Lily Ng

TARGETED DISRUPTION OF NMDA RECEPTOR 1 GENE ABOLISHES NMDA RESPONSE AND RESULTS IN NEONATAL DEATH

In vitro studies have suggested that the NMDA receptor consists of an essential subunit, NR1, and various modulatory NR2 subunits. To test this hypothesis directly in vivo, we generated mice carrying a disrupted NR1 allele. NMDA-inducible increases in intracellular calcium and membrane currents were abolished in neurons from homozygous null mutants (NR1-/-). Thus, NR1 has a unique role, which cannot be substituted by any other subunit, in determining the activity of the endogenous NMDA receptor. A concomitant reduction in levels of NR2B but not NR2A occurred in NR1-/- mice, demonstrating that there is an interdependence of subunit expression. NR1-/- mice died 8-15 hr after birth, indicating a vital neonatal function for the NMDA receptor. Although the NMDA receptor has been implicated in several aspects of neurodevelopment, overall neuroanatomy of NR1-/- mice appeared normal. Pathological evidence suggested that respiratory failure was the ultimate cause of death.

Retinoid-related orphan nuclear receptor RORβ is an early-acting factor in rod photoreceptor development

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 β 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.

The Thyroid Hormone Receptor β Gene: Structure and Functions in the Brain and Sensory Systems

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.

Making sense with thyroid hormone—the role of T 3 in auditory development

The senses are our window to the world, our interface with the habitat in which we live in and the basis for our communication with each other. Although sensory systems are not generally viewed as major targets of endocrine regulation, sensory development is profoundly influenced by thyroid hormone (T3) signalling. In this article, we discuss this developmental role of T3 and highlight the auditory system as the best-studied example of the interplay between systemic and local tissue mechanisms by which T3 stimulates the onset of sensory function. Several genes that mediate the action of T3 are known to promote sensory development in mice, including genes that encode T3 receptors and deiodinase enzymes that amplify or deplete levels of T3. We also discuss the current knowledge of sensory defects in human genetic disorders in which T3 signalling is impaired. As sensory input provides the only means of acquiring information from the environment, the stimulation of sensory development is one of the most fundamental functions of T3 signalling.

Developmental expression of thyroid hormone receptor β2 protein in cone photoreceptors in the mouse

Thyroid hormone receptor &bgr;2 (TR&bgr;2) controls the patterning of cone opsin photopigments that mediate colour vision. We raised an antiserum against TR&bgr;2 to study cone photoreceptor development by western blot and immunostaining analyses. TR&bgr;2-positive cells first appeared between embryonic day 10 (E10) and E12. Numbers increased until near birth, correlating with generation of the cone population. At birth, signals decreased until postnatal day 10, then declined to very low levels in adulthood. TR&bgr;2-positive cells were initially dispersed but became aligned at the edge of the outer neuroblastic layer by E15. Postnatally, these cells migrated inwardly until postnatal day 10, then outwardly to the edge of the outer nuclear layer, the location of mature cones. TR&bgr;2 represents a functionally unique marker for cone development.

An isoform of retinoid-related orphan receptor β directs differentiation of retinal amacrine and horizontal interneurons

Amacrine and horizontal interneurons integrate visual information as it is relayed through the retina from the photoreceptors to the ganglion cells. The early steps that generate these interneuron networks remain unclear. Here we show that a distinct RORβ1 isoform encoded by the retinoid-related orphan nuclear receptor β gene (Rorb) is critical for both amacrine and horizontal cell differentiation in mice. A fluorescent protein cassette targeted into Rorb revealed RORβ1 as a novel marker of immature amacrine and horizontal cells and of undifferentiated, dividing progenitor cells. RORβ1-deficient mice lose expression of pancreas-specific transcription factor 1a (Ptf1a) but retain forkhead box n4 factor (Foxn4), two early-acting factors necessary for amacrine and horizontal cell generation. RORβ1 and Foxn4 synergistically induce Ptf1a expression, suggesting a central role for RORβ1 in a transcriptional hierarchy that directs this interneuron differentiation pathway. Moreover, ectopic RORβ1 expression in neonatal retina promotes amacrine cell differentiation.

Deletion of the Thyroid Hormone–Activating Type 2 Deiodinase Rescues Cone Photoreceptor Degeneration but Not Deafness in Mice Lacking Type 3 Deiodinase

Type 2 deiodinase amplifies and type 3 deiodinase depletes levels of the active form of thyroid hormone, triiodothyronine. Given the opposing activities of these enzymes, we tested the hypothesis that they counteract each other’s developmental functions by investigating whether deletion of type 2 deiodinase (encoded by Dio2) modifies sensory phenotypes in type 3 deiodinase-deficient (Dio3−/−) mice. Dio3−/− mice display degeneration of retinal cones, the photoreceptors that mediate daylight and color vision. In Dio2−/− mice, cone function was largely normal but deletion of Dio2 in Dio3−/− mice markedly recovered cone numbers and electroretinogram responses, suggesting counterbalancing roles for both enzymes in cone survival. Both Dio3−/− and Dio2−/− strains exhibit deafness with cochlear abnormalities. In Dio3−/−;Dio2−/− mice, deafness was exacerbated rather than alleviated, suggesting unevenly balanced actions by these enzymes during auditory development. Dio3−/− mice also exhibit an atrophic thyroid gland, low thyroxine, and high triiodothyronine levels, but this phenotype was ameliorated in Dio3−/−;Dio2−/− mice, indicating counterbalancing roles for the enzymes in determining the thyroid hormone status. The results suggest that the composite action of these two enzymes is a critical determinant in visual and auditory development and in setting the systemic thyroid hormone status.

Analysis of Nuclear Receptor Function in the Mouse Auditory System

Publisher Summary This chapter focuses on the analysis of nuclear receptor function in the mouse auditory system. Hearing is a prominent example of a sensory function that is regulated by nuclear receptors. Thyroid hormone receptors are essential for the development of hearing and other receptors. Although the requirement for a nuclear receptor gene may be suggested in some human inherited diseases with an incidence of hearing loss, the investigation of the underlying mechanisms requires a suitable model species. The mouse provides a valuable model because of the availability of spontaneous mutants and because it is amenable to genetic manipulation. Selected methods are presented in the chapter for studying the mouse auditory system, with a focus on the cochlea that contains the sensory hair cells. The cochlea within the inner ear houses a microcosm of cellular and physiological machinery that mediates auditory transduction. The overall function of the cochlea is highly dependent on its biomechanical structure. The easiest access to the cochlea is gained from the dorsal or upper surface of the base of the skull. Although increasing numbers of genes involved in cochlear function are being identified, the analysis of the regulation of these genes by nuclear receptors or other factors is still at an elementary stage.