Functional diversity on synaptic plasticity mediated by endocannabinoids

Endocannabinoids (eCBs) act as modulators of synaptic transmission through activation of a number of receptors, including, but not limited to, cannabinoid receptor 1 (CB1). eCBs share CB1 receptors as a common target with Δ⁹-tetrahydrocannabinol (THC), the main psychoactive ingredient in marijuana. Although THC has been used for recreational and medicinal purposes for thousands of years, little was known about its effects at the cellular level or on neuronal circuits. Identification of CB1 receptors and the subsequent development of its specific ligands has therefore enhanced our ability to study and bring together a substantial amount of knowledge regarding how marijuana and eCBs modify interneuronal communication. To date, the eCB system, composed of cannabinoid receptors, ligands and the relevant enzymes, is recognized as the best-described retrograde signalling system in the brain. Its impact on synaptic transmission is widespread and more diverse than initially thought. The aim of this review is to succinctly present the most common forms of eCB-mediated modulation of synaptic transmission, while also illustrating the multiplicity of effects resulting from specializations of this signalling system at the circuital level.     

Endocannabinoid signaling in adult hippocampal neurogenesis: A mechanistic and integrated perspective

Dentate gyrus of the hippocampus continuously gives rise to new neurons, namely, adult-born granule cells, which contribute to conferring plasticity to the mature brain throughout life. Within this neurogenic region, the fate and behavior of neural stem cells (NSCs) and their progeny result from a complex balance and integration of a variety of cell-autonomous and cell-to-cell-interaction signals and underlying pathways. Among these structurally and functionally diverse signals, there are endocannabinoids (eCBs), the main brain retrograde messengers. These pleiotropic bioactive lipids can directly and/or indirectly influence adult hippocampal neurogenesis (AHN) by modulating, both positively and negatively, multiple molecular and cellular processes in the hippocampal niche, depending on the cell type or stage of differentiation. Firstly, eCBs act directly as cell-intrinsic factors, cell-autonomously produced by NSCs following their stimulation. Secondly, in many, if not all, niche-associated cells, including some local neuronal and nonneuronal elements, the eCB system indirectly modulates the neurogenesis, linking neuronal and glial activity to regulating distinct stages of AHN. Herein, we discuss the crosstalk of the eCB system with other neurogenesis-relevant signal pathways and speculate how the hippocampus-dependent neurobehavioral effects elicited by (endo)cannabinergic medications are interpretable in light of the key regulatory role that eCBs play on AHN.     

Endocannabinoids generated by Ca2+ or by metabotropic glutamate receptors appear to arise from different pools of diacylglycerol lipase

The identity and subcellular sources of endocannabinoids (eCBs) will shape their ability to affect synaptic transmission and, ultimately, behavior. Recent discoveries support the conclusion that 2-arachidonoyl glycerol, 2-AG, is the major signaling eCB, however, some important issues remain open. 2-AG can be synthesized by a mechanism that is strictly Ca(2+)-dependent, and another that is initiated by G-protein coupled receptors (GPCRs) and facilitated by Ca(2+). An important question is whether or not the 2-AG in these cases is synthesized by the same pool of diacylglycerol lipase alpha (DAGLα). Using whole-cell voltage-clamp techniques in CA1 pyramidal cells in acute in vitro rat hippocampal slices, we investigated two mechanistically distinct eCB-mediated responses to address this issue. We now report that pharmacological inhibitors of DGLα have quantitatively different effects on eCB-mediated responses triggered by different stimuli, suggesting that functional, and perhaps physical, distinctions among pools of DAGLα exist.     

Developmental regulation of fear learning and anxiety behavior by endocannabinoids

The developing brain undergoes substantial maturation into adulthood and the development of specific neural structures occurs on differing timelines. Transient imbalances between developmental trajectories of corticolimbic structures, which are known to contribute to regulation over fear learning and anxiety, can leave an individual susceptible to mental illness, particularly anxiety disorders. There is a substantial body of literature indicating that the endocannabinoid (eCB) system critically regulates stress responsivity and emotional behavior throughout the life span, making this system a novel therapeutic target for stress‐ and anxiety‐related disorders. During early life and adolescence, corticolimbic eCB signaling changes dynamically and coincides with different sensitive periods of fear learning, suggesting that eCB signaling underlies age‐specific fear learning responses. Moreover, perturbations to these normative fluctuations in corticolimbic eCB signaling, such as stress or cannabinoid exposure, could serve as a neural substrate contributing to alterations to the normative developmental trajectory of neural structures governing emotional behavior and fear learning. In this review, we first introduce the components of the eCB system and discuss clinical and rodent models showing eCB regulation of fear learning and anxiety in adulthood. Next, we highlight distinct fear learning and regulation profiles throughout development and discuss the ontogeny of the eCB system in the central nervous system, and models of pharmacological augmentation of eCB signaling during development in the context of fear learning and anxiety.     

