An ex vivo preparation of the intact mouse vomeronasal organ and accessory olfactory bulb

Department of Anatomy and Neurobiology, Washington University School of Medicine, Campus Box 8108, 660 S. Euclid Ave, Saint Louis, MO 63110, United States
Journal of Neuroscience Methods (Impact Factor: 1.96). 03/2009; DOI: 10.1016/j.jneumeth.2008.11.013

ABSTRACT The accessory olfactory system (AOS) in mammals detects and processes information from liquid-phase environmental odorants, including pheromones. The AOS carries out tasks such as individual recognition, learning, and decision-making with relatively few stages of neural processing; it thus represents an attractive system for investigating the neural circuits that carry out these functions. Progress in understanding the AOS has long been impeded by its relative inaccessibility to standard physiological approaches. In this report, we detail a novel dissection and tissue perfusion strategy that improves access to the accessory olfactory bulb (AOB) while maintaining afferent connections from sensory neurons in the vomeronasal organ (VNO). Mitral cells demonstrated spontaneous and evoked firing patterns consistent with recent in vivo reports. We assayed cell degradation in the AOB tissue using Fluoro-Jade C and found that the VNO and AOB glomerular, external plexiform, and mitral cell layers showed minimal signs of degeneration for up to 6 h. Whereas histology indicated some degeneration in the deep inhibitory granule cell layer over time, electrophysiological assays demonstrated intact inhibitory function on mitral cells. Pharmacological blockade of GABAA receptors with 3 M SR95531 (gabazine) resulted in increased evoked mitral cell activity. Furthermore, mitral cells displayed suppression of responses to preferred urine stimuli when preferred and non-preferred stimuli were mixed, an effect thought to involve functional laterally connected inhibition. These results demonstrate the utility of whole mount ex vivo preparations for studying sensory processing in the AOS, and suggest that similar strategies may improve experimental access to other difficult-to-study neural circuits.

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    ABSTRACT: The mammalian accessory olfactory system extracts information about species, sex and individual identity from social odors, but its functional organization remains unclear. We imaged presynaptic Ca(2+) signals in vomeronasal inputs to the accessory olfactory bulb (AOB) during peripheral stimulation using light sheet microscopy. Urine- and steroid-responsive glomeruli densely innervated the anterior AOB. Glomerular activity maps for sexually mature female mouse urine overlapped maps for juvenile and/or gonadectomized urine of both sexes, whereas maps for sexually mature male urine were highly distinct. Further spatial analysis revealed a complicated organization involving selective juxtaposition and dispersal of functionally grouped glomerular classes. Glomeruli that were similarly tuned to urines were often closely associated, whereas more disparately tuned glomeruli were selectively dispersed. Maps to a panel of sulfated steroid odorants identified tightly juxtaposed groups that were disparately tuned and dispersed groups that were similarly tuned. These results reveal a modular, nonchemotopic spatial organization in the AOB.
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    ABSTRACT: In mice, nonvolatile social cues are detected and analyzed by the accessory olfactory system (AOS). Here we provide a first view of information processing in the AOS with respect to individual chemical cues. 12 sulfated steroids, recently discovered mouse AOS ligands, caused widespread activity among vomeronasal sensory neurons (VSNs), yet VSN responses clustered into a small number of repeated functional patterns or processing streams. Downstream neurons in the accessory olfactory bulb (AOB) responded to these ligands with enhanced signal/noise compared to VSNs. Although the dendritic connectivity of AOB mitral cells suggests the capacity for broad integration, most sulfated steroid responses were well-modeled by linear excitatory drive from just one VSN processing stream. However, a substantial minority demonstrated multi-stream integration. Most VSN excitation patterns were also observed in the AOB, but excitation by estradiol sulfate processing streams was rare, suggesting AOB circuit organization is specific to the biological relevance of sensed cues.
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