfactory bulb was often disrupted in the bulbar remnant; the large dendrites of the deafferented mitral cells bent toward the ectopically located glomerular structures establishing numerous synaptic contacts. The results from this study indicate that the olfactory input plays an important role in the reorganization of the damaged olfactory bulb. Behavioral experiments in partially bulbectomized animals should provide essential information about the importance of a topological map in the processing of olfactory cues.     

N-acetyl-lactosamine in the rat olfactory system: Expression and potential role in neurite growth

Primary sensory olfactory neurons exhibit a mosaic topographical projection from the olfactory neuroepithelium in the nasal cavity to the olfactory bulb formation of the telencephalon. Axons from primary neurons that are widely scattered in the epithelium terminate in discrete regions of the olfactory bulb. It has been hypothesised that carbohydrates present on the surface of primary olfactory axons mediate selective fasciculation and the formation of the topographical pathway. We examined the expression of the disaccharide N-acetyl-lactosamine in both the developing and the adult rat olfactory system. N-acetyl-lactosamine was expressed by all primary sensory olfactory neurons and by their terminations in the olfactory bulb throughout embryonic development and early postnatal life. In adults, N-acetyl-lactosamine was restricted to a subpopulation of primary sensory olfactory neurons that were dispersed throughout the neuroepithelium but that projected predominantly to the ventrolateral and ventromedial surfaces of the olfactory bulb. The axons of these neurons sort out in the outer layer of the bulb and preferentially self-fasciculate to form distinct axon bundles that terminate within select glomeruli. The role of N-acetyl-lactosamine in axon growth was tested by culturing primary sensory olfactory neurons on substrate-bound carbohydrates. Olfactory neuroepithelium cultures from both embryonic and postnatal rats revealed that substrate-bound N-acetyl-lactosamine was a strong and specific neurite growth-promoting agent. These data suggest that, during development of the olfactory projection, N-acetyl-lactosamine, which is present on all olfactory axons, acts as a nonselective permissive substrate for axon growth. In adults, however, the restricted distribution of N-acetyl-lactosamine on a subpopulation of axons may facilitate sorting out and self-fasciculation, which is necessary for preserving the mosaic nature of the olfactory pathway in this highly plastic region of the nervous system. These results support the hypothesis that cell surface carbohydrates are involved in axon growth in the olfactory system. © 1996 Wiley-Liss, Inc.     

Adult-derived neural precursors transplanted into multiple regions in the adult brain

Neural stem cells persist in the adult brain subventricular zone (SVZ). These cells generate a large number of new neurons that migrate to the olfactory bulb, where they complete their differentiation. Here, we transplanted cells carrying beta-galactosidase under the control of neuron-specific enolase promoter (NSE::LacZ) from the SVZ of adult mice into the striatum cortex and olfactory bulb, with or without an excitotoxin lesion. Between 2 and 8 weeks after transplantation, grafted cells were present in the recipient regions, but extensive migration and differentiation into mature neurons of grafted cells were only observed in the olfactory bulb. Clusters of graft-derived neuroblasts forming chain-like structures were observed within or close to the grated sites in the cortex and striatum; electron microscopy confirmed that graft-derived cells in the olfactory bulb and a small number in the striatum were neurons. Surprisingly, most of the cells expressing NSE::LacZ outside the olfactory bulb were astrocytes. We conclude that primary precursors from the SVZ migrate and differentiate effectively only within the environment of the olfactory bulb. Only limited survival and differentiation were observed in other brain regions studied.     

Distinct subsets of sensory olfactory neurons in mouse: Possible role in the formation of the mosaic olfactory projection

