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Bullfrog tadpoles, Aquarana catesbeiana, were exposed to settleable particulate matter (SePM), (1 g L-1, 96 h) and their organs were collected for analysis of metal/metalloid, oxidative stress and neurotoxicity in liver, muscle, kidney and brain. The SePM water of the exposed groups contained 18 of the 28 metals/metalloids detected in ambient particulate matter (APM). Fe56 and Al were those that presented the highest concentrations, Cr, Mn, Pb and Cu increased from 10 to 20 times and Ti, V, Sr, Rb, Cd, Sn and Ni increased from 1 to 3 times compared to the control. Bioaccumulation of metals/metalloids in the exposure water varied significantly between organs, with the muscle and liver showing the highest concentrations of metals, followed by the brain. Lipoperoxidation and malondialdehyde increased only in muscle, while carbonyl proteins increased only in the liver and brain. Regarding nitric oxide synthase, there was an increase in the liver and brain in the group exposed to SePM. Catalase activity decreased in the liver and muscle, while the activity of glutathione peroxidase, increased in the liver and kidney and decreased in muscle. Glutathione S-transferase, which is mainly responsible for detoxification, increased in the liver and decreased in muscle and the kidney. Cholinesterase activity increased only in the muscle. The results indicate oxidative stress, due to oxidation catalyzed by metals, components of SePM. Thus, the results contribute to the understanding that SePM has a deleterious effect on the aquatic environment, negatively affecting bullfrog tadpoles, in different ways and levels in relation to the analyzed organs.

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Carnivorous reptiles exhibit an intense metabolic increment during digestion, which is accompanied by several cardiovascular adjustments responsible for meeting the physiological demands of the gastrointestinal system. Postprandial tachycardia, a well-documented phenomenon in these animals, is mediated by the withdrawal of vagal tone associated with the chronotropic effects of non-adrenergic and non-cholinergic (NANC) factors. However, herbivorous reptiles exhibit a modest metabolic increment during digestion and there is no information about postprandial cardiovascular adjustments. Considering the significant impact of feeding characteristics on physiological responses, we investigated cardiovascular and metabolic responses, as well as the neurohumoral mechanisms of cardiac control, in the herbivorous lizard Iguana iguana during digestion. We measured oxygen consumption rate (O2), heart rate (fH), mean arterial blood pressure (MAP), myocardial activity, cardiac autonomic tone, fH/MAP variability and baroreflex efficiency in both fasting and digesting animals before and after parasympathetic blockade with atropine followed by double autonomic blockade with atropine and propranolol. Our results revealed that the peak of O2 in iguanas was reached 24 h after feeding, accompanied by an increase in myocardial activity and a subtle tachycardia mediated exclusively by a reduction in cardiac parasympathetic activity. This represents the first reported case of postprandial tachycardia in digesting reptiles without the involvement of NANC factors. Furthermore, this withdrawal of vagal stimulation during digestion may reduce the regulatory range for short-term fH adjustments, subsequently intensifying the blood pressure variability as a consequence of limiting baroreflex efficiency.

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In addition to their well-known classical effects, cannabinoid CB1 and CB2 receptors have also been involvement in both deleterious and protective actions on the heart under various pathological conditions. While the potential therapeutic applications of the endocannabinoid system in the context of cardiovascular function are indeed a viable prospect, significant debate exists within the literature regarding whether CB1, CB2, or a combination of both receptors exert a favorable influence on cardiac function. Hence, the aim of this study was to investigate the effects of CB1 + CB2 or CB2 agonists on cardiac excitation-contraction (E-C) coupling, utilizing fish (Brycon amazonicus) as an experimental model. The CB2 agonist elicited marked positive inotropic and lusitropic responses in isolated ventricular myocardium, induced cyclic adenosine 3',5'-monophosphate (cAMP) production, and upregulated critical Ca2+ handling proteins, such as sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) and Na+/Ca2+ exchanger (NCX). Our current study demonstrated, for the first time, that CB2 receptor activation-induced effects improved the efficiency of Ca2+ cycling, excitation-contraction coupling (E-C coupling), and cardiac performance in under physiological conditions. Hence, CB2 receptors could be considered a potential therapeutic target for modulating cardiac contractile dysfunctions.

