The calcium dependence of pigment translocation in freshwater shrimp red ovarian chromatophores

The roles of calcium in cell signaling consequent to chromatophorotropin action and as an activator of mechanochemical transport proteins responsible for pigment granule translocation were investigated in the red ovarian chromatosomes of the freshwater shrimp Macrobrachium olfersii. Chromatosomes were perfused with known concentrations of free Ca++ (10(-3) to 10(-9) M) prepared in Mg(++)-EGTA-buffered physiological saline after selectively permeabilizing with 25 microM calcium ionophore A23187 or with 10(-8) M red pigment concentrating hormone (RPCH). The degree of pigment aggregation and the translocation velocity of the leading edges of the pigment mass were recorded in individual chromatosomes during aggregation induced by RPCH or A23187 and dispersion induced by low Ca++. Aggregation is Ca++ dependent, showing a dual extracellular and intracellular requirement. After perfusion with reduced Ca++ (10(-4) to 10(-9) M), RPCH triggers partial aggregation (approximately 65%), although the maximum translocation velocities (approximately 16.5 microns/min) and velocity profiles are unaffected. After aggregation induced at or below 10(-5) M Ca++, spontaneous pigment dispersion ensues, suggesting a Ca++ requirement for RPCH coupling to its receptor, or a concentration-dependent, Ca(++)-induced Ca(++)-release mechanism. The Ca(++)-channel blockers Mn++ (5 mM) and verapamil (50 microM) have no effect on RPCH-triggered aggregation. An intracellular Ca++ requirement for aggregation was demonstrated in chromatosomes in which the Ca++ gradient across the cell membrane was dissipated with A23187. At free [Ca++] above 10(-3) M, aggregation is complete; at 10(-4) M, aggregation is partial, followed by spontaneous dispersion; below 10(-5) M Ca++, pigments do not aggregate but disperse slightly. Aggregation velocities diminish from 11.6 +/- 1.2 microns/min at 5.5 mM Ca++ to 7.4 +/- 1.3 microns/min at 10(-4) M Ca++. Half-maximum aggregation occurs at 3.2 x 10(-5) M Ca++ and half-maximum translocation velocity at 4.8 x 10(-5) M Ca++. Pigment redispersion after 5.5 mM Ca(++)-A23187-induced aggregation is initiated by reducing extracellular Ca++: slight dispersion begins at 10(-7) M, complete dispersion being attained at 10(-9) M Ca++. Dispersion velocities increase from 0.6 +/- 0.2 to 3.1 +/- 0.5 microns/min. Half-maximum dispersion occurs at 7.6 x 10(-9) M Ca++ and half-maximum translocation velocity at 2.9 x 10(-9) M Ca++. These data reveal an extracellular and an intracellular Ca++ requirement for RPCH action, and demonstrate that the centripetal or centrifugal direction of pigment movement, the translocation velocity, and the degree of pigment aggregation or dispersion attained are calcium-dependent properties of the granule translocation apparatus.     

Early embryonic development of the freshwater shrimp Caridina multidentata (Crustacea,Decapoda, Atyidae)

Decapod crustaceans show a great diversity of developmental modes at all levels. In particular, early cleavage varies from total via mixed to superficial modes and from determinate cleavage with a stereotyped pattern to indeterminate cleavage. However, the ground pattern of early decapod development is not clear. To address this problem, we studied the early embryonic development of the caridean shrimp Caridina multidentata with a combination of confocal laser scanning microscopy, scanning electron microscopy, 4D microscopy and 3-D reconstruction software. Despite a yolk-rich egg, the cleavage is holoblastic and shows a distinct pattern of blastomere arrangement, characterized by two interlocking cell bands. This resembles the conditions in dendrobranchiate shrimps, which most likely are the sister group to Pleocyemata to which C. multidentata belongs. Hence, our results offer the possibility to assume total cleavage with blastomeres arranged in two interlocking cell bands as ancestral cleavage mode for Decapoda.     

