Biochemical and developmental effects of thyroid and anti-thyroid drugs on different early life stages of Xenopus laevis.

The effects of two drugs containing the synthetic thyroid hormone levothyroxine (LEV) and an anti-thyroid drug containing propylthiouracil (PTU) on the three early life stages of Xenopus laevis were evaluated with the Frog Embryo Teratogenesis Assay-Xenopus, Tadpole Toxicity Test, and Amphibian Metamorphosis Assay using biochemical and morphological markers. Tested drugs caused more effective growth retardation in stage 8 embryos than stage 46 tadpoles. Significant inhibition of biomarker enzymes has been identified in stage 46 tadpoles for both drugs. AMA test results showed that LEV-I caused progression in the developmental stage and an increase in thyroxine level in 7 days exposure and growth retardation in 21 days exposure in stage 51 tadpoles. On the other hand, increases in lactate dehydrogenase activity for both drugs in the AMA test may be due to impacted energy metabolism during sub-chronic exposure. These results also show that the sensitivity and responses of Xenopus laevis at different early developmental stages may be different when exposed to drugs.

Thyroid disruption and oxidative stress in American kestrels following embryonic exposure to the alternative flame retardants, EHTBB and TBPH.

Brominated flame retardant chemicals, such as 2-ethylhexyl-2,3,4,5-tetrabromobenzoate (EHTBB) (CAS #: 183658-27-7) and bis(2-ethylhexyl)-2,3,4,5-tetrabromophthalate (TBPH) (CAS #: 26040-51-7), have been detected in avian tissues and eggs from remote regions. Exposure to EHTBB and TBPH has been shown to cause oxidative stress and altered thyroid function in rodents and fish, yet no controlled studies have examined potential adverse effects of exposure in birds. Because flame retardants have been detected in wild raptors, we used American kestrels (Falco sparverius) as a model raptor to determine whether in ovo exposure to EHTBB or TBPH affected growth, hatching success, oxidative stress, or thyroid function. We exposed kestrel embryos to nominal concentrations (10, 50, or 100 ng g-1 egg weight) of EHTBB and TBPH via egg-injection on embryonic day 5. Embryonic exposure (~23 d) to EHTBB increased thyroid gland mass, reduced glandular colloid and total thyroxine (T4) in hatchling males and females, whereas deiodinase enzyme activity increased in males but decreased in females. Hatchlings exposed to TBPH in ovo exhibited reduced colloid and increased oxidative stress. Although exposure to EHTBB and TBPH caused several physiological effects (e.g., heart and brain mass), only exposure to 50 ng g-1 EHTBB appeared to reduce hatching success. Our results suggest these flame retardants may be hazardous for predatory birds. Future research should evaluate long-term survival and fitness consequences in birds exposed to these chemicals.

Effects of sodium perchlorate and exogenous L-thyroxine on growth, development and leptin signaling pathway of Bufo gargarizans tadpoles during metamorphosis.

Sodium perchlorate (NaClO4) and exogenous L-thyroxine (T4), two kinds of endocrine-disrupting chemicals (EDCs), mainly affect the circulating thyroid hormones, which regulate the initiation and rate of metamorphosis in amphibian. The aim of this study is to evaluate the potential role of EDCs in regulating the development of tadpoles and leptin signaling pathway of liver during the metamorphosis of Bufo gargarizans. There was completely opposite result of average development stage of tadpoles and morphological parameters between the NaClO4 and T4 exposure groups. Histological analysis revealed that NaClO4 and T4 exposure both caused liver injury, such as the decreased size of hepatocytes, atrophy of nucleus, increased melanomacrophage centres and disappearance of hepatocyte membranes. In addition, the results of RT-qPCR revealed that NaClO4 treatment significantly inhibited the transcript levels of genes related to thyroid hormone (D2, TRα and TRß) and leptin signaling pathway (LepR, JAK1, JAK2, and TYK2), while there was an increase of mRNA expression of these genes in the liver of tadpoles administrated with T4 compared with control. This work lays an important foundation for assessing the risk of EDCs in relation to amphibian development during metamorphosis.

The effect of thyroid dysfunction and treatment on adropin, asprosin and preptin levels in rats.

