Color vision is facilitated by distinct populations of cone photoreceptors in the retina. In rodents, cones expressing different opsin photopigments are sensitive to middle (M, 'green') and short (S, 'blue') wavelengths, and are differentially distributed across the retina. The mechanisms that control which opsin is expressed in a particular cone are poorly understood, but previous in vitro studies implicated thyroid hormone in cone differentiation. Thyroid hormone receptor beta 2 (TR beta 2) is a ligand-activated transcription factor that is expressed in the outer nuclear layer of the embryonic retina. Here we delete Thrb (encoding Tr beta 2) in mice, causing the selective loss of M-cones and a concomitant increase in S-opsin immunoreactive cones. Moreover, the gradient of cone distribution is disturbed, with S-cones becoming widespread across the retina. The results indicate that cone photoreceptors throughout the retina have the potential to follow a default S-cone pathway and reveal an essential role for Tr beta 2 in the commitment to an M-cone identity. Our findings raise the possibility that Thrb mutations may be associated with human cone disorders.

Most mammals have two types of cone photoreceptors, which contain either medium wavelength (M) or short wavelength (S) opsin. The number and spatial organization of cone types varies dramatically among species, presumably to fine-tune the retina for different visual environments. In the mouse, S-and M-opsin are expressed in an opposing dorsal-ventral gradient. We previously reported that cone opsin patterning requires thyroid hormone ␤2, a nuclear hormone receptor that regulates transcription in conjunction with its ligand, thyroid hormone (TH). Here we show that exogenous TH inhibits S-opsin expression, but activates M-opsin expression. Binding of endogenous TH to TR␤2 is required to inhibit S-opsin and to activate M-opsin. TH is symmetrically distributed in the retina at birth as S-opsin expression begins, but becomes elevated in the dorsal retina at the time of M-opsin onset (postnatal day 10). Our results show that TH is a critical regulator of both S-opsin and M-opsin, and suggest that a TH gradient may play a role in establishing the gradient of M-opsin. These results also suggest that the ratio and patterning of cone types may be determined by TH availability during retinal development.photoreceptor ͉ retinal development ͉ nuclear hormone receptor T hyroid hormone (TH) is secreted mostly as thyroxine (T4) from the thyroid gland and converted locally to a transcriptionally active form, 3,5,3Ј-triiodothyronine (T3), by iodothyronine deiodinating enzymes (1). T3 receptors bind to DNA regulatory elements, often as heterodimers with retinoid X receptors (RXRs), to activate or repress transcription of target genes (for review, see refs. 2 and 3). T3 regulates diverse developmental processes in multiple regions of the central nervous system including the hippocampus, cerebral cortex, inner ear, and cerebellum (4-8). Numerous developmental processes are TH-dependent, including cell differentiation, migration, and dendritic growth (2, 3, 9-11). Perturbations in maternal or fetal TH in humans can lead to clinical syndromes ranging from severe mental retardation and hearing loss to mild or moderate deficits in motor skills, language, memory, and attention (12)(13)(14).In the retina, the TH ␤2 receptor isoform (TR␤2) is restricted to cone photoreceptors (15, 16), and we previously showed that it is a critical regulator of late-stage cone differentiation. In the first stage of cone differentiation, cones exit the cell cycle and begin to express specific transcription factors, including cone rod homeobox (CRX), TR␤2,. In the second stage of differentiation, cones express opsin and form specific synaptic connections. Most mammalian cones express short wavelength (S-opsin) or medium wavelength (M-opsin) opsin. However, the spatial arrangement of cone types varies considerably among species. In mice, S-opsin expression begins just before birth and is expressed predominantly in ventral cones. By contrast, M-opsin expression begins at the end of the first postnatal week and is expressed predominantly in dorsal cones (21, 22...

