Functional confocal live cell imaging of the pulmonary neuroepithelial body microenvironment in GAD67‐GFP mice

Abstract

As integrated components of the airway epithelium, pulmonary neuroepithelial bodies (NEBs), composed of clusters of neuroendocrine cells, are ideally located to sense changes in the airway environment. Connections of NEBs with different populations of mainly vagal sensory nerve terminals allow fast transduction of information to the CNS. NEBs are dispersed in the airway epithelium and inaccessible in vivo. The selective identification of NEBs is a prerequisite to enable functional molecular live cell imaging in our ex vivo lung slice model, but the vital dye 4‐Di‐2‐ASP that we used so far has been criticized. In this study the lung slice model was further strengthened using knock‐in mice that express GFP driven by the promoter of glutamate decarboxylase 67 (GAD67), the rate‐limiting enzyme of GABA synthesis. Highly selective expression of GFP in NEB cells in mouse lungs allows unequivocal selection. Since ex vivo vibratome slices of GAD67‐GFP mouse lungs could be loaded with red‐fluorescent functional probes, and proof‐of‐concept experiments were successful, this model will certainly boost future functional studies of NEBs in health and disease. Simultaneously the model enables to explore the significance of a GABAergic signaling system in the NEB microenvironment.Support: IWT fellowship SB/81162 (RL); FWO grants G.0081.08 (DA) and G.0589.11N (DA, JPT); UA grants GOA BOF 2007 (DA) and KP BOF 2006 (IB)