Collaborative Research: Glial regulation of extracellular potassium as an underlying developmental mechanism for the male-female difference in pacemaker neuron firing frequency

在许多物种中，雄性和雌性在体型和行为上的差异已经得到了很好的证实，但人们对它们在发育过程中是如何组织起来的却知之甚少。棕色鬼刀鱼的电子通讯为阐明介导这种差异的细胞机制提供了一个独特的机会。在这种弱电鱼中，雄鱼产生的电器官放电频率比雌鱼高。放电频率的差异是由起搏器核控制的，这是脑干中的一个神经回路。使用跨学科的策略，一个新的机制假设将被测试：星形胶质细胞（一种胶质细胞类型）调节（缓冲）细胞外钾离子在男性和女性不同，这是负责他们的电器官放电的差异。除了提供新的见解，在分子和细胞机制的基础上，一种特定的行为是男性和女性之间的二态，这项研究也将推进知识星形胶质细胞在钾缓冲在神经系统中的作用，并通过这些离子调节神经网络。后两个主题在涉及模式生成神经回路的病理状况的病因学中发挥重要作用（例如，高频癫痫发作被认为是由钾缓冲功能受损引起的，钾缓冲功能受损会增加此类癫痫发作的易感性）。由于这项研究的跨学科性质，也为培养不同学科的学生提供了多种机会，并为生物学和其他STEM学科之间的接口课程开发提供了机会。本科生将通过东北大学和弗吉尼亚大学提供的指导学习课程参与这项研究项目；东北大学的学生也将获得为期6个月的实习机会，参与该项目和弗吉尼亚大学的部分项目。项目负责人和联合项目负责人还将把部分研究成果用于他们在各自机构教授的以实验室为基础的高级本科课程，以及在当地高中开展外展活动和一系列面向公众的STEM活动。弱电鱼leptorhynchus的电器官放电是一种强健且特征明确的行为系统。这些放电的频率在男性和女性中确实不同，并由起搏器核的神经活动控制，起搏器核是延髓中的一种内源性振荡器。这些振荡产生的关键神经元是星形细胞合胞体包裹的起搏器细胞和中继细胞。这些星形胶质细胞的形态、胶质特异性蛋白的表达以及合胞体内单个细胞的间隙连接偶联存在显著的雌雄差异。本研究将验证这样的假设：雄性雌性电器官放电频率差异的形成是通过起搏器核内细胞外钾离子的星形细胞缓冲能力来调节的，从而导致细胞外钾浓度的变化，最终改变由起搏器和中继细胞组成的神经网络的振荡频率。为了实现这一目标，将采用多学科交叉的方法，包括星形细胞合胞体与起搏器和中继细胞的形态学和分子分析；神经胶质功能的药理作用及其行为效应的研究星形胶质细胞控制起搏器核放电频率的离子机制的电生理特性计算模型(i)星形细胞合胞体对钾离子的摄取和再分配，（ii）这种作用对起搏器核神经振荡频率的影响。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Differences in body form and behavior between males and females are well established in numerous species, but little is known about how they are organized during development. Electric communication by the brown ghost knifefish provides a unique opportunity for elucidating the cellular mechanisms mediating one such difference. In this weakly electric fish, males produce electric organ discharges that contain higher frequencies than those of females. The difference in discharge frequencies is controlled by the pacemaker nucleus, a neural circuit in the brainstem. Using an interdisciplinary strategy, a novel mechanistic hypothesis will be tested: astrocytes (a glial cell type) regulate (buffer) extracellular potassium ions differently in males and females, and this is responsible for the difference in their electric organ discharges. Besides providing new insights into the molecular and cellular mechanisms that underlie one specific behavior that is dimorphic between males and females, this research will also advance knowledge about the role of astrocytes in potassium buffering in the nervous system, and about the regulation of neural networks by these ions. The latter two topics play an important role in the etiology of pathological conditions involving pattern-generating neural circuits (for example, high-frequency epileptic seizures are thought to result from an impairment in potassium buffering that enhances the susceptibility for such seizures). Because of the interdisciplinary nature of this research, multiple opportunities are also provided for training students in diverse disciplines, and for curriculum development at the interface between biology and other STEM disciplines. Undergraduates will be involved in carrying out this research project via directed study courses being offered at both Northeastern University and the University of Virginia; 6-month internships will also be offered for students from Northeastern University to participate in the parts of the project conducted and at the University of Virginia. The PI and co-PI will also use parts of this research in laboratory-based upper-level undergraduate courses that they teach at their respective institutions, as well as engaging in outreach activities at local high schools and a series of STEM events for the general public. The electric organ discharge of the weakly electric fish Apteronotus leptorhynchus is a robust and well-characterized behavioral system. The frequency of these discharges is reliably different in males and females, and controlled by the neural activity of the pacemaker nucleus, an endogenous oscillator in the medulla oblongata. The neurons critically involved in the generation of these oscillations are the pacemaker and relay cells, which are enveloped by astrocytic syncytia. Significant male-female differences exist in the morphology of these astrocytes, their expression of glia-specific proteins, and the gap-junction coupling of individuals cells within the syncytium. The study will test the hypothesis that the development of the male-female difference in the frequency of the electric organ discharge is regulated through modulation by the astrocytic buffering capacity of extracellular potassium ions in the pacemaker nucleus, thereby resulting in changes in the extracellular potassium concentration and ultimately in the oscillation frequency of the neural network composed of pacemaker and relay cells. To achieve this objective, a multi- and interdisciplinary approach will be employed, including morphological and molecular analysis of the association of the astrocytic syncytium with the pacemaker and relay cells; pharmacological manipulation of glial function and examination of its behavioral effect; electrophysiological characterization of the ionic mechanisms of astrocytes that control the firing frequency of the pacemaker nucleus; and computational modeling of (i) the uptake and redistribution of potassium ions by the astrocytic syncytium, (ii) the effect of this action on the frequency of the neural oscillations of the pacemaker nucleus.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.