OBJECTIVES: Thyroid hormones have important roles in normal development and energy regulating mechanisms as well as signaling mechanisms that affect energy consumption through central and peripheral pathways. The aim of this study was to determine the effects of thyroid dysfunction on adropin, asprosin and preptin levels in rat. METHODS: The study was performed on the 38 male Wistar-albino rats. Experiment groups were designed as follows. 1-Control, 2-Hypothyroidism; To induce hypothyroidism PTU was applied by intraperitoneal as 10 mg/kg/day for 2 weeks. 3-Hypothyroidism + Thyroxine; Previously animals were made with hypothyroidism by 1 week PTU application and then 1 week l-thyroxine was given by intraperitoneal as 1.5 mg/kg/day. 4-Hyperthyroidism; Rats were made with hyperthyroidism by 3 weeks l-thyroxine (0.3 mg/kg/day). 5-Hyperthyroidism + PTU; Animals were made hyperthyroisim by l-thyroxine as groups 4, then 1 week PTU was applied to treatment of hiperthyrodism. At the end of supplementation animals were sacrificed and blood samples were collected for FT3, FT4, adropin, asprosin, preptin analysis. RESULTS: FT3 ve FT4 levels were reduced significantly in hypothyroidism while increased in hyperthyroidism (p<0.001). Hipothyrodism led to reduces adropin, asprosin and preptin levels. And also hyperthyroidism reduced adropin and preptin levels (p<0.001). CONCLUSIONS: The results of study show that experimental hypothyroidism and hyperthyroidism lead to significantly change to adropin, asprosin and preptin levels. However, correction of thyroid function caused to normals levels in asprosin and preptin.

The Impact of Spironolactone on Markers of Myocardial Oxidative Status, Inflammation and Remodeling in Hyperthyroid Rats.

BACKGROUND: Hyperthyroidism promotes the development and progression of cardiovascular diseases (CVD). Aldosterone, a key mediator of myocardial inflammation, oxidative stress and fibrosis, may be activated in hyperthyroidism. OBJECTIVE: To assess the impact of hyperthyroidism on aldosterone levels and myocardial oxidative status, inflammatory and fibrotic markers in hyperthyroid rats, and to test if the use of spironolactone (an aldosterone antagonist) attenuates these changes. METHODS: Adult Wistar rats were randomly distributed into 4 groups; controls, spironolactone treated rats (Spir, 50mg/kg/day), hyperthyroid rats (Hyper, daily intraperitoneal levothyroxine 0.3mg/kg/day), and spironolactone treated hyperthyroid rats (Hyper+Spir) for 4 weeks. Blood pressure (Bp), and levels of serum and myocardial aldosterone, oxidants/antioxidants, inflammatory and fibrotic markers were measured. RESULTS: Levothyroxine increased serum thyroid hormones and increased Bp, heart rate and heart to bodyweight ratio. Relative to control, serum aldosterone levels were increased in Hyper and Hyper+ Spir groups. In parallel, cardiac lipid peroxides and serum endothelin-1 were increased whereas cardiac superoxide dismutase, catalase, glutathione, and matrix metalloproteinase -2 were reduced in the Hyper group. Spironolactone decreased serum thyroid hormones and improved cardiac lipid peroxides and metalloproteinase -2 levels. The use of spironolactone decreased serum nitrite levels and increased cardiac SOD and glutathione. Cardiac levels of aldosterone, endothelin-1, transforming growth factor-beta and nitrite were similar among all groups. CONCLUSION: Hyperthyroid status was associated with an increase in aldosterone and oxidant/ inflammatory biomarkers. The use of spironolactone enhanced antioxidant defenses. Aldosterone antagonists may serve as potential drugs to attenuate the development of cardiac disease in hyperthyroidism.

Changes in thyroid hormone activity disrupt photomotor behavior of larval zebrafish.

