Astrocyte-mediated Blood Flow Autoregulation as a Disease Mechanism in Glaucoma

描述（申请人提供）：血流自动调节受损被认为是青光眼中BF减少和视网膜神经节细胞损伤的重要病理机制之一。在这种情况下，当眼内压（IOP）和/或血压（BP）波动时，眼睛的血流量（BF），特别是视神经头的血流量（BF）不能再保持在正常范围内。临床和实验室实验证明，这种血流动力学变化导致眼组织内血流量减少，并通过视网膜神经纤维层厚度和功能丧失来测量相关的神经节细胞损伤。因此，针对导致自身调节功能障碍的特定途径的早期干预可以减轻青光眼的血流动力学变化并保护视网膜神经节细胞免受损伤。然而，目前关于与疾病机制相关的眼BF自动调节的知识不足以解释在临床和实验中观察到的许多现象。星形胶质细胞在大脑和视网膜中以前馈机制将神经元活动与血流耦合（称为神经血管耦合）是必不可少的。一系列研究表明，星形胶质细胞也是参与血流自动调节的重要细胞成分。这具有临床意义，因为确定星形胶质细胞在BF自动调节中的作用可能导致一种新的病理

DESCRIPTION (provided by applicant): Compromised blood flow autoregulation has been proposed as one of important pathological mechanisms contributing to decreased BF and retinal ganglion cell damage in glaucoma. Within this paradigm, blood flow (BF) in the eye, particularly in the optic nerve head, can no longer be held within a normal range when intraocular pressure (IOP) and/or blood pressure (BP) fluctuates. As evidenced in clinical and laboratory experiments, such hemodynamic changes result in a blood flow decrease within the ocular tissues and associated ganglion cell damage measured by retinal nerve fiber layer thickness and functional loss. Thus, early intervention targeting specific pathways leading to autoregulation dysfunction could mitigate the hemodynamic changes and protect retinal ganglion cells from damage in glaucoma. However, the current knowledge of ocular BF autoregulation associated with the mechanism of the disease is insufficient to interpret many of the phenomena observed in clinics and in experiments. Astrocytes are essential to couple neuronal activities with blood flow (known as neurovascular coupling) in both brain and retina in a feed forward mechanism. A line of studies have suggested that astrocyte is also an essential cellular element involved in blood flow autoregulation. This has clinical implications because identification of the astrocyte role in BF autoregulation is likely to lead to a novel pathological mechanism of hemodynamic change and subsequently innovative pharmacological interventions for glaucoma. This study proposes to develop new techniques to examine peri-arterial astrocyte bioactivities and vascular response in relation to intra and extravascular pressure alteration. This initiative will lead future studies by utilizing these techniques to examine hypothesized mechanisms of BF autoregulation in normal and in glaucomatous eyes. In Specific Aim 1: (a) To develop and validate a system to set and manipulate both intravascular pressure (simulating BP) and extravascular pressure (simulating IOP) to different level and, simultaneously, image calcium signals (labeled with Fluo-4AM) in peri-arterial astrocytes and vessel diameter (labeled with Dil). (b) Compare the calcium signals within the peri-arterial astrocytes and vessel diameter changes against the magnitude of mechanical stimuli induced by changes in intra- and/ or extravascular pressure in isolated pig retina. In Specific Aim 2: To develop a novel in vivo calcium imaging technique to monitor calcium changes within peri-arterial astrocytes while BP is changed. This will be achieved by intravitreal injection of a calcium indicator (Fluo-4AM). Utilizing fluorescence mode in confocal scanning laser ophthalmoscopy and spectral domain optical coherence tomography, the rat fundus is imaged continuously to monitor the calcium signals in peri-arterial astrocytes and vessel diameter, whilst the rat BP is deterministically increased and decreased. Proving the involvement of astrocytes in BF autoregulation, identification of specific targets for therapeutic intervention may protect te ocular tissue from ischemic insult and subsequent progressive RGC damage in human glaucoma.