Effects of prenatal alcohol exposure on the olfactory system development.

Fetal Alcohol Spectrum Disorders (FASD), resulting from maternal alcohol consumption during pregnancy, are a prominent non-genetic cause of physical disabilities and brain damage in children. Alongside common symptoms like distinct facial features and neurocognitive deficits, sensory anomalies, including olfactory dysfunction, are frequently noted in FASD-afflicted children. However, the precise mechanisms underpinning the olfactory abnormalities induced by prenatal alcohol exposure (PAE) remain elusive. Utilizing rodents as a model organism with varying timing, duration, dosage, and administration routes of alcohol exposure, prior studies have documented impairments in olfactory system development caused by PAE. Many reported a reduction in the olfactory bulb (OB) volume accompanied by reduced OB neuron counts, suggesting the OB is a brain region vulnerable to PAE. In contrast, no significant olfactory system defects were observed in some studies, though subtle alterations might exist. These findings suggest that the timing, duration, and extent of fetal alcohol exposure can yield diverse effects on olfactory system development. To enhance comprehension of PAE-induced olfactory dysfunctions, this review summarizes key findings from previous research on the olfactory systems of offspring prenatally exposed to alcohol.

Developmental and evolutionary constraints on olfactory circuit selection.

SignificanceIn this work, we explore the hypothesis that biological neural networks optimize their architecture, through evolution, for learning. We study early olfactory circuits of mammals and insects, which have relatively similar structure but a huge diversity in size. We approximate these circuits as three-layer networks and estimate, analytically, the scaling of the optimal hidden-layer size with input-layer size. We find that both longevity and information in the genome constrain the hidden-layer size, so a range of allometric scalings is possible. However, the experimentally observed allometric scalings in mammals and insects are consistent with biologically plausible values. This analysis should pave the way for a deeper understanding of both biological and artificial networks.

The maturational characteristics of the GABA input in the anterior piriform cortex may also contribute to the rapid learning of the maternal odor during the sensitive period.

During the first ten postnatal days (P), infant rodents can learn olfactory preferences for novel odors if they are paired with thermo-tactile stimuli that mimic components of maternal care. After P10, the thermo-tactile pairing becomes ineffective for conditioning. The current explanation for this change in associative learning is the alteration in the norepinephrine (NE) inputs from the locus coeruleus (LC) to the olfactory bulb (OB) and the anterior piriform cortex (aPC). By combining patch-clamp electrophysiology and computational simulations, we showed in a recent work that a transitory high responsiveness of the OB-aPC circuit to the maternal odor is an alternative mechanism that could also explain early olfactory preference learning and its cessation after P10. That result relied solely on the maturational properties of the aPC pyramidal cells. However, the GABAergic system undergoes important changes during the same period. To address the importance of the maturation of the GABAergic system for early olfactory learning, we incorporated data from the GABA inputs, obtained from in vitro patch-clamp experiment in the aPC of rat pups aged P5-P7 reported here, to the model proposed in our previous publication. In the younger than P10 OB-aPC circuit with GABA synaptic input, the number of responsive aPC pyramidal cells to the conditioned maternal odor was amplified in 30% compared to the circuit without GABAergic input. When compared with the circuit with other younger than P10 OB-aPC circuit with adult GABAergic input profile, this amplification was 88%. Together, our results suggest that during the olfactory preference learning in younger than P10, the GABAergic synaptic input presumably acts by depolarizing the aPC pyramidal neurons in such a way that it leads to the amplification of the pyramidal neurons response to the conditioned maternal odor. Furthermore, our results suggest that during this developmental period, the aPC pyramidal cells themselves seem to resolve the apparent lack of GABAergic synaptic inhibition by a strong firing adaptation in response to increased depolarizing inputs.

Morphogenesis of Olfactory Organ of Bushymouth Catfish Ancistrus dolichopterus (Teleostei: Loricariidae) before Switching to Exogenous Feeding.

