Electron microscopic autoradiography of serotonin uptake in the ganglia of the fresh-water mussel (Anodonta cygneak L.).

After in vitro incubation, the uptake of labelled serotonin was investigated by electron microscopic autoradiography in the ganglia of fresh-water mussel (Anodonta cygnea L.) The labelled serotonin was primarily taken up by the axons of the neurons. The silver grains could always be localized over axons containing eccentric dense-core vesicles with a diameter of 100--200 nm. The results suggest (a) the possibility of the electron microscopic identification of serotonin-containing neurons, and (b) the direct role of the eccentric dense-core vesicles in the storage of serotonin.     

Ultrastructural effects of 6-hydroxy-dopamine and 5, 6-dihydroxytryptamine on the central nervous system of fresh-water mussel, Anodonta cygnea L

Ultrastructural effects of 6-hydroxydopamine and 5, 6-dihydroxytryptamine treatments were investigated in the central nervous system of fresh-water mussel. Two days after the treatments, the following characteristic find-structural alterations could be observed in the neuropil of the ganglia: frequent occurrence of multilamellar bodies, lysosomatic structures and elongated tubular forms; shrinking of varicose axon profiles with an enchancement of the density of the axoplasm and clumping of its content; abnormal swelling of certain axons in the neuropil. This degenerative process was accompanied by an intense phagocytosis. The damages evoked by the employed "false transmitters" in the mussel ganglia were, in general, similar to those found in vertebrates. Statistical analysis of the vesicle population of ganglia suggests the intragranular uptake of 6-hydroxydopamine and 5, 6-dihydroxytryptamine and, in addition, the role of dense-core vesicles of different types in the storage of both serotonin and catecholamines. Perikarya composing the cortical layer of the ganglia were not affected by the "false transmitters". This shows that different parts of a mussel neuron are differently sensitive to 6-hydroxydopamine and 5, 6-dihydroxytryptamine.     

Distinct glutathione-dependent enzyme activities and a verapamil-sensitive binding of xenobiotics in a fresh-water mussel Anodonta cygnea

A fresh-water mussel Anodonta cygnea, an aquatic invertebrate resistant to pollution, possesses an inherent high potential to bind 2-acetylaminofluorene onto membrane vesicles. This binding is saturable and trypsin- and verapamil-sensitive. Simultaneously, this mussel reveals a relatively high inherent activity of glutathione-dependent enzyme activities with a distinct spectrum of substrate affinities. Both these activities are similar to the elements of the molecular mechanism involved in the acquired multi-drug resistance phenomenon described in tumor cell-lines. The recognition that in organisms exposed to polluted waters a multi-xenobiotic resistance mechanism may be involved is essential for understanding both the biological impact of pollution and the development of methods for rational risk assessment in regulatory policy.     

Effect of cadmium on ciliary and ATPase activity in the gills of freshwater mussel Anodonta cygnea

1. Cadmium (≤ 50 μM) decreases the heat resistance (39°C) of the activity of frontal cilia in the Anodonta cygnea gills incubated in dechlorinated tap water, while in the presence of added 2 mM Ca²⁺ the minimal acting concentration of cadmium rises up to 100 μM.2. The inhibitory effect of Cd²⁺ (1.5 mM) on the ATPase activity measured in the gill microsomal fraction is temperature dependent and increases as follows: ouabain insensitive Na²⁺- or K⁺-ATPase (no inhibition), Ca²⁺-ATPase (50% inhibition), Mg²⁺-ATPase (100% inhibition).3. Cadmium itself (≤ 50 μM) added to microsomal suspension stimulates the H⁺-sensitive ATP hydrolysis resembling on its pH-dependence the Mg²⁺- but not Ca²⁺-ATPase activity.4. Cd²⁺ can mimic the effect of Mg²⁺ as a cofactor required for activation of the ouabain-insensitive Na⁺- or K⁺-ATPase. Monovalent cations fail to activate the ATPase when Mg²⁺ is substituted by Ca²⁺.5. One of the mechanisms underlying the toxicity of Cd²⁺ to Anodonta gills could be based upon an interaction of Cd²⁺ with Mg²⁺-ATPase followed by suppression of the ciliary activity.     

