Daniel A. Lee

Tanycytes of the hypothalamic median eminence form a diet-responsive neurogenic niche.

Adult hypothalamic neurogenesis has recently been reported, but the cell of origin and the function of these newborn neurons are unknown. Using genetic fate mapping, we found that median eminence tanycytes generate newborn neurons. Blocking this neurogenesis altered the weight and metabolic activity of adult mice. These findings reveal a previously unreported neurogenic niche in the mammalian hypothalamus with important implications for metabolism.

A genomic atlas of mouse hypothalamic development

The hypothalamus is a central regulator of many behaviors that are essential for survival, such as temperature regulation, food intake and circadian rhythms. However, the molecular pathways that mediate hypothalamic development are largely unknown. To identify genes expressed in developing mouse hypothalamus, we performed microarray analysis at 12 different developmental time points. We then conducted developmental in situ hybridization for 1,045 genes that were dynamically expressed over the course of hypothalamic neurogenesis. We identified markers that stably labeled each major hypothalamic nucleus over the entire course of neurogenesis and constructed a detailed molecular atlas of the developing hypothalamus. As a proof of concept of the utility of these data, we used these markers to analyze the phenotype of mice in which Sonic Hedgehog (Shh) was selectively deleted from hypothalamic neuroepithelium and found that Shh is essential for anterior hypothalamic patterning. Our results serve as a resource for functional investigations of hypothalamic development, connectivity, physiology and dysfunction.

A large-scale in vivo analysis reveals that TALENs are significantly more mutagenic than ZFNs generated using context-dependent assembly

Zinc-finger nucleases (ZFNs) and TAL effector nucleases (TALENs) have been shown to induce targeted mutations, but they have not been extensively tested in any animal model. Here, we describe a large-scale comparison of ZFN and TALEN mutagenicity in zebrafish. Using deep sequencing, we found that TALENs are significantly more likely to be mutagenic and induce an average of 10-fold more mutations than ZFNs. We observed a strong correlation between somatic and germ-line mutagenicity, and identified germ line mutations using ZFNs whose somatic mutations rates are well below the commonly used threshold of 1%. Guidelines that have previously been proposed to predict optimal ZFN and TALEN target sites did not predict mutagenicity in vivo. However, we observed a significant negative correlation between TALEN mutagenicity and the number of CpG repeats in TALEN target sites, suggesting that target site methylation may explain the poor mutagenicity of some TALENs in vivo. The higher mutation rates and ability to target essentially any sequence make TALENs the superior technology for targeted mutagenesis in zebrafish, and likely other animal models.

Functional implications of hypothalamic neurogenesis in the adult mammalian brain

Adult neurogenesis represents a striking example of structural plasticity in the mature brain. Research on adult mammalian neurogenesis today focuses almost exclusively on two areas: the subgranular zone (SGZ) in the dentate gyrus of the hippocampus, and the subventricular zone (SVZ) of the lateral ventricles. Numerous studies, however, have also reported adult neurogenesis in the hypothalamus, a brain structure that serves as a central homeostatic regulator of numerous physiological and behavioral functions, such as feeding, metabolism, body temperature, thirst, fatigue, aggression, sleep, circadian rhythms, and sexual behavior. Recent studies on hypothalamic neurogenesis have identified a progenitor population within a dedicated hypothalamic neurogenic zone. Furthermore, adult born hypothalamic neurons appear to play a role in the regulation of metabolism, weight, and energy balance. It remains to be seen what other functional roles adult hypothalamic neurogenesis may play. This review summarizes studies on the identification and characterization of neural stem/progenitor cells in the mammalian hypothalamus, in what contexts these stem/progenitor cells engage in neurogenesis, and potential functions of postnatally generated hypothalamic neurons.

The orphan nuclear hormone receptor ERRβ controls rod photoreceptor survival

Mutation of rod photoreceptor-enriched transcription factors is a major cause of inherited blindness. We identified the orphan nuclear hormone receptor estrogen-related receptor β (ERRβ) as selectively expressed in rod photoreceptors. Overexpression of ERRβ induces expression of rod-specific genes in retinas of wild-type as well as Nrl−/− mice, which lack rod photoreceptors. Mutation of ERRβ results in dysfunction and degeneration of rods, whereas inverse agonists of ERRβ trigger rapid rod degeneration, which is rescued by constitutively active mutants of ERRβ. ERRβ coordinates expression of multiple genes that are rate-limiting regulators of ATP generation and consumption in photoreceptors. Furthermore, enhancing ERRβ activity rescues photoreceptor defects that result from loss of the photoreceptor-specific transcription factor Crx. Our findings demonstrate that ERRβ is a critical regulator of rod photoreceptor function and survival, and suggest that ERRβ agonists may be useful in the treatment of certain retinal dystrophies.

