Short-term exposure to dietary Pb and/or DMSA affects dopamine and dopamine metabolite levels in the medulla, optic tectum, and cerebellum of rainbow trout (Oncorhynchus mykiss).

Rainbow trout (Oncorhynchus mykiss) were randomly assigned to one of the following dietary exposure conditions: lead (Pb) solvent (2% nitric acid), meso-2,3-dimercaptosuccinic acid (DMSA) solvent (0.1 N NaOH), Pb, DMSA, Pb followed by Pb solvent, or Pb followed by DMSA. Medulla, cerebellum, and optic tectum homogenates were analyzed for dopamine (DA), homovanillic acid (HVA), and 3,4-dihydroxyphenylacetic acid (DOPAC). DA levels in all brain regions tended to be highest for trout exposed to dietary Pb followed by dietary DMSA. DA levels were elevated for trout exposed to dietary DMSA and Pb followed by Pb solvent. DA levels were below control levels for trout exposed to Pb only. HVA levels varied across brain regions. However, HVA levels in all brain regions tended to be elevated for trout exposed to dietary DMSA and Pb followed by Pb solvent. DOPAC levels across all brain regions were below control levels for trout dietary exposed to DMSA, Pb only, Pb followed by Pb solvent, and Pb followed by DMSA. These data indicate that Pb and/or DMSA have the potential of altering DA, HVA, and DOPAC levels in the medulla, cerebellum, and optic tectum. The animal model of short-term dietary exposure to Pb and DMSA, both alone and sequentially, to mimic dietary exposure to Pb and the oral delivery of DMSA, that our laboratory has developed, may be useful in future studies aimed at characterizing the neurobiological mechanisms by which Pb and/or DMSA alter neurotransmitter levels and behavior.

Exposure to sublethal levels of waterborne lead alters reproductive behavior patterns in fathead minnows (Pimephales promelas).

Lead (Pb) caused multiple effects on reproductive behavior and overall reproductive success. Adult fathead minnows (Pimephales promelas) were acclimated at a 16L:8D photoperiod to stimulate reproductive development. Reproductively mature adults were separated as male-female pairs and maintained for 4 weeks in either 0.0 or 0.5 ppm Pb. High lead accumulations occurred in testes and ovaries of treated fish; lead concentrations in control fish gonads were not detectable. Lead suppressed spermatocyte production and retarded ovarian development, although no lead-induced gonadosomatic index changes for either sex were noted. Lead decreased the number of eggs oviposited, increased interspawn periods and suppressed embryo development. Control males displayed maximum secondary sex characteristic development (banding, tubercle formation, head and eye darkening); lead-exposed fish displayed less. Control males spent more time in ceiling-directed behaviors associated with nest preparation and maintenance than lead-exposed. These variables were affected differentially with respect to stage of reproductive maturity at time of lead exposure, i.e., fish displaying greater secondary sex characteristic development before exposure were less affected by lead than those fish that showed less development.

Metal redistribution in largemouth bass (Micropterus salmoides) in response to restrainment stress and dietary cadmium: role of metallothionein and other metal-binding proteins.

1. Fish stressed by restrainment displayed elevated serum cortisol, copper and zinc levels; dietary cadmium had no effect. 2. Stress/dietary cadmium increased liver copper levels in a metal pool containing metallothionein and non-metallothionein proteins but decreased intestinal zinc bound as low molecular weight forms. 3. After restrainment, zinc losses occurred in dorsal skeletal muscle, ovary and spleen: copper decreased in intestine and pyloric caecum. 4. Dietary cadmium altered intestinal zinc distribution and raised hepatic Cu-binding protein levels but did not alter plasma zinc, copper or cortisol levels. 5. Alterations in zinc and copper concentrations during stress contrast with mammalian models.

Immunochemical and immunohistochemical studies of cadmium associated proteins in Rana tadpoles.

Previous observations suggested that Rana tadpoles treated with aqueous cadmium (Cd) accumulate Cd in their liver and mesonephros. In order to study the response to Cd in these tissues we (a) exposed tadpoles in mid-limb bud stages to sublethal quantities of Cd, (b) isolated Cd-associated protein (CAP) from a liver cytosol fraction, (c) prepared a heterologous rabbit antiserum against glutaraldehyde-treated CAP (G-CAP), (d) used the rabbit anti-G-CAP antiserum in order to assess the tissue distribution of CAP in Cd-treated and untreated tadpoles, and (e) assessed species cross-reactivities of our anti-G-CAP with CAPs and metallothioneins (MTs) isolated from Cd-treated vertebrate liver cytosol fractions. We found that (a) CAP was present in higher quantities in liver cytosol obtained from Cd-treated tadpoles compared to liver cytosol obtained from untreated control tadpoles, (b) indirect immunofluorescent analysis revealed that CAP was localized in liver hepatocytes and kidney tubule epithelial cells in Cd-treated tadpoles, and (c) the anti-G-CAP crossreacted with rodent and fish CAP. These observations suggest that the developing liver and mesonephros are involved in responses to toxic metals and that our anti G-CAP antiserum may be used to gauge exposure to environmental Cd.

