Helen Nilsson Sköld

Rapid color change in fish and amphibians – function, regulation, and emerging applications

Physiological color change is important for background matching, thermoregulation as well as signaling and is in vertebrates mediated by synchronous intracellular transport of pigmented organelles in chromatophores. We describe functions of and animal situations where color change occurs. A summary of endogenous and external factors that regulate this color change in fish and amphibians is provided, with special emphasis on extracellular stimuli. We describe not only color change in skin, but also highlight studies on color change that occurs using chromatophores in other areas such as iris and on the inside of the body. In addition, we discuss the growing field that applies melanophores and skin color in toxicology and as biosensors, and point out research areas with future potential.

Chapter 6 New Insights into Melanosome Transport in Vertebrate Pigment Cells

Pigment cells of lower vertebrates provide an excellent model to study organelle transport as they specialize in the translocation of pigment granules in response to defined chemical cues. This review will focus on the well-studied melanophore/melanocyte systems in fish, amphibians, and mammals. We will describe the roles of melanin, melanophores, and melanocytes in animals, current views on how the three motor proteins dynein, kinesin, and myosin-V are involved in melanosome transport along microtubules and actin filaments, and how signal transduction pathways regulate the activities of the motors to achieve aggregation and dispersion of melanosomes. We will also describe how melanosomes are transferred to surrounding skin cells in amphibians and mammals. Comparative studies have revealed that the ability of physiological color change is lost during evolution while the importance of morphological color change, mainly via transfer of pigment to surrounding skin cells, increases. In humans, pigment mainly has a role in protection against ultraviolet radiation, but also perhaps in the immune system.

Chromatic interaction between egg pigmentation and skin chromatophores in the nuptial coloration of female two-spotted gobies

SUMMARY In two-spotted gobies (Gobiusculus flavescens Fabricius 1779), females develop an orange belly as they approach sexual maturity. Bright belly coloration is preferred by males and has been suggested to act as a female ornament. This coloration is unusual in that it originates partly from pigmentation of the abdominal skin but also from strongly pigmented gonads directly visible through the skin. In addition, females have been observed to temporarily become more colourful during courtship and competition. To understand how gonad and skin pigmentation interact in this nuptial coloration, the potential for colour modification via regulation of skin chromatophores was investigated. Noradrenaline caused aggregation of chromatophore pigment and was used to experimentally reduce the contribution of skin chromatophores to the nuptial coloration. Chromatophore pigment aggregation caused bellies to become less colourful and abdominal skin biopsies to become less colourful and more transparent. There was a strong positive relationship between belly coloration and the coloration of the underlying gonads. This shows that belly coloration honestly reflects egg pigmentation, mainly because the transparency of the abdominal skin allows other fish to see the gonads directly. Interestingly, when noradrenaline caused pigment to aggregate and thereby increased the transparency of the skin, the relationship between belly and gonad coloration weakened. We conclude that female G. flavescens have a potential to use skin chromatophores to rapidly alter their nuptial coloration, thereby affecting the efficacy with which information about gonad coloration is conveyed.

Induced cell proliferation in putative haematopoietic tissues of the sea star, Asterias rubens (L.)

SUMMARY The coelomic fluid of the echinoderm Asterias rubens possesses large populations of circulating coelomocytes. This study aimed to expand the knowledge about the haematopoietic sources of these cells. Injection of the immune-stimulating molecules lipopolysaccharide (LPS) and concanavalin A (ConA) resulted in an increase in coelomocytes. To investigate if these molecules induce cell proliferation in putative haematopoietic tissues (HPTs), short-term exposure of the substitute nucleotide 5-bromo-2′-deoxyuridine (BrdU) was conducted. Immunohistochemical analysis, using fluorescein-labelled antibodies to trace BrdU, showed pronounced cell division in the coelomic epithelium and axial organ. In the pyloric caeca, not considered as an HPT, proliferation was not detected. BrdU labelling of monolayers of cells obtained by collagenase treatment of coelomic epithelium, axial organ and Tiedemann body revealed induced cell proliferation in response to both LPS and ConA while proliferation of pyloric caeca and circulating coelomocytes remained sparse. By using confocal microscopy it was observed that both the morphology and functional behaviour of cells released from explants of coelomic epithelium showed high similarity to those of circulating phagocytes. It was concluded that the increased coelomocyte numbers observed in response to LPS and ConA were reflected in an induced cell proliferation in coelomic epithelium, axial organ and Tiedemann body, which reinforces the idea that these organs are HPTs and the sources of coelomocyte renewal.

