Sarah K. McMenamin

Thyroid hormone–dependent adult pigment cell lineage and pattern in zebrafish

Origin of fish pigment cell for pattern Zebrafish stripes arise from the interactions of pigment cells: black melanophores, iridescent iridophores, and yellow-orange xanthophores. Melanophores and iridophores develop from nerve-associated stem cells, but the origin of xanthophores is unclear. Two studies now reveal that adult xanthophores originate from xanthophores in embryonic and larval fish, when they proliferate to cover the skin before the arrival of black and silver cells in a striped arrangement. Mahalwar et al. show that xanthophores change their final shape and color depending on their location. In black cells, xanthophores appear faint and stellate, but in silver cells, they are bright and compact. Precise superposition creates the blue and golden colors. McMenamin et al. observe the loss of pigment in embryonic xanthophores and the later reappearance in the adult. They show that redifferentiation depends on the thyroid hormone that also limits melanophore population expansion. Science, this issue p. 1362 and p. 1358 Zebrafish adult pigment cells arise from several lineages and require thyroid hormone for pattern development. Pigment patterns are useful for elucidating fundamental mechanisms of pattern formation and how these mechanisms evolve. In zebrafish, several pigment cell classes interact to generate stripes, yet the developmental requirements and origins of these cells remain poorly understood. Using zebrafish and a related species, we identified roles for thyroid hormone (TH) in pigment cell development and patterning, and in postembryonic development more generally. We show that adult pigment cells arise from distinct lineages having distinct requirements for TH and that differential TH dependence can evolve within lineages. Our findings demonstrate critical functions for TH in determining pigment pattern phenotype and highlight the potential for evolutionary diversification at the intersection of developmental and endocrine mechanisms.

Dwarfism and increased adiposity in the gh1 mutant zebrafish vizzini.

Somatic growth and adipogenesis are closely associated with the development of obesity in humans. In this study, we identify a zebrafish mutant, vizzini, that exhibits both a severe defect in somatic growth and increased accumulation of adipose tissue. Positional cloning of vizzini revealed a premature stop codon in gh1. Although the effects of GH are largely through igfs in mammals, we found no decrease in the expression of igf transcripts in gh1 mutants during larval development. As development progressed, however, we found overall growth to be progressively retarded and the attainment of specific developmental stages to occur at abnormally small body sizes relative to wild type. Moreover, both subcutaneous (sc) and visceral adipose tissues underwent precocious development in vizzini mutants, and at maturity, the sizes of different fat deposits were greatly expanded relative to wild type. In vivo confocal imaging of sc adipose tissue (SAT) expansion revealed that vizzini mutants exhibit extreme enlargement of adipocyte lipid droplets without a corresponding increase in lipid droplet number. These findings suggest that GH1 signaling restricts SAT hypertrophy in zebrafish. Finally, nutrient deprivation of vizzini mutants revealed that SAT mobilization was greatly diminished during caloric restriction, further implicating GH1 signaling in adipose tissue homeostasis. Overall, the zebrafish gh1 mutant, vizzini, exhibits decreased somatic growth, increased adipose tissue accumulation, and disrupted adipose plasticity after nutrient deprivation and represents a novel model to investigate the in vivo dynamics of vertebrate obesity.

