Development of pacemaker properties and rhythmogenic mechanisms in the mouse embryonic respiratory network.

Marc Chevalier,Natalia Toporikova,Muriel Thoby-Brisson,John Simmers

doi:10.7554/elife.16125

Abstract

Breathing is a vital rhythmic behavior generated by hindbrain neuronal circuitry, including the preBötzinger complex network (preBötC) that controls inspiration. The emergence of preBötC network activity during prenatal development has been described, but little is known regarding inspiratory neurons expressing pacemaker properties at embryonic stages. Here, we combined calcium imaging and electrophysiological recordings in mouse embryo brainstem slices together with computational modeling to reveal the existence of heterogeneous pacemaker oscillatory properties relying on distinct combinations of burst-generating INaP and ICAN conductances. The respective proportion of the different inspiratory pacemaker subtypes changes during prenatal development. Concomitantly, network rhythmogenesis switches from a purely INaP/ICAN-dependent mechanism at E16.5 to a combined pacemaker/network-driven process at E18.5. Our results provide the first description of pacemaker bursting properties in embryonic preBötC neurons and indicate that network rhythmogenesis undergoes important changes during prenatal development through alterations in both circuit properties and the biophysical characteristics of pacemaker neurons.

Highlights

Rhythmic motor activities are generated and controlled by neuronal networks organized as central pattern generators (CPG) (Marder and Bucher, 2001; Harris-Warrick, 2010)
Heterogeneous discharge patterns of embryonic inspiratory pacemaker neurons To identify pacemaker neurons in preBotzinger complex network (preBotC) respiratory circuitry of mouse embryos between E16.5 and E18.5, we combined electrophysiological recordings of population rhythmic activity on one side with individual cell calcium imaging on the contralateral side of brainstem slice preparations (Figure 1A)
Slices were previously incubated en bloc with the Calcium Green 1-AM indicator, allowing fluorescence fluctuations due to somatic Ca2+ fluxes resulting from spontaneous impulse burst generation to be monitored

Summary

Introduction

Rhythmic motor activities are generated and controlled by neuronal networks organized as central pattern generators (CPG) (Marder and Bucher, 2001; Harris-Warrick, 2010). Two main processes that may operate in varying combinations underlie motor rhythm generation: (i) the CPG network in question contains endogenously oscillatory neurons, so-called pacemakers, which drive the wider circuit cell population, and/or (ii), the rhythm emerges from the pattern of synaptic connections within the network. In addition to these intrinsic rhythmogenic mechanisms, the dynamics of network function can be conferred by extrinsic neuromodulatory actions. The first is the parafacial respiratory group (RTN/pFRG; Onimaru and Homma, 2003) that appears to generate preinspiratory activity in neonates in vitro, active expiration in adults

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