Maturation of the mammalian nervous system requires adequate provision of thyroid hormone and mechanisms that enhance tissue responses to the hormone. Here, we report that the development of cones, the photoreceptors for daylight and color vision, requires protection from thyroid hormone by type 3 deiodinase, a thyroid hormone-inactivating enzyme. Type 3 deiodinase, encoded by Dio3, is expressed in the immature mouse retina. In Dio3 −/− mice, ~80% of cones are lost through neonatal cell death. Cones that express opsin photopigments for response to both short (S) and medium-long (M) wavelength light are lost. Rod photoreceptors, which mediate dim light vision, remain largely intact. Excessive thyroid hormone in wild type pups also eliminates cones. Cone loss is mediated by cone-specific thyroid hormone receptor β2 (TRβ2) as deletion of TRβ2 rescues cones in Dio3 −/− mice. However, rescued cones respond to short but not longer wavelength light because TRβ2 under moderate hormonal stimulation normally induces M opsin and controls the patterning of M and S opsins over the retina. The results suggest that type 3 deiodinase limits hormonal exposure of the cone to levels that safeguard both cone survival and the patterning of opsins that is required for cone function.

Thyroid hormone is necessary for cochlear development and auditory function, but the factors that control these processes are poorly understood. Previous evidence indicated that in mice, the serum supply of thyroid hormone is augmented within the cochlea itself by type 2 deiodinase, which amplifies the level of T(3), the active form of thyroid hormone, before the onset of hearing. We now report that type 3 deiodinase, a thyroid hormone-inactivating enzyme encoded by Dio3, is expressed in the immature cochlea before type 2 deiodinase. Dio3-/- mice display auditory deficits and accelerated cochlear differentiation, contrasting with the retardation caused by deletion of type 2 deiodinase. The Dio3 mRNA expression pattern in the greater epithelial ridge, stria vascularis, and spiral ganglion partly overlaps with that of thyroid hormone receptor beta (TRbeta), the T(3) receptor that is primarily responsible for auditory development. The proposal that type 3 deiodinase prevents premature stimulation of TRbeta was supported by deleting TRbeta, which converted the Dio3-/- cochlear phenotype from one of accelerated to one of delayed differentiation. The results indicate a protective role for type 3 deiodinase in hearing. The auditory system illustrates the considerable extent to which tissues can autoregulate their developmental response to thyroid hormone through both type 2 and 3 deiodinases.

Color vision requires the expression of opsin photopigments with different wavelength sensitivities in retinal cone photoreceptors. The basic color visual system of mammals is dichromatic, involving differential expression in the cone population of two opsins with sensitivity to short (S, blue) or medium (M, green) wavelengths. However, little is known of the factors that directly activate these opsin genes and thereby contribute to the S or M opsin identity of the cone. We report that the orphan nuclear receptor RORbeta (retinoid-related orphan receptor beta) activates the S opsin gene (Opn1sw) through binding sites upstream of the gene. RORbeta lacks a known physiological ligand and activates the Opn1sw promoter modestly alone but strongly in synergy with the retinal cone-rod homeobox factor (CRX), suggesting a cooperative means of enhancing RORbeta activity. Comparison of wild-type and mutant lacZ reporter transgenes showed that the RORbeta-binding sites in Opn1sw are required for expression in mouse retina. RORbeta-deficient mice fail to induce S opsin appropriately during postnatal cone development. Photoreceptors in these mice also lack outer segments, indicating additional functions for RORbeta in photoreceptor morphological maturation. The results identify Opn1sw as a target gene for RORbeta and suggest a key role for RORbeta in regulating opsin expression in the color visual system.