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Fipronil is widely used as a broad-spectrum insecticide in agriculture, urban environments, and veterinary medicine. Fipronil can enter aquatic ecosystems and spread to sediment and organic matter, representing a risk to non-target species. This study aimed to evaluate the effects of short-term (96 h) exposure to a low and realistic concentration of sediment-associated fipronil (4.2 µg.kg-1 of Regent® 800 WG) on myocardial contractility of armored catfish Hypostomus regain, a benthic fish species. Fipronil exposure induced increased inotropism and acceleration of contractile kinetics, although no alterations in the relative ventricular mass were observed. This better cardiac function was associated with an elevated expression and/or function of the Na+/Ca2+ exchanger and its marked contribution to contraction and relaxation, probably due to a stress-induced adrenergic stimulation. Ventricle strips of exposed fish also exhibited a faster relaxation and a higher cardiac pumping capacity, indicating that armored catfish were able to perform cardiac adjustments to face the exposure. However, a high energetic cost to maintain an increased cardiac performance can make fish more susceptible to other stressors, impairing developmental processes and/or survival. These findings highlight the need for regulations of emerging contaminants, such as fipronil, to ensure adequate protection of the aquatic system.

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Iron and steel industries discharge a large amount of atmospheric particulate matter (PM) containing metals and metallic nanoparticles (NPs) that contaminate not only the air, but also settle into the aquatic environments. However, the effects of settleable atmospheric particulate matter (SePM) on aquatic fauna are still poorly understood. This study aimed to evaluate the sublethal effects of a short-term exposure to a realistic concentration of SePM on Nile tilapia (Oreochromis niloticus) using a multi-biomarker approach: relative ventricular mass (RVM) and heart function, blood oxidative stress, stress indicators, hemoglobin concentration, metallic NPs internalization, and metal bioaccumulation. Exposed fish exhibited reduced hemoglobin content and elevated plasma cortisol and glucose levels, reflecting stressed states. Furthermore, SePM caused blood oxidative stress increasing lipid and protein oxidation, decreasing glutathione levels, and inhibiting superoxide and glutathione reductase activities. SePM exposure also increased RVM and improved cardiac performance, increasing myocardial contractile force and rates of contraction and relaxation. In the heart tissue there was a significant accumulation of Fe > Zn > > Cr > Cu > Rb > Ni > V > Mn > Se > Mo > As. On the other hand, in the erythrocytes there was significant accumulation of Sn > Zn > > Cr > Ti > Mn = Ni > Nb > As > Bi. The highest bioaccumulation factors were found for Cr, Zn and Ni in both tissues. NPs (Ti, Sn, Al, Fe, Cu, Si, Zn) were also detected in ventricular myocardium of fish exposed and nanocrystallographic analysis revealed a predominance of anatase phase of TiO2-NP, which is regarded to be more cytotoxic. The association between blood oxidative stress and energy expenditure to sustain increased cardiac pumping capacity under stress condition suggests that SePM has negative impacts on fish physiological performance, threatening their survival, growth rate and/or population establishment.

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The aim of this study was to evaluate the effects of different doses of alternagin-C, a disintegrin-like protein from Rhinocerophis alternatus venom, on myocardial contractility of the freshwater fish Hoplias malabaricus, an alternative model to contractile function studies. Alternagin-C treatment exhibited a hormetic-like dose-response curve with a strong positive inotropism and enhanced cardiac pumping capacity at low dose, whereas a modest inotropism and a left shift in the force-frequency relationship was registered at high dose.