Olfaction plays an important role in a fish's life. Its value may differ at different developmental stages, depending on the feeding style of the species. The goal of the present study was to investigate the olfactory organ of a species that feeds mainly on algae- the bushymouth catfish, Ancistrus dolichopterus-at developmental stages from olfactory placode to the definitive olfactory chamber. For this study, we used light and electron (scanning) microscopy. The topography of the olfactory placode of A. dolichopterus is typical for teleostei. Formation of olfactory pit takes place at the same time as rostral elevation formation. Rostral elevation participates in the formation of the nasal bridge and anterior tubular nostril. It was found out that the anlage of olfactory rosette in A. dolichopterus arises earlier than in most teleostei. However, the number of lamellae does not increase until switching to exogenous feeding. We suppose that the early development of olfactory organ is necessary for intraspecific communication, not just for finding food.

Structured spike series specify gene expression patterns for olfactory circuit formation.

Neural circuits emerge through the interplay of genetic programming and activity-dependent processes. During the development of the mouse olfactory map, axons segregate into distinct glomeruli in an olfactory receptor (OR)-dependent manner. ORs generate a combinatorial code of axon-sorting molecules whose expression is regulated by neural activity. However, it remains unclear how neural activity induces OR-specific expression patterns of axon-sorting molecules. We found that the temporal patterns of spontaneous neuronal spikes were not spatially organized but were correlated with the OR types. Receptor substitution experiments demonstrated that ORs determine spontaneous activity patterns. Moreover, optogenetically differentiated patterns of neuronal activity induced specific expression of the corresponding axon-sorting molecules and regulated axonal segregation. Thus, OR-dependent temporal patterns of spontaneous activity play instructive roles in generating the combinatorial code of axon-sorting molecules during olfactory map formation.

Lack of MeCP2 leads to region-specific increase of doublecortin in the olfactory system.

The protein doublecortin is mainly expressed in migrating neuroblasts and immature neurons. The X-linked gene MECP2, associated to several neurodevelopmental disorders such as Rett syndrome, encodes the protein methyl-CpG-binding protein 2 (MeCP2), a regulatory protein that has been implicated in neuronal maturation and refinement of olfactory circuits. Here, we explored doublecortin immunoreactivity in the brain of young adult female Mecp2-heterozygous and male Mecp2-null mice and their wild-type littermates. The distribution of doublecortin-immunoreactive somata in neurogenic brain regions was consistent with previous reports in rodents, and no qualitative differences were found between genotypes or sexes. Quantitatively, we found a significant increase in doublecortin cell density in the piriform cortex of Mecp2-null males as compared to WT littermates. A similar increase was seen in a newly identified population of doublecortin cells in the olfactory tubercle. In these olfactory structures, however, the percentage of doublecortin immature neurons that also expressed NeuN was not different between genotypes. By contrast, we found no significant differences between genotypes in doublecortin immunoreactivity in the olfactory bulbs. Nonetheless, in the periglomerular layer of Mecp2-null males, we observed a specific decrease of immature neurons co-expressing doublecortin and NeuN. Overall, no differences were evident between Mecp2-heterozygous and WT females. In addition, no differences could be detected between genotypes in the density of doublecortin-immunoreactive cells in the hippocampus or striatum of either males or females. Our results suggest that MeCP2 is involved in neuronal maturation in a region-dependent manner.

A Population of Navigator Neurons Is Essential for Olfactory Map Formation during the Critical Period.

In the developing brain, heightened plasticity during the critical period enables the proper formation of neural circuits. Here, we identify the "navigator" neurons, a group of perinatally born olfactory sensory neurons, as playing an essential role in establishing the olfactory map during the critical period. The navigator axons project circuitously in the olfactory bulb and traverse multiple glomeruli before terminating in perspective glomeruli. These neurons undergo a phase of exuberant axon growth and exhibit a shortened lifespan. Single-cell transcriptome analyses reveal distinct molecular signatures for the navigators. Extending their lifespan prolongs the period of exuberant growth and perturbs axon convergence. Conversely, a genetic ablation experiment indicates that, despite postnatal neurogenesis, only the navigators are endowed with the ability to establish a convergent map. The presence and the proper removal of the navigator neurons are both required to establish tight axon convergence into the glomeruli.

