Biomedical subjects

Pamela A Raymond

Publications and source records attributed to Pamela A Raymond.

8 recordsLinked to original sources

PubMed2006

Molecular characterization of retinal stem cells and their niches in adult zebrafish.

BACKGROUND: The persistence in adult teleost fish of retinal stem cells that exhibit all of the features of true 'adult stem cells'--self-renewal, multipotency, and the capacity to respond to injury by mitotic activation with the ability to regenerate differentiated tissues--has been known for several decades. However, the specialized cellular and molecular characteristics of these adult retinal stem cells and the microenvironmental niches that support their maintenance in the differentiated retina and regulate their activity during growth and regeneration have not yet been elucidated. RESULTS: Our data show that the zebrafish retina has two kinds of specialized niches that sustain retinal stem cells: 1) a neuroepithelial germinal zone at the interface between neural retina and ciliary epithelium, called the ciliary marginal zone (CMZ), a continuous annulus around the retinal circumference, and 2) the microenvironment around some Müller glia in the differentiated retina. In the uninjured retina, scattered Müller glia (more frequently those in peripheral retina) are associated with clusters of proliferating retinal progenitors that are restricted to the rod photoreceptor lineage, but following injury, the Müller-associated retinal progenitors can function as multipotent retinal stem cells to regenerate other types of retinal neurons. The CMZ has several features in common with the neurogenic niches in the adult mammalian brain, including access to the apical epithelial surface and a close association with blood vessels. Müller glia in the teleost retina have a complex response to local injury that includes some features of reactive gliosis (up-regulation of glial fibrillary acidic protein, GFAP, and re-entry into the cell cycle) together with dedifferentiation and re-acquisition of phenotypic and molecular characteristics of multipotent retinal progenitors in the CMZ (diffuse distribution of N-cadherin, activation of Notch-Delta signaling, and expression of rx1, vsx2/Chx10, and pax6a) along with characteristics associated with radial glia (expression of brain lipid binding protein, BLBP). We also describe a novel specific marker for Müller glia, apoE. CONCLUSION: The stem cell niches that support multi-lineage retinal progenitors in the intact, growing and regenerating teleost retina have properties characteristic of neuroepithelia and neurogenic radial glia. The regenerative capacity of the adult zebrafish retina with its ability to replace lost retinal neurons provides an opportunity to discover the molecular regulators that lead to functional repair of damaged neural tissue.

Aging↗

PubMed2006

GFAP transgenic zebrafish.

We have generated transgenic zebrafish that express green fluorescent protein (GFP) in glial cells driven by the zebrafish glial fibrillary acidic protein (GFAP) regulatory elements. Transgenic lines Tg(gfap:GFP) were generated from three founders; the results presented here are from the mi2001 line. GFP expression was first visible in the living embryo at the tail bud-stage, then in the developing brain by the 5-somite-stage ( approximately 12 h post-fertilization, hpf) and then spreading posteriorly along the developing spinal cord by the 12-somite stage (approximately 15 hpf). At 24 hpf GFP-expressing cells were in the retina and lens. By 72 hpf GFP expression levels were strong and localized to the glia of the brain, neural retina, spinal cord, and ventral spinal nerves, with moderate expression in the enteric nervous system and weaker levels in the olfactory sensory placode and otic capsule. GFP expression in glia co-localized with anti-GFAP antibodies, but did not co-localize with the neuronal antibodies HuC/D or calretinin in mature neurons.

Animals↗

PubMed2005

Genetic dissection of the zebrafish retinal stem-cell compartment.

In a large-scale forward-genetic screen, we discovered that a limited number of genes are required for the regulation of retinal stem cells after embryogenesis in zebrafish. In 18 mutants out of almost 2000 F2 families screened, the eye undergoes normal embryonic development, but fails to continue growth from the ciliary marginal zone (CMZ), the post-embryonic stem-cell niche. Class I-A mutants (5 loci) display lower amounts of proliferation in the CMZ, while nearly all cells in the retina appear differentiated. Class I-B mutants (2 loci) have a reduced CMZ with a concomitant expansion in the retinal pigmented epithelium (RPE), suggesting a common post-embryonic stem cell is the source for these neighboring cell types. Class II encompasses three distinct types of mutants (11 loci) with expanded CMZ, in which the progenitor population is arrested in the cell cycle. We also show that in at least one combination, the reduced CMZ phenotype is genetically epistatic to the expanded CMZ phenotype, suggesting that Class I genes are more likely to affect the stem cells and Class II the progenitor cells. Finally, a comparative mapping analysis demonstrates that the new genes isolated do not correspond to genes previously implicated in stem-cell regulation. Our study suggests that embryonic and post-embryonic stem cells utilize separable genetic programs in the zebrafish retina.