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This study investigated the dependence of contraction from extracellular Ca2+, the presence of a functional sarcoplasmic reticulum (SR), and the effects of ß-adrenergic stimulation using isometric cardiac muscle preparations. Moreover, the expression of Ca2+-handling proteins such as SR-Ca2+-ATPase (SERCA), phospholamban (PLN), and Na+/Ca2+ exchanger (NCX) were also evaluated in the ventricular tissue of adult African sharptooth catfish, Clarias gariepinus, a facultative air-breathing fish. In summary, we observed that (1) contractility was strongly regulated by extracellular Ca2+; (2) inhibition of SR Ca2+-release by application of ryanodine reduced steady-state force production; (3) ventricular myocardium exhibited clear post-rest decay, even in the presence of ryanodine, indicating a decrease in SR Ca2+ content and NCX as the main pathway for Ca2+ extrusion; (4) a positive force-frequency relationship was observed above 60 bpm (1.0 Hz); (5) ventricular tissue was responsive to ß-adrenergic stimulation, which caused significant increases in twitch force, kept a linear force-frequency relationship from 12 to 96 bpm (0.2 to Hz), and improved the cardiac pumping capacity (CPC); and (6) African catfish myocardium exhibited similar expression patterns of NCX, SERCA, and PLN, corroborating our findings that both mechanisms for Ca2+ transport across the SR and sarcolemma contribute to Ca2+ activator. In conclusion, this fish species displays great physiological plasticity of E-C coupling, able to improve the ability to maintain cardiac performance under physiological conditions to ecological and/or adverse environmental conditions, such as hypoxic air-breathing activity.

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Pythons are important models of studies on postprandial metabolism because their physiological responses are exacerbated when digesting large prey. Prior studies of these animals have shown hypertrophy of the cardiac tissue 2 to 3 days after feeding, coinciding with the peak of the specific dynamic action (SDA), but the consequences of this remodeling in myocardial contractility have not been studied, which is the purpose of this work. Specimens of Python molurus were divided into two groups: a Digesting group (2 days after feeding, at the peak of SDA), and a Fasting group (28 days after feeding). When compared to the Fasting group, the Digesting group showed higher relative ventricular mass and calcium-handling protein expression such as sarcoplasmic reticulum Ca2+-ATPase (SERCA), phospholamban (PLB), and the Na+/Ca2+ exchanger (NCX). Digesting pythons also exhibited significant increases in the cardiac contraction force (Fc), rates of force development and relaxation, and cardiac pumping capacity. Therefore, the higher SERCA, PLB and NCX expression levels increased cytosolic Ca2+ transient amplitude, improving myofilament force. These changes are crucial to maintain cardiac output and a relatively high and continuous blood flow required by metabolic expenditure that occurs in postprandial animals.

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This study investigated the effect of microcystin-LR (MC-LR) on in vivo cardiorespiratory function and on tissue biomarkers of oxidative stress in gills and liver of the trahira, a neotropical freshwater fish. Trahira were treated with an intraperitoneal injection of 100 µg MC-LR.kg-1 body mass or a saline, with the toxic effects of MC-LR then evaluated after 48 h. Rates of oxygen uptake (V̇O2) did not differ significantly between Control and the exposed group (Mcys), but exposure to MC-LR significantly reduced O2 extraction in the Mcys group at all O2 tensions. This was associated with higher gill ventilation volume (V̇G) in the Mcys group at all O2 tensions except 140 and 120 mmHg, and a higher tidal volume (VT) of the Mcys group at all tensions except 140 mmHg. Heart rate was also higher in the Mcys group, significantly so at an O2 tension of 40 mmHg. In the liver of trahira, exposure to MC-LR has significant effects on antioxidant defense systems, inducing a significant increase in the activity of the (GPx) glutathione peroxidase enzyme (100%) and in the reduced glutathione (GSH) content (70%) compared to the control group, but no effects on superoxide dismutase (SOD), catalase (CAT) and glutathione S-transferase (GST) enzymes. The liver showed no oxidative damage, when measured as lipid peroxidation (LPO) levels and protein carbonyl (PC) content. In the gills SOD and GPx enzyme activity increased significantly in the Mcys group (98% and 73% respectively) compared to the controls, although GSH, CAT and GST did not differ between groups. There was also no significant difference in GSH in this tissue. Levels of lipid peroxidation in the gills were 53% higher in the Mcys group, although carbonyl protein levels did not differ. In conclusion, these data show that MC-LR leads to development of hyperventilation and increased activity of the detoxification system and that this species was able to compensate the deleterious effects of microcystin on its vital functions. The antioxidant defense in the liver was able to contain the propagation of LPO and prevent the oxidation of proteins, although the gills of the fishes exposed to MC-LR were not able to contain the formation of reactive oxygen species and LPO, which led to the establishment of oxidative stress which impaired gill function.

