Direct measurements of neurotransmitter concentrations in vivo

描述(申请人提供)：在两个神经元相遇的地方，它们可以形成突触，可以进行化学交流。这是大脑的功能单位，正是在这个层面上，变化等同于学习。这项拟议的研究将发展并使用电化学方法来测量体内易被氧化的神经递质的绝对浓度--这是目前的方法所不可能做到的。神经递质细胞外浓度的快速变化可由神经元的时相放电引起。出于这个原因，化学传感器应该能够在广泛的时间范围内工作。理想的神经递质检测传感器具有高灵敏度、能区分化合物、响应时间快等特点。电化学方法为容易氧化的神经递质提供了一种方法，通过在神经递质释放部位旁边使用电极来实现这一点。背景减去循环伏安法是一种广泛使用的方法，能够在亚秒的时间尺度上测量神经递质浓度的变化。这种方法已被证明是测量浓度的有价值的工具；然而，它不能测量神经递质的绝对浓度。绝对浓度是至关重要的，因为它影响受体的占有率，并可以改变释放的神经递质对受体的影响。一种在单秒时间尺度上测量易氧化神经递质绝对浓度的新方法将被开发出来。这种方法依赖于这些分子在传感表面的吸附。背景-减去循环伏安法通常在动力学受限的状态下进行。在这个实验中，我们允许通过改变检测参数来改变吸附到表面的量。通过允许系统接近平衡，然后将其推得更远，可以测量存在的神经递质的绝对浓度。该方法将通过使用特征良好的药物进行验证。可卡因和精神刺激剂哌醋甲酯(利他林)对大鼠纹状体多巴胺绝对浓度的影响将被确定。哌醋甲酯被批准用于治疗注意力缺陷多动障碍、体位性心动过速综合征和发作性睡病。像其他兴奋剂一样，哌醋甲酯会增加大脑中的多巴胺浓度；然而，哌醋甲酯是如何改变这些浓度的，还没有用精确的时间分辨率来测量。

DESCRIPTION (provided by applicant): Where two neurons meet they can form a synapse and chemical communication can occur. This is the functional unit of the brain, and it is at this level where changes equate to learning. The proposed research will develop and then use electrochemical methods to measure the absolute concentrations of easily oxidized neurotransmitters in vivo - something not possible with current methodology. Fast changes in the extracellular concentration of neurotransmitters can arise from phasic neuronal firing. For this reason, chemical sensors should be able to operate on a wide range of time scales. An ideal sensor for the detection of neurotransmitters has high sensitivity, can distinguish between compounds, and has a fast response time. Electrochemical approaches offer a way to accomplish this for easily oxidized neurotransmitters by using an electrode next to sites where the neurotransmitter is released. Background-subtracted cyclic voltammetry, one widely used method is capable of measuring changes in neurotransmitter concentrations on a sub-second timescale. This methodology has proved to be a valuable tool to measure the concentrations; however, it cannot measure the absolute concentrations of neurotransmitters. The absolute concentration is of critical importance as it affects the receptor occupancy and can change the effect of released neurotransmitters on the receptors. A novel method to measure the absolute concentrations of easily oxidizable neurotransmitters on a single-second timescale will be developed. This method relies on the adsorption of these molecules to the sensing surface. Background-subtracted cyclic voltammetry is typically performed in a kinetically limited regime. In this experiment, we allow the amount adsorbed to the surface to vary by changing detection parameters. By allowing the system to come close to equilibrium, and then pushing it farther away, the absolute concentration of neurotransmitter present can be measured. The method will be validated by using well-characterized drugs. The effects of cocaine and methylphenidate (Ritalin), a psychostimulant drug on the absolute dopamine concentration in the rat striatum will be determined. Methylphenidate is approved for the treatment of attention-deficit hyperactivity disorder, postural orthostatic tachycardia syndrome, and narcolepsy. Methylphenidate, like other stimulants, increases the concentration of dopamine in the brain; however how methylphenidate changes these concentrations has not been measured with exquisite temporal resolution.