The axons of the primary sensory olfactory neurons project from the olfactory neuroepithelium lining the nasal cavity, onto glomeruli covering the surface of the olfactory bulb. Neuroanatomical studies have shown previously that individual olfactory glomeruli are innervated by neurons that are dispersed widely within the nasal cavity. The aim of the present study was to test the hypothesis that phenotypically unique subsets of primary sensory olfactory neurons, scattered throughout the nasal cavity, project to a subset of glomeruli in specific olfactory bulb loci. Immunochemical and histochemical analyses in neonatal mice revealed that the plant lectin, Dolichos biflorus agglutinin, bound to a subset of mature primary sensory olfactory neurons which express the olfactory marker protein. This subset of neurons was principally located in the rostromedial and dorsal portions of the nasal cavity and projected specifically to a subset of glomeruli in the rostromedial and caudodorsal portions of the olfactory bulb. Analysis of Dolichos biflorus-reactive axons revealed that these axons coursed randomly, with no evidence of their selective fasciculation, within the olfactory nerve. It was only at the level of the rostral olfactory bulb that a significant reorganisation of their trajectory was observed. Within the outer fibre layer of the bulb, discrete bundles of lectin-reactive axons began to coalesce selectively into fascicles which preferentially oriented toward the medial side of the olfactory bulb. These data demonstrated that a phenotypically distinct subset of primary sensory olfactory neurons exhibits a topographical projection from the olfactory epithelium to the olfactory bulb, and suggests that these, and other subsets, may form the basis of the mosaic nature of this pathway. Moreover, it appears that the outer nerve fibre layer in the rostral olfactory bulb plays an important instructive role in the guidance and fasciculation of olfactory sensory axons. © 1993 Wiley-Liss, Inc.     

Expression of Bcl-2 extends the survival of olfactory receptor neurons in the absence of an olfactory bulb

In the olfactory neuroepithelium, the number of olfactory receptor neurons (ORNs) is maintained at a relatively constant level by a precise balance between the elimination of mature receptors and proliferation of their precursors. However, little is known of the mechanisms that couple alterations in receptor death rates to changes in precursor proliferation. To investigate this relationship, we generated a line of mice expressing Bcl-2, a protein with anti-apoptotic properties, in mature olfactory receptor neurons using the Olfactory Marker Protein (OMP) promoter. OMP-bcl-2 transgenic mice showed selective expression of Bcl-2 in mature sensory neurons of the olfactory neuroepithelium (ONE) and vomeronasal organ. Olfactory bulbectomy (OBX) resulted in the death of mature receptor neurons followed by the sustained proliferation of their precursors in wild-type and OMP-bcl-2 transgenic mice. The persistently enhanced proliferation of olfactory neuroblasts that followed bulbectomy was indistinguishable between transgenic and non-transgenic mice. However, receptor neurons that were subsequently born in the absence of the bulb had longer life spans in OMP-bcl-2 mice. The increased proliferation of neuroblasts and extended life spans combined to restore near normal numbers of olfactory receptors in bulbectomized OMP-bcl-2 mice. A model is proposed to explain the dissociation of death and proliferation in OMP-bcl-2 transgenic mice.     

Olfactory sensory neurons are trophically dependent on the olfactory bulb for their prolonged survival.

In most neural systems, developing neurons are trophically dependent on contact with their synaptic target for their survival and for some features of their differentiation. However, in the olfactory system, it is unclear whether or not the survival and differentiation of olfactory sensory neurons depend on contact with the olfactory bulb (normally the sole synaptic target for these neurons). In order to address this issue, we examined neuronal life-span and differentiation in adult rats subjected to unilateral olfactory bulb ablation at least 1 month prior to use. Life-span of a newly generated cohort of olfactory neurons was determined by labeling them at their "birth" via the incorporation of 3H-thymidine. In the absence of the bulb, neurons are continually produced at a twofold greater rate. However, the epithelium on the ablated side is thinner, indicating that average neuronal life-span must be reduced in the targetless epithelium. Indeed, nearly 90% of the labeled neurons disappear from the bulbectomized side between 5 d and 2 weeks of neuronal age. Moreover, on electron microscopic examination, olfactory axons are degenerating in large numbers on the ablated side. Since labeled neurons migrate apically through the width of the epithelium during this same period, it appears that most, if not all, neurons on the ablated side have a life-span on the order of 2 weeks or less. In contrast, there is a more moderate degree of neuronal loss on the unoperated side of the same animals during the first 2 weeks after tracer injection, and that occurs while the neurons are concentrated in the deeper half of the epithelium, suggesting that there is a preexisting population of neurons in the control epithelium that does not die during this period. Likewise, degenerating axons are much less frequent on the unoperated side. We conclude that life-span is significantly shorter for olfactory neurons born in the targetless epithelium and that olfactory neurons are trophically dependent on the presence of the bulb for their prolonged survival. Neuronal differentiation in the absence of the bulb was assessed according to ultrastructural criteria and the pattern of protein expression using antisera to the growth associated protein GAP-43 and the olfactory marker protein. By both measures, most neurons in the epithelium on the bulbectomized side, but not all, are immature.(ABSTRACT TRUNCATED AT 400 WORDS)     