High throughput in vitro, in silico, and computational approaches have identified numerous environmental chemicals that interfere with thyroid hormone (TH) activity, and it is posited that human exposures to such chemicals are a contributing factor to neurodevelopmental disorders. However, whether hits in screens of TH activity are predictive of developmental neurotoxicity (DNT) has yet to be systematically addressed. The zebrafish has been proposed as a second tier model for assessing the in vivo DNT potential of TH active chemicals. As an initial evaluation of the feasibility of this proposal, we determined whether an endpoint often used to assess DNT in larval zebrafish, specifically photomotor behavior, is altered by experimentally induced hyper- and hypothyroidism. Developmental hyperthyroidism was simulated by static waterborne exposure of zebrafish to varying concentrations (3-300 nM) of thyroxine (T4) or triiodothyronine (T3) beginning at 6 h post-fertilization (hpf) and continuing through 5 days post-fertilization (dpf). Teratogenic effects and lethality were observed at 4 and 5 dpf in fish exposed to T4 or T3 at concentrations >30 nM. However, as early as 3 dpf, T4 (> 3 nM) and T3 (> 10 nM) significantly increased swimming activity triggered by sudden changes from light to dark, particularly during the second dark period (Dark 2). Conversely, developmental hypothyroidism, which was induced by treatment with 6-propyl-2-thiouracil (PTU), morpholino knockdown of the TH transporter mct8, or ablation of thyroid follicles in adult females prior to spawning, generally decreased swimming activity during dark periods, although effects did vary across test days. All effects of developmental hypothyroidism on photomotor behavior occurred independent of teratogenic effects and were most robust during Dark 2. Treatment with the T4 analog, Tetrac, restored photomotor response in mct8 morphants to control levels. Collectively, these findings suggest that while the sensitivity of photomotor behavior in larval zebrafish to detect TH disruption is influenced by test parameters, this test can distinguish between TH promoting and TH blocking activity and may be useful for assessing the DNT potential of TH-active chemicals.

Thyroxine, levothyroxine, and thyroxine complexed into cyclodextrin changed animal behavior, oxygen consumption, and photopic electroretinogram of Colossoma macropomum.

The toxic effects of thyroxine (T4F), levothyroxine (L-T4), and thyroxine complexed into ß-cyclodextrin (ß-CD-T4) on the biological parameters of tambaqui (Colossoma macropomum) were evaluated. The animals were exposed to a chronic toxicity test based on concentrations of influent (60 ng/L) for 2 months. Weight, total length, animal behavior, oxygen consumption, photopic electroretinogram (ERG), and the Flicker exam were evaluated. No significant differences were observed (p > 0.05) on the weight and total length measurements between all groups studied. Behavioral observations of the animals exposed to L-T4 and ß-CD-T4 complex showed a reduction (p < 0.05) in slow swimming and an increase in staying motionless events. The animals exposed to the ß-CD-T4 complex presented the highest O2 consumption. L-T4 and ß-CD-T4 promoted a reduction in the ability of the animals to respond to stimuli in the photoreceptors according to the photopic ERG examination. Data from the experimental Flicker exam showed no significant differences (p > 0.05) in all groups studied. It can be concluded that the complexation of T4 into ß-CD and L-T4 modified the toxicity of this hormone, promoting changes in the behavior, oxygen consumption, and electrophysiological responses of the exposed animals, suggesting that inclusion complexes should be submitted to new toxicity tests to ensure higher safety.

Thyroxine promotes lung cancer growth in an orthotopic mouse model.

Thyroid hormones are important for physiology and homeostasis. In addition to nuclear thyroid hormone receptors, the plasma membrane protein integrin αvß3 has been recognized as a receptor for both thyroxine (T4) and triiodothyronine (T3). Here, we studied whether thyroid hormone promotes growth of murine lung cancer via αvß3 in vivo. Murine Lewis lung carcinoma cells (3LL), stably transfected with luciferase, were injected into mouse lungs. Tumor growth in untreated mice was compared to hypothyroid mice and hypothyroid mice treated with T3 or T4 with or without the αvß3 inhibitor 3,5,3',5'-tetraiodothyroacetic acid (Tetrac). Tumor progression was determined by serial in vivo imaging of bioluminescence emitted from the tumor. Tumor weight was recorded at the end of the experiment. Neoangiogenesis was determined by immunohistochemistry for CD31. Tumor growth was reduced in hypothyroidism and increased by T4 treatment. Strikingly, only T4 but not T3 treatment promoted tumor growth. This T4 effect was abrogated by the αvß3 inhibitor Tetrac. Tumor weight and neoangiogenesis were also significantly increased only in T4-treated mice. The T4 effect on tumor weight and neoangiogenesis was abolished by Tetrac. In vitro, T4 did not stimulate 3LL cell proliferation or signaling pathway activation. We conclude that T4 promotes lung cancer growth in this orthotopic mouse model. The tumor-promoting effect is mediated via the plasma membrane integrin αvß3 and increased neoangiogenesis rather than direct stimulation of 3LL cells. These data suggest that such effects of levothyroxine may need to be considered in cancer patients on T4 substitution.

Altered thyroid hormone levels affect body condition at metamorphosis in larvae of Xenopus laevis.