The role of diacylglycerol lipase in constitutive and angiotensin AT1 receptor-stimulated cannabinoid CB1 receptor activity

The cannabinoid CB1 receptor (CB1R) is a G protein-coupled receptor, which couples to the Gi/o family of heterotrimeric G proteins. The receptor displays both basal and agonist-induced signaling and internalization. Although basal activity of CB1Rs is attributed to constitutive (agonist-independent) receptor activity, studies in neurons suggested a role of postsynaptic endocannabinoid (eCB) release in the persistent activity of presynaptic CB1Rs. To elucidate the role of eCBs in basal CB1R activity, we have investigated the role of diacylglycerol lipase (DAGL) in this process in Chinese hamster ovary (CHO) cells, which are not targeted specifically with eCBs. Agonist-induced G protein activation was determined by detecting dissociation G protein subunits expressed in CHO cells with bioluminescence resonance energy transfer (BRET), after labeling the alpha and beta subunits with Renilla luciferase and enhanced yellow fluorescent protein (EYFP), respectively. Preincubation of the cells with tetrahydrolipstatin (THL), a known inhibitor of DAGLs, caused inhibition of the basal activity of CB1R. Moreover, preincubation of CHO and cultured hippocampal neurons with THL increased the number of CB1Rs on the cell membrane, which reflects its inhibitory action on CB1R internalization in non-simulated cells. In CHO cells co-expressing CB1R and angiotensin AT1 receptors, angiotensin II-induced Go protein activation that was blocked by both a CB1R antagonist and THL. These data indicate that cell-derived eCB mediators have a general role in the basal activity of CB1Rs in non-neural cells and neurons, and that this mechanism can be stimulated by AT1 receptor activation.     

Site-specific and time-dependent activation of the endocannabinoid system after transection of long-range projections

Background

After focal neuronal injury the endocannabinioid system becomes activated and protects or harms neurons depending on cannabinoid derivates and receptor subtypes. Endocannabinoids (eCBs) play a central role in controlling local responses and influencing neural plasticity and survival. However, little is known about the functional relevance of eCBs in long-range projection damage as observed in stroke or spinal cord injury (SCI).

Methods

In rat organotypic entorhino-hippocampal slice cultures (OHSC) as a relevant and suitable model for investigating projection fibers in the CNS we performed perforant pathway transection (PPT) and subsequently analyzed the spatial and temporal dynamics of eCB levels. This approach allows proper distinction of responses in originating neurons (entorhinal cortex), areas of deafferentiation/anterograde axonal degeneration (dentate gyrus) and putative changes in more distant but synaptically connected subfields (cornu ammonis (CA) 1 region).

Results

Using LC-MS/MS, we measured a strong increase in arachidonoylethanolamide (AEA), oleoylethanolamide (OEA) and palmitoylethanolamide (PEA) levels in the denervation zone (dentate gyrus) 24 hours post lesion (hpl), whereas entorhinal cortex and CA1 region exhibited little if any changes. NAPE-PLD, responsible for biosynthesis of eCBs, was increased early, whereas FAAH, a catabolizing enzyme, was up-regulated 48hpl.

Conclusion

Neuronal damage as assessed by transection of long-range projections apparently provides a strong time-dependent and area-confined signal for de novo synthesis of eCB, presumably to restrict neuronal damage. The present data underlines the importance of activation of the eCB system in CNS pathologies and identifies a novel site-specific intrinsic regulation of eCBs after long-range projection damage.     