Intraocular transplants of olfactory neuroepithelium in rat

Olfactory neuroepithelium of neonatal rat pups has been transplanted in the anterior chamber of the eye of adult rats. Structural and ultrastructural observations at 5, 10, 30, 50, 90 and 120 days show that mature neurons degenerate rapidly in the expiant (before 5 days) while the basal elements proliferate and produce a new population of young neurons between 1 and 10 days. At longer survivals (30–120 days) it is seen that the neurons acquire morphological maturity, and positivity to the olfactory marker protein, as demonstrated by immunohistochemical methods. Our observations show that new neurons can be generated in the transplanted neuroepithelium and that their generation and maturation occurs in the absence of connections with a brain target.     

Long‐term survival of olfactory sensory neurons after target depletion

Life‐long addition and elimination of neurons within the adult olfactory epithelium and olfactory bulb allows for adaptive structural responses to sensory experience, learning, and recovery after injury. The interdependence of the two structures is highlighted by the shortened life span of sensory neurons deprived of bulb contact, and has prompted the hypothesis that trophic cues from the bulb contribute to their survival. The specific identity and source of these signals remain unknown. To investigate the potential role of target neurons in this support, we employed a neurotoxic lesion to selectively remove them while preserving the remaining nerve projection pathway, and examined the dynamics of sensory neuron proliferation and survival. Pulse‐labeling of progenitors with bromodeoxyuridine showed that, as with surgical bulb removal, increased apoptosis in the epithelium triggered accelerated production of new neurons after chemical depletion of target cells. Rather than undergoing premature death, a large subpopulation of these neurons survived long term. The combination of increased proliferation and extended survival resulted in essentially normal numbers of new sensory neurons surviving for as long as 5 weeks, with an accompanying restoration of olfactory marker protein expression. Changes in neurotrophic factor expression levels as measured by quantitative polymerase chain reaction (Q‐PCR), and in bulb cell populations, including the addition of new neurons generated in the subventricular zone, were observed in the injured bulb. These data indicate that olfactory sensory neurons can adapt to reductions in their normal target field by obtaining sufficient support from remaining or alternative cell sources to survive and maintain their projections. J. Comp. Neurol. 515:696–710, 2009. © 2009 Wiley‐Liss, Inc.     

Expression of the intermediate filament protein nestin by sustentacular cells in mature olfactory neuroepithelium

The intermediate filament protein nestin has been widely used as a marker for proliferating neural progenitor cells in the nervous system. The mammalian olfactory neuroepithelium is a region of the nervous system that robustly supports ongoing neurogenesis, yet where nestin has not been reported to mark proliferating progenitors. Using immunohistochemistry, we examined nestin expression in the mature olfactory neuroepithelium and found it to be tightly restricted to the basal compartment where the olfactory neuronal progenitor cell population resides. The pattern of nestin immunoreactivity was consistent with expression by the endfeet and inferior processes of sustentacular cells rather than basal cells. Using a bank of defined antibody markers, we confirmed nestin's pattern of distribution to be different from that of cytokeratin, vimentin, GBC-1, GAP43, and carnosine. It was highly similar to the pattern of SUS-4 immunoreactivity in the basal region of the neuroepithelium. Following surgical bulbectomy, nestin expression was up-regulated and became evident in the cell bodies of sustentacular cells situated more apically in the neuroepithelium. We have shown nestin to be present in the basal region of the adult olfactory neuroepithelium in the zone that supports ongoing neurogenesis in the adult, but its expression is restricted to the inferior parts of sustentacular cells rather than the neuronal progenitor cells. Nestin may play a potential role in the migration of recently proliferated olfactory neurons on the scaffolding of sustentacular cells in a manner analogous to its proposed role in radial glia during embryonic development of the central nervous system.     