A spring-matrix model for pigment translocation in the red ovarian chromatophores of the freshwater shrimp Macrobrachium olfersi (Crustacea, Decapoda)

A model for intracellular transport of pigment granules in the red ovarian chromatophores of the freshwater shrimp Macrobrachium olfersi is proposed on the basis of shifts in the equilibrium of resting forces acting on an elastic pigment matrix. The model describes a pigment-transport mechanism in which mechanochemical protein motors like kinesin and myosin alternately stretch and compress a structurally unified, elastic pigment matrix. Quantifiable properties of the spring-matrix obey Hooke's Law during the rapid phases of pigment aggregation and dispersion. The spring-like response of the pigment mass is estimated from previous kinetic experiments on pigment translocation induced by red pigment concentrating hormone, or by the calcium ionophore A23187. Both translocation effectors trigger an initial phase of rapid pigment aggregation, and their removal or washout after complete aggregation produces a phase of rapid pigment dispersion, followed by slow pigment translocation. The rapid-phase kinetics of pigment transport are in reasonable agreement with Hooke's Law, suggesting that such phases represent the release of kinetic energy, probably produced by the mechanochemical protein motors and stored in the form of matrix deformation during the slow phases of translocation. This semiquantitative model should aid in analyzing intracellular transport systems that incorporate an elastic component.     

The effects of ionic lanthanum and hypertonic physiological salines on the nervous systems of larval and adult stick insects.

The effects of the electron-opaque tracer ionic lanthanum in various concentrations and of hyperosmotic physiological salines on the nervous system of the stick insect, Carausius morosus, have been studied. Examination of the experimentally treated tissues revealed that the diffusion barrier to the exogenous tracer was maintained in all cases in the adult central and peripheral nervous systems, but not in the hatchling. When hatchling nervous tissues were incubated in 50 mM ionic lanthanum in phyerosmotic physiological saline, the tracer readily infiltrated all the extracellular spaces between axons and glia of all components of the nervous system examined. No difference was noted in this regard between fed and unfed hatchlings, Further, in all cases examined of adults and hatchlings, lanthanum readily surrounded those neurosecretory axons which are found in the neutral lamella, or extracellular nerve sheath, of the insect. The possible meanings of these variations in hatchling and adult nervous systems and in the accessibility of different elements of the nervous system to exogenous ionic lanthanum are discussed.     

The toxicity and physiological effects of copper on the freshwater pulmonate snail, Lymnaea stagnalis

Several recent studies have demonstrated that the freshwater pulmonate snail Lymnaea stagnalis is extremely sensitive to metals (Co, Ni, Pb) in chronic exposures. The objective of the current study was to evaluate the acute and chronic sensitivity of L. stagnalis to Cu and investigate the underlying mechanism(s) of toxic action. A 96-h LC50 of 31μg L(-1) Cu was estimated indicating L. stagnalis was moderately acutely sensitive to Cu relative to other aquatic organisms. However, in a 30-day chronic exposure using juvenile snails an EC20 of 1.8μg L(-1) Cu was estimated for snail growth making L. stagnalis the most sensitive organism tested to date for Cu. Hardness-based and BLM-based water quality criteria for Cu at the water quality conditions used in this study were 7.8 and 1.5μg L(-1), respectively, indicating L. stagnalis is significantly under-protected by hardness-based WQC. Investigations into the mechanism(s) of toxic action for Cu were conducted on young adult snails necessitating higher Cu exposures. Exposure to Cu at 12μg L(-1) resulted in no detectable effects on hemolymph osmolality, net Ca(2+) uptake, titratable acid excretion, or ammonia excretion. Exposure to 48μg L(-1) Cu was shown to significantly reduce (91%) net Ca(2+) uptake which is strongly correlated with shell deposition and corresponding snail growth. Snails exposed to 48μg L(-1) Cu also exhibited reduced ammonia excretion, a marked hemolymph acidosis, and a compensatory increase in titratable acid excretion. The reduction in net Ca(2+) uptake was hypothesized to be a secondary effect of Cu-induced inhibition of carbonic anhydrase, but no reduction in carbonic anhydrase activity was detected. Overall, it remains unclear whether inhibition of Ca(2+) uptake is a direct result of Cu exposure or, along with the other observed physiological effects, is secondary to an unidentified primary mode of toxic action. Given the hypersensitivity of L. stagnalis to Cu, further study into the mechanisms of action and effects of varying water chemistry on Cu toxicity is clearly warranted.     

