Association of kinesin and myosin with pigment granules in crustacean chromatophores

Chromatic adaptation in crustaceans results from the differential distribution of colored pigment granules within their chromatophores consequent to cell signaling by neurosecretory peptides. However, the force transducing, mechanochemical protein motors responsible for granule translocation, and their molecular mechanisms of action, are not well understood. The present study uses immunocytochemical techniques and a motility assay in vitro to demonstrate that protein motors from the kinesin and myosin superfamilies are stably associated with membrane-bounded pigment granules in the red, ovarian chromatophores of the freshwater, palaemonid shrimp, Macrobrachium olfersii. Monoclonal antibodies against conventional kinesin heavy chain, and an anti-myosin whole serum, labeled pigment-containing fragments prepared from homogenates of chromatophores with fully dispersed or aggregated pigments: this finding infers a permanent association between the protein motors and the pigment granules, and suggests that such motors may be regulated while bound to their cargos. The pigment aggregator appears to be a myosin since the anti-myosin whole serum attenuated hormonally triggered pigment aggregation in the motility assay in vitro, and induced pigment hyper-dispersion in some chromatophores. Western blots of the chromatophore-containing, ovarian tissue homogenate demonstrated protein bands consistent with myosin II and myosin XII, either of which may be the pigment aggregator. This study provides the first direct evidence for myosin and kinesin protein motors directly and stably associated with pigment granules in crustacean chromatophores, and may represent the first successful isolation of myosin class XII.     

Interval timing in Siamese fighting fish (Betta splendens)

The present study evaluated the temporal performance of Siamese fighting fish (Betta splendens) given short-term exposure to four fixed interval (FI) schedules of reinforcement, FI 30, 60, 120, and 240 s, during which a reinforcer (mirror image) was given for the first response (swimming through a hoop) after the interval requirement had elapsed. Response levels were generally low early in an interval and increased as the interval elapsed; wait times and break points in an interval increased with increases in the FI requirement. The results were similar to that obtained with other species and different types of responses and reinforcers, and demonstrate that the procedure is a feasible method for studying interval timing in fish.     

Mitotic and pigment-translocating activities of cultured chromatophores of the guppy, Lebistes reticulatus

Using Ham's F-12 medium, an in vitro culture system permitting cellular survival for over 6 months has been developed for the chromatophores of the guppy. In this culture system, the various types of chromatophores (melanophores, erythrophores and xanthophores) migrated out of the explanted tail fin tissue, retained their pigmentation, and displayed both mitotic and pigment-translocating activities. The mitotic activity was evident during the first 3 or 4 weeks in culture, whereas the pigment-translocating ability persisted for 16 weeks. The cultured chromatophores of male fish displayed pigment aggregation in response to adrenergic agents (epinephrine and norepinephrine) and pigment dispersion in response to alpha-melanocyte stimulating hormone (alpha-MSH), cyclic AMP and dibutyryl cyclic AMP. Cyclic GMP did not elicit pigment-translocating responses in any of the chromatophores.     

Calcium movements during pigment aggregation in freshwater shrimp chromatophores

Pigment granule migration within crustacean chromatophores provides an excellent model with which to investigate cytoplasmic movements, given the antagonistic, neurosecretory peptide regulation of granule translocation, and the absence of innervation in these large, brightly colored cells. Red pigment-concentrating hormone (RPCH) induces pigment aggregation in shrimp chromatophores via an increase in intracellular Ca2+; however, how this increase is brought about is not known. To examine the putative Ca2+ movements leading to pigment translocation in red, ovarian chromatophores of the freshwater shrimp, Macrobrachium olfersii, this study manipulates intra- and extracellular Ca2+ employing ER Ca2+-ATPase inhibitors, ryanodine-sensitive, ER Ca2+ channel blockers, and EDTA/EGTA-buffered A23187/Ca2+-containing salines. Our findings reveal that during pigment aggregation, cytosolic Ca2+ apparently increases from an intracellular source, the abundant SER, loaded by the SERCA and released through ryanodine-sensitive receptor/channels, triggered by capacitative calcium influx and/or calcium-induced calcium release mechanisms. Aggregation also depends on external calcium, which may modulate RPCH/receptor coupling. Such calcium-regulated pigment movements form the basis of a complex system of chromatic adaptation, which confers selective advantages like camouflage and protection against ultra-violet radiation to this palaemonid shrimp.     