Chemical, physical and biological environmental stressors may affect the endocrine system, such as the thyroid hormone (TH) axis in larval amphibians with consequences for energy partitioning among development, growth and metabolism. We studied the effects of two TH level affecting compounds, exogenous l-thyroxine (T4 ) and sodium perchlorate (SP), on various measures of development and body condition in larvae of the African clawed frog (Xenopus laevis). We calculated the scaled mass index, hepatosomatic index and relative tail muscle mass as body condition indices to estimate fitness. Altered TH levels significantly altered the growth, development, survival and body condition in metamorphic larvae in different directions. While exogeno us T4 reduced growth and accelerated development, SP treatment increased growth but slowed down development. Altered TH levels improved body conditions in both treatments and particularly in larvae of the SP treatment but to the detriment of lower survival rates in both TH level altering treatments. The hepatosomatic index was negatively affected by exogenous T4 , but not by SP treatment indicating a lower lipid reserve in the liver in larvae of T4 treatment. These altered TH levels as caused by several environmental stressors may have an influence on individual fitness across life, as body condition at the onset of metamorphosis determines metamorphic and juvenile survival. Further research is needed to determine synergetic effects of environmental stressors on TH levels and its effects on physiological traits such as metabolic rate.

Transcriptomics investigation of thyroid hormone disruption in the olfactory system of the Rana [Lithobates] catesbeiana tadpole.

Thyroid hormones (THs) regulate vertebrate growth, development, and metabolism. Despite their importance, there is a need for effective detection of TH-disruption by endocrine disrupting chemicals (EDCs). The frog olfactory system substantially remodels during TH-dependent metamorphosis and the objective of the present study is to examine olfactory system gene expression for TH biomarkers that can evaluate the biological effects of complex mixtures such as municipal wastewater. We first examine classic TH-response gene transcripts using reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR) in the olfactory epithelium (OE) and olfactory bulb (OB) of premetamorphic Rana (Lithobates) catesbeiana tadpoles after 48 h exposure to biologically-relevant concentrations of the THs, 3,5,3'-triiodothyronine (T3) and L-thyroxine (T4), or 17-beta estradiol (E2); a hormone that can crosstalk with THs. As the OE was particularly sensitive to THs, further RNA-seq analysis found >30,000 TH-responsive contigs. In contrast, E2 affected 267 contigs of which only 57 overlapped with THs suggesting that E2 has limited effect on the OE at this developmental phase. Gene ontology enrichment analyses identified sensory perception and nucleoside diphosphate phosphorylation as the top affected terms for THs and E2, respectively. Using classic and additional RNA-seq-derived TH-response gene transcripts, we queried TH-disrupting activity in municipal wastewater effluent from two different treatment systems: anaerobic membrane bioreactor (AnMBR) and membrane enhanced biological phosphorous removal (MEBPR). While we observed physical EDC removal in both systems, some TH disruption activity was retained in the effluents. This work lays an important foundation for linking TH-dependent gene expression with olfactory system function in amphibians.

No obvious sympathetic excitation after massive levothyroxine overdose: A case report.

RATIONALE: Thyrotoxicosis from an overdose of medicinal thyroid hormone is a condition that may be associated with a significant delay in onset of toxicity. However, limited literature is available regarding thyrotoxicosis attributed to excessive ingestion of exogenous thyroid hormone and most cases described were pediatric clinical researches. Herein, we presented the course of a patient who ingested a massive amount of levothyroxine with no obvious sympathetic excited symptoms exhibited and reviewed feasible treatment options for such overdoses. PATIENT CONCERNS: A 41-year-old woman patient with ureteral calculus ingested a massive amount of levothyroxine (120 tablets, equal to 6 mg in total) during her hospitalization. Her transient vital signs were unremarkable after ingestion except for significantly accelerated breathing rate of 45 times per minute. Initial laboratory findings revealed evidently elevated serum levels of thyroxine (T4) >320 nmol/L, free triiodothyronine (fT3) 10.44 pmol/L, and free thyroxine (fT4) >100 pmol/L. The patient had a history of hypothyroidism, which was managed with thyroid hormone replacement (levothyroxine 100 µg per day). Besides, she also suffered from systemic lupus erythematosus and chronic pancreatitis. DIAGNOSES: This is a case of excessive ingestion of exogenous thyroid hormone in an adult. INTERVENTIONS: The interventions included use propranolol to prevent heart failure; utilize hemodialysis to remove redundant thyroid hormone from blood; closely monitor the vital signs, basal metabolic rate, blood biochemical indicators, and serum levels of thyroid hormone. OUTCOMES: The woman had no obvious symptoms of thyrotoxicosis. After 4 weeks, the results of thyroid function indicated that serum thyroid hormone levels were completely recovered to pre-ingestion levels. Accordingly, the levothyroxine was used again as before. LESSONS: Adults often exhibit more severe symptoms after intaking overdose levothyroxine due to their complex medical history and comorbidities than children. As for them, hemodialysis should be considered as soon as possible. Besides, diverse treatments according to specific symptoms and continuously monitoring were indispensable.

