Stage-dependent sequential organization of nascent smooth muscle cells and its implications for the gut coiling morphogenesis in Xenopus larva

• Published in: Zoology (Jena) 2021-06, Vol.146, p.125905-125905, Article 125905
• Main Authors: Akinaga, Kaoru, Azumi, Yoshitaka, Mogi, Kazue, Toyoizumi, Ryuji
• Format: Article
• Language: English
• Subjects: actomyosin; cell orientation; explant culture; intestinal smooth muscle; morphogenesis
• ISSN: 0944-2006; 1873-2720
• DOI: 10.1016/j.zool.2021.125905

[Display omitted] •In Xenopus larvae, stage-dependent pattern of the coiling morphogenesis of the nascent gut tube has long been known.•Spatiotemporally regulated expression pattern of the smooth muscle-specific actin and its expansion is first evident.•Furthermore, sequential organization of the meshwork of the smooth muscle cells and its anisotropic twisting is revealed.•Using pharmacological approach coupled with gut explant culture, contractile force by actomyosin is suggested to be essential for the coiling. In vertebrates, gut coiling proceeds left-right asymmetrically during expansion of the gastrointestinal tract with highly organized muscular structures facilitating peristalsis. In this report, we explored the mechanisms of larval gut coiling morphogenesis relevant to its nascent smooth muscle cells using highly transparent Xenopus early larvae. First, to visualize the dynamics of intestinal smooth muscle cells, whole-mount specimens were immunostained with anti-smooth muscle-specific actin (SM-actin) antibody. We found that the nascent gut of Xenopus early larvae gradually expands the SM-actin-positive region in a stage-dependent manner. Transverse orientation of smooth muscle cells was first established, and next, the cellular longitudinal orientation along the gut axis was followed to make a meshwork of the contractile cells. Finally, anisotropic torsion by the smooth muscle cells was generated in the center of gut coiling, suggesting that twisting force might be involved in the late phase of coiling morphogenesis of the gut. Administration of S-(–)-Blebbistatin to attenuate the actomyosin contraction in vivo resulted in cancellation of coiling of the gut. Development of decapitation embryos, trunk ‘torso’ explants, and gut-only explants revealed that initial coiling of the gut proceeds without interactions with the other parts of the body including the central nervous system. We newly developed an in vitro model to assess the gut coiling morphogenesis, indicating that coiling pattern of the nascent Xenopus gut is partially gut-autonomous. Using this gut explant culture technique, inhibition of actomyosin contraction was performed by administrating either actin polymerization inhibitor, myosin light chain kinase inhibitor, or calmodulin antagonist. All of these reagents decreased the extent of gut coiling morphogenesis in vitro. Taken together, these results suggest that the contraction force generated by actomyosin-rich intestinal smooth muscl