Effects of prolonged low-level cadmium exposure on the tadpole immune system.

Uninjected (Group I) and sheep erythrocyte (SRBC)-injected (Group II) Rana tadpoles were exposed to varying sublethal concentrations of cadmium (Cd) for 6 weeks. In order to assess possible effects on the tadpole immune system we determined pre-B, B mu, and plasma cell (PC mu) frequencies in liver and mesonephros of Group I larvae, and hemagglutinating antibody (HA) titers of Group II animals. Group I and Group II control animals were cultivated in water with no added Cd (0 ppm), while treatments were set at 0.1, 0.2, 0.4 and 0.8 ppm Cd. We found that (a) Cd appeared to stimulate a significant increase in the frequency of B mu cells in animals treated at 0.4 and 0.8 ppm, (b) certain treated Group II larvae contained significantly increased amounts of HA in their serum, while their serum protein concentrations were not significantly different, and (c) there was a dose-related increase in tissue Cd levels in treated Group II larvae. Our observations suggest that chronic low-level exposure to Cd may (a) result in a slight increases in the frequency of B mu cells in unimmunized animals, (b) increase immune responsiveness of immunized larvae, and (c) increase liver and mesonephros accumulations of Cd. Preliminary studies indicated that cytosolic Cd is associated with a protein which appears to be similar to mammalian metallothionein.

Effects of the light-dark cycle and scheduled feeding on behavioral and reproductive rhythms of the cyprinodont fish, Medaka, Oryzias latipes.

Medaka were maintained on a 16:8 light-dark cycle and fed once daily on one of 5 different feeding schedules. The daily rhythm of agonistic behavior rapidly entrained to the scheduled feeding time and maintained this entrainment during a 3-day starvation period. In contrast the daily rhythms of egg laying and courtship stayed entrained to the L:D cycle regardless of the feeding schedule. Thus, temporal integration of this fish with its daily environment can involve multistimuli which concurrently and differentially entrain externally expressed circadian systems.

Euglena gracilis cadmium-binding protein-II contains sulfide ion.

Sulfide ions are a constituent of the cadmium-binding protein-II in the alga Euglena gracilis. Their presence was demonstrated by the methylene blue assay, by acid labilization induced reductions in the Cd-S charge transfer band at 254 nm and by reactions with the thiol reagent, 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB). Direct reduction of DTNB by sulfide and precipitation of CdS yield a complex stoichiometry for the DTNB reaction. The S2-/Cd2+ ratios determined, 1.25 +/- 0.10 (methylene blue) and 1.37 +/- 0.16 (DTNB), are in good agreement.

Characterization of a highly negative and labile binding protein induced in Euglena gracilis by cadmium.

The physiochemical properties and physiological significance of the cadmium-binding protein (CdBP) of the algae Euglena gracilis have been studied. Following in vivo exposure of cells to 0.4 or 1.3 micrograms/mL of Cd2+, all the cytosolic Cd is bound to high molecular weight species. At 4.7 micrograms/mL, appreciable CdBP has formed in cells grown under illumination or in the dark. An analogous ZnBP could not be isolated from control or Zn-exposed (20 micrograms/mL) cells, but zinc and a trace of copper were bound to the CdBP when 2-mercaptoethanol (2-ME) is added to the homogenates of Cd-treated cells and the buffers used during isolation. The large pool of very low molecular weight zinc species previously reported is increased when cells are exposed to high cadmium levels. Two distinct species, BP-1 and BP-2 are resolved by ion-exchange chromatography on DEAE-Sephadex. Unusually high conductivities (25 and 40 mSiemen) are required to displace them, indicating that they are very negatively charged proteins at pH 8.6. The pH for half-titration of bound Cd2+ is between 5 and 6. EDTA (0.4 M) and the CdBP isolated by gel-exclusion chromatography react biphasically with pseudo-first-order rate constants of 4 +/- 3 X 10(-4) sec-1 and 7 +/- 2 X 10(-5) sec-1. Neither form of the CdBP cross-reacts with antibodies to rat liver metallothionein (MT) antibodies. The structural, chemical, and functional differences between the Euglena CdBPs and mammalian MTs are discussed.(ABSTRACT TRUNCATED AT 250 WORDS)