Caridina bruneiana, a new species of freshwater shrimp (Decapoda, Caridea, Atyidae) from Negara Brunei Darussalam, Borneo

A new species of the freshwater atyid shrimp, Caridina bruneiana from Negara Brunei Darussalam, Borneo is described. Caridina bruneiana is characterized by its moderately long and deep rostrum, pereiopod shape, spination and segmental ratios, the shape and spination of its posterior telsonic margin, egg size and the presence of an appendix interna on the endopod of the first male pleopod. The distribution of atyid shrimps in forested streams and their possible use as bioindicators of stream water quality in tropical rainforests are briefly discussed.     

Geographical distributions of mitochondrial DNA lineages reflect ancient directions of river flow: a case study of the Japanese freshwater shrimp Neocaridina denticulata denticulata (Decapoda: Atyidae)

The mitochondrial DNA phylogeographical structure of the freshwater atyid shrimp Neocaridina denticulata denticulata was investigated near the Yura and Kako Rivers in western Japan. To assess the biological significance of drainage evolution, fragments of 390 base pairs (bp) extending from the NADH dehydrogenase subunit 2 gene to the tryptophan transfer RNA gene were sequenced for 246 specimens from twenty locations. The phylogenetic tree detected two distinct clades corresponding to the region along the Sea of Japan and Seto Inland Sea, respectively. Geographical mapping of the two clades well reflected the river capture, in which the upper reaches of the Yura River draining into the Sea of Japan had been captured from the Kako River into the Seto Inland Sea. The clear pattern of mitochondrial DNA distribution may be accounted for by the ecological characteristics of species that minimize passive downstream drift, local adaptation, and mountain topography.     

Population dynamics and production of the tropical freshwater shrimp Caridina nilotica (Decapoda: Atyidae) in the littoral of Lake Sibaya

SUMMARY. Seasonal changes in population structure, standing stock levels and production of Caridina nilotica were studied at three sites in the littoral margins of subtropical Lake Sibaya between January 1975 and March 1976. Average population density at these sites declined from a maximum of c. 1400 to a minimum of c. 350 individuals per m² (3.4–0.4 g m^?2 dry wt) during the study, possibly as a result of emigration into peripheral vegetation inundated by rising lake levels. Shrimps bred perennially and, although egg stocks and instantaneous birth rates (b) were highest during summer, no corresponding increases in populaton density were observed, suggesting that the seasonally higher birth rates were offset by higher mortality rates. Population size structure and size-specific sex ratios did not change seasonally to any marked extent. Relative abundance declined with size and females grew larger than males. Clutch size increased linearly as a function of female carapace length. Estimates of overall mean annual somatic production (g m^?2 year^?1 dry wt) for the three sites between January 1975 and January 1976 ranged between c. 132 (egg-ratio method), 37.5 (summation of growth increments) and 24 (Hynes-Hamilton method) at an annual mean standing stock level of 2.7 g m^?2 dry wt (calorific value, 20.34 kJ g^?1 dry wt). Production at sites 1, 2 and 3 decreased in line with declining annual mean standing stocks (5.32, 3.67 and 0.23 g m^?2, respectively). The growth increment method gave an overall mean annual P/B value of 13.9. Egg production amounted to a further 5.6, 3.6 and 0.1 g m^?2 year^?1 dry wt (calorific value, 28.01 kJ g^?1) at sites 1, 2 and 3, or 2.7 g m^?2 year^?1 on average.     

Physiological salines and the mechanical properties of trout red blood cells

Suspensions of rainbow trout erythrocytes in different physiological salines were compared with respect to their haematological and filtration properties.
A method is described for the suspension of erythrocytes in Cortland saline which has proved suitable for studies of their mechanical properties over periods of several hours.
Significant differences were found between whole blood samples taken during cannulation and after several days recovery, particularly mean cell volume, frequency distribution of red cell volumes and the pore passage time through nucleopore filters. These differences were also found using red cell suspensions of the same bloods. The pore passage time of whole blood sampled during cannulation or its suspensions is less than that of recovery blood although its mean cell volume is greater.     