FGF2 promotes neuronal differentiation in explant cultures of adult and embryonic mouse olfactory epithelium

Neurogenesis in the adult olfactory epithelium is highly regulated in vivo. Little is known of the molecular signals which control this process, although contact with the olfactory bulb or with astrocytes has been implicated. Explants of mouse olfactory epithelium were grown in the presence or absence of several peptide growth factors. Basic fibroblast growth factor (FGF2) stimulated differentiation of sensory neurons in adult and embryonic olfactory epithelium. Other growth factors tested were ineffective. FGF2-stimulated neurons were born in vitro and expressed neurofilament, neural cell adhesion molecule, and β-tubulin. The cells also expressed olfactory marker protein, a marker for mature olfactory sensory neurons in vivo. These bipolar neurons did not express glial fibrillary acidic protein or low-affinity nerve growth factor receptor. These results indicate that neither astrocytes nor olfactory bulb are necessary for differentiation of olfactory sensory neurons in vitro. © 1996 Wiley-Liss, Inc.     

Chemically and morphologically identifiable glomeruli in the rat olfactory bulb.

Primary olfactory neurons that express the same odorant receptor are distributed mosaically throughout the olfactory neuroepithelium lining the nasal cavity, yet their axons converge and form discrete glomeruli in the olfactory bulb. We previously proposed that cell surface carbohydrates mediate the sorting out and selective fasciculation of primary olfactory axons en route to glomeruli. If this were the case, then axons that terminate in the same glomerulus would express the same complement of cell surface carbohydrates. In this study, we examined the expression of a novel carbohydrate (NOC-3) on neural cell adhesion molecule in the adult rat olfactory system. NOC-3 was expressed by a subset of neurons distributed throughout the olfactory neuroepithelium. The axons of these neurons entered the nerve fiber layer and terminated in a subset of glomeruli. It is interesting to note that we identified three unusually large glomeruli in the lateral, ventrolateral, and ventromedial olfactory bulb that were innervated by axons expressing NOC-3. NOC-3-expressing axons sorted out and fasciculated into discrete fascicles prior to entering these glomeruli. Each of these glomeruli was in a topographically fixed position in the olfactory bulbs of the same animal as well as in different animals, and their lengths were approximately 10% of the total length of the bulb. They could be identified reliably by both their topographical position and their unique morphology. These results reveal that axons expressing the same cell surface carbohydrates consistently target the same topographically fixed glomeruli, which supports a role for these molecules in axon navigation in the primary olfactory nerve pathway.     

Differential expression of G proteins in the mouse olfactory system

Transmembrane signaling events at the dendrites and axons of olfactory receptor neurons mediate distinct functions. Whereas odorant recognition and chemosensory transduction occur at the dendritic membranes of olfactory neurons, signal propagation, axon sorting and target innervation are functions of their axons. The roles of G proteins in transmembrane signaling at the dendrites have been studied extensively, but axonal G proteins have not been investigated in detail. We used immunohistochemistry to visualize expression of alpha subunits of G(o) and G(i2) in the mouse olfactory system. G(o) is expressed ubiquitously on axons of olfactory receptor neurons throughout the olfactory neuroepithelium and in virtually all glomeruli in the main olfactory bulb. In contrast, expression of G(i2) is restricted to a sub-population of olfactory neurons, along the dorsal septum and the dorsal recess of the nasal cavity, which projects primarily to medial regions of the olfactory bulb, with the exception of glomeruli adjacent to the pathway of the vomeronasal nerve. In contrast to the overlapping expression patterns of G(o) and G(i2) in the main olfactory system, neurons expressing G(o) and those expressing G(i2) in the accessory olfactory bulb are more clearly separated, in agreement with previous studies. Vomeronasal axons terminating in glomeruli in the rostral region of the accessory olfactory bulb express G(i2), whereas those projecting to the caudal region express G(o). Characterization of the expression patterns of G(i2) and G(o) in the olfactory projection is essential for future studies aimed at relating transmembrane signaling events to signal propagation, axon sorting and target innervation.