Maternal environment and craniofacial growth: geometric morphometric analysis of mandibular shape changes with in utero thyroxine overexposure in mice.

An estimated 3% of US pregnancies are affected by maternal thyroid dysfunction, with between one and three of every 1000 pregnancies being complicated by overactive maternal thyroid levels. Excess thyroid hormones are linked to neurological impairment and excessive craniofacial variation, affecting both endochondral and intramembranous bone. Using a geometric morphometric approach, this study evaluates the role of in utero thyroxine overexposure on the growth of offspring mandibles in a sample of 241 mice. Canonical variate analysis utilized 16 unilateral mandibular landmarks obtained from 3D micro-computed tomography to assess shape changes between unexposed controls (n = 63) and exposed mice (n = 178). By evaluating shape changes in the mandible among three age groups (15, 20 and 25 days postnatal) and different dosage levels (low, medium and high), this study found that excess maternal thyroxine alters offspring mandibular shape in both age- and dosage-dependent manners. Group differences in overall shape were significant (P < 0.001), and showed major changes in regions of the mandible associated with muscle attachment (coronoid process, gonial angle) and regions of growth largely governed by articulation with the cranial base (condyle) and occlusion (alveolus). These results compliment recent studies demonstrating that maternal thyroxine levels can alter the cranial base and cranial vault of offspring, contributing to a better understanding of both normal and abnormal mandibular development, as well as the medical implications of craniofacial growth and development.

Memantine, an NMDA Receptor Antagonist, Prevents Thyroxin-induced Hypertension, but Not Cardiac Remodeling.

Stimulation of glutamatergic tone has been causally linked to myocardial pathogenesis and amplified systemic blood pressure (BP). Memantine, a noncompetitive N-methyl-D-aspartate glutamatergic receptor (NMDA-R) antagonist, has been proposed to be an active cardioprotective drug. However, the efficacy of memantine and subsequently the possible involvement of the NMDA-R in the thyroxin (T4)-induced cardiovascular complications have never been investigated. We examined the effect of memantine (30 mg·kg·d) on the T4 (500 µg·kg·d)-provoked increase in mouse BP, cardiac hypertrophy indicated by enlarged overall myocardial mass, and reformed reactions of the contractile myocardium both in vivo and ex vivo after 2 weeks of treatment. Memantine alone did not result in any cardiovascular pathology in mice. Instead, memantine significantly prevented the T4-triggered systemic hypertension. But, it did not reverse cardiac hypertrophy, coupled in vivo left ventricular dysfunction (LV) or ex vivo right ventricular (RV) papillary muscle contractile alterations of the T4-treated mice. Our results openly direct the cardiovascular safety and tolerability of memantine therapy. Yet, extra research is necessary to endorse these prospective advantageous outcomes. Also, we believe that this is the first study to inspect the possible role of NMDA-R in the T4-stimulated cardiovascular disorders and concluded that NMDA-R could play a key role in the T4-induced hypertension.

Thyroxine Exposure Effects on the Cranial Base.

Thyroid hormone is important for skull bone growth, which primarily occurs at the cranial sutures and synchondroses. Thyroid hormones regulate metabolism and act in all stages of cartilage and bone development and maintenance by interacting with growth hormone and regulating insulin-like growth factor. Aberrant thyroid hormone levels and exposure during development are exogenous factors that may exacerbate susceptibility to craniofacial abnormalities potentially through changes in growth at the synchondroses of the cranial base. To elucidate the direct effect of in utero therapeutic thyroxine exposure on the synchondroses in developing mice, we provided scaled doses of the thyroid replacement drug, levothyroxine, in drinking water to pregnant C57BL6 wild-type dams. The skulls of resulting pups were subjected to micro-computed tomography analysis revealing less bone volume relative to tissue volume in the synchondroses of mouse pups exposed in utero to levothyroxine. Histological assessment of the cranial base area indicated more active synchondroses as measured by metabolic factors including Igf1. The cranial base of the pups exposed to high levels of levothyroxine also contained more collagen fiber matrix and an increase in markers of bone formation. Such changes due to exposure to exogenous thyroid hormone may drive overall morphological changes. Thus, excess thyroid hormone exposure to the fetus during pregnancy may lead to altered craniofacial growth and increased risk of anomalies in offspring.

