Genetic identification of an embryonic parafacial oscillator coupling to the preBötzinger complex

Abstract

The hindbrain transcription factors Phox2b and Egr2 (also known as Krox20) are linked to the development of the autonomic nervous system and rhombomere-related regulation of breathing, respectively. Mutations in these proteins can lead to abnormal breathing behavior as a result of an alteration in an unidentified neuronal system. We characterized a bilateral embryonic parafacial (e-pF) population of rhythmically bursting neurons at embryonic day (E) 14.5 in mice. These cells expressed Phox2b, were derived from Egr2-expressing precursors and their development was dependent on the integrity of the Egr2 gene. Silencing or eliminating the e-pF oscillator, but not the putative inspiratory oscillator (preBötzinger complex, preBötC), led to an abnormally slow rhythm, demonstrating that the e-pF controls the respiratory rhythm. The e-pF oscillator, the only one active at E14.5, entrained and then coupled with the preBötC, which emerged independently at E15.5. These data establish the dual organization of the respiratory rhythm generator at the time of its inception, when it begins to drive fetal breathing.

Defective Respiratory Rhythmogenesis and Loss of Central Chemosensitivity in Phox2b Mutants Targeting Retrotrapezoid Nucleus Neurons

“…By genetic tracing, partnering either the Lbx1 cre or the Egr2 cre allele with the Tau GFPnLacZ reporter, we first confirmed published evidence (Pagliardini et al, 2008;Thoby-Brisson et al, 2009) that the majority of RTN neurons arise from the Lbx1 cre -expressing domain of the alar plate (Sieber et al, 2007), in the Egr2 creexpressing r3 or r5 (Voiculescu et al, 2001) (Fig. 3A-J (Fig.…”

“…Moreover, a population of neurons termed e-pF for embryonic parafacial neurons that display respiratorylike rhythmic activity are already detected at E14.5 in the parafacial region. They are intrinsically rhythmic, express NK1R and Phox2b (Thoby-Brisson et al, 2009), and as shown here, respond to a pH challenge, as do pFRG neurons in the newborn rat (Onimaru et al, 2008). They thus likely represent the embryonic forerunner of the pFRG.…”

“…We will thus use the term RTN neurons to refer to the neurons located ventral and immediately lateral and caudal to nVII that are Phox2b ϩ ;Vglut2 ϩ , NK1R ϩ , and TH-negative, adding the criterion of Atoh1 expression (see below). Their precise electrophysiological properties, however, appear to change between fetal, neonatal, and adult stages (Mulkey et al, 2004;Stornetta et al, 2006;Onimaru et al, 2008;Thoby-Brisson et al, 2009), and we will use the terms e-pF or pFRG when referring explicitly to the electrophysiological properties of these neurons in fetal and newborn rodents.…”

Neural mechanisms underlying respiratory rhythm generation in the lamprey

“…the hypothesized rostral expiratory oscillator of mammals (see e.g. Onimaru et al, 2009;Thoby-Brisson et al, 2009;Guyenet and Mulkey, 2010;Feldman et al, 2013;Smith et al, 2013). This oscillator may display burst activity involving endogenous I NaP -dependent properties, as it occurs in the preBötC ThobyBrisson et al, 2009;Molkov et al, 2010).…”

“…This oscillator may display burst activity involving endogenous I NaP -dependent properties, as it occurs in the preBötC ThobyBrisson et al, 2009;Molkov et al, 2010). In addition, it contains glutamatergic neurons that express NK1 receptors, but it is not sensitive to opioids (Mulkey et al, 2004;Onimaru et al, 2008;Takakura et al, 2008;Lazarenko et al, 2009;Thoby-Brisson et al, 2009; for reviews, see Feldman et al, 2013;Guyenet and Mulkey, 2010). In conclusion, similarly to other neurophysiological features (Ericsson et al, , 2013Stephenson-Jones et al, 2011, 2012a, 2012b, the basic oscillatory and neuromodulatory mechanisms of the respiratory network seem to be highly evolutionary conserved regardless of their location and their inspiratory or expiratory function.…”