Pigment biogenesis in freshwater shrimp ventral nerve chord chromatophores

Summary The possible biogenesis of two pigment granule types present in the monochromatic, brown chromatosomes enveloping the ventral nerve chord of the freshwater palaemonid shrimps Macrobrachium acanthurus, M. heterochirus and M. olfersii is examined by transmission electron microscopy in thin section and freeze fracture replicas. Prominent, membrane limited granules are suggested to have their origin in a complex, juxtanuclear, smooth endoplasmic reticulum labyrinth, continuous with the nuclear envelope. Amembranous, lipocarotenoid granules possibly derive from the external surface of the smooth endoplasmic reticulum. Nuclear envelope and SER membranes contain numerous 11 nm diameter intramembranous particles while pigment granule membranes exhibit fewer particles. A dictyosomal origin for the lipocarotenoid granules is discounted. Granulogenesis is suggested to be a continuous process in crustacean chromatophores.     

The novel application of an immunological technique reveals the immunosuppressive effect of phytoestrogens in Betta splendens

The novel application of a standard technique to assess cell‐mediated immune response to phytohaemagglutinin (PHA) injected in the caudal peduncle in a small fish (<2 g) to test the immunosuppressive effect of three phytoestrogens: genistein, equol and β‐sitosterol is described. Individual Betta splendens exposed to these oestrogenic chemicals produced weaker inflammatory responses to PHA than did control individuals, suggesting that phytoestrogens are immunosuppressive. This technique should enable immune function in fish species, too small to provide sufficient blood for conventional immunological measures, to be assessed.     

The influence of long-term chromatic adaptation on pigment cells and striped pigment patterns in the skin of the zebrafish, Danio rerio

The striped pigment patterns in the flanks of zebrafish result from chromatophores deep within the dermis or hypodermis, while superficial melanophores associated with dermal scales add a dark tint to the dorsal coloration. The responses of these chromatophores were compared during the long-term adaptation of zebrafish to a white or a black background. In superficial skin, melanophores, xanthophores, and two types of iridophores are distributed in a gradient along the dorso-ventral axis independent of the hypodermal pigment patterns. Within one week the superficial melanophores and iridophores changed their density and/or areas of distribution, which adopted the dorsal skin color and the hue of the flank to the background, but did not affect the striped pattern. The increases or decreases in superficial melanophores are thought to be caused by apoptosis or by differentiation, respectively. When the adaptation period was prolonged for more than several months, the striped color pattern was also affected by changes in the width of the black stripes. Some black stripes disappeared and interstripe areas were emphasized with a yellow color within one year on a white background. Such long-term alteration in the pigment pattern was caused by a decrease in the distribution of melanophores and a concomitant increase in xanthophores in the hypodermis. These results indicate that morphological responses of superficial chromatophores contribute to the effective and rapid background adaptation of dorsal skin and while prolonged adaptation also affects hypodermal chromatophores in the flank to alter the striped pigment patterns.     

Bubble nest habitat characteristics of wild Siamese fighting fish

Siamese fighting fish Betta splendens in Thailand inhabit shallow water amongst dense emergent vegetation near the margin of rice paddy fields. Nesting water was low in dissolved oxygen, pH, salinity but high in free carbon dioxide and temperature. This fish aggregated with a mean population density of 1·7 fish m^-2 and an equal adult sex ratio. Males were heavier, longer and wider than females. The size of their bubble nest is in proportion to their weight and length.     

Instrumental conditioning of choice behavior in male Siamese fighting fish (Betta splendens)

Despite the differences in the response of male Betta splendens toward various stimuli, no research has attempted to determine the preference for a live conspecific versus a mirror presentation. A submerged T-maze was used to present both stimuli to healthy male B. splendens (N=16). The results indicated that subjects' start box and swimway latencies decreased significantly over the 30 trials. Moreover, the analysis of choices demonstrated a modest, but statistically significant, preference for the live conspecific over the mirror presentation. The results are discussed in terms of the stimuli qualities that elicit an aggressive response in B. splendens and the implications for common experimental procedures.     

Social partner preferences of male and female fighting fish (Betta splendens)

While the social interactions of Betta splendens have been studied in the contexts of dominance hierarchies, mate choice and communication networks, the social partner preferences of Betta have been largely overlooked. In this study, we presented male and female Betta with a single male, a single female, and a group of three females in dichotomous choice tests in order to better understand basic social interactions in this largely nonsocial species. The highly territorial Betta preferred associating with conspecifics in nearly every configuration we tested, with exceptions noted when single females were given the choice between a lone male and an empty chamber, and when males were presented with a single female and an empty chamber. Also, in most tests, the fish chose to spend more time with the larger group of females. The motivation for this preference certainly varied from reproductive to anti-predator. While such behavior might not suggest true shoaling behavior, it does demonstrate a subtle degree of sociality.     