A Lower Olfactory Capacity Is Related to Higher Circulating Concentrations of Endocannabinoid 2-Arachidonoylglycerol and Higher Body Mass Index in Women

The endocannabinoid (eCB) system can promote food intake by increasing odor detection in mice. The eCB system is over-active in human obesity. Our aim is to measure circulating eCB concentrations and olfactory capacity in a human sample that includes people with obesity and explore the possible interaction between olfaction, obesity and the eCB system. The study sample was made up of 161 females with five groups of body mass index sub-categories ranging from under-weight to morbidly obese. We assessed olfactory capacity with the “Sniffin´Sticks” test, which measures olfactory threshold-discrimination-identification (TDI) capacity. We measured plasma concentrations of the eCBs 2-arachidonoylglycerol (2-AG) and N-arachidonoylethanolamine or anandamide (AEA), and several eCB-related compounds, 2-acylglycerols and N-acylethanolamines. 2-AG and other 2-acylglycerols fasting plasma circulating plasma concentrations were higher in obese and morbidly obese subjects. AEA and other N-acylethanolamine circulating concentrations were lower in under-weight subjects. Olfactory TDI scores were lower in obese and morbidly obese subjects. Lower TDI scores were independently associated with higher 2-AG fasting plasma circulating concentrations, higher %body fat, and higher body mass index, after controlling for age, smoking, menstruation, and use of contraceptives. Our results show that obese subjects have a lower olfactory capacity than non-obese ones and that elevated fasting plasma circulating 2-AG concentrations in obesity are linked to a lower olfactory capacity. In agreement with previous studies we show that eCBs AEA and 2-AG, and their respective congeners have a distinct profile in relation to body mass index. The present report is the first study in humans in which olfactory capacity and circulating eCB concentrations have been measured in the same subjects.     

Wiring and firing neuronal networks: endocannabinoids take center stage

Endocannabinoids (eCBs) function as retrograde messengers at both excitatory and inhibitory synapses, and control various forms of synaptic plasticity in the adult brain. The molecular machinery required for specific eCB functions during synaptic plasticity is well established. However, eCB signaling plays surprisingly fundamental roles in controlling the acquisition of neuronal identity during CNS development. Recent work suggests that selective recruitment of regulatory signaling networks to CB1 cannabinoid receptors dictates neuronal state-change decisions. In addition, the spatial localization and temporal precision of eCB actions emerges as a novel organizer in developing neuronal networks. Current challenges include fitting novel molecular candidates into regulatory eCB signaling pathways, and defining the temporal dynamics of context-dependent signaling mechanisms underpinning particular neuronal specification events.     

Cannabinoid-mediated short-term plasticity in hippocampus

Endocannabinoids (eCBs) modulate both excitatory and inhibitory neurotransmission in hippocampus via activation of pre-synaptic cannabinoid receptors. Here, we present a model for cannabinoid mediated short-term depression of excitation (DSE) based on our recently developed model for the equivalent phenomenon of suppressing inhibition (DSI). Furthermore, we derive a simplified formulation of the calcium-mediated endocannabinoid synthesis that underlies short-term modulation of neurotransmission in hippocampus. The simplified model describes cannabinoid-mediated short-term modulation of both hippocampal inhibition and excitation and is ideally suited for large network studies. Moreover, the implementation of the simplified DSI/DSE model provides predictions on how both phenomena are modulated by the magnitude of the pre-synaptic cell’s activity. In addition we demonstrate the role of DSE in shaping the post-synaptic cell’s firing behaviour qualitatively and quantitatively in dependence on eCB availability and the pre-synaptic cell’s activity. Finally, we explore under which conditions the combination of DSI and DSE can temporarily shift the fine balance between excitation and inhibition. This highlights a mechanism by which eCBs might act in a neuro-protective manner during high neural activity.     

Exercise training and high-fat diet elicit endocannabinoid system modifications in the rat hypothalamus and hippocampus

The purpose of the present study was to examine the effect of chronic exercise on the hypothalamus and hippocampus levels of the endocannabinoids (eCBs) anandamide (AEA) and 2-arachidonoylglycerol (2-AG) and of two AEA congeners and on the expression of genes coding for CB1, CB2 receptors (Cnr1 and Cnr2, respectively), and the enzymes responsible for eCB biosynthesis and degradation, in rats fed with a standard or high-fat diet. Male Wistar rats (n = 28) were placed on a 12-week high-fat (HFD) or standard diet period, followed by 12 weeks of exercise training for half of each group. Tissue levels of eCBs and related lipids were measured by liquid chromatography mass spectrometry, and expression of genes coding for CB1 and CB2 receptors and eCB metabolic enzymes was measured by quantitative real-time polymerase chain reaction (qPCR). HFD induced a significant increase in 2-AG (p < 0.01) in hypothalamus. High-fat diet paired with exercise training had no effect on AEA, 2-AG, and AEA congener levels in the hypothalamus and hippocampus. Cnr1 expression levels were significantly increased in the hippocampus in response to HFD, exercise, and the combination of both (p < 0.05). Our results indicate that eCB signaling in the CNS is sensitive to diet and/or exercise.     

