The development and maintenance of a spontaneous respiratory rhythm is essential for the survival of all vertebrates. The precise location of neurons responsible for respiratory rhythmogenesis , as well as the afferent and efferent connections of the respiratory Sudden Infant Death Syndrome (SIDS) and sleep apnea, disorders which afflict both ends of the developmental spectrum, An in vitro brainstem preparation of the bullfrog tadpole Rana catesbeiana has been developed in order to investigate the development of the neural substrates essential to central respiratory pattern generation and chemoreception. The proposed research will further the understanding of the neural components necessary to produce and maintain spontaneous, rhythmic respiratory motor output. In addition, the significance of both central and peripheral chemoreceptor input to the respiratory central pattern generator (CPG) during development will be described. Specifically, experiments have been designed: 1. To determine during development the relative contribution both of central and peripheral respiratory chemoreceptor input on central respiratory pattern generation and motor output. 2. To define the precise location of neurons responsible for both gill and lung respiratory pattern generation in the developing vertebrate. 3. To begin to determine, in the developing vertebrate, the cellular, ionic, biophysical properties and network connections underlying gill and lung respiratory pattern generation. Both whole nerve and intracellular recordings from respiratory-related neurons will be employed in semi-intact and in vitro superfused tadpole brainstem preparations. A semi-intact brainstem/gill arch preparation will be used to examine the influence of both central and peripheral chemoreceptor stimulation; all other studies will use the in vitro brainstem preparation. lontophoresis of fluorescent dyes will be used to create maps of the projections of respiratory motor neurons, interneurons, and candidate central pattern generation neurons. Bath application of various neurotransmitter agonists and antagonists documented to be responsible for respiratory rhythmicity in vertebrates will be performed in order to characterize their effects on gill and lung motor patterns. Thin slice preparations of the brain stem will be performed to elucidate the precise location of gill and lung pattern generating neurons. The developmental aspect of each study will be addressed by performing the above experiments on tadpoles from all stages of development.
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