Two types of sensory proliferation patterns underlie the formation of spatially tuned olfactory receptive fields in the cockroach Periplaneta americana.

In the cockroach Periplaneta americana, to represent pheromone source in the receptive space, axon terminals of sex pheromone-receptive olfactory sensory neurons (pSNs) are topographically organized within the primary center, the macroglomerulus, according to the peripheral locations of sex pheromone-receptive single walled (sw)-B sensilla. In this study, we sought to determine when and where pSNs emerge in the nymphal antenna. We revealed two different pSN proliferation patterns that underlie the formation of topographic organization in the macroglomerulus. In nymphal antennae, which lack sw-B sensilla, pSNs are identified in the shorter sensilla, termed sw-A sensilla. Because new sw-A sensilla emerge on the proximal antenna at every molt, topographic organization in the macroglomerulus must be formed by adding axon terminals of newly emerged pSNs to the lateral region in the macroglomerulus at each molt. At the final molt, a huge number of new sw-B sensilla appeared throughout the whole antenna. Sw-B sensilla in the proximal part of the adult antenna were newly formed during the last instar stage, whereas those located in the distal antenna were transformed from sw-A sensilla. This transformation was accompanied by an increase in the number of pSNs. Axon terminals of newborn pSNs in new sw-B sensilla were recruited to the lateral part of the macroglomerulus, whereas those of newborn pSNs in transformed sw-B sensilla were recruited to the macroglomerulus according to the sensillar location. These mechanisms enable an increase in sensitivity to sex pheromone in adulthood while retaining the topographic map formed during the postembryonic development.

Diverse populations of local interneurons integrate into the Drosophila adult olfactory circuit.

Drosophila olfactory local interneurons (LNs) in the antennal lobe are highly diverse and variable. How and when distinct types of LNs emerge, differentiate, and integrate into the olfactory circuit is unknown. Through systematic developmental analyses, we found that LNs are recruited to the adult olfactory circuit in three groups. Group 1 LNs are residual larval LNs. Group 2 are adult-specific LNs that emerge before cognate sensory and projection neurons establish synaptic specificity, and Group 3 LNs emerge after synaptic specificity is established. Group 1 larval LNs are selectively reintegrated into the adult circuit through pruning and re-extension of processes to distinct regions of the antennal lobe, while others die during metamorphosis. Precise temporal control of this pruning and cell death shapes the global organization of the adult antennal lobe. Our findings provide a road map to understand how LNs develop and contribute to constructing the olfactory circuit.

Light Microscopy of Development of the Olfactory Organ of European Weatherfish Misgurnus fossilis (Тeleostei: Cobitidae).

We studied the formation of the olfactory organ of European weatherfish Misgurnus fossilis (Linnaeus, 1758) from the formation of its placodes to the formation of lamellae using light microscopy. The aim of the present investigation was to determine which morphogenetic features of the olfactory organ ate associated with the demersal lifestyle. The olfactory organ of European weatherfish goes through such developmental stages as olfactory placode, olfactory pit, and olfactory chamber with lamellae. Formation of the olfactory pit occurs in prolarvae, but a considerable increase in size takes place during the switch to exogenous feeding. In late larva, lamellae are formed in the aboral part olfactory chamber as an outward fold at its bottom. At the same time, incurrent and excurrent nostrils start being formed as the result of approximation of lateral and medial margins in the rostal part above the olfactory cavity. In contrast to other fishes, the peculiarity of morphogenesis of olfactory organ of M. fossilis is its late formation followed by intensive postembryonic development.

Immunohistochemical Approach to Understanding the Organization of the Olfactory System in the Cuttlefish, Sepia officinalis.