Effect of sub-lethal concentrations of aluminium on the filtration activity of the freshwater mussel Anodonta cygnea L. at neutral pH

Significant amounts of aluminium (Al) are commonly present in rivers and lakes, largely in particulate form in neutral waters. Freshwater bivalves, as filter feeders are therefore exposed to both particulate and dissolved metal and are potentially vulnerable to Al. The effect of Al on filtering behaviour of the freshwater mussel Anodonta cygnea L. was investigated during short (1 hour) and long-term (15 days) exposure to environmentally relevant concentrations (250 and 500 microg l(-1)) at neutral pH. Water flow through the outflow siphon was monitored as an indicator of pumping capacity. Short-term (1 hour) exposure to 500 microg l(-1) added Al produced an irreversible decrease in the duration of filtering periods, presumably as an avoidance response to the toxicant. One-hour exposure 250 microg l(-1) Al had no detectable effect. When mussels were exposed to 250 or 500 microg l(-1) added Al for 15 days, siphon activity measured in days 11-15 of exposure was inhibited by 50% and 65%, respectively, compared to pre-exposure levels. Recovery occurred following transfer of mussels to uncontaminated water. Interaction between Al and freshwater bivalves at neutral pH may affect both the performance of the mussels and the chemical speciation of the metal in the natural environment.     

Factors affecting the heart activity and blood pressure of the swan mussel Anodonta cygnea.

1. The rate of heart beat increased with temperature and was three times as high in the active as in the inactive animal. 2. The rate of shell valve movement rose and the rate of heart beat fell when the foot was extended. 3. The rates of heart beat and shell valve movement decreased when the water was saturated with carbon dioxide. This heart response remained when the visceral ganglion was destroyed. 4. Ventricular contraction occurred simultaneously over the shole chamber. The passage of blood into the posterior aorta could be restricted by the protuberances on its wall. 5. Pericardial cavity pressure rose by about 5 cm H2O at shell valve adduction and 0-25--0-6 cm H2O at ventricular diastole. 6. Pulse pressure changes of 0-25--0-6 cm H2O occurred in the auricle and 1--3 cm H2O in the ventricle and anterior aorta.     

In vitro culture of glochidia from the freshwater mussel Anodonta cygnea

Abstract. Larvae of the freshwater swan mussel, Anodonta cygnea , were cultured in artificial media at the controlled temperature of 23°±2°C, with successful metamorphosis for the first time. The artificial medium contained a mixture of M199, common carp plasma, and antibiotics/antimycotics. Glochidia were reared to the juvenile stage in the medium after 10–11 d of culture. After 15 d of controlled feeding with phytoplankton, the juveniles showed an elongated shell with several growth lines. Larval survival was 34.3±9.3%, whereas the proportion undergoing metamorphosis was ≤60.8±4.2%. The ultrastructure of early developmental stages was observed by scanning electron microscopy, from the glochidial to the juvenile stage. Glochidia had a hooked shell, with two equal triangular valves formed by a calcareous layer with numerous pores and covered by a thin cuticle of chitin–keratin. The appearance of the complete foot within 11 d of in vitro culture was considered the final feature of metamorphosis to the juvenile stage. The main alteration during juvenile development was the formation, under the glochidial shell, of a new periostracum with growth lines. The prominent foot, gradually covered by long, dense cilia, showed rhythmical movements involved in the capture of particulate matter. Similarly, cilia and microvilli present in the mantle also performed the same role. Longer cilia, sparsely distributed in the mantle, may function as chemotactile sensors.     