Phenotypic Plasticity Confers Multiple Fitness Benefits to a Mimic

Animal communication is often deceptive; however, such dishonesty can become ineffective if it is used too often, is used out of context, or is too easy to detect [1-3]. Mimicry is a common form of deception, and most mimics gain the greatest fitness benefits when they are rare compared to their models [3, 4]. If mimics are encountered too frequently or if their model is absent, avoidance learning of noxious models is disrupted (Batesian mimicry [3]), or receivers become more vigilant and learn to avoid perilous mimics (aggressive mimicry [4]). Mimics can moderate this selective constraint by imperfectly resembling multiple models [5], through polymorphisms [6], or by opportunistically deploying mimetic signals [1, 7]. Here we uncover a novel mechanism to escape the constraints of deceptive signaling: phenotypic plasticity allows mimics to deceive targets using multiple guises. Using a combination of behavioral, cell histological, and molecular methods, we show that a coral reef fish, the dusky dottyback (Pseudochromis fuscus), flexibly adapts its body coloration to mimic differently colored reef fishes and in doing so gains multiple fitness benefits. We find that by matching the color of other reef fish, dottybacks increase their success of predation upon juvenile fish prey and are therefore able to deceive their victims by resembling multiple models. Furthermore, we demonstrate that changing color also increases habitat-associated crypsis that decreases the risk of being detected by predators. Hence, when mimics and models share common selective pressures, flexible imitation of models might inherently confer secondary benefits to mimics. Our results show that phenotypic plasticity can act as a mechanism to ease constraints that are typically associated with deception. VIDEO ABSTRACT.

Embryonic response to long-term exposure of the marine crustacean Nephrops norvegicus to ocean acidification and elevated temperature.

Due to anthropogenic CO2 emissions, our oceans have gradually become warmer and more acidic. To better understand the consequences of this, there is a need for long-term (months) and multistressor experiments. Earlier research demonstrates that the effects of global climate change are specific to species and life stages. We exposed berried Norway lobsters (Nephrops norvegicus), during 4 months to the combination of six ecologically relevant temperatures (5–18°C) and reduced pH (by 0.4 units). Embryonic responses were investigated by quantifying proxies for development rate and fitness including: % yolk consumption, mean heart rate, rate of oxygen consumption, and oxidative stress. We found no interactions between temperature and pH, and reduced pH only affected the level of oxidative stress significantly, with a higher level of oxidative stress in the controls. Increased temperature and % yolk consumed had positive effects on all parameters except on oxidative stress, which did not change in response to temperature. There was a difference in development rate between the ranges of 5–10°C (Q10: 5.4) and 10–18°C (Q10: 2.9), implicating a thermal break point at 10°C or below. No thermal limit to a further increased development rate was found. The insensitivity of N. norvegicus embryos to low pH might be explained by adaptation to a pH-reduced external habitat and/or internal hypercapnia during incubation. Our results thus indicate that this species would benefit from global warming and be able to withstand the predicted decrease in ocean pH in the next century during their earliest life stages. However, future studies need to combine low pH and elevated temperature treatments with hypoxia as hypoxic events are frequently and increasingly occurring in the habitat of benthic species.

Stem Cells in Asexual Reproduction of Marine Invertebrates

While sexual reproduction is conserved and almost ubiquitous, asexual reproduction in forms of parthenogenesis or agametic cloning from somatic tissue is less conserved. The phylogeny shows that agametic cloning is widespread but scattered with many different modes for asexual formation of a new animal. This suggests that independent forms of cloning have evolved later from sexual ancestors between and within different phyla. Here, we present an overview of agametic cloning in the marine animal kingdom and discuss molecular and evolutionary aspects of somatic stem cell usage for asexual cloning. The molecular tissue characterizations and the relative role of different stem cells involved in agametic cloning are only at its beginning with whole phyla largely uncovered. An emerging hypothesis is that the first somatic stem cells used in cloning were also able to form a germ-line and that the more limited lineage specific stem cells are derived. We discuss advantages and problems with agametic cloning from somatic tissue and propose that the levels of stem cell potential held in the tissue can have large consequences for the reproductive life cycle strategies and long-term fitness in clonal animals and strains. We finally describe suitable molecular experimental approaches for future research on this topic.

Potential for clonal animals in longevity and ageing studies.

Ageing is defined as a decline in reproductive and/or somatic performance over time, and as such is experienced by most organisms. Evolutionary theories explain ageing as a consequence of reduced selection pressure against mutations and reduced allocation to somatic maintenance in post-reproductive individuals. In addition, the fecundity of younger age-groups makes a more significant contribution than infinite maintenance of the parental body to the production of subsequent generations. However, in clonal animals, as well as in plants that reproduce by agametic cloning, the adult body is itself a reproductive unit that increases its fitness as a function of genet size. Given the apparent longevity of many such clonal organisms, species undergoing agametic cloning are often assumed to be non-ageing and even potentially immortal. Here, we present a brief overview of ageing in organisms undergoing agametic cloning, focusing on animals and molecular investigation. We discuss molecular and evolutionary aspects of ageing or non-ageing with respect to selection in clonal species. Of particular relevance to the search for potential mechanistic processes behind longevity is the notion that clonal organisms are frequently smaller than their obligate sexual counterparts. In conclusion, we find that while clonal animals also commonly age, evolutionary arguments together with empirical evidence suggest that they are likely to be long-lived and stress resistant at the genet level. However, theoretical modeling continues to predict the possibility of immortality, if the contribution from sexual reproduction is low. Future in-depth study of long-lived clones should present an excellent opportunity to discover novel mechanisms for renewal and long-term somatic maintenance and health.

