CENTRAL RESPIRATORY CHEMORECEPTION IN AMPHIBIANS

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 A.具体目标 应用中的具体目标没有改变，并根据这些目标进行了实验。在自发节律的体外蝌蚪脑干制备中，我们已经在执行细胞内pH(Phi)的光学记录方面取得了进展。 B.研究和成果 在BCECF校准曲线完成后，我们能够同时记录从化学和非化学敏感区域发射的荧光定量，同时记录整个神经呼吸活动。我们报告了位于已知化疗敏感区的神经元和来自非化疗敏感区的神经元在静息PHI上的显著差异。此外，我们利用氯化铵预脉冲技术来诱导细胞内酸化，同时将控制的浴液pH值维持在7.8。最后，我们能够证明化疗敏感神经元PHI的降低和肺爆发频率的增加之间存在显著的负相关。我们已经成功地证明了PHI和通风之间的相关性，正如我们的具体目标所建议的那样，并正在提交这些数据供同行评审发表。我们还探索了一种潜在的新技术来诱导细胞内酸化，而不是使用氯化铵预脉冲技术，因为氯化铵预脉冲技术会诱导所有脑干神经元的细胞内酸化。我们希望通过暴露在紫外光下使用H的“去化”，从而允许我们仅在感兴趣的细胞中集中诱导PHI的降低。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. A. Specific Aims The Specific Aims in the application have not changed and experiments have been conducted in accordance with these aims. We have progressed in our abilities to perform optical recordings of intracellular pH (pHi) from neurons in the spontaneously rhythmic in vitro tadpole brainstem preparation. B. Studies and Results Upon the completion of the BCECF calibration curve, we were then able to record emitted fluorescence rations from both chemo- and non-chemosensitive regions while simultaneously recording whole nerve respiratory activities. We report significant differences in resting pHi between neurons located in known chemosensitive areas and neurons from non-chemosensitive areas. In addition, we utilized the ammonium chloride prepulse technique to induce intracellular acidification while maintaining a control bath pH of 7.8. Finally, we were able to demonstrate a significant negative correlation between decreases in pHi in chemosensitive neurons and increases in lung burst frequency. We have successfully demonstrated a correlation between pHi and ventilation, as proposed in our Specific Aims, and are in the process of submitting these data for peer review publication. We also explored a potential new technique to induce intracellular acidosis rather than use the ammonium chloride prepulse technique, as the ammonium chloride prepulse technique induces intracellular acidification to all brainstem neurons. We hoped to use "uncaging" of H+ via exposure to ultraviolet light, thereby allowing us to focally induce decreases in pHi in only those cells of interest.