Opposing Effects of Maternal Hypo- and Hyperthyroidism on the Stability of Thalamocortical Synapses in the Visual Cortex of Adult Offspring.

Insufficient or excessive thyroid hormone (TH) levels during fetal development can cause long-term neurological and cognitive problems. Studies in animal models of perinatal hypo- and hyperthyroidism suggest that these problems may be a consequence of the formation of maladaptive circuitry in the cerebral cortex, which can persist into adulthood. Here we used mouse models of maternal hypo- and hyperthyroidism to investigate the long-term effects of altering thyroxine (T4) levels during pregnancy (corresponding to embryonic days 6.5-18.5) on thalamocortical (TC) axon dynamics in adult offspring. Because perinatal hypothyroidism has been linked to visual processing deficits in humans, we performed chronic two-photon imaging of TC axons and boutons in primary visual cortex (V1). We found that a decrease or increase in maternal serum T4 levels was associated with atypical steady-state dynamics of TC axons and boutons in V1 of adult offspring. Hypothyroid offspring exhibited axonal branch and bouton dynamics indicative of an abnormal increase in TC connectivity, whereas changes in hyperthyroid offspring were indicative of an abnormal decrease in TC connectivity. Collectively, our data suggest that alterations to prenatal T4 levels can cause long-term synaptic instability in TC circuits, which could impair early stages of visual processing.

Determinação de fármacos em mananciais do estado de São Paulo e estudo da sua ecotoxicidade sobre a cianobactéria Microcystis aeruginosa / Pharmaceuticals determination in São Paulo stare springs and evaluation of their toxicity in cyanobacterium Microcystis aeruginosa

A contaminação de corpos d'água por fármacos é um tema de extrema relevância, tendo em vista problemas como a escassez de água, florações de cianobactérias tóxicas e lançamentos clandestinos de efluentes domésticos. Sendo assim, este trabalho teve como objetivo determinar a presença de cafeína (CAF), fluoxetina (FLX), levotiroxina (LVX) e bezafibrato (BZF) em mananciais do estado de São Paulo, bem como avaliar a toxicidade desses compostos à cianobactéria Microcystis aeruginosa LTPNA 08. Um método por LC-MS/MS foi desenvolvido e validado, de acordo com a RDC nº 166 da ANVISA, para a detecção de CAF, FLX, LVX e BZF em amostras ambientais. As represas Guarapiranga e Billings, bem como os rios Taiçupeba, Sorocaba, Baixo Cotia, Grande e Paraíba foram monitorados de abril a setembro de 2017. A toxicidade dos fármacos foi avaliada por meio do monitoramento do crescimento, produção de microcistinas e viabilidade celular da cianobactéria M. aeruginosa LTPNA 08. CAF foi detectada em todas as amostras analisadas, com concentrações que variaram de 6,6 ng.L-1 a 16,47 µg.L-1. No Rio Cotia foram verificadas as maiores concentrações de CAF, FLX e BZF (16,47 µg.L-1; 3,5 ng.L-1 e 322 ng.L-1, respectivamente). A LVX, cujos produtos de biotransformação não foram monitorados, não foi detectada em nenhuma amostra analisada. A concentração de 50 µg.L-1 de FLX inibiu o crescimento da cianobactéria em 82,3% (CE50: 31,4 µg.L-1). Em relação à produção de microcistinas totais, os fármacos inibiram a liberação da fração extracelular para a maior concentração testada ao longo do tempo de monitoramento, embora não tenham demonstrado efeito sobre a viabilidade celular. Sendo assim, considerando-se que fármacos estão presentes nos mananciais monitorados no estado de São Paulo e que a FLX pode causar efeito sobre a M. aeruginosa, os efeitos decorrentes da exposição a concentrações ambientais contínuas e cumulativas de fármacos em corpos d'água devem ser estudados. Além disso, uma vez que a ocorrência destas substâncias e outros contaminantes antropogênicos no ambiente aquático natural é uma questão emergente devido aos efeitos adversos potenciais que estes compostos representam para a vida aquática e os seres humanos, os tipos e níveis destes compostos, que têm um impacto maior na qualidade da água, deve ser constantemente monitorada. Práticas de gestão que investem em saneamento e na redução da descarga de efluentes não tratados, e um plano de proteção de recursos hídricos com o objetivo de garantir a segurança da água seriam medidas essenciais para reduzir o aporte de contaminantes nos corpos d'água do estado de São Paulo

