RESUMO
Operant conditioning is a crucial tool in neuroscience research for probing brain function. While molecular, anatomical and even physiological techniques have seen radical increases in throughput, efficiency, and reproducibility in recent years, behavioural tools have somewhat lagged behind. Here we present a fully automated, high-throughput system for self-initiated conditioning of up to 25 group-housed, radio-frequency identification (RFID) tagged mice over periods of several months and >106 trials. We validate this "AutonoMouse" system in a series of olfactory behavioural tasks and show that acquired data is comparable to previous semi-manual approaches. Furthermore, we use AutonoMouse to systematically probe the impact of graded olfactory bulb lesions on olfactory behaviour, demonstrating that while odour discrimination in general is robust to even most extensive disruptions, small olfactory bulb lesions already impair odour detection. Discrimination learning of similar mixtures as well as learning speed are in turn reliably impacted by medium lesion sizes. The modular nature and open-source design of AutonoMouse should allow for similar robust and systematic assessments across neuroscience research areas.
Assuntos
Condicionamento Operante/fisiologia , Aprendizagem por Discriminação/fisiologia , Bulbo Olfatório/fisiologia , Olfato/fisiologia , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Odorantes , Reprodutibilidade dos TestesRESUMO
The raphe nuclei provide serotonergic innervation widely in the brain, thought to mediate a variety of neuromodulatory effects. The mammalian olfactory bulb (OB) is a prominent recipient of serotonergic fibers, particularly in the glomerular layer (GL), where they are thought to gate incoming signals from the olfactory nerve. The dorsal raphe nucleus (DRN) and the median raphe nucleus (MRN) are known to densely innervate the OB. The majority of such projections are thought to terminate in the GL, but this has not been explicitly tested. We sought to investigate this using recombinant adeno-associated viruses (rAAV)-mediated expression of green fluorescent protein (GFP)-synaptophysin targeted specifically to neurons of the DRN or the MRN. With DRN injections, labeled fibers were found mostly in the granule cell layer (GCL), not the GL. Conversely, dense labeling in the GL was observed with MRN injections, suggesting that the source of GL innervation is the MRN, not the DRN, as previously thought. The two raphe nuclei thus give dual innervation within the OB, with distinct innervation patterns.
Assuntos
Bulbo Olfatório/anatomia & histologia , Núcleos da Rafe/anatomia & histologia , Animais , Dependovirus/genética , Vetores Genéticos , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Camundongos Endogâmicos C57BL , Microscopia Confocal , Vias Neurais/anatomia & histologia , Vias Neurais/metabolismo , Técnicas de Rastreamento Neuroanatômico , Bulbo Olfatório/metabolismo , Núcleos da Rafe/metabolismo , Neurônios Serotoninérgicos/citologia , Neurônios Serotoninérgicos/metabolismo , Serotonina/metabolismo , Sinaptofisina/genética , Sinaptofisina/metabolismoRESUMO
Circuits in the brain possess the ability to orchestrate activities on different timescales, but the manner in which distinct circuits interact to sculpt diverse rhythms remains unresolved. The olfactory bulb is a classic example of a place in which slow theta and fast gamma rhythms coexist. Furthermore, inhibitory interneurons that are generally implicated in rhythm generation are segregated into distinct layers, neatly separating local and global motifs. We combined intracellular recordings in vivo with circuit-specific optogenetic interference to examine the contribution of inhibition to rhythmic activity in the mouse olfactory bulb. We found that the two inhibitory circuits controlled rhythms on distinct timescales: local, glomerular networks coordinated theta activity, regulating baseline and odor-evoked inhibition, whereas granule cells orchestrated gamma synchrony and spike timing. Notably, granule cells did not contribute to baseline rhythms or sniff-coupled odor-evoked inhibition. Thus, activities on theta and gamma timescales are controlled by separate, dissociable inhibitory networks in the olfactory bulb.
Assuntos
Ritmo Gama/fisiologia , Interneurônios/fisiologia , Bulbo Olfatório/citologia , Bulbo Olfatório/fisiologia , Ritmo Teta/fisiologia , Animais , Potenciais Evocados/fisiologia , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Inibição Neural/fisiologia , Odorantes , Optogenética , Técnicas de Patch-Clamp , Olfato/fisiologia , VigíliaRESUMO
Multiphoton imaging (MPI) is widely used for recording activity simultaneously from many neurons in superficial cortical layers in vivo. We combined regenerative amplification multiphoton microscopy (RAMM) with genetically encoded calcium indicators to extend MPI of neuronal population activity into layer 5 (L5) of adult mouse somatosensory cortex. We found that this approach could be used to record and quantify spontaneous and sensory-evoked activity in populations of L5 neuronal somata located as much as 800 µm below the pia. In addition, we found that RAMM could be used to simultaneously image activity from large (80) populations of apical dendrites and follow these dendrites down to their somata of origin.