Physiological colour changes in Odonata eyes. A comparison between eye and epidermal chromatophore pigment migrations

Pigment migration in the eyes of Austrolestes annulosus and Ischnura heterosticta cause pronounced colour changes which superficially resemble those of Odonata epidermal chromatophores. In both species, the migratory pigment is confined to the distal pigment cells of dorsal ommatidia. When the pigment is concentrated around the base of the crystalline cones, a dense layer of Tyndall blue bodies produce bright ‘blue phase’ colours. Distal migration of the pigment disrupts the Tyndall effect and produces ‘dark phase’ (grey-brown) colours. As in chromatophores, eye pigments consist of a mixture of xanthommatin and dihydroxanthommatin together with an additional pigment, possibly ommin A, not found in chromatophores.As with chromatophores, eye pigments respond to change in temperature only, change in light intensity having no effect. The change from blue to dark phase (at 8°C) occurs at the same rate as in chromatophores, whereas the reverse change (at 20°C) is significantly slower. Equilibrium colours at constant temperature are variable but significantly different from those of chromatophores at 12°C and above. There is no diurnal variation in responsiveness as is found in chromatophores.Isolated dark phase eyes or undamaged pieces of eye are able to change to blue phase after temperature increase. Isolated blue phase eyes show little response to temperature decrease, isolated undamaged pieces show no response. A temperature difference between the eyes of the same intact insect may result in minor colour differences. Ablation of the optic tract or of tissue posterior to the optic tract prevents normal colour change from blue to dark phase. The above results indicate that eye pigment cells are structurally similar to Odonata chromatophores and are under similar environmental and physiological control.     

α-肾上能受体兴奋剂及阻断剂对米虾棕色载色细胞内色素聚散的效应

萘唑啉或肾上腺素能使米虾离体棕色胞内色素集中,此作用被酚妥拉明短时所阻断。异丙肾上腺素对棕色胞无影响。作者认为棕色胞上只具有α受体,可以作为筛选α受体兴奋剂及阻断剂的模型。     

Vertebrate melanophores as potential model for drug discovery and development: A review

Drug discovery in skin pharmacotherapy is an enormous, continually expanding field. Researchers are developing novel and sensitive pharmaceutical products and drugs that target specific receptors to elicit concerted and appropriate responses. The pigment-bearing cells called melanophores have a significant contribution to make in this field. Melanophores, which contain the dark brown or black pigment melanin, constitute an important class of chromatophores. They are highly specialized in the bidirectional and coordinated translocation of pigment granules when given an appropriate stimulus. The pigment granules can be stimulated to undergo rapid dispersion throughout the melanophores, making the cell appear dark, or to aggregate at the center, making the cell appear light. The major signals involved in pigment transport within the melanophores are dependent on a special class of cell surface receptors called G-protein-coupled receptors (GPCRs). Many of these receptors of adrenaline, acetylcholine, histamine, serotonin, endothelin and melatonin have been found on melanophores. They are believed to have clinical relevance to skin-related ailments and therefore have become targets for high throughput screening projects. The selective screening of these receptors requires the recognition of particular ligands, agonists and antagonists and the characterization of their effects on pigment motility within the cells. The mechanism of skin pigmentation is incredibly intricate, but it would be a considerable step forward to unravel its underlying physiological mechanism. This would provide an experimental basis for new pharmacotherapies for dermatological anomalies. The discernible stimuli that can trigger a variety of intracellular signals affecting pigment granule movement primarily include neurotransmitters and hormones. This review focuses on the role of the hormone and neurotransmitter signals involved in pigment movement in terms of the pharmacology of the specific receptors.     

Cyt toxins produced by Bacillus thuringiensis: A protein fold conserved in several pathogenic microorganisms

Bacillus thuringiensis bacteria produce different insecticidal proteins known as Cry and Cyt toxins. Among them the Cyt toxins represent a special and interesting group of proteins. Cyt toxins are able to affect insect midgut cells but also are able to increase the insecticidal damage of certain Cry toxins. Furthermore, the Cyt toxins are able to overcome resistance to Cry toxins in mosquitoes. There is an increasing potential for the use of Cyt toxins in insect control. However, we still need to learn more about its mechanism of action in order to define it at the molecular level. In this review we summarize important aspects of Cyt toxins produced by Bacillus thuringiensis, including current knowledge of their mechanism of action against mosquitoes and also we will present a primary sequence and structural comparison with related proteins found in other pathogenic bacteria and fungus that may indicate that Cyt toxins have been selected by several pathogenic organisms to exert their virulence phenotypes.     