Developmental dynamics of Ambystoma tigrinum in a changing landscape

BackgroundLoss of pond habitat is catastrophic to aquatic larval amphibians, but even reduction in the amount of time a breeding site holds water (hydroperiod) can influence amphibian development and limit reproductive success. Using the landscape variation of a glacial valley in the Greater Yellowstone Ecosystem as the context for a natural experiment, we examined variation in growth pattern and life history of the salamander Ambystoma tigrinum melanostictum and determined how these developmental characteristics varied with hydroperiod over several summers.ResultsIn ponds that dried early in the season, maximum larval size was reduced relative to the sizes achieved in permanent ponds. Ephemeral ponds were associated with early metamorphosis at small body sizes, while permanent ponds facilitated longer larval periods and later metamorphosis. Paedomorphosis resulted from indefinite metamorphic postponement, and was identified only in the most permanent environments. Patterns of growth and allometry were similar between ponds with different hydroperiods, but considerable life history variation was derived from modulating the timing of and size at metamorphosis. Considering maximum rates of growth and inferring the minimum size at metamorphosis across 25 ponds over the course of three years, we calculated that hydroperiods longer than three months are necessary to support these populations through metamorphosis and/or reproductive maturity.ConclusionsLandscape heterogeneity fosters life history variation in this natural population. Modulation of the complex ambystomatid life cycle allows this species to survive in unpredictable environments, but current trends towards rapid pond drying will promote metamorphosis at smaller sizes and could eliminate the paedomorphic phenotype from this region. Metamorphosis at small size is has been linked to altered fitness traits, including reduced survival and fecundity. Thus, widespread environmental truncation of larval periods may lead to decreased population persistence. We found that the hydroperiods of many ponds in this region are now shorter than the developmental period required for larvae to reach the minimum size for metamorphosis; these locations serve as reproductive sinks that may be detrimental for persistence of the species in the region.

Working with zebrafish at postembryonic stages.

As the processes of embryogenesis become increasingly well understood, there is growing interest in the development that occurs at later, postembryonic stages. Postembryonic development holds tremendous potential for discoveries of both fundamental and translational importance. Zebrafish, which are small, rapidly and externally developing, and which boast a wealth of genetic resources, are an outstanding model of vertebrate postembryonic development. Nonetheless, there are specific challenges posed by working with zebrafish at these stages, and this chapter is meant to serve as a primer for those working with larval and juvenile zebrafish. Since accurate staging is critical for high-quality results and experimental reproducibility, we outline best practices for reporting postembryonic developmental progress. Emphasizing the importance of accurate staging, we present new data showing that rates of growth and size-stage relationships can differ even between wild-type strains. Finally, since rapid and uniform development is particularly critical when working at postembryonic stages, we briefly describe methods that we use to achieve high rates of growth and developmental uniformity through postembryonic stages in both wild-type and growth-compromised zebrafish.

MicroCT-based phenomics in the zebrafish skeleton reveals virtues of deep phenotyping in a distributed organ system

Phenomics, which ideally involves in-depth phenotyping at the whole-organism scale, may enhance our functional understanding of genetic variation. Here, we demonstrate methods to profile hundreds of phenotypic measures comprised of morphological and densitometric traits at a large number of sites within the axial skeleton of adult zebrafish. We show the potential for vertebral patterns to confer heightened sensitivity, with similar specificity, in discriminating mutant populations compared to analyzing individual vertebrae in isolation. We identify phenotypes associated with human brittle bone disease and thyroid stimulating hormone receptor hyperactivity. Finally, we develop allometric models and show their potential to aid in the discrimination of mutant phenotypes masked by alterations in growth. Our studies demonstrate virtues of deep phenotyping in a spatially distributed organ system. Analyzing phenotypic patterns may increase productivity in genetic screens, and facilitate the study of genetic variants associated with smaller effect sizes, such as those that underlie complex diseases.

Ancient DNA assessment of tiger salamander population in Yellowstone National Park.

Recent data indicates that blotched tiger salamanders (Ambystoma tigrinum melanostictum) in northern regions of Yellowstone National Park are declining due to climate-related habitat changes. In this study, we used ancient and modern mitochondrial haplotype diversity to model the effective size of this amphibian population through recent geological time and to assess past responses to climatic changes in the region. Using subfossils collected from a cave in northern Yellowstone, we analyzed >700 base pairs of mitochondrial sequence from 16 samples ranging in age from 100 to 3300 years old and found that all shared an identical haplotype. Although mitochondrial diversity was extremely low within the living population, we still were able to detect geographic subdivision within the local area. Using serial coalescent modelling with Bayesian priors from both modern and ancient genetic data we simulated a range of probable population sizes and mutation rates through time. Our simulations suggest that regional mitochondrial diversity has remained relatively constant even through climatic fluctuations of recent millennia.