Rods and cones are morphologically and developmentally distinct photoreceptor types with different functions in vision. Cones mediate daylight and color vision and in most mammals express M and S opsin photopigments for sensitivity to medium-long and short light wavelengths, respectively. Rods mediate dim light vision and express rhodopsin photopigment. The transcription factor networks that direct differentiation of each photoreceptor type are incompletely defined. Here, we report that Rorb ؊/؊ mice lacking retinoid-related orphan nuclear receptor ␤ lose rods but overproduce primitive S cones that lack outer segments. The phenotype reflects pronounced plasticity between rod and cone lineages and resembles that described for Nrl ؊/؊ mice lacking neural retina leucine zipper factor. Rorb ؊/؊ mice lack Nrl expression and reexpression of Nrl in Rorb ؊/؊ mice converts cones to rod-like cells. Thus, Rorb directs rod development and does so at least in part by inducing the Nrl-mediated pathway of rod differentiation.cone ͉ differentiation R ods and cones are distinct receptor cell types that mediate dim and bright light vision, respectively. In the mouse, cones are generated between midgestation and birth (1) and subpopulations differentially express M and S opsin photopigments for sensitivity to medium-long and short light wavelengths, respectively (2). Rods express rhodopsin and greatly outnumber cones in mice. Rod generation lags behind that of cones and is more protracted, lasting until about a week after birth. Rods, cones, and other retinal cell types are generated in a stereotypical order from multipotent progenitors, and it has been proposed that a combination of transcription factor activities and external signals at a given developmental stage prompts progenitors to enter specific differentiation pathways (3, 4).The transcription factors that direct the generation of rod and cone precursors and the terminal differentiation of these cell types are incompletely defined. During terminal differentiation of cones, thyroid hormone receptor TR␤2 is required for M opsin induction such that without TR␤2, cones express only S opsin (5). Factors that promote rod differentiation and survival include leucine zipper protein Nrl (6), orphan nuclear receptor Nr2e3 (7, 8), homeodomain proteins Crx and Otx2 (9-11), and retinoblastoma protein Rb (12,13). Nrl induces Nr2e3 expression and these two genes define a transcriptional hierarchy for rod differentiation. Nrl Ϫ/Ϫ mice overproduce S cones at the expense of rods (6) whereas ectopic Nrl expression converts cones to rods (14). Nr2e3 deficiency causes an enhanced cone phenotype and misexpression of cone genes in rods, suggesting that Nr2e3 represses cone genes to maintain the rod phenotype (15-19). Human NRL and NR2E3 mutations result in retinopathy phenotypes (8,20). The above findings suggest that rod and cone precursors share a default differentiation program as cones and that rod differentiation requires the action of additional transcription factors.The Rorb gene encodin...

AimsRecombinant Neuregulin (NRG)-1β has multiple beneficial effects on cardiac myocytes in culture, and has potential as a clinical therapy for heart failure (HF). A number of factors may influence the effect of NRG-1β on cardiac function via ErbB receptor coupling and expression. We examined the effect of the NRG-1β isoform, glial growth factor 2 (GGF2), in rats with myocardial infarction (MI) and determined the impact of high-fat diet as well as chronicity of disease on GGF2 induced improvement in left ventricular systolic function. Potential mechanisms for GGF2 effects on the remote myocardium were explored using microarray and proteomic analysis.Methods and ResultsRats with MI were randomized to receive vehicle, 0.625 mg/kg, or 3.25 mg/kg GGF2 in the presence and absence of high-fat feeding beginning at day 7 post-MI and continuing for 4 weeks. Residual left ventricular (LV) function was improved in both of the GGF2 treatment groups compared with the vehicle treated MI group at 4 weeks of treatment as assessed by echocardiography. High-fat diet did not prevent the effects of high dose GGF2. In experiments where treatment was delayed until 8 weeks after MI, high but not low dose GGF2 treatment was associated with improved systolic function. mRNA and protein expression analysis of remote left ventricular tissue revealed a number of changes in myocardial gene and protein expression altered by MI that were normalized by GGF2 treatment, many of which are involved in energy production.ConclusionsThis study demonstrates that in rats with MI induced systolic dysfunction, GGF2 treatment improves cardiac function. There are differences in sensitivity of the myocardium to GGF2 effects when administered early vs. late post-MI that may be important to consider in the development of GGF2 in humans.

Thyroid hormone profoundly influences the development of the vertebrate nervous system. The thyroid hormone receptor beta gene (Thrb) is a key mediator of many of these actions. The Thrb gene is complex, spanning up to 400 kb in mammals, and differentially expresses distinct receptor subtypes through independent tissue-specific promoters and alternative splicing. These receptors serve a range of functions in the brain as well as particularly sensitive functions in the auditory and visual sensory systems. The Thrb gene illustrates how versatility in neurodevelopmental control can be achieved at the receptor level.