Dual inhibition of alpha/beta-hydrolase domain 6 and fatty acid amide hydrolase increases endocannabinoid levels in neurons

Agonists at cannabinoid receptors, such as the phytocannabinoid Δ(9)-tetrahydrocannabinol, exert a remarkable array of therapeutic effects but are also associated with undesirable psychoactive side effects. Conversely, targeting enzymes that hydrolyze endocannabinoids (eCBs) allows for more precise fine-tuning of cannabinoid receptor signaling, thus providing therapeutic relief with reduced side effects. Here, we report the development and characterization of an inhibitor of eCB hydrolysis, UCM710, which augments both N-arachidonoylethanolamine and 2-arachidonoylglycerol levels in neurons. This compound displays a unique pharmacological profile in that it inhibits fatty acid amide hydrolase and α/β-hydrolase domain 6 but not monoacylglycerol lipase. Thus, UCM710 represents a novel tool to delineate the therapeutic potential of compounds that manipulate a subset of enzymes that control eCB signaling.     

Inhibition of the adrenergic phenotype in cultured neural crest cells by norepinephrine uptake inhibitors

Tricyclic antidepressants in combination with in vitro clonal analysis of quail neural crest cells were used to examine the role the norepinephrine uptake mechanism might play in the development of adrenergic neural crest derivatives. Norepinephrine (NE) uptake inhibitors blocked expression of the adrenergic phenotype by neural crest cells. The degree of inhibition of phenotypic expression correlated with the potency and specificity of the uptake inhibitors. The drugs acted during the early phase of in vitro development, i.e., several days before overt expression of the adrenergic phenotype in clonal culture. They were nontoxic, and a chronic exposure of the cells to NE uptake inhibitors was necessary to cause an effect. These observations suggest that norepinephrine and possibly related neurotransmitters play a direct or indirect role in the expression of the adrenergic phenotype by neural crest cells and that tricyclic antidepressants may affect neurogenesis during sensitive stages of embryonic development. The data may reflect in vivo mechanisms, since there are neurotransmitters present in the migratory pathway of presumptive sympathetic neurons and the norepinephrine uptake system is expressed in the embryo by these cells before they synthesize and accumulate catecholamines.     

Wnt/β‐catenin signalling pathway mediated aberrant hippocampal neurogenesis in kainic acid‐induced epilepsy

Temporal lobe epilepsy is a chronic disorder of nerve system, mainly characterized by hippocampal sclerosis with massive neuronal loss and severe gliosis. Aberrant neurogenesis has been shown in the epileptogenesis process of temporal lobe epilepsy. However, the molecular mechanisms underlying aberrant neurogenesis remain unclear. The roles of Wnt signalling cascade have been well established in neurogenesis during multiple aspects. Here, we used kainic acid‐induced rat epilepsy model to investigate whether Wnt/β‐catenin signalling pathway is involved in the aberrant neurogenesis in temporal lobe epilepsy. Immunostaining and western blotting results showed that the expression levels of β‐catenin, Wnt3a, and cyclin D1, the key regulators in Wnt signalling pathway, were up‐regulated during acute epilepsy induced by the injection of kainic acids, indicating that Wnt signalling pathway was activated in kainic acid‐induced temporal lobe epilepsy. Moreover, BrdU labelling results showed that blockade of the Wnt signalling by knocking down β‐catenin attenuated aberrant neurogenesis induced by kainic acids injection. Altogether, Wnt/β‐catenin signalling pathway mediated hippocampal neurogenesis during epilepsy, which might provide new strategies for clinical treatment of temporal lobe epilepsy. Temporal lobe epilepsy is a chronic disorder of nerve system, mainly characterized by hippocampal sclerosis. Aberrant neurogenesis has been shown to involve in the epileptogenesis process of temporal lobe epilepsy. In the present study, we discovered that Wnt3a/β‐catenin signalling pathway serves as a link between aberrant neurogenesis and underlying remodelling in the hippocampus, leading to temporal lobe epilepsy, which might provide new strategies for clinical treatment of temporal lobe epilepsy.     