Animals↗

PubMed2005

Nephrocystin-5, a ciliary IQ domain protein, is mutated in Senior-Loken syndrome and interacts with RPGR and calmodulin.

Nephronophthisis (NPHP) is the most frequent genetic cause of chronic renal failure in children. Identification of four genes mutated in NPHP subtypes 1-4 (refs. 4-9) has linked the pathogenesis of NPHP to ciliary functions. Ten percent of affected individuals have retinitis pigmentosa, constituting the renal-retinal Senior-Loken syndrome (SLSN). Here we identify, by positional cloning, mutations in an evolutionarily conserved gene, IQCB1 (also called NPHP5), as the most frequent cause of SLSN. IQCB1 encodes an IQ-domain protein, nephrocystin-5. All individuals with IQCB1 mutations have retinitis pigmentosa. Hence, we examined the interaction of nephrocystin-5 with RPGR (retinitis pigmentosa GTPase regulator), which is expressed in photoreceptor cilia and associated with 10-20% of retinitis pigmentosa. We show that nephrocystin-5, RPGR and calmodulin can be coimmunoprecipitated from retinal extracts, and that these proteins localize to connecting cilia of photoreceptors and to primary cilia of renal epithelial cells. Our studies emphasize the central role of ciliary dysfunction in the pathogenesis of SLSN.

Amino Acid Sequence↗

PubMed2004

Zebrafish cone-rod (crx) homeobox gene promotes retinogenesis.

The mammalian Cone-rod homeobox (Crx) gene is a divergent member of the Otx gene family known to be involved in differentiation and survival of retinal photoreceptors and photoentrainment of circadian rhythms. Zebrafish have two genes in the Otx5/crx orthology class, and we previously showed that crx can transactivate rhodopsin expression in vitro, and that otx5 (orthodenticle-related gene), but not crx, regulates expression of circadian genes in the pineal. Here, we show that zebrafish crx does not regulate expression of opsins and other photoreceptor-specific genes in the pineal. We further show that crx is expressed in proliferating retinal progenitors and may be involved in patterning the early optic primordium and in promoting the differentiation of retinal progenitors, including photoreceptors. These results suggest novel functions for zebrafish crx during retinal specification and differentiation.

Animals↗

PubMed2004

A moving wave patterns the cone photoreceptor mosaic array in the zebrafish retina.

In this paper, we describe the embryonic origin and patterning of the planar mosaic array of cone photoreceptor spectral subtypes in the zebrafish retina. A discussion of possible molecular mechanisms that might generate the cone mosaic array considers but discards a model that accounts for formation of neuronal mosaics in the inner retina and discusses limitations of mathematical simulations that reproduce the zebrafish cone mosaic pattern. The formation and organization of photoreceptors in the ommatidia of the compound eye of Drosophila is compared with similar features in the developing zebrafish cone mosaic, and a model is proposed that invokes spatiotemporally coordinated cell-cell interactions among cone progenitors to determine the identity and positioning of cone spectral subtypes.

Animals↗

PubMed2002

Embryonic origin of the eyes in teleost fish.

The developmental history of the vertebrate eye begins at an early embryonic stage, with the formation of the body axes and induction of neural tissue. Several recent experimental embryological and genetic studies in teleost fish have produced new insights into the morphogenetic and molecular regulation of eye formation. Molecular signaling pathways and patterned expression of transcription factors implicated in eye determination are discussed, and the importance of morphogenetic cell movements is emphasized.

Animals↗

PubMed2001

Otx5 regulates genes that show circadian expression in the zebrafish pineal complex.

The photoneuroendocrine system translates environmental light conditions into the circadian production of endocrine and neuroendocrine signals. Central to this process is the pineal organ, which has a conserved role in the cyclical synthesis and release of melatonin to influence sleep patterns and seasonal reproduction. In lower vertebrates, the pineal organ contains photoreceptors whose activity entrains an endogenous circadian clock and regulates transcription in pinealocytes. In mammals, pineal function is influenced by retinal photoreceptors that project to the suprachiasmatic nucleus-the site of the endogenous circadian clock. A multisynaptic pathway then relays information about circadian rhythmicity and photoperiod to the pineal organ. The gene cone rod homeobox (crx), a member of the orthodenticle homeobox (otx) family, is thought to regulate pineal circadian activity. In the mouse, targeted inactivation of Crx causes a reduction in pineal gene expression and attenuated entrainment to light/dark cycles. Here we show that crx and otx5 orthologs are expressed in both the pineal organ and the asymmetrically positioned parapineal of larval zebrafish. Circadian gene expression is unaffected by a reduction in Crx expression but is inhibited specifically by depletion of Otx5. Our results indicate that Otx5 rather than Crx regulates genes that show circadian expression in the zebrafish pineal complex.

Amino Acid Sequence↗