Gamma amino butyric acid (GABA) is a major retinal neurotransmitter than can function at a variety of retinal receptors. Some of these receptors are ligand-gated ion channels, probably similar to the GABA/A receptors prominently expressed in the brain. Complementary DNAs encoding 14 different GABA/A receptor subunits arranged in 4 major classes (alpha, beta, gamma, and delta) have been cloned from the brain. We have recently described a GABA rho subunit class discovered through cDNA cloning. GABA rho1 expression in retina is at least 20-fold greater than its expression in any other tissue. This retina-specific receptor subunit has distinctive structural features and avidly associates into homo-oligomeric receptors displaying unique pharmacologic and physiologic characteristics. A second rho subunit isolated from a retinal cDNA library is expressed at lower levels in retina RNA but displays structural and functional features similar to the rho1 subunit. This application proposes to investigate the molecular biology of this distinct class of retinal GABA receptor subunits and to explore their possible roles in retinal pathology by pursuit of the following aims: 1) Analyzing the genomic organization and relative orientation of the rho genes including detailed examination of the putative promoter regions of each gene. 2) Determining whether mutations in GABA rho1 or rho2 cause an autosomal dominant form of macular dystrophy which has recently been mapped to the same chromosomal region (6q13-q21) as the rho genes. 3) Ascertaining whether additional members of alternatively-spliced forms of the GABA rho subunit family are expressed in retina by PCR amplification of retinal cDNA using primers derived from rho cDNA sequences. 4) Performing structure/function analysis to determine which portions of the GABA rho cDNA sequences confer the pharmacologic and physiologic properties characteristic of this subunit class. 5) Ascertaining which retinal cell types express the GABA rho subunits using Northern and PCR analysis of RNA, in situ hybridization and immunohistochemical techniques.