Cephalopods are nontraditional but captivating models of invertebrate neurobiology, particularly in evolutionary comparisons. Cephalopod olfactory systems have striking similarities and fundamental differences with vertebrates, arthropods, and gastropods, raising questions about the ancestral origins of those systems. We describe here the organization and development of the olfactory system of the common cuttlefish, Sepia officinalis, using immunohistochemistry and in situ hybridization. FMRFamide and/or related peptides and histamine are putative neurotransmitters in olfactory sensory neurons. Other neurotransmitters, including serotonin and APGWamide within the olfactory and other brain lobes, suggest efferent control of olfactory input and/or roles in the processing of olfactory information. The distributions of neurotransmitters, along with staining patterns of phalloidin, anti-acetylated α-tubulin, and a synaptotagmin riboprobe, help to clarify the structure of the olfactory lobe. We discuss a key difference, the lack of identifiable olfactory glomeruli, in cuttlefish in comparison to other models, and suggest its implications for the evolution of olfaction.

Chronic perinatal odour exposure with heptaldehyde affects odour sensitivity and olfactory system homeostasis in preweaning mice.

Exposure to specific odorants in the womb during pregnancy or in the milk during early nursing is known to impact morpho-functional development of the olfactory circuitry of pups. This can be associated with a modification in olfactory sensitivity and behavioural olfactory-based preferences to the perinatally encountered odorants measured at birth, weaning or adult stage. Effects depend on a multitude of factors, such as odorant type, concentration, administration mode and frequency, as well as timing and mice strain. Here, we examined the effect of perinatal exposure to heptaldehyde on the neuro-anatomical development of the olfactory receptor Olfr2 circuitry, olfactory sensitivity and odour preferences of preweaning pups using mI7-IRES-tau-green fluorescent protein mice. We found that perinatal odour exposure through the feed of the dam reduces the response to heptaldehyde and modulates transcript levels of neuronal transduction proteins in the olfactory epithelium of the pups. Furthermore, the number of I7 glomeruli related to Olfr2-expressing OSN is altered in a way similar to that seen with restricted post-natal exposure, in an age-dependent way. These variations are associated with a modification of olfactory behaviours associated with early post-natal odour preferences at weaning.

Structure and development of the subesophageal zone of the Drosophila brain. II. Sensory compartments.

The subesophageal zone (SEZ) of the Drosophila brain processes mechanosensory and gustatory sensory input from sensilla located on the head, mouth cavity and trunk. Motor output from the SEZ directly controls the movements involved in feeding behavior. In an accompanying paper (Hartenstein et al., ), we analyzed the systems of fiber tracts and secondary lineages to establish reliable criteria for defining boundaries between the four neuromeres of the SEZ, as well as discrete longitudinal neuropil domains within each SEZ neuromere. Here we use this anatomical framework to systematically map the sensory projections entering the SEZ throughout development. Our findings show continuity between larval and adult sensory neuropils. Gustatory axons from internal and external taste sensilla of the larva and adult form two closely related sensory projections, (a) the anterior central sensory center located deep in the ventromedial neuropil of the tritocerebrum and mandibular neuromere, and (b) the anterior ventral sensory center (AVSC), occupying a superficial layer within the ventromedial tritocerebrum. Additional, presumed mechanosensory terminal axons entering via the labial nerve define the ventromedial sensory center (VMSC) in the maxilla and labium. Mechanosensory afferents of the massive array of chordotonal organs (Johnston's organ) of the adult antenna project into the centrolateral neuropil column of the anterior SEZ, creating the antenno-mechanosensory and motor center (AMMC). Dendritic projections of dye back-filled motor neurons extend throughout a ventral layer of the SEZ, overlapping widely with the AVSC and VMSC. Our findings elucidate fundamental structural aspects of the developing sensory systems in Drosophila.

Olfactory Development, Part 2: Neuroanatomic Maturation and Dysgeneses.

Olfactory axons project from nasal epithelium to the primitive telencephalon before olfactory bulbs form. Olfactory bulb neurons do not differentiate in situ but arrive via the rostral migratory stream. Synaptic glomeruli and concentric laminar architecture are unlike other cortices. Fetal olfactory maturation of neuronal differentiation, synaptogenesis, and myelination remains incomplete at term and have a protracted course of postnatal development. The olfactory ventricular recess involutes postnatally but dilates in congenital hydrocephalus. Olfactory bulb, tract and epithelium are repositories of progenitor stem cells in fetal and adult life. Diverse malformations of the olfactory bulb can be diagnosed by clinical examination, imaging, and neuropathologically. Cellular markers of neuronal differentiation and synaptogenesis demonstrate immaturity of the olfactory system at birth, previously believed by histology alone to occur early in fetal life. Immaturity does not preclude function.

