ROD AND CONE SIGNALING PATHWAYS IN MAMMALIAN RETINA

描述(申请人提供)：这一新研究项目的总体目标是了解哺乳动物视网膜中视杆细胞和视锥细胞信号通路的详细突触机制。解剖学研究表明，哺乳动物视网膜中的视杆细胞通道和视锥细胞通道遵循一套特定的电路规则：视杆细胞仅在视杆去极化双极细胞(DBCRS)上形成突触，而DBCRS不直接在神经节细胞(GC)上产生输出突触，而是间接通过AII无长突细胞(AIIAC)产生突触。AIIAC通过在分别与ON和OFF GC突触的锥体去极化和超极化双极细胞(DBCCS和HBCCS)上“搭载”，向ON和OFF GC发送杆介导的信号。在这项研究计划中，我们计划以小鼠视网膜为模型系统，系统地研究这些哺乳动物特有的突触回路。除了使用全细胞电压钳(带有染料填充)、解剖学和药理学技术外，我们还将利用四个途径特定的突变小鼠品系来阐明哺乳动物的视杆细胞和视锥信号是如何传递到BCS、ACS和GC的，并确定哺乳动物特有的电路规则是否完全有效，因为最近的证据和我们的初步结果表明，严格的视杆/视锥输入规则可能不适用于所有哺乳动物的视网膜神经元。我们将检验“小鼠DBCRS的亚群接受锥体的直接突触输入，小鼠DBCCS和HBCCS的亚群接受杆的直接突触输入，杆和锥介导的信号在发送到GC之前被AIIAC进一步混合，AC介导的抑制性突触输入到BCS和GC携带混合的杆/锥信号”的总体假设。本应用有3个具体目标，重点研究视杆细胞和视锥细胞对光诱发的阳离子电流和氯电流(分别代表谷氨酸能和GABA能/甘氨酸能突触输入)的贡献：(1)七种类型的双极细胞(HBCMC/RS、HBCMCS、HBCSCS、DBCC2/MCS、DBCC1/MC/RS、DBCR2S、DBCR1S)；(2)AII无长突细胞；以及(3)三种类型的阿尔法神经节细胞(ON、SOIF和TOF1GC)。所获得的结果将增加我们对平行通道，如视杆/视锥信号通路，如何在眼睛和大脑中处理、分离和整合信息的理解。由于许多视觉障碍与视杆细胞和视锥细胞信号通路的异常有关，本研究项目将有助于确定导致这些眼病发病的细胞和突触部位。 与公共健康相关：眼睛是大脑的“窗口”，通过将工作分成两部分，人眼可以记录到巨大的光强度(从星光下的物体到阳光下的雪)：杆状感光器编码暗淡的图像，锥形感光器编码明亮和彩色的图像。了解视杆细胞和视锥细胞信号是如何被视网膜突触通路处理的，是解开视觉感知和脑运作机制的基础和关键步骤。某些形式的黄斑变性、视网膜色素变性、先天性静止性夜盲和青光眼与视杆和视锥信号通路的功能障碍有关，因此，该项目的结果将为视杆和视锥突触通路中的特定缺陷如何调节这些眼部疾病提供关键信息。

DESCRIPTION (provided by applicant): The overall objective of this new research project is to understand detailed synaptic mechanisms underlying rod and cone signaling pathways in the mammalian retina. Anatomical studies have suggested that rod and cone channels in the mammalian retina follow a specific set of circuitry rules: rods make synapses only on rod depolarizing bipolar cells (DBCRS), and DBCRS do not make output synapses directly on ganglion cells (GCs), but indirectly through AII amacrine cells (AIIACs). AIIACs send rod-mediated signals to ON and OFF GCs by "piggybacking" on the cone depolarizing and hyperpolarizing bipolar cells (DBCCS and HBCCS), which synapse on ON and OFF GCs, respectively. In this research proposal, we plan to systematically investigate these mammalian-specific synaptic circuits by using the mouse retina as a model system. In addition to using whole-cell voltage clamp (with dye-filling), anatomical and pharmacological techniques, we will take advantage of four strains of pathway-specific mutant mice to elucidate how mammalian rod and cone signals are transmitted to BCs, ACs and GCs, and to determine whether the mammalian-specific circuitry rules are totally valid, as recent evidence and our preliminary results suggest that the strict rod/cone input rules may not hold for all mammalian retinal neurons. We will test the overall hypothesis that "subpopulations of mouse DBCRS receive direct synaptic inputs from cones and subpopulations of mouse DBCCS and HBCCS receive direct synaptic inputs from rods, the rod- and cone-mediated signals are further mixed by AIIACs before sending to GCs, and the AC-mediated inhibitory synaptic inputs to BCs and GCs carry mixed rod/cone signals". This application has 3 specific aims focused on studying rod and cone contributions to light-evoked cation and chloride currents (?Ic and ?Ic1, representing glutamatergic and GABAergic/glycinergic synaptic inputs, respectively) in: (1) seven types of bipolar cells (HBCMC/RS, HBCMCS, HBCSCS, DBCC2/MCS, DBCC1/MC/RS, DBCR2S, DBCR1S); (2) AII amacrine cells; and (3) three types of alpha ganglion cells (ON, sOFF and tOFF 1GCs) in the dark-adapted mouse retina. Results obtained will increase our understanding of how parallel channels, such as the rod/cone signaling pathways, process, segregate and integrate information in the eye and in the brain. Since many visual disorders are associated with abnormalities in the rod and cone signaling pathways, this research project will help to identify cellular and synaptic sites responsible for the pathogenesis of these eye diseases. PUBLIC HEALTH RELEVANCE: The eye is the "window" of the brain and human eyes can register an enormous range of light intensities (from objects under starlit sky to snow under bright sun light) by dividing the labor into two parts: rod photoreceptors encode dim images and cone photoreceptors encode bright and color images. Understanding how rod and cone signals are processed by retinal synaptic pathways is a fundamental and essential step for unraveling mechanisms of visual perception and brain operation. Certain forms of macular degeneration, retinitis pigmentosa, congenital stationary night blindness and glaucoma are associated with dysfunction of the rod and cone signaling pathways, and thus results obtained from this project will provide crucial information on how specific defects in the rod and cone synaptic pathways mediate these eye disorders.