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Microcystin's (MCs) are toxins produced by several groups of cyanobacteria, in water bodies throughout the world, in a process which is being intensified by human action. Among the variants of MCs, MC-LR stands out for its distribution and toxicity. MCs are potent inhibitors of protein phosphatases 1 and 2 A, which causes disruption of the cytoskeleton and consequent cell death. They can also alter the antioxidant system and induce oxidative stress in various organs of many species. There is, however, a lack of information about the effects of MCs on the antioxidant system and oxidative damage in Brazilian fishes. This study evaluated the effect of microcystin-LR on the antioxidant system in liver and gills of the Brazilian fish Brycon amazonicus, after 48 h of i.p injection of 100 µg MC-LR.kg-1 body mass. The liver exhibited increases in the activity of GST (74%) and GPx (217%), and a 47% decrease in SOD activity, with no changes in CAT values. In the gills of fish exposed to MC-LR, CAT and GPx activities did not show significant changes, while SOD and GST activity decreased by 66% and 37%, respectively. The GSH content did not change significantly in the liver, however, a decrease of 43% was observed in the gills. Oxidative damage measured by protein oxidation (PC) and lipoperoxidation (LPO) showed significant effects in both tissues. In hepatic tissue, there was no change in PC levels but LPO increased by 116%. Conversely, in the gills LPO levels did not change but PC increased by 317%. In conclusion, these data show that MC-LR induces oxidative damage in both tissues but in different ways, with being liver most sensitive to LPO and gills to PC. This also suggests that the gills are most sensitive to oxidative stress than liver, due to the inhibition of its antioxidant responses following MC-LR exposure.

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Alternagin-C (ALT-C) is a disintegrin-like protein isolated from Rhinocerophis alternatus snake venom, which induces endothelial cell proliferation and angiogenesis. The aim of this study was to evaluate the systemic effects of a single dose of alternagin-C (0.5 mg·kg-1, via intra-arterial) on oxidative stress biomarkers, histological alterations, vascular endothelial growth factor (VEGF) production, and the degree of vascularization in the liver of the freshwater fish traíra, Hoplias malabaricus, seven days after the initiation of therapy. ALT-C treatment increased VEGF levels and hepatic angiogenesis. ALT-C also enhanced hepatic antioxidant enzymes activities such as superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase, decreasing the basal oxidative damage to lipids and proteins in the fish liver. These results indicate that ALT-C improved hepatic tissue and may play a crucial role in tissue regeneration mechanisms.

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The growing Hg input in aquatic environments results in high accumulation of mercury in fish tissue and their consumers, which poses a serious risk to humans and ecosystems. The aim of this study was to evaluate the effects of the inorganic mercury exposure on cardiorespiratory responses in two species of neotropical fish ecologically distinct, matrinxã (Brycon amazonicus) and traíra (Hoplias malabaricus). Matrinxãs were exposed to a nominal and sublethal concentration of 0.15 mgL(-1) of HgCl2 for 96 h. Traíras were exposed to trophic doses (each 4 days, during 30 days) of inorganic Hg (0.45 mg as total Hg) using juvenile B. amazonicus as prey vehicle. The metabolic rate (VO2), critical oxygen tensions (PcO2), gill ventilation (VG), tidal volume (VT), respiratory frequency (fR), O2 extraction from the ventilatory current (EO2), and heart rate (fH) were measured under normoxia (140 mm Hg) and graded hypoxia (120, 100, 80, 60, 40, 20, and 10 mm Hg). Regarding matrinxã specifically, the critical point highlighted was tachypnea. In traíras, bradypnea, decreased metabolic rate and O2 extraction, severe bradycardia, and elevated tidal volume were observed in normoxia. Both acute and sub-chronic exposures increased the critical tension of O2 values in more than 100%. In addition, Hg exposures modulated hypoxia-induced responses resulting in impairment of cardio-respiratory system of both species. Thus, mercury, via food or water, decreases the plasticity of the cardiorespiratory responses reducing the survival chances of B. amazonicus and H. malabaricus under hypoxic conditions frequently observed in theirs wild habitats.