Impact of organophosphates on the ovarian changes in freshwater caridean prawn Caridina weberi

The toxicity of the pesticides (organophosphates) malathion, dimecron and monochrotophos was tested on the freshwater caridean prawn, Caridina weberi. The lethal concentration was determined in experiments carried out up to 96 hrs. The effect of organophosphates on the reproduction was shown by observing the histological changes in the ovary. The tissues were highly damaged, with loss of ooplasmic material, lack of distinct nucleus, nucleolus and chromatin material.     

The effects of oestrogenic substances on two components of freshwater ecosystems

The effects of an oestrogenic compound, similar to those recently linked with falling human sperm counts, were investigated in two common freshwater organisms commonly used in education

Oestrogenic compounds have recently been linked with falling sperm counts in human males and also with cancer of the testes and breast. If these compounds can have such dangerous and dramatic effects in humans, as well as affecting sexual behaviour and physiology of fish, what effects do they have on other members of freshwater food chains? The aim of this investigation was to perform some simple experiments using basic laboratory facilities for student research projects. Daphnia, an important primary consumer and food source for fish, was exposed to increasing amounts of ethynyloestradiol, an oestrogen commonly used in contraceptive tablets. The higher concentrations of the oestrogen (0.1 to 1.0 ppm) were found to significantly reduce the heart rate of the Daphnia after approximately 30 minutes' exposure. It is proposed that such a reduction in the activity of oestrogen exposed Daphnia would make them more prone to predation and therefore increase the accumulation of oestrogens by fish. A further experiment investigated the effects of the oestrogen on photosynthesis in Elodea, a common pond weed. The same concentrations of ethynyloestradiol increased photosynthetic rates which could lead to a proliferation in growth. This again would promote the transfer of these compounds if they accumulate in the tissues for aquatic plants. These experiments on a currently significant topic were performed using readily available organisms and a small capital expenditure, thus making them a pertinent and interesting practical for both Alevel and undergraduate students.     

The effects of salines and fixatives upon the size of an identified neuron

Because accurate neuronal dimensions are essential for mathematical modeling of neuronal properties, the effects of a number of salines and fixative procedures on neuronal size were compared, including the non-chemical, freeze substitution method. Using an identified neuron we compared diameters and found some of the fixative-saline combinations caused shrinkage by as much as a factor of four from our best estimates of the in vivo size from the quick frozen preparations. A glutaraldehyde based fixation procedure was found which gives results in good agreement with the frozen tissue.     

An island in the stream: Australia's place in the cosmopolitan world of Indo-West Pacific freshwater shrimp (Decapoda: Atyidae: Caridina)

Mitochondrial DNA sequences were used to investigate phylogenetic and biogeographic relationships among Australian freshwater shrimp from the genus Caridina H. Milne Edwards, 1837 (Atyidae) and congeners from potential source populations throughout the Indo-West Pacific region. Numerous Australian taxa have close evolutionary relationships with non-Australian taxa from locations throughout the region, indicating a diverse origin of the Australian freshwater fauna. This implies many colonisations to or from Australia over a long period, and thus highlights the surprising adeptness of freshwater shrimp in dispersal across ocean barriers and the unity of much of the region's freshwater biota. Interestingly, a study on Australia's other main genus of atyid shrimp, Paratya Miers, 1882, inferred only a single colonisation. A number of potential species radiations within Australia were also identified. This agrees with patterns detected for a large number of Australian freshwater taxa, and so implies a vicariant explanation due to the development of colder, dryer climates during the late Miocene/early Pliocene.     