Body Condition in Male Betta splendens Does Not Predict Their Ability to Perform Opercular Displays Under Hypoxic Conditions

Many fish, including the fighting fish Betta splendens , perform a display in which the opercula are extended away from the head and gills. Previous work has shown that opercular display rates by male B. splendens decrease under conditions of reduced dissolved oxygen (hypoxia). We tested the hypothesis that the ability to maintain opercular display rates under hypoxic conditions is related to body condition in male B. splendens . We also tested the hypothesis that females would show a greater preference for males performing this display under hypoxic conditions, when the display should be a more reliable indicator of male phenotypic quality. We found no evidence to support either hypothesis. Male opercular display rate in hypoxic conditions was unrelated to natural or experimentally induced variation in body condition. Female B. splendens showed no differential preference for the opercular display, assessed through the use of computer animated male stimuli, in either acute or chronic hypoxia. We conclude that the presence of an air-breathing organ in this species makes the opercular display an unreliable signal of male quality as measured by body condition.     

Photosensitivity of Chromatophores

Almost all major phyla of invertebrates and lower vertebratesdisplay a direct sensitivity of their chromatophores to lightby either dispersing or—in rare cases—aggregatingthe pigment granules within the cell. This "primary response"of color change is an accessory component of the "dermal lightsense" and characterizes the chromatophores as an independentreceptor for light and effector of the chromomotor response.Photosensitivity does not seem to be restricted to certain partsof the pigment cell but is rather supposed to be a ubiquitousproperty of the chromatophore. Experiments on partially illuminatedchromatophores show that the photopigment, whose chemical compositionis still unknown, is localised within the plasma membrane orthe cytoplasmic ground substance. Threshold responses for ajust visible reaction are much higher than for background responsesand have for some pigment cells been determined to be in a rangebetween 0.2 and 0.5 erg/sec/cm². The physiological significanceof the photosensory reaction is closely related with thermoregulationand the protection of underlying tissue against harmful radiation.The chain of events involved in photosensory transduction remainsto be further studied and can at present be interpreted onlyon the basis of related phenomena like retinal pigment migrationand the light-sensitivity of simple non-pigmented contractilesystems.     

The fine structure of odonata chromatophores

The appearance, fine structure and pigment composition of the epidermal chromatophores of mature Austrolestes annulosus (Lestidae) are described and compared with the developing chromatophores of teneral Austrolestes and the mature chromatophores of Diphlebia lestoides (Amphipterygidae) and Ischnura heterosticta (Caenagrionidae). Mature chromatophores contain masses of near spherical light-scattering bodies and larger irregularly shaped pigment vesicles. These effect colour change by migrating in opposite directions, through a system of interconnecting granular endoplasmic reticulum tubules. The pigment, a mixture of xanthommatin and dihydroxanthommatin, has a liquid or gelatinous consistency. Developing chromatophores of teneral insects lack light-scattering bodies and well-defined migratory pigment vesicles, but contain irregular masses of pigment of similar chemical composition.     

Role of MAPK p38 in the cellular responses to pore-forming toxins

Understanding the mechanism of action of pore-forming toxins (PFTs) produced by different bacteria, as well as the host responses to toxin action, would provide ways to deal with these pathogenic bacteria. PFTs affect the permeability of target cells by forming pores in their plasma membrane. Target organisms may overcome these effects by triggering intracellular responses that have evolved as defense mechanisms to PFT. Among them it is well documented that stress-activated protein kinases, and specially MAPK p38 pathway, play a crucial role triggering defense responses to several PFTs in different eukaryotic cells. In this review we describe different intracellular effects induced by PFTs in eukaryotic cells and highlight diverse responses activated by p38 pathway.     

Ultrastructure of the chromatophores of Palaemon affinis Heilprin (crustacea: decapoda). The structural basis of pigment migration

The relationships between pigment granules and the prominent smooth endoplasmic reticulum in the chromatophores of the shrimp, Palaemon affinis Heilprin, were investigated by transmission electron microscopy. Different types of pigment granules within the chromatophores were found to exhibit a close structural continuity with the cisternal membranes. The membranes of membrane-bound pigment granules were seen to be continuous with those of the ER cisternae, while pigment granules lacking membranes appear to adhere to the external cisternal surfaces. The reticulum, which seems to form a network enmeshing the pigment granules, is proposed to be part of a continuum linking these granules with their translocating force.     

The involvement of calmodulin in motile activities of fish chromatophores

1. Effects of W-7 and W-5, calmodulin antagonists, on the pigment aggregation within melanophores and coloring response of iridophores were examined in the blue damselfish, Chrysiptera cyanea.2. W-7 was found to antagonize norepinephrine-induced responses of the chromatophores, whereas W-5 had only a slight effect on inhibition of the responses.3. H-7, a specific antagonist of protein kinase C, did not arrest the responses of melanophores and iridophores at all.4. The chromatophores responded normally to norepinephrine in Ca²⁺, Mg²⁺-free saline solution.5. These results indicate that it is a Ca²⁺/calmodulin-regulated enzyme and not protein kinase C that is involved in motile activities of fish chromatophores. Ca²⁺ may be supplied from an intracellular store.