Development of respiratory centers in the bullfrog tadpole brainstem

Abstract

Among vertebrates, rhythmic motor behaviors such as breathing, swallowing, and sucking are controlled by rhythm generators or neural oscillators located at various sites in the medulla of the brainstem. That all vertebrates exhibit these behaviors, leads investigators to hypothesize common ancestry for the cellular networks responsible for homeostatic rhythm generation in the brainstem. While the locations and functions of rhythm generating sites controlling some of these behaviors have been well investigated, details regarding the development of these sites remain largely unknown. Recent work has suggested that neural oscillators in the rostral and caudal medulla, which contribute to ventilation in amphibians, may be homologous with those controlling breathing in mammals. I first investigated the developmental contributions of these regions to CO₂ sensitivity and rhythm generation in bullfrog tadpoles at different stages of metamorphosis. I then characterized the function and structure of a neural oscillator essential for lung rhythmogenesis in the tadpoles and compared it to similar oscillators in mammals. To investigate functional aspects of brainstem, I used a combination of single-unit and whole-nerve electrophysiology in the presence of pharmacological agents (neuronal receptor agonists and antagonists) or following removal of portions of the isolated brainstem of bullfrog tadpoles at different stages of metamorphosis. Structural studies were accomplished using immunohistochemistry, staining for phenotypic markers common to mammalian rhythmogenic sites, and assessing the difference between early and late metamorphic bullfrog tadpoles. Taken together, my results suggest that amphibians may indeed have a rhythmogenic site in the rostral medulla that is homologous to a mammalian rhythmogenic site; it is both structurally and functionally similar to the mammalian parafacial respiratory group/retrotrapezoid nucleus complex. This region undergoes structural and functional changes as tadpoles develop through metamorphosis. Understanding the development of respiration in amphibians may provide clues into the evolution and development of breathing in mammals.