The effects of salinity,and calcium and potassium variations on the growth of two estuarine red algae

The effects of varying salinity and calcium and potassium concentrations on the growth of two species of estuarine Rhodophyta, Bostrychia radicans Montagne and Caloglossa lepricurii (Montagne) J. Agardh, were examined in unialgal culture. Inocula of settled tetraspores on glass coverslips were incubated in six concentrations of potassium (0. 100, 200, 300, 400 and 500 mg/l) at five salinities (0, 5, 15, 25, and 35%) in the presence of calcium. Growth responses of each alga were determined from the average cell number of 75 tetrasporelings after 4 days in Ott's synthetic sea-water medium. The concentrations of dissolved potassium and calcium in sea water along a salinity gradient in an estuary were determined with atomic absorption spectrophotometry. Four-day-old tetrasporeling of Bostrychia and Caloglossa demonstrated growth over wide ranges of potassium concentrations with growth maxima at 400 500 and 200 400 mg/l, at the optimal salinities for growth of 25 and 15%, respectively. These studies indicate also that the presence of calcium is essential for appreciable growth of both species at each salinity and the effects of variations in potassium are dependent upon the presence of calcium. The abundance of both species in the Mullica River estuary. New Jersey, appears to be a response to a total osmotic effect along the salinity gradient because sufficient levels of the major cations are present. However, the upper limit of both species towards the head of the Mullica River estuary may be determined by combinations of low salinity and low levels of dissolved calcium and potassium rather than by low salinity alone.     

Ultrastructural and histological study of degenerative changes in the antennal glands, hepatopancreas, and midgut of grass shrimp exposed to two dithiocarbamate biocides

The histological and ultrastructural alterations observed in the antennal glands, hepatopancreas, and midgut of grass shrimp exposed to either a 50% potassium dimethyldithiocarbamate biocide (Busan-85; 5–60 ppb) for 14 days, or to a different biocide, composed of 15% sodium dimethyldithiocarbamate and 15% disodium ethylene bisdithiocarbamate (Aquatreat DNM-30), for 3–4 days (60–140 ppb) and 28–35 days (40–120 ppb), were compared and contrasted with the normal morphological features in control shrimp. Only those experimental shrimp that exhibited various degrees of branchial abnormality were examined. Although the alterations in Busan-exposed shrimp were generally more pronounced, the antennal glands of 32 out of 36 experimental shrimp exhibited abnormalities that were manifested primarily as increased secretory activity by the labyrinth cells. In dithiocarbamate-exposed shrimp with “black gills”, the labyrinth epithelium exhibited moderate nuclear hypertrophy, apparent cell sloughing, intense secretory activity, and occasional melanized lesions; alterations in the antennal gland coelomosac included nuclear pyknosis, a general deterioration of podocyte organization, and an unusual increase in hemolymph density adjacent to affected tissues. Although there was an apparent increase in mitotic activity in the hepatopancreatic tubules of shrimp exposed to Aquatreat for 28–35 days, degenerative changes were most frequent and extensive in the hepatopancreas and midgut of dithiocarbamate-exposed shrimp with “black gills”. These observed changes included the diminution of the basal midgut and hepatopancreatic tubular system, moderate midgut hypertrophy, pronounced activity by the hepatopancreatic fixed phagocytes, development of mitochondrial inclusions and megamitochondria, loss of cytoplasmic density, hepatopancreatic nuclear pyknosis, and irreversible degeneration of hepatopancreatic tubule apices. This study suggests that some of the observed abnormal/pathological changes are the indirect consequence of branchial degeneration. A number of possible defensive reactions to dithiocarbamate poisoning, including heterostasis, phagocytosis, encapsulation, and the possible participation of reserve inclusion cells are proposed.     

Electron microscopic study of the open circulatory system of the shrimp,Caridina japonica. I. Gill capillaries

The ultrastructure of the phyllobranchiate type gill of the shrimp, Caridina japonica, was studied. The most characteristic feature of the open circulatory system of Cardina is the vascular lumen of the gill capillaries which is considered to be the interstitial space. The following observations substantiate this view: (1) a thin fibrous layer forms the innermost structure of the walls of gill capillaries and is in direct contact with the blood stream; (2) filaments in the fibrous layer are assumed to correspond to the reticular fibers in the interstitial space of the alveolar wall of mammals; (3) the absence of the endothelium as well as the endothelial basal lamina which are the essential structural components of the closed circulatory system in vertebrates. The gill epithelium contains intermediate, septate and tight junctions. The first two form a junctional complex near the apical cell border and may function as a permeability barrier by occluding the intercellular space as well as functioning in electrical coupling and cellular adhesion. The tight junction is spot-like and may serve no role in the function of the permeability barrier.