Low sensitivity of ciliary activity in the gills of Anodonta cygnea to some ecotoxicals

1. The effects of some ecotoxical model substances on the activity of frontal gill cilia of freshwater mussel Anodonta cygnea were studied in 1 and 24 hr experiments with the particle transport method. The effects were compared with the results of standard Daphnia magna ec₅₀ tests with the same substances.2. Variations of pH between 4.8 and 8.1 affected the ciliary activity by less than 30%.3. Ciliary activity was less sensitive to heavy metal salts, sodium lauryl sulphate and to two pulp industry waste model substances than D. magna.4. Compared to D. magna, the ciliary activity was more sensitive to formalin and 2,4-dinitrophenol, and as sensitive to ethanol.5. Ciliary activity in bivalve gills is a well regulated and/or a relative insensitive function, but it may be used as a rapid screening test for some cell ecotoxicals.     

Light and Electron microscopic autoradiography of 3H-dopamine uptake in the ganglia of the freshwater mussel, Anodonta cygnea L

In the central nervous system of the freshwater mussel, Anodonta cygnea, uptake of 3H-dopamine has been investigated by light and electron microscopic autoradiography following in vitro incubation and topical application, respectively. The autoradiographic reaction observed over the ganglia can be taken as a morphological evidence of the existence of a specific uptake mechanism for dopamine. It has been found that labeled dopamine was taken up by the axon profiles and terminals, while perikarya remained free of activity, indicating a basic difference between the perkaryal and axonal membrane in respect of the 3H-dopamine uptake. According to the electron microscopic autoradiography, labeled terminals and axon profiles contained both normal and accentric dense-core vesicles and they proved to be similar to those seen after 3H-serotonin in vitro incubation. Therefore, it seems possible that certain Anodonta central neurons contain both dopamine and serotonin.     

Growth and age structure of the swan mussel Anodonta cygnea (L.) at different depths in lake Mattsee (Salzburg, Austria)

Unionid clams were collected at 1–2 m, 3–4 m and 6–7 m depth in lake Mattsee, a moderately mesotrophic lake, to investigate the effect of depth on clam growth and age structure. No significant differences in age structure of Anodonta cygnea were found (p=0.65). Three and ten years old clams were present at all depths, but in different percentages. Whereas at 1–2 m 13.3% of the collected clams were <4 years old, this percentage was 4.4% at 6–7 m and 7.1% at 3–4 m. A greater percentage (6.7%) of older mussels (9, 10 years) were collected at 6–7 m than at 1–2 m (2.2%). Growth declined with depth. Total length at a given age of clams at 1–2 m and 3–4 m did not differ (p=0.54), whereas differences were significant between clams at 1–2 m and 6–7 m (p<0.05) as well as between 3–4 m and 6–7 m (p<0.05). The Growth constant k was highest at 1–2 m depth.     

The role of abiotic factors and pesticide levels on enzymatic activity in the freshwater mussel Anodonta cygnea at three different exposure sites

Natural variation in abiotic factors, such as temperature and pH, probably influence the activity of enzymes used as potential biomarkers in bivalve mollusks to assess environmental contamination in the field. Changes in levels of an enzymatic biomarker may thus merely reflect natural variation in the annual physiological cycle of a species rather than exposure to contaminants. To investigate this issue, we documented the relationship between pesticide levels in water and three different enzymatic biomarkers over 1 year in enclosed populations of the freshwater unionid mussel Anodonta cygnea at three different sites of exposure. We considered the natural variation in temperature, pH and dissolved oxygen over the year and across the different sites as a potential correlate of enzymatic activity to disentangle the relative contribution of abiotic factors and pesticide levels. Pesticide levels varied among the three sites and over the course of the year. Catalase (CAT) and acetylcholinesterase activity (AChE) varied as a function of abiotic factors but showed no relation to pesticide levels. Glutathione S-transferase (GST) activity was also related to abiotic factors but also decreased with increases in total pesticide levels. The lack of activity induction or inhibition by pesticides and the natural variation in abiotic factors among sites and across time limits the use of CAT and AChE to assess environmental contamination in this species.