RESUMEN
During development of the nervous system, neurons connect to one another in a precisely organized manner. Sensory systems provide a good example of this organization, whereby the composition of the outside world is represented in the brain by neuronal maps. Establishing correct patterns of neural circuitry is crucial, as inaccurate map formation can lead to severe disruptions in sensory processing. In rodents, olfactory stimuli modulate a wide variety of behaviors essential for survival. The formation of the olfactory glomerular map is dependent on molecular cues that guide olfactory receptor neuron axons to broad regions of the olfactory bulb and on cell adhesion molecules that promote axonal sorting into specific synaptic units in this structure. Here, we demonstrate that the cell adhesion molecule Amigo1 is expressed in a subpopulation of olfactory receptor neurons, and we investigate its role in the precise targeting of olfactory receptor neuron axons to the olfactory bulb using a genetic loss-of-function approach in mice. While ablation of Amigo1 did not lead to alterations in olfactory sensory neuron axonal targeting, our experiments revealed that the presence of a neomycin resistance selection cassette in the Amigo1 locus can lead to off-target effects that are not due to loss of Amigo1 expression, including unexpected altered gene expression in olfactory receptor neurons and reduced glomerular size in the ventral region of the olfactory bulb. Our results demonstrate that insertion of a neomycin selection cassette into the mouse genome can have specific deleterious effects on the development of the olfactory system and highlight the importance of removing antibiotic resistance cassettes from genetic loss-of-function mouse models when studying olfactory system development.
Asunto(s)
Neuronas Receptoras Olfatorias , Animales , Ratones , Neuronas Receptoras Olfatorias/metabolismo , Mucosa Olfatoria , Bulbo Olfatorio , Axones/metabolismo , Expresión GénicaRESUMEN
Biomineralization is a crucial process whereby organisms produce mineralized tissues such as teeth for mastication, bones for support, and shells for protection. Mineralized tissues are composed of hierarchically organized hydroxyapatite crystals, with a limited capacity to regenerate when demineralized or damaged past a critical size. Thus, the development of protein-based materials that act as artificial scaffolds to guide hydroxyapatite growth is an attractive goal both for the design of ordered nanomaterials and for tissue regeneration. In particular, amelogenin, which is the main protein that scaffolds the hierarchical organization of hydroxyapatite crystals in enamel, amelogenin recombinamers, and amelogenin-derived peptide scaffolds have all been investigated for in vitro mineral growth. Here, we describe uniaxial hydroxyapatite growth on a nanoengineered amelogenin scaffold in combination with amelotin, a mineral promoting protein present during enamel formation. This bio-inspired approach for hydroxyapatite growth may inform the molecular mechanism of hydroxyapatite formation in vitro as well as possible mechanisms at play during mineralized tissue formation.
Asunto(s)
Amelogenina/química , Materiales Biomiméticos/química , Biomineralización/genética , Proteínas del Esmalte Dental/química , Durapatita/química , Nanoestructuras/química , Amelogenina/genética , Biomimética/métodos , Cristalización , Esmalte Dental/química , Proteínas del Esmalte Dental/genética , Humanos , Nanotecnología/métodos , Ingeniería de Proteínas/métodos , Pliegue de Proteína , Proteínas Recombinantes/química , Diente/químicaRESUMEN
The formation of olfactory maps in the olfactory bulb (OB) is crucial for the control of innate and learned mouse behaviors. Olfactory sensory neurons (OSNs) expressing a specific odorant receptor project axons into spatially conserved glomeruli within the OB and synapse onto mitral cell dendrites. Combinatorial expression of members of the Kirrel family of cell adhesion molecules has been proposed to regulate OSN axonal coalescence; however, loss-of-function experiments have yet to establish their requirement in this process. We examined projections of several OSN populations in mice that lacked either Kirrel2 alone, or both Kirrel2 and Kirrel3. Our results show that Kirrel2 and Kirrel3 are dispensable for the coalescence of MOR1-3-expressing OSN axons to the most dorsal region (DI) of the OB. In contrast, loss of Kirrel2 caused MOR174-9- and M72-expressing OSN axons, projecting to the DII region, to target ectopic glomeruli. Our loss-of-function approach demonstrates that Kirrel2 is required for axonal coalescence in subsets of OSNs that project axons to the DII region and reveals that Kirrel2/3-independent mechanisms also control OSN axonal coalescence in certain regions of the OB.
Asunto(s)
Axones/fisiología , Inmunoglobulinas/fisiología , Proteínas de la Membrana/fisiología , Bulbo Olfatorio/metabolismo , Neuronas Receptoras Olfatorias/metabolismo , Sinapsis/genética , Animales , Femenino , Regulación del Desarrollo de la Expresión Génica , Inmunoglobulinas/genética , Masculino , Proteínas de la Membrana/genética , Ratones , Ratones Transgénicos , Bulbo Olfatorio/citología , Bulbo Olfatorio/fisiología , Neuronas Receptoras Olfatorias/citología , Neuronas Receptoras Olfatorias/fisiología , Sinapsis/metabolismo , Transmisión Sináptica/genéticaRESUMEN
The accessory olfactory system controls social and sexual behaviours in mice, both of which are critical for their survival. Vomeronasal sensory neuron (VSN) axons form synapses with mitral cell dendrites in glomeruli of the accessory olfactory bulb (AOB). Axons of VSNs expressing the same vomeronasal receptor (VR) converge into multiple glomeruli within spatially conserved regions of the AOB. Here, we have examined the role of the cell adhesion molecule Kirrel2 in the formation of glomeruli within the AOB. We find that Kirrel2 expression is dispensable for early axonal guidance events, such as fasciculation of the vomeronasal tract and segregation of apical and basal VSN axons into the anterior and posterior regions of the AOB, but is necessary for glomeruli formation. Specific ablation of Kirrel2 expression in VSN axons results in the disorganization of the glomerular layer of the posterior AOB and in the formation of fewer and larger glomeruli. Furthermore, simultaneous ablation of Kirrel2 and Kirrel3 expression leads to a loss of morphologically identifiable glomeruli in the AOB, reduced excitatory synapse numbers, and larger presynaptic terminals. Taken together, our results demonstrate that Kirrel2 and Kirrel3 are essential for the formation of glomeruli and suggest they contribute to synaptogenesis in the AOB.