Olfactory Development, Part 1: Function, From Fetal Perception to Adult Wine-Tasting.

Discrimination of odorous molecules in amniotic fluid occur after 30 weeks' gestation; fetuses exhibit differential responses to maternal diet. Olfactory reflexes enable reliable neonatal testing. Olfactory bulbs can be demonstrated reliably by MRI after 30 weeks' gestation, and their hypoplasia or aplasia also documented by late prenatal and postnatal MRI. Olfactory axons project from nasal epithelium to telencephalon before olfactory bulbs form. Fetal olfactory maturation remains incomplete at term for neuronal differentiation, synaptogenesis, myelination, and persistence of the transitory fetal ventricular recess. Immaturity does not signify nonfunction. Olfaction is the only sensory system without thalamic projection because of its own intrinsic thalamic equivalent. Diverse malformations of the olfactory bulb can be diagnosed by clinical examination, imaging, and neuropathology. Some epileptic auras might be primarily generated in the olfactory bulb. Cranial nerve 1 should be tested in all neonates and especially in patients with brain malformations, endocrinopathies, chromosomopathies, and genetic/metabolic diseases.

Semaphorin-1a prevents Drosophila olfactory projection neuron dendrites from mis-targeting into select antennal lobe regions.

Elucidating how appropriate neurite patterns are generated in neurons of the olfactory system is crucial for comprehending the construction of the olfactory map. In the Drosophila olfactory system, projection neurons (PNs), primarily derived from four neural stem cells (called neuroblasts), populate their cell bodies surrounding to and distribute their dendrites in distinct but overlapping patterns within the primary olfactory center of the brain, the antennal lobe (AL). However, it remains unclear whether the same molecular mechanisms are employed to generate the appropriate dendritic patterns in discrete AL glomeruli among PNs produced from different neuroblasts. Here, by examining a previously explored transmembrane protein Semaphorin-1a (Sema-1a) which was proposed to globally control initial PN dendritic targeting along the dorsolateral-to-ventromedial axis of the AL, we discover a new role for Sema-1a in preventing dendrites of both uni-glomerular and poly-glomerular PNs from aberrant invasion into select AL regions and, intriguingly, this Sema-1a-deficient dendritic mis-targeting phenotype seems to associate with the origins of PNs from which they are derived. Further, ectopic expression of Sema-1a resulted in PN dendritic mis-projection from a select AL region into adjacent glomeruli, strengthening the idea that Sema-1a plays an essential role in preventing abnormal dendritic accumulation in select AL regions. Taken together, these results demonstrate that Sema-1a repulsion keeps dendrites of different types of PNs away from each other, enabling the same types of PN dendrites to be sorted into destined AL glomeruli and permitting for functional assembly of olfactory circuitry.

Ontogenetic Development of the Derived Olfactory System of the Mantellid Frog Mantidactylus betsileanus.

The nasal cavity of Mantidactylus betsileanus, a frog of the Madagascar-Comoroan endemic family Mantellidae, is characterized by a unique internal architecture. Unlike the state commonly observed in anurans, the two discernible olfactory subsystems of M. betsileanus (the main olfactory organ and the vomeronasal organ) are anatomically separated from each other, suggesting an enhanced functional differentiation. Here we evaluate the ontogenetic formation of this extraordinary anatomical state based on a histological study of a developmental series of M. betsileanus. The olfactory system of premetamorphic tadpoles, and most of its changes during metamorphosis, resembles that of other anurans. At the end of metamorphosis however, a growing obstruction of the passage between main olfactory organ and vomeronasal organ takes place, leading to the deviant morphological state previously described for adults. The late appearance of this atypical anatomical feature in the course of ontogeny agrees with the phylogenetic hypothesis of the observed obstruction representing a derived state for these frogs. From a functional point of view, the apparent autonomy of the vomeronasal organ is possibly linked to the presence of clade-specific femoral glands that are known to produce pheromones and that likewise are fully expressed in adults only. Anat Rec, 299:943-950, 2016. © 2016 Wiley Periodicals, Inc.