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This study evaluated the effects of trophic and subchronic exposure to inorganic mercury (Hg) on the oxidative stress biomarkers and its bioaccumulation potential in the liver, gills, white muscle and heart of the freshwater top predator fish, Hoplias malabaricus, fed with contaminated live juveniles of matrinxã, Brycon amazonicus, as prey vehicle. Inorganic mercury increased superoxide dismutase, catalase (CAT), glutathione peroxidase (GPx), glutathione S-transferase, and glutathione reductase (GR) activities in the liver, white muscle and heart. Gills CAT activity remained unchanged while GPx and GR values showed a significant decrease. In the liver and gills, Hg induced significant increase in the reduced (GSH) and oxidized (GSSG) glutathione content, concomitantly with a significant decrease in [GSH]/[GSSG] ratio. Differently, in cardiac tissue, the Hg caused an increase in GSH level and increase in [GSH]/[GSSG] ratio. Lipid and protein oxidation and metallothionein levels were significantly higher after Hg trophic exposure in the liver, gills and heart, but remained at control values in the white muscle. Tissue-specific responses against oxidative stress were observed, and the liver and gills were the most sensitive organs, showing signs of redox homeostasis failure. At the end of the experiment, dietary inorganic mercury accumulated through food chain levels. In order, Hg bioaccumulation was: gills > liver >> white muscle = heart. These results pointed out the potential of inorganic Hg to bioaccumulate in aquatic systems. Taken together, our findings suggest that Hg, even in the inorganic form and sublethal amounts, is a risk factor for aquatic biota.

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Alterations in the antioxidant cellular system have often been proposed as biomarkers of pollutant-mediated toxicity. This study evaluated the effects of mercury on oxidative stress biomarkers and bioaccumulation in the liver, gills, white muscle and heart of the freshwater fish matrinxã, Brycon amazonicus, exposed to a nominal and sub-lethal concentration (~20% of 96 h-LC(50)) of 0.15 mg L(-1) of mercury chloride (HgCl(2)) for 96 h in a static system. Increases in superoxide dismutase, catalase, glutathione peroxidase (GPx), glutathione S-transferase (GST) and glutathione reductase (GR) were observed in all tissues after HgCl(2) exposure, except for white muscle GR activity and hepatic GPx. In the liver and gills, the exposure to HgCl(2) also induced significant increases in reduced glutathione (GSH). Conversely, exposure to HgCl(2) caused a significant decrease in the GSH levels and an increase in the oxidized glutathione (GSSG) content in the white muscle, while both GSH and GSSG levels increased significantly in the heart muscle. Metallothionein concentrations were significantly high after HgCl(2) exposure in the liver, gills and heart, but remained at control values in the white muscle. HgCl(2) exposure induced oxidative damage, increasing the lipid peroxidation and protein carbonyl content in all tissues. Mercury accumulated significantly in all the fish tissue. The pattern of accumulation follows the order gills > liver >> heart > white muscle. In conclusion, these data suggest that oxidative stress in response to inorganic mercury exposure could be the main pathway of toxicity induced by this metal in fish.

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The cardio-respiratory function, oxidative stress and fish antioxidants were analyzed in juvenile Nile tilapia exposed for 96 h to a sublethal trichlorfon (TRC-Neguvon, Bayer) concentration of 0.5 mg L(-1). The exposure to TRC induced oxidative stress in the heart, as manifested by the glutathione S-transferase depletion and hydroperoxide elevation, and was the most sensitive organ when compared to the liver and gills, in which the antioxidant mechanisms against TRC exposure were sufficient to remove reactive oxygen species (ROS), preventing the increase of lipid peroxidation. TRC exposure also reduced O(2) uptake (V O(2)) and increased the critical oxygen tension (PcO(2)), reducing the species capacity to survive prolonged hypoxic conditions. The heart rate and force contraction were significantly impaired, making the heart the most sensitive organ when exposed to the TRC.

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Methyl parathion (MP), an organophosphate widely applied in agriculture and aquaculture, induces oxidative stress due to free radical generation and changes in the antioxidant defense system. The antioxidant roles of selenium (Se) were evaluated in Brycon cephalus exposed to 2 mg L(-1) of Folisuper 600 BR (MP commercial formulation - MPc, 600 g L(-1)) for 96 h. Catalase (CAT), glutathione peroxidase (GPx), superoxide dismutase (SOD), glutathione S-transferase (GST), reduced glutathione (GSH) and lipid peroxidation (LPO) levels in the gills, white muscle and liver were evaluated in fish fed on diets containing 0 or 1.5 mg Se kg(-1) for 8 weeks. In fish treated with a Se-free diet, the MPc exposure increased SOD and CAT activities in all tissues. However, the GPx activity decreased in white muscle and gills whereas no alterations were observed in the liver. MPc also increased GST activity in all tissues with a concurrent decrease in GSH levels. LPO values increased in white muscle and gills and did not change in liver after MPc exposure. A Se-supplemented diet reversed these findings, preventing increases in LPO levels and concurrent decreases in GPx activity in gills and white muscle. Similarly, GSH levels were maintained in all tissue after MPc exposure. These results suggest that dietary Se supplementation protects cells against MPc-induced oxidative stress.