Contamination of water bodies by drugs is a subject of extreme relevance considering related problems such as water scarcity, harmful cyanobacterial blooms and discharge of untreated domestic effluents. Therefore, the aim of this work was to determine the presence of caffeine (CAF), fluoxetine (FLX), levothyroxine (LVX) and bezafibrate (BZF) in springs in the State of São Paulo, and to evaluate the toxicity of these compounds in cyanobacteria Microcystis aeruginosa LTPNA 08. A LC-MS/MS method was developed and validated according to RDC nº 166 of ANVISA to assess the concentration of CAF, FLX, LVX and BZF in environmental samples. Guarapiranga and Billings reservoirs, as well as the Taiçupeba, Sorocaba, Baixo Cotia, Grande and Paraíba rivers were monitored from April to September 2017.The drugs toxicity in M. aeruginosa LTPNA 08 was assessed by monitoring their effects on cyanobacterial growth, microcystins production and cell viabilityby flow cytometry. CAF was detected in all analyzed samples at concentrations ranging from 6.6 ng to 16.47 µg.L-1.Among studied sites, Cotia river showed the highest concentrations of CAF, FLX and BZF (16.47 µg.L-1, 3.5 ng.L-1 and 322 ng.L-1, respectively). LVX, which biotransformation products were not monitored, was not detected in any of the analyzed samples. Regarding the drugs toxicity, 50 µg.L-1 of FLX inhibited the cyanobacterial grow thin 82.3% (EC50 of 31.4 µg.L-1). Although no effect on cell viability was seen by flow cytometry, the highest concentrations of all compounds tested were able to inhibit the release of microcystins. Therefore, considering that some of the drugs monitored showed to be present in water sources in São Paulo State and that FLX affects cyanobacteria M. aeruginosa growth, the effects of continuous and cumulative exposure at environmental drug concentrations of in water bodies should be evaluated. Also, since the occurrence of these substances and other anthropogenic contaminants in the natural aquatic environment is an emerging issue due to the potential adverse effects these compounds pose to aquatic life and humans, thet ypes and levels of these compounds, which have a greater impact on water quality, should be constantly monitored. Management practices investing in sanitation and in reducing discharge of untreated effluents, as well as a plan for water resources protection with the goal of ensuring water security would be essential measures in reducing drugs loading into water bodies situated in São Paulo State

Adult-onset hyperthyroidism impairs spatial learning: possible involvement of mitogen-activated protein kinase signaling pathways.

Given evidence that mitogen-activated protein kinase (MAPK) activation is part of the nongenomic actions of thyroid hormones, we investigated the possible consequences of hyperthyroidism for the cognitive functioning of adult rats. Young adult rats were treated with L-thyroxine or saline. Twenty rats in each group were exposed to Morris water maze testing, measuring their performance in a hidden-platform spatial task. In a separate set of rats not exposed to Morris water maze testing (untrained rats), the expression and phosphorylated levels of p38-MAPK and of its two downstream effectors, Elk-1 and cAMP response element-binding protein, were evaluated using quantitative reverse transcriptase-PCR and western blotting. Rats with hyperthyroidism showed delayed acquisition of learning compared with their wild-type counterparts, as shown by increased escape latencies and distance moved on the last two trials of daily training in the water maze. The hyperthyroid rats, however, showed no difference during probe trials. Western blot analyses of the hippocampus showed that hyperthyroidism increased phosphorylated p38-MAPK levels in untrained rats. Although our study is correlative in nature and does not exclude the contribution of other molecular targets, our findings suggest that the observed impairments in acquisition during actual learning in rats with hyperthyroidism may result from the increased phosphorylation of p38-MAPK.

Low-frequency stimulation induces a durable long-term depression in young adult hyperthyroid rats: the role of p38 mitogen-activated protein kinase and protein phosphatase 1.

Long-term potentiation and long-term depression (LTD) are cellular