The capacity for internal colour change is related to body transparency in fishes

Dear Sir, Chromatophores are large, stellate and spectacular pigment bearing cells, typically located in the skin, that generate body colouration. In many animals, melanocytes ⁄melanophores, the melanin containing chromatophores, are also present in various tissues inside the body, for instance in the ear, brain, abdominal cavity, around internal organs and skeleton. The presence of such internal pigmentation in puzzling as it is hidden from sight. While there is an enormous amount of studies and data on skin chromatophores, from the fine details of the motile machinery to animal behaviour (Aspengren et al., 2009), internal melanocytes have historically been largely ignored, until recently (Aoki et al., 2009; Brito and Kos, 2008; Randhawa et al., 2009; Yajima and Larue, 2008). Remarkably little is still known about their possible functions, and this uncertainty is problematic for the more general question of the role(s) of melanin in itself (Aspengren et al., 2009; Boissy and Hornyak, 2006; Braasch et al., 2009; Hill, 2000; Ito, 2009). While internal melanocytes are prevalent, internal erythrophores and xanthophores appear more uncommon. In fish, however, such cells can be found interspersed with melanocytes and reflective chromatophores in the highly pigmented peritoneum (the endothelium that covers the abdominal cavity) (Nilsson Skold et al., 2008). In juveniles, as well as in adults of species with relatively transparent bodies, internal chromatophores may actually contribute to the overall body colouration, as shown in two-spotted goby females, where abdominal trunk biopsies were analysed (Nilsson Skold et al., 2008). In fishes, skin patterns can be rapidly modified by aggregation or dispersion of the pigment-containing organelles inside the chromatophores present in the skin (i.e. physiological colour change) for background adaptation or signalling displays (Aspengren et al., 2009; Fujii and Oshima, 1994). In comparison, colour change in internal chromatophores has been largely ignored as it has been generally considered that they are not responsive (Boissy and Hornyak, 2006). However, melanocytes in the peritoneum and around the skeleton of the ice goby, Leucopsarion petersii, do indeed adapt to the background by pigment translocations in vivo and in biopsies, thus providing the first evidence for internal colour change (Goda and Fujii, 1996). Recent work on biopsies from the two-spotted goby showed that also internal erythrophores and xanthophores can be responsive (Nilsson Skold et al., 2008). As these examples come from relatively transparent species, it is possible that this phenomenon is more common than earlier believed, especially in species with some degree of body transparency. In order to test if a capacity for internal colour change was related to the degree of body transparency, and to reveal a possible function of these cells, we analysed the regulatory capability of peritoneal melanocytes in eight different teleost species, representing five different orders within the large super order Acanthopterygii and in one member of the super order Clupeomorpha, all with different degrees of body transparency. We used epinephrine and melatonin as potential pigment aggregating agents (see Appendix S1 for methodology). A positive relationship between body transparency and rate of internal colour change would suggest a special adaptive role for internal melanocytes in transparent fish species, and thus constitute a novel function for internal pigments. Our results showed that peritoneal melanocytes were present in all investigated fish species. Especially high densities were found in the gobies and in pipefish, plaice and herring (Figure 1A, B). Internal erythrophores and xanthophores were also observed in the gobies, pipefish and in plaice, but not in the other species. A capacity to regulate peritoneal melanocytes by melatonin and epinephrine was detected in six out of the eight species (Figure 1A, B and Table S1). In all species tested, peritoneal melanocytes had dispersed pigment from the start of the experiment. The epidermal melanocytes of sand goby, black goby and wrasse responded within 30 min, whereas plaice and pipefish only responded appreciably after 2 h. In herring, both controls and treated peritoneum biopsies showed a response after 30 min with an only moderate further

Telomerase deficiency in a colonial ascidian after prolonged asexual propagation.

In organisms that propagate by agametic cloning, the parental body is the reproductive unit and fitness increases with clonal size, so that colonial metazoans, despite lack of experimental data, have been considered potentially immortal. Using asexual propagation rate as a measure of somatic performance, and telomerase activity and relative telomere length as molecular markers of senescence, old (7-12 years) asexual strains of a colonial ascidian, Diplosoma listerianum, were compared with their recent sexually produced progeny. We report for the first time evidence for long-term molecular senescence in asexual lineages of a metazoan, and that only passage between sexual generations provides total rejuvenation permitting indefinite propagation and growth. Thus, this colonial ascidian has not fully escaped ageing. The possibility of somatic replicative senescence also potentially helps to explain why metazoans, with the capacity for asexual propagation through agametic cloning, commonly undergo cycles of sexual reproduction in the wild.