U-46619 but not serotonin increases endocannabinoid content in middle cerebral artery: evidence for functional relevance

Cerebral vascular smooth muscle cells express the CB(1) cannabinoid receptor, and CB(1) receptor agonists produce vasodilation of cerebral arteries. The purpose of this study was to determine whether vasoconstriction of rat middle cerebral artery (MCA) results in the local formation of endocannabinoids (eCBs), which, via activation of CB(1) receptors, oppose the vasoconstriction in a feedback manner. The thromboxane A(2) (TXA(2)) mimetic U-46619 significantly increased N-arachidonylethanolamine (AEA) and 2-arachidonylglycerol (2-AG) content of isolated MCA, whereas 5-hydroxytrypamine (5-HT) decreased AEA and 2-AG content. If eCBs play a feedback role in the regulation of MCA tone, then CB(1) receptor antagonists should enhance the constriction of MCA produced by U-46619 but not 5-HT. U-46619 caused concentration-dependent constrictions of endothelium-denuded MCA. Two CB(1) receptor antagonists SR-141716 and AM-251 decreased the EC(50) value for U-46619 to constrict endothelium-denuded MCA without affecting the maximal effect. A low concentration of CB(1) receptor agonist Win-55212-2 (30 nM) produced vasodilation of MCAs constricted with low but not saturating concentrations of U-46619. SR-141716 had no effect on the 5-HT concentration-contraction relationship. These data suggest that TXA(2) receptor activation increases MCA eCB content, which, via activation of CB(1) receptors, reduces the constriction produced by moderate concentrations of the TXA(2) agonist. Although 5-HT-induced vasoconstriction is reduced by exogenous CB(1) receptor agonist, activation of 5-HT receptors does not increase eCB content. These results suggest that MCA production of eCBs is not regulated by constriction per se but likely via a signaling pathway that is specific for TXA(2) receptors and not 5-HT receptors.     

Sterol Carrier Protein-2: Binding Protein for Endocannabinoids

The endocannabinoid (eCB) system, consisting of eCB ligands and the type 1 cannabinoid receptor (CB1R), subserves retrograde, activity-dependent synaptic plasticity in the brain. eCB signaling occurs “on-demand,” thus the processes regulating synthesis, mobilization and degradation of eCBs are also primary mechanisms for the regulation of CB1R activity. The eCBs, N-arachidonylethanolamine (AEA) and 2-arachidonoylglycerol (2-AG), are poorly soluble in water. We hypothesize that their aqueous solubility, and, therefore, their intracellular and transcellular distribution, are facilitated by protein binding. Using in silico docking studies, we have identified the nonspecific lipid binding protein, sterol carrier protein 2 (SCP-2), as a potential AEA binding protein. The docking studies predict that AEA and AM404 associate with SCP-2 at a putative cholesterol binding pocket with ?G values of ?3.6 and ?4.6 kcal/mol, respectively. These values are considerably higher than cholesterol (?6.62 kcal/mol) but consistent with a favorable binding interaction. In support of the docking studies, SCP-2-mediated transfer of cholesterol in vitro is inhibited by micromolar concentrations of AEA; and heterologous expression of SCP-2 in HEK 293 cells increases time-related accumulation of AEA in a temperature-dependent fashion. These results suggest that SCP-2 facilitates cellular uptake of AEA. However, there is no effect of SCP-2 transfection on the cellular accumulation of AEA determined at equilibrium or the IC50 values for AEA, AM404 or 2-AG to inhibit steady state accumulation of radiolabelled AEA. We conclude that SCP-2 is a low affinity binding protein for AEA that can facilitate its cellular uptake but does not contribute significantly to intracellular sequestration of AEA.     

