Functional role of vesicular zinc in synaptic mechanisms in the amygdala

描述(由申请人提供)：本提案的目标是建立和表征突触可释放锌在杏仁核回路谷氨酸能突触中的功能作用，这些锌包含在突触小泡(囊泡锌)中，与听觉恐惧条件反射有关。我们最近的发现表明，锌转运蛋白-3(ZNT-3)作为含锌神经元的标志物，表达于杏仁外侧核和听觉皮质的TE3区，在恐惧条件反射过程中将听觉条件刺激(CS)信息传递到杏仁外侧核(LA)，提示囊泡锌在恐惧条件反射通路的突触功能调节中具有潜在的作用。我们用锌离子螯合剂的实验结果表明，囊泡锌离子可以控制皮层-杏仁核突触的棘波时序依赖性长时程增强(LTP)的诱导。我们现在提出了一个详细的电生理学分析，在突触激活过程中释放的囊泡锌在突触传递和大脑皮质和丘脑传入LA的可塑性中所起的作用，在恐惧条件作用期间将听觉CS信息传递到杏仁核，从ZNT-3基因敲除的小鼠和对照小鼠的切片中。这些定向破坏ZNT-3基因的转基因小鼠缺乏水泡锌。用全细胞膜片钳技术记录复合突触后反应和单位突触后反应，并对其进行分析，以评估去锌对杏仁核突触传递和可塑性的影响。我们还将探索ZNT-3基因敲除小鼠是否通过减少LA中主要神经元的GABA能抑制来弥补囊泡性锌的缺乏，并分析对照组和ZNT-3基因敲除小鼠脑片中的抑制性神经传递。我们的假设是，在恐惧条件反射通路中，谷氨酸能突触经历LTP的能力可以由LTP诱导刺激过程中激活的突触释放的囊泡锌2控制。拟议的研究将提高我们对突触可塑性机制的理解，突触可塑性可能是获得恐惧记忆的基础。更好地了解恐惧相关行为的细胞机制将有助于合理开发治疗创伤后应激障碍、广泛性焦虑和其他涉及大脑恐惧系统的疾病的新疗法。 公共卫生相关性：拟议的研究将提高我们对突触可塑性机制的理解，突触可塑性可能是获得恐惧记忆的基础。更好地了解恐惧相关行为的细胞机制将有助于合理开发治疗创伤后应激障碍、广泛性焦虑和其他涉及大脑恐惧系统的疾病的新疗法。

DESCRIPTION (provided by applicant): The objective of this proposal is to establish and characterize the functional role of synaptically-releasable zinc, contained in synaptic vesicles (vesicular zinc), at glutamatergic synapses of the amygdala circuitry implicated in auditory fear conditioning. Our recent findings indicate that zinc transporter-3 (ZnT-3), serving as a marker for zinc-containing neurons, is expressed in the lateral amygdala and the TE3 area of the auditory cortex, conveying the auditory conditioned stimulus (CS) information to the lateral nucleus of the amygdala (LA) during fear conditioning, thus suggesting a potential role for vesicular zinc in regulation of synaptic functions in fear conditioning pathways. Results of our experiments with Zn2+ chelators imply that vesicular Zn2+ can control the induction of spike timing-dependent long-term potentiation (LTP) at cortico-amygdala synapses. We now propose a detailed electrophysiological analysis of the role of vesicular Zn2+, releasable in the course of synaptic activation, in synaptic transmission and plasticity in cortical and thalamic inputs to the LA, delivering auditory CS information to the amygdala during fear conditioning, in slices from ZnT-3 knockout and control mice. These genetically modified mice with targeted disruption of the ZnT-3 gene lack vesicular zinc. Both the compound and unitary postsynaptic responses will be recorded with whole-cell patch-clamp techniques and analyzed to estimate the effects of ZnT3 ablation on synaptic transmission and plasticity in the amygdala. We will also explore whether ZnT-3 knockout mice compensate for the lack of vesicular zinc by decreased GABAergic inhibition of principal neurons in the LA, assaying inhibitory neurotransmission in slices from control and ZnT-3 knockout mice. Our hypothesis is that the ability of glutamatergic synapses in fear conditioning pathways to undergo LTP can be controlled by vesicular Zn2+, released at activated synapses during LTP- inducing stimulation. The proposed studies will improve our understanding of the mechanisms of synaptic plasticity which could underlie the acquisition of fear memory. A better knowledge of the cellular mechanisms of fear-related behaviors will permit the rational development of novel therapeutic treatments for posttraumatic stress disorder, generalized anxiety, and other disorders implicating the fear system of the brain. PUBLIC HEALTH RELEVANCE: The proposed studies will improve our understanding of the mechanisms of synaptic plasticity which may underlie the acquisition of fear memory. A better knowledge of the cellular mechanisms of fear-related behaviors will permit the rational development of novel therapeutic treatments for posttraumatic stress disorder, generalized anxiety, and other disorders implicating the fear system of the brain.