KCNQ5/K(v)7.5 potassium channel expression and subcellular localization in primate retinal pigment epithelium and neural retina.
ABSTRACT Previous studies identified in retinal pigment epithelial (RPE) cells an M-type K(+) current, which in many other cell types is mediated by channels encoded by KCNQ genes. The aim of this study was to assess the expression of KCNQ genes in the monkey RPE and neural retina. Application of the specific KCNQ channel blocker XE991 eliminated the M-type current in freshly isolated monkey RPE cells, indicating that KCNQ subunits contribute to the underlying channels. RT-PCR analysis revealed the expression of KCNQ1, KCNQ4, and KCNQ5 transcripts in the RPE and all five KCNQ transcripts in the neural retina. At the protein level, KCNQ5 was detected in the RPE, whereas both KCNQ4 and KCNQ5 were found in neural retina. In situ hybridization in frozen monkey retinal sections revealed KCNQ5 gene expression in the ganglion cell layer and the inner and outer nuclear layers of the neural retina, but results in the RPE were inconclusive due to the presence of melanin. Immunohistochemistry revealed KCNQ5 in the inner and outer plexiform layers, in cone and rod photoreceptor inner segments, and near the basal membrane of the RPE. The data suggest that KCNQ5 channels contribute to the RPE basal membrane K(+) conductance and, thus, likely play an important role in active K(+) absorption. The distribution of KCNQ5 in neural retina suggests that these channels may function in the shaping of the photoresponses of cone and rod photoreceptors and the processing of visual information by retinal neurons.
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ABSTRACT: Ion channels and ion transporters play essential roles in the function of the retinal pigment epithelium (RPE). The use of cell cultures has been exploited as a key method for successfully identifying and studying ion channels and transporters of the RPE. Cultured RPE cells enable robust and long-lasting patch-clamp recordings, Ussing chamber investigations of the transepithelial transport within the isolated RPE, and analyses of the intracellular Ca2+ or pH with fluorescent probes. Furthermore, cultured RPE can be transfected at high success rates, permitting the easy use of siRNA to study the involvement of ion channels on the molecular level. However, the expression patterns of the ion channels in the RPE appear to be a very sensitive marker reflecting the extent of RPE differentiation in vitro. Having originated from the neuroectoderm, cultured RPE cells seem to retain some capacity to change into a more neuronal phenotype expressing TTX-blockable Na+ channels or synaptic Ca2+ channels. Therefore, the identification of ion channels and transporters in cultured cells should be verified in freshly isolated RPE cells and in situ preparations of the RPE, via immunohistochemistry and the analysis of RPE-specific signals in the electroretinogram from transgenic animals.Experimental Eye Research 09/2014; 126:27–37. · 3.02 Impact Factor
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ABSTRACT: Refractive error (RE) is a complex, multifactorial disorder characterized by a mismatch between the optical power of the eye and its axial length that causes object images to be focused off the retina. The two major subtypes of RE are myopia (nearsightedness) and hyperopia (farsightedness), which represent opposite ends of the distribution of the quantitative measure of spherical refraction. We performed a fixed effects meta-analysis of genome-wide association results of myopia and hyperopia from 9 studies of European-derived populations: AREDS, KORA, FES, OGP-Talana, MESA, RSI, RSII, RSIII and ERF. One genome-wide significant region was observed for myopia, corresponding to a previously identified myopia locus on 8q12 (p = 1.25610 28), which has been reported by Kiefer et al. as significantly associated with myopia age at onset and Verhoeven et al. as significantly associated to mean spherical-equivalent (MSE) refractive error. We observed two genome-wide significant associations with hyperopia. These regions overlapped with loci on 15q14 (minimum p value = 9.11610 211) and 8q12 (minimum p value 1.82610 211) previously reported for MSE and myopia age at onset. We also used an intermarker linkage-disequilibrium-based method for calculating the effective number of tests in targeted regional replication analyses. We analyzed myopia (which represents the closest phenotype in our data to the one used by Kiefer et al.) and showed replication of 10 additional loci associated with myopia previously reported by Kiefer et al. This is the first replication of these loci using myopia as the trait under analysis. ''Replication-level'' association was also seen between hyperopia and 12 of Kiefer et al.'s published loci. For the loci that show evidence of association to both myopia and hyperopia, the estimated effect of the risk alleles were in opposite directions for the two traits. This suggests that these loci are important contributors to variation of refractive error across the distribution.PLoS ONE 09/2014; 9(9). · 3.53 Impact Factor
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ABSTRACT: Human, monkey, and bovine retinal pigment epithelial (RPE) cells exhibit an M-type K+ current, which in many other cell types is mediated by channels composed of KCNQ α-subunits and KCNE auxiliary subunits. Recently, we demonstrated the expression of KCNQ1, KCNQ4, and KCNQ5 in the monkey RPE. Here, we investigated the expression of KCNQ and KCNE subunits in native bovine RPE. RT-PCR analysis revealed the expression of KCNQ1, KCNQ4, and KCNQ5 transcripts in the RPE, but, in Western blot analysis of RPE plasma membranes, only KCNQ5 was detected. Among the five members of the KCNE gene family, transcripts for KCNE1, KCNE2, KCNE3, and KCNE4 were detected in bovine RPE, but only KCNE1 and KCNE2 proteins were detected. Immunohistochemistry of frozen bovine retinal sections revealed KCNE1 expression near the apical and basal membranes of the RPE, in cone outer segments, in the outer nuclear layer, and throughout the inner retina. The localization of KCNE1 in the RPE basal membrane, where KCNQ5 was previously found to be present, suggests that this β-subunit may contribute to M-type K(+) channels in this membrane.Experimental Eye Research 11/2013; 116:424-32. · 3.02 Impact Factor