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Os benzodiazepínicos estão entre as drogas mais freqüentemente prescritas em razão de suas propriedades ansiolíticas. O objetivo deste trabalho foi avaliar a influência do diazepam sobre a resposta inflamatória peritoneal aguda induzida por lipopolissacarídeo. Para tanto, camundongos Swiss foram tratados com diazepam (1 ou 10 mg/kg de peso), em dose única, por via subcutânea, uma hora antes do desafio intraperitoneal com lipopolissacarídeo bacteriano. Após 16 horas do desafio, os animais foram sacrificados, coletando-se os lavados peritoneais para determinação do número total de células e das subpopulações de mononucleares e polimorfonucleares, além da atividade de TNF-α e da porcentagem de macrófagos espraiados. Observou-se que o tratamento com diazepam, nas doses de 1 ou 10 mg/kg, reduziu significativamente a porcentagem de macrófagos estimulados por LPS e a liberação de TNF-α independente de estímulo. Houve também significativa redução da migração de leucócitos nos animais estimulados com LPS e tratados com 10 mg/kg de diazepam em relação aqueles não tratados. Concluímos que a administração do diazepam, em dose única, pode influenciar significativamente o influxo celular, a estimulação de macrófagos e a atividade de TNF-α na resposta inflamatória aguda induzida por LPS em camundongos, com possíveis implicações na eficiência da resposta anti-infecciosa.

Benzodiazepines are one of the most frequently prescribed drugs due to their anxiolytic properties. The aim of this study was to evaluate the effects of diazepam on lipopolysaccharide-induced peritoneal acute inflammatory responses. Swiss mice were treated with diazepam in a single dose of 1 or 10 mg/kg- subcutaneously 1 h before an intraperitoneal injection of lipopolysaccharide or sterile saline solution. The mice were killed 16 h after and the cells were washed from the peritoneal cavity to determine the total number of cells and the mononuclear and polimorfonuclear subpopulations, as well as the TNF-alpha activity and percentage of spread macrophages. Our results showed that the diazepam treatment (1 and 10 mg/kg) induced a significant reduction in the LPS-induced macrophage stimulation and TNF-α activity. Diazepam (10 mg/kg) also reduced the inflammatory cellular migration when compared to the control. It can be concluded that the diazepam treatment in a single dose is able to influence the inflammatory cellular influx, macrophage stimulation and TNF-α activity in the acute inflammatory response in mice, having possible implications on the anti-infectious response efficiency.

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Methyl parathion (MP) is an organophosphorus insecticide used worldwide in agriculture and aquaculture due to its high activity against a broad spectrum of insect pests. The effect of a single exposure to 2 mg L(- 1) of a commercial formulation of MP (MPc: Folisuper 600(R), MP 600 g L(- 1)) on catalase (CAT), glutathione peroxidase (GPx), superoxide dismutase (SOD), glutathione S-transferase (GST), reduced glutathione (GSH) and lipid peroxidation (LPO) of the liver, white muscle and gills of Brycon cephalus was evaluated after 96 h of treatment. MPc exposure resulted in a significant induction of SOD, CAT and GST activity in all tissues. However, the GPx activity decreased significantly in white muscle and gills, whereas no alterations were observed in hepatic GPx activity. MPc also induced a significant increase in LPO values in the white muscle and gills, while hepatic LPO levels did not show any significant alteration. The current data suggest that MPc has oxidative-stress-inducing potential in fish, and that gills and white muscle are the most sensitive organs of B. cephalus, with poor antioxidant potentials. The various parameters studied in this investigation can also be used as biomarkers of exposure to MPc.

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