Airway branching has conserved needs for local parasympathetic innervation but not neurotransmission

Danielle V Bower,Kai Zinn,Scott E Fraser,Rusty Lansford,Edwin C Jesudason,David Warburton,Hyung-Kook Lee

doi:10.1186/preaccept-1492846834139911

Abstract

BackgroundParasympathetic signaling has been inferred to regulate epithelial branching as well as organ regeneration and tumor development. However, the relative contribution of local nerve contact versus secreted signals remains unclear. Here, we show a conserved (vertebrates to invertebrates) requirement for intact local nerves in airway branching, persisting even when cholinergic neurotransmission is blocked.ResultsIn the vertebrate lung, deleting enhanced green fluorescent protein (eGFP)-labeled intrinsic neurons using a two-photon laser leaves adjacent cells intact, but abolishes branching. Branching is unaffected by similar laser power delivered to the immediately adjacent non-neural mesodermal tissue, by blocking cholinergic receptors or by blocking synaptic transmission with botulinum toxin A. Because adjacent vasculature and epithelial proliferation also contribute to branching in the vertebrate lung, the direct dependence on nerves for airway branching was tested by deleting neurons in Drosophila embryos. A specific deletion of neurons in the Drosophila embryo by driving cell-autonomous RicinA under the pan-neuronal elav enhancer perturbed Drosophila airway development. This system confirmed that even in the absence of a vasculature or epithelial proliferation, airway branching is still disrupted by neural lesioning.ConclusionsTogether, this shows that airway morphogenesis requires local innervation in vertebrates and invertebrates, yet neurotransmission is dispensable. The need for innervation persists in the fly, wherein adjacent vasculature and epithelial proliferation are absent. Our novel, targeted laser ablation technique permitted the local function of parasympathetic innervation to be distinguished from neurotransmission.

Highlights

Parasympathetic signaling has been inferred to regulate epithelial branching as well as organ regeneration and tumor development
Avoiding the off-target effects of wholesale ganglion excision, we developed a novel technique for highly precise laser denervation of the lung to show that this halts branching and reduces both epithelial and endothelial proliferation, but in a manner that cannot be reproduced by parasympathetic neurotransmitter blockade
Contrary to reports in the salivary gland, we found that neural regulation of airway morphogenesis is not mediated by cholinergic neurotransmitter discharge at this phase of development during the early ramification of the bronchial tree

Summary

Introduction

Parasympathetic signaling has been inferred to regulate epithelial branching as well as organ regeneration and tumor development. Parasympathetic (cholinergic) signaling is reported to regulate morphogenesis and organ regeneration via effects on epithelial stem cells [1,2]. The neurogenic regulation of tissue biology, To date, these wide-reaching roles of neural regulation of epithelial biology have been attributed to neurotransmitter signaling. It is unclear whether the nerves serve other purposes, such as provision of important structural cues and secreted factors. Distinguishing a role for direct interactions between nerves and epithelium versus neurotransmitter-mediated signaling or intermediary guidance via the vasculature is important in understanding the mechanism of neural regulation of epithelial tissue morphogenesis

Methods

Results

Discussion

Conclusion