Nuclear diacylglycerol lipase‐α in rat brain cortical neurons: evidence of 2‐arachidonoylglycerol production in concert with phospholipase C‐β activity

In this report, we describe the localization of diacylglycerol lipase‐α (DAGLα) in nuclei from adult cortical neurons, as assessed by double‐immunofluorescence staining of rat brain cortical sections and purified intact nuclei and by western blot analysis of subnuclear fractions. Double‐labeling assays using the anti‐DAGLα antibody and NeuN combined with Hoechst staining showed that only nuclei of neuronal origin were DAGLα positive. At high resolution, DAGLα‐signal displayed a punctate pattern in nuclear subdomains poor in Hoechst's chromatin and lamin B1 staining. In contrast, SC‐35‐ and NeuN‐signals (markers of the nuclear speckles) showed a high overlap with DAGLα within specific subdomains of the nuclear matrix. Among the members of the phospholipase C‐β (PLCβ) family, PLCβ1, PLCβ2, and PLCβ4 exhibited the same distribution with respect to chromatin, lamin B1, SC‐35, and NeuN as that described for DAGLα. Furthermore, by quantifying the basal levels of 2‐arachidonoylglycerol (2‐AG) by liquid chromatography and mass spectrometry (LC‐MS), and by characterizing the pharmacology of its accumulation, we describe the presence of a mechanism for 2‐AG production, and its PLCβ/DAGLα‐dependent biosynthesis in isolated nuclei. These results extend our knowledge about subcellular distribution of neuronal DAGLα, providing biochemical grounds to hypothesize a role for 2‐AG locally produced within the neuronal nucleus.

     

Impaired cognitive function and reduced anxiety‐related behavior in a promyelocytic leukemia (PML) tumor suppressor protein‐deficient mouse

The promyelocytic leukemia (PML) protein is a tumor suppressor factor mostly known by its involvement in acute promyelocytic leukemia (APL). Interestingly, recent studies have provided evidence that, in the central nervous system, PML is involved in neurogenesis. However, prospective studies of PML in brain are lacking. To further understand the role of PML in the mammalian brain, we studied plasticity and behavioral changes in PML knockout mice. If PML is involved in neurogenesis, and neurogenesis is an important process for proper brain development as well as learning and memory functions, we hypothesized that PML might have a role in plasticity and cognition. Behavioral studies demonstrated that PML knockout mice present abnormalities in conditioned learning and spatial memory, as determined by fear conditioning and Morris water maze tasks. Experiments to determine normal exploratory behavior interestingly revealed that PML knockout mice present reduced anxiety‐related responses as compared to control animals. This was confirmed when PML knockout mice spent more time in the open arms of an elevated plus‐maze, which is an indication of decreased anxiety. Additionally, impairments in hippocampus‐dependent learning were mirrored by altered long‐term plasticity at Schaffer collateral‐CA1 synapses. We now provide the first evidence for an important role of PML in the brain, indicating that PML might have a role in synaptic plasticity and associated behavioral processes.     

Progressive effects of N‐myc deficiency on proliferation,neurogenesis, and morphogenesis in the olfactory epithelium

N‐myc belongs to the myc proto‐oncogene family, which is involved in numerous cellular processes such as proliferation, growth, apoptosis, and differentiation. Conditional deletion of N‐myc in the mouse nervous system disrupted brain development, indicating that N‐myc plays an essential role during neural development. How the development of the olfactory epithelium and neurogenesis within are affected by the loss of N‐myc has, however, not been determined. To address these issues, we examined an N‐myc^Foxg1Cre conditional mouse line, in which N‐myc is depleted in the olfactory epithelium. First changes in N‐myc mutants were detected at E11.5, with reduced proliferation and neurogenesis in a slightly smaller olfactory epithelium. The phenotype was more pronounced at E13.5, with a complete lack of Hes5‐positive progenitor cells, decreased proliferation, and neurogenesis. In addition, stereological analyses revealed reduced cell size of post‐mitotic neurons in the olfactory epithelium, which contributed to a smaller olfactory pit. Furthermore, we observed diminished proliferation and neurogenesis also in the vomeronasal organ, which likewise was reduced in size. In addition, the generation of gonadotropin‐releasing hormone neurons was severely reduced in N‐myc mutants. Thus, diminished neurogenesis and proliferation in combination with smaller neurons might explain the morphological defects in the N‐myc depleted olfactory structures. Moreover, our results suggest an important role for N‐myc in regulating ongoing neurogenesis, in part by maintaining the Hes5‐positive progenitor pool. In summary, our results provide evidence that N‐myc deficiency in the olfactory epithelium progressively diminishes proliferation and neurogenesis with negative consequences at structural and cellular levels. © 2013 The Authors. Developmental Neurobiology Published by Wiley Periodicals, Inc. Develop Neurobiol 74: 643–656, 2014