A Phox2b BAC Transgenic Rat Line Useful for Understanding Respiratory Rhythm Generator Neural Circuitry.

Keiko Ikeda,Hiroshi Onimaru,Satoru Arata,Toru Ishizuka,Hiromu Yawo,Masanori Takahashi,Shigeru Sato,E Michelle Southard-Smith,Hiroyuki Igarashi,Kiyoshi Kawakami,Branden Nelson

doi:10.1371/journal.pone.0132475

Abstract

The key role of the respiratory neural center is respiratory rhythm generation to maintain homeostasis through the control of arterial blood pCO2/pH and pO2 levels. The neuronal network responsible for respiratory rhythm generation in neonatal rat resides in the ventral side of the medulla and is composed of two groups; the parafacial respiratory group (pFRG) and the pre-Bötzinger complex group (preBötC). The pFRG partially overlaps in the retrotrapezoid nucleus (RTN), which was originally identified in adult cats and rats. Part of the pre-inspiratory (Pre-I) neurons in the RTN/pFRG serves as central chemoreceptor neurons and the CO2 sensitive Pre-I neurons express homeobox gene Phox2b. Phox2b encodes a transcription factor and is essential for the development of the sensory-motor visceral circuits. Mutations in human PHOX2B cause congenital hypoventilation syndrome, which is characterized by blunted ventilatory response to hypercapnia. Here we describe the generation of a novel transgenic (Tg) rat harboring fluorescently labeled Pre-I neurons in the RTN/pFRG. In addition, the Tg rat showed fluorescent signals in autonomic enteric neurons and carotid bodies. Because the Tg rat expresses inducible Cre recombinase in PHOX2B-positive cells during development, it is a potentially powerful tool for dissecting the entire picture of the respiratory neural network during development and for identifying the CO2/O2 sensor molecules in the adult central and peripheral nervous systems.

Highlights

Respiration is an important function in animals and controlled by two main respiratory rhythm generators, the parafacial respiratory group and the pre-Bötzinger complex inspiratory group, which reside in the ventral side of the medulla in rodents [1,2,3,4]
The clinical symptoms of disease caused by mutations in the coding sequence of human PHOX2B are consistent with the expression patterns observed in rodents [31]
Since evaluation of the RP24-95M11 bacterial artificial chromosome (BAC) clone could be useful for driving expression in transgenic rats, we compared the evolutionarily conserved non-coding sequences (CNSs) surrounding the paired-like homeobox 2b gene (Phox2b) exons in mammals, together with related species, such as painted turtle, chicken, Xenopus, coelacanth, spotted gar, and zebrafish using mVISTA (Fig 1)

Summary

Introduction

Respiration is an important function in animals and controlled by two main respiratory rhythm generators, the parafacial respiratory group (pFRG) and the pre-Bötzinger complex inspiratory group (preBötC), which reside in the ventral side of the medulla in rodents [1,2,3,4]. The ventral part of the pFRG was identified electrophysiologically in a region ventral to the facial nucleus (nVII) and close to the ventral surface in neonatal rats [4] This part of the pFRG overlaps with the retrotrapezoid nucleus (RTN), the latter was first identified in the adult cat by retrograde tracing analysis as a candidate site for respiratory rhythmogenesis and later in the adult rat [9,10,11,12,13,14]. Unlike the RTN/pFRG, neurons of the preBötC do not express Phox2b (Ikeda and Onimaru, unpublished results)

Methods

Results

Conclusion