First Extinctions on Land

The golden toad (Bufo periglenes) disappeared from Costa Rica in 1989 and became the first terrestrial extinction to be linked to climate change. Like the first marine extinction attributed to climate change (see chapter 7), the extinction of the golden toad was linked to El Nino events. The marine extinction is irrefutably linked to coral bleaching, but the causes of the golden toad extinction are far more controversial. Golden toad sightings have been reported in Guatemala since the 1980s, but these sightings have never been confirmed. Although there is some hope that residual populations still survive, Bufo periglenes is currently listed as extinct in the International Union for Conservation of Nature (IUCN) Red List, and the cause of the extinction is hotly debated.

Thyroid hormone modulation during zebrafish development recapitulates evolved diversity in danionin jaw protrusion mechanics

One of three vertebrates belongs to a fish lineage for which protrusile jaws are a synapomorphy. Identifying the developmental determinants of protrusion ability will improve our understanding of an important area of evolutionary diversification. The high water viscosities experienced by tiny fish larvae inhibit the viability of protrusile jaws. In the zebrafish protrusion does not arise until after metamorphosis. Fish metamorphosis typically includes significant changes in trophic morphology, accompanies a shift in feeding niche and coincides with increased thyroid hormone production. We tested whether thyroid hormone affects the development of zebrafish feeding mechanics. We found that it affected all developmental stages examined, but that these effects were most pronounced after metamorphosis. Thyroid hormone levels affected the development of jaw morphology, feeding mechanics, shape variation and cranial ossification. Adult zebrafish utilize protrusile jaws, but an absence of thyroid hormone eliminated postmetamorphic remodeling of the premaxilla and the premaxillary structure that permits protrusion never formed. The premaxillae of late juvenile and adult zebrafish are similar to those found in the adults of other Danio species. Premaxillae from early juvenile zebrafish and hypothyroid adult zebrafish resemble those from adults in the genera Danionella, Devario and Microdevario that show little to no jaw protrusion.

microCT-Based Skeletal Phenomics in Zebrafish Reveals Virtues of Deep Phenotyping at the Whole-Organism Scale

Phenomics, which ideally involves in-depth phenotyping at the whole-organism scale, may enhance our functional understanding of genetic variation. Here, we demonstrate methods to profile hundreds of measures comprised of morphological and densitometric traits from a large number sites in the axial skeleton of adult zebrafish. We show the potential for vertebral patterns to confer heightened sensitivity, with similar specificity, in discriminating mutant populations compared to analyzing individual vertebrae in isolation. We identify phenotypes associated with human brittle bone disease and thyroid stimulating hormone receptor hyperactivity. Finally, we develop allometric models and show their potential to aid in the discrimination of mutant phenotypes masked by alterations in growth. Our studies demonstrate virtues of deep phenotyping in a spatially distributed organ. Analyzing phenotypic patterns may increase productivity in genetic screens, and could facilitate the study of genetic variants associated with smaller effect sizes, such as those that underlie complex diseases.Phenomics—in-depth phenotyping at the whole-organism scale—holds promise to enhance our fundamental understanding of genes and genomic variation, yet methods in vertebrates are limited. Here, we demonstrate rapid whole-body profiling of hundreds of traits in the axial skeleton of adult zebrafish. We show the potential for vertebral patterns to confer heightened sensitivity, with similar specificity, in discriminating mutant populations compared to analyzing individual vertebrae in isolation, even when the latter is performed at higher resolution. We identify phenotypes associated with human brittle bone disease and thyroid stimulating hormone receptor hyperactivity. Finally, we develop allometric models and show their potential to discriminate mutant phenotypes masked by growth alterations in growth. Our studies demonstrate virtues of whole-body phenomic pattern analysis in a single organ system. The high sensitivity may increase productivity in genetic screens, and facilitate the study genetic variants of smaller effect size, such as those that underlie complex diseases.

Reply to Patla et al.: Amphibian habitat and populations in Yellowstone damaged by drought and global warming

Patla et al. (1) suggest that loss of amphibian habitat is distinct from amphibian status, and they claim that we (2) wrongly define populations. Ours is a functional definition of population that serves as a biologically relevant metric of species presence. We demonstrate that this amphibian metapopulation is being destabilized as habitat is lost.