Effects of induced systemic hypothyroidism upon the retina: regulation of thyroid hormone receptor alpha and photoreceptor production.

PURPOSE: Investigate the effects of systemic hypothyroidism upon the differentiated, growing, and regenerating retina of postmetamorphic winter flounder, a vertebrate that experiences a thyroid hormone (TH) induced metamorphosis during development. METHODS: A loss-of-signal strategy was utilized in which TH signaling was disrupted by inhibiting TH synthesis. Induced hypothyroidism was confirmed by radioimmunoassay. Reverse transcriptase PCR (RT-PCR), real-time quantitative PCR (qPCR), molecular cloning, non-isotopic in situ hybridization, western blot analysis, and indirect immunohistochemistry techniques were performed to analyze retinal thyroid hormone receptors (TR), photoreceptor production, and the phenotypic repertoire of differentiated retinal cells as a function of TH signaling status. RESULTS: Molecular bases for TH signaling were supported by retinal expression of TH receptors alpha and beta. TH-dependent transcriptional regulation of TRalpha but not TRbeta was indicated, with induced hypothyroidism producing an increase in TRalpha expression. Evidence for post-transcriptional regulation of retinal TRalpha was observed. The repertoire of inner retinal cell types in premetamorphic fish (a naturally low TH condition) matched that observed in the central retinas of both normal postmetamorphic fish (a naturally elevated TH condition) and postmetamorphic fish rendered hypothyroidic. In differentiated postmetamorphic retina there was no evidence for significant differences in opsin expression between normal and hypothyroidic animals. Induced hypothyroidism did, however, significantly affect the types of photoreceptors that were produced in postmetamorphic retina: as a hypothyroidic postmetamorphic retina grew or regenerated following injury, the phenotypic repertoire of newly-produced photoreceptors matched that observed for premetamorphic retina, in which rods, SWS2-expressing "blue" cones, and LWS-expressing "red" cones are absent, and only the RH2-expressing "green" cone type is present. The effects of induced hypothyroidism upon photoreceptor specification (manifestation of the rod lineage) and differentiation (expression of a particular opsin by specified cones) were apparently reversible. CONCLUSIONS: The results suggest a TH-dependent regulation of retinal TRalpha, a lack of TH-dependent regulation of the phenotypic identity of differentiated retinal cells, and the operation of similar cytogenic mechanisms during retinal growth and regeneration. The principal conclusion is that TH signaling significantly affects, in a targeted manner, the production of both rod and cone photoreceptors during retinal growth and regeneration.

Photoreceptor differentiation during retinal development, growth, and regeneration in a metamorphic vertebrate.

To test the hypothesis that growth and regeneration of the adult retina involves a mechanistic recapitulation of retinal development, the patterns of photoreceptor differentiation were investigated in the developing retina, as well as growing and regenerating adult retina, of a metamorphic vertebrate, the winter flounder. Only one opsin, of type RH2 (a "green" cone opsin), was expressed in premetamorphic (developing) retina, and a corresponding middle-wavelength visual pigment was observed. In premetamorphic retinas there was no evidence for any other cone opsins or pigments, rods, rod opsin expression, or rod visual pigment. In contrast, a rod opsin (RH1) and three cone opsins (SWS2, RH2, and LWS) were expressed in postmetamorphic (adult) retina, and these opsins were consistent with the observed repertoire of visual pigments. During postmetamorphic retinal growth and regeneration, cones were always produced before rods, but the different cone types were apparently produced simultaneously, suggesting that cone differentiation mechanisms might change after metamorphosis. The results support the hypothesis that photoreceptor differentiation during growth and regeneration of the adult retina involves a recapitulation of mechanisms that control the sequence of photoreceptor production during retinal development.

Flagellar quiescence in Chlamydomonas: Characterization and defective quiescence in cells carrying sup-pf-1 and sup-pf-2 outer dynein arm mutations.

Chlamydomonas reinhardtii can use their flagella for two distinct types of movement: swimming through liquid or gliding on a solid substrate. Cells switching from swimming to gliding motility undergo a reversible flagellar quiescence. This phenomenon appears to involve the outer dynein arms, since mutants having altered outer arm beta and gamma dyneins (sup-pf-1 and sup-pf-2) show a diminished ability to quiesce. Sup-pf-1 and sup-pf-2 were originally isolated as gain-of-function mutations that suppress the flagellar paralysis resulting from radial spoke or central pair defects. Defective quiescence is also a gain-of-function phenomenon, as cells completely lacking outer arm heavy chains show a normal quiescence phenotype. These data suggest that regulation of outer arm dynein activity is essential for flagellar quiescence and furthermore that regulation of quiescence involves a signal transduction pathway that shares elements with the radial spoke/central pair system.