Shaping the Reproductive System: Role of Semaphorins in Gonadotropin-Releasing Hormone Development and Function.

The semaphorin proteins, which contribute to the morphogenesis and homeostasis of a wide range of systems, are among the best-studied families of guidance cues. Much recent research has focused on the role of semaphorins in the development and adult activity of hormone systems and, reciprocally, how circulating reproductive hormones regulate their expression and function. Specifically, several reports have focused on the molecular mechanisms underlying the effects of semaphorins on the migration, survival and structural and functional plasticity of neurons that secrete gonadotropin-releasing hormone (GnRH), essential for the acquisition and maintenance of reproductive competence in mammals. Alterations in the development of this neuroendocrine system lead to anomalous or absent GnRH secretion, resulting in heterogeneous reproductive disorders such as congenital hypogonadotropic hypogonadism (CHH) or other conditions characterized by infertility or subfertility. This review summarizes current knowledge of the role of semaphorins and their receptors on the development, differentiation and plasticity of the GnRH system. In addition, the involvement of genetic deficits in semaphorin signaling in some forms of CHH in humans is discussed.

Developmental regulation of neural map formation in the mouse olfactory system.

In the mouse olfactory system, various odorants are detected by approximately 1000 different odorant receptors (ORs) expressed in the olfactory sensory neurons (OSNs). It is well established that each OSN expresses only one functional OR gene in a monoallelic manner. Furthermore, OSN axons expressing the same OR converge to a set of glomeruli in the olfactory bulb (OB). During embryonic development, a coarse map is formed by the combination of two genetically programmed processes. One is OR-independent axonal projection along the dorsal-ventral (D-V) axis, and the other is OR-dependent projection along the anterior-posterior (A-P) axis. D-V projection is regulated by the anatomical location of OSNs within the olfactory epithelium (OE), whereas A-P projection is instructed by expressed OR molecules using cyclic adenosine monophosphate (cAMP) signals. After birth, the map is further refined in an activity-dependent manner by its conversion from a continuous to a discrete map through segregation of glomerular structures. Here, we summarize recent progress from our laboratory in understanding neural map formation in the mouse olfactory system.

Olfactory ensheathing cells are the main phagocytic cells that remove axon debris during early development of the olfactory system.

During development of the primary olfactory system, axon targeting is inaccurate and axons inappropriately project within the target layer or overproject into the deeper layers of the olfactory bulb. As a consequence there is considerable apoptosis of primary olfactory neurons during embryonic and postnatal development and axons of the degraded neurons need to be removed. Olfactory ensheathing cells (OECs) are the glia of the primary olfactory nerve and are known to phagocytose axon debris in the adult and postnatal animal. However, it is unclear when phagocytosis by OECs first commences. We investigated the onset of phagocytosis by OECs in the developing mouse olfactory system by utilizing two transgenic reporter lines: OMP-ZsGreen mice which express bright green fluorescent protein in primary olfactory neurons, and S100ß-DsRed mice which express red fluorescent protein in OECs. In crosses of these mice, the fate of the degraded axon debris is easily visualized. We found evidence of axon degradation at embryonic day (E)13.5. Phagocytosis of the primary olfactory axon debris by OECs was first detected at E14.5. Phagocytosis of axon debris continued into the postnatal animal during the period when there was extensive mistargeting of olfactory axons. Macrophages were often present in close proximity to OECs but they contributed only a minor role to clearing the axon debris, even after widespread degeneration of olfactory neurons by unilateral bulbectomy and methimazole treatment. These results demonstrate that from early in embryonic development OECs are the primary phagocytic cells of the primary olfactory nerve.