Protocol

Sagittal Explant Culture for 3D Confocal Time-Lapse Analysis of Chick Peripheral Nervous System Formation

¹Stowers Institute for Medical Research, Kansas City, MO 64110, USA
²Montana State University, Department of Cell Biology and Neuroscience, Bozeman, MT 59717, USA

³Corresponding author (pmk{at}stowers-institute.org)

INTRODUCTION

The peripheral nervous system (PNS) regulates key events within the body including breathing, heart rate, and pain and temperature sensation. In the trunk of the developing embryo, neural crest (NC) cells that follow a ventro-medial pathway form the dorsal root ganglia (DRG) and sympathetic ganglia (SG) of the PNS. The cellular and molecular mechanisms that mediate the formation of the PNS are not completely understood due to the lack of a model system to monitor NC cell migratory behaviors deep within the embryo. Here, we describe a unique sagittal explant culture technique that allows for visualization of DRG and SG formation in the relatively intact chick embryo to assess potential molecules and signals influencing the formation of the PNS. By making a midline cut down the chick antero-posterior axis, each half of the embryo is an explant that can be imaged either from the medial perspective, to reveal events occurring deep within the embryo (SG formation), or from the lateral surface, to image events occurring in the dorso-lateral plane (DRG formation).