Dietary and sex-specific factors regulate hypothalamic neurogenesis in young adult mice

The hypothalamus is the central regulator of a broad range of homeostatic and instinctive physiological processes, such as the sleep-wake cycle, food intake, and sexually dimorphic behaviors. These behaviors can be modified by various environmental and physiological cues, although the molecular and cellular mechanisms that mediate these effects remain poorly understood. Recently, it has become clear that both the juvenile and adult hypothalamus exhibit ongoing neurogenesis, which serve to modify homeostatic neural circuitry. In this report, we share new findings on the contributions of sex-specific and dietary factors to regulating neurogenesis in the hypothalamic mediobasal hypothalamus, a recently identified neurogenic niche. We report that high fat diet (HFD) selectively activates neurogenesis in the median eminence (ME) of young adult female but not male mice, and that focal irradiation of the ME in HFD-fed mice reduces weight gain in females but not males. These results suggest that some physiological effects of high fat diet are mediated by the stimulation of ME neurogenesis in a sexually dimorphic manner. We discuss these results in the context of recent advances in understanding the cellular and molecular mechanisms that regulate neurogenesis in postnatal and adult hypothalamus.

The LIM Homeodomain Factor Lhx2 Is Required for Hypothalamic Tanycyte Specification and Differentiation

Hypothalamic tanycytes, a radial glial-like ependymal cell population that expresses numerous genes selectively enriched in embryonic hypothalamic progenitors and adult neural stem cells, have recently been observed to serve as a source of adult-born neurons in the mammalian brain. The genetic mechanisms that regulate the specification and maintenance of tanycyte identity are unknown, but are critical for understanding how these cells can act as adult neural progenitor cells. We observe that LIM (Lin-11, Isl-1, Mec-3)-homeodomain gene Lhx2 is selectively expressed in hypothalamic progenitor cells and tanycytes. To test the function of Lhx2 in tanycyte development, we used an intersectional genetic strategy to conditionally delete Lhx2 in posteroventral hypothalamic neuroepithelium, both embryonically and postnatally. We observed that tanycyte development was severely disrupted when Lhx2 function was ablated during embryonic development. Lhx2-deficient tanycytes lost expression of tanycyte-specific genes, such as Rax, while also displaying ectopic expression of genes specific to cuboid ependymal cells, such as Rarres2. Ultrastructural analysis revealed that mutant tanycytes exhibited a hybrid identity, retaining radial morphology while becoming multiciliated. In contrast, postnatal loss of function of Lhx2 resulted only in loss of expression of tanycyte-specific genes. Using chromatin immunoprecipitation, we further showed that Lhx2 directly regulated expression of Rax, an essential homeodomain factor for tanycyte development. This study identifies Lhx2 as a key intrinsic regulator of tanycyte differentiation, sustaining Rax-dependent activation of tanycyte-specific genes while also inhibiting expression of ependymal cell-specific genes. These findings provide key insights into the transcriptional regulatory network specifying this still poorly characterized cell type.

A Zebrafish Genetic Screen Identifies Neuromedin U as a Regulator of Sleep/Wake States.

Neuromodulation of arousal states ensures that an animal appropriately responds to its environment and engages in behaviors necessary for survival. However, the molecular and circuit properties underlying neuromodulation of arousal states such as sleep and wakefulness remain unclear. To tackle this challenge in a systematic and unbiased manner, we performed a genetic overexpression screen to identify genes that affect larval zebrafish arousal. We found that the neuropeptide neuromedin U (Nmu) promotes hyperactivity and inhibits sleep in zebrafish larvae, whereas nmu mutant animals are hypoactive. We show that Nmu-induced arousal requires Nmu receptor 2 and signaling via corticotropin releasing hormone (Crh) receptor 1. In contrast to previously proposed models, we find that Nmu does not promote arousal via the hypothalamic-pituitary-adrenal axis, but rather probably acts via brainstem crh-expressing neurons. These results reveal an unexpected functional and anatomical interface between the Nmu system and brainstem arousal systems that represents a novel wake-promoting pathway.

Feed Your Head: Neurodevelopmental Control of Feeding and Metabolism

During critical periods of development early in life, excessive or scarce nutritional environments can disrupt the development of central feeding and metabolic neural circuitry, leading to obesity and metabolic disorders in adulthood. A better understanding of the genetic networks that control the development of feeding and metabolic neural circuits, along with knowledge of how and where dietary signals disrupt this process, can serve as the basis for future therapies aimed at reversing the public health crisis that is now building as a result of the global obesity epidemic. This review of animal and human studies highlights recent insights into the molecular mechanisms that regulate the development of central feeding circuitries, the mechanisms by which gestational and early postnatal nutritional status affects this process, and approaches aimed at counteracting the deleterious effects of early over- and underfeeding.

D-Aspartate and D-aspartate oxidase show selective and developmentally dynamic localization in mouse retina

D-Enantiomer amino acids, endogenously synthesized and degraded in mammals, participate in multiple developmental and physiological processes. We characterize both d-aspartate and its degradative enzyme (d-aspartate oxidase) in post-natal mouse retina. d-Aspartate attains a developmental peak of 886 nmol/g dry weight at 1-2 weeks post-natal while l-aspartate transiently declines, consistent with the de novo synthesis of d-aspartate. d-Aspartate is localized in many cell-types in different retinal layers, with a notable shift from outer retina to inner retina with time. d-Aspartate oxidase is localized to horizontal cells by in situ hybridization, immunohistochemistry, and histochemical staining methods. Given that final cell specification, differentiation, and synaptogenesis occur during the immediate 2-week post-natal period in the rodent retina and the dynamic appearance of d-aspartate and d-aspartate oxidase, we propose that d-aspartate plays a role in mammalian retinal development.