Abstract
Purpose: :
In vertebrates, eye morphogenesis begins with the bilateral evagination of tissue from the forebrain to form the optic vesicles, which then undergo dramatic invagination to form the optic cup. During this process, the neuroectodermal layers of the optic cup must fuse ventrally at the choroid fissure in order to contain the retina and RPE. Failure of choroid fissure closure results in a coloboma. The molecular and cellular mechanisms facilitating choroid fissure closure and neuroepithelial cell fusion in the choroid fissure are unknown. Utilizing the zebrafish as a model, we sought to characterize choroid fissure closure in vivo and to identify gene products required for the process.
Methods: :
Immunohistochemical studies and confocal imaging assays were performed to visualize choroid fissure closure. In situ hybridization was performed to identify gene products spatially and temporally associated with choroid fissure closure. Genetic mutants and morpholinos were utilized to identify gene products required for choroid fissure closure and neuroepithelial fusion within the fissure.
Results: :
Temporal and spatial aspects of choroid fissure closure were determined; neuroepithelial fusion within the fissure initiates at approximately 36 hours post fertilization (hpf) and is completed between 48 and 60hpf. Fusion initiates in the center of the fissure, and it proceeds bidirectionally. Numerous gene products were identified to be expressed in or around the fissure during its closure, and these include netrin1a, integrin alpha5, integrin-linked kinase (ILK), talin and kindlin. Loss of netrin1a, ILK or talin function resulted in colobomas.
Conclusions: :
Zebrafish provide an excellent in vivo model in which the cellular and molecular underpinnings of choroid fissure closure can be analyzed, and we have identified spatial and temporal aspects of the process, as well as morphological characteristics of cells that occupy the fissure. Loss-of-function studies demonstrate that Netrin1a, ILK and talin function are required for choroid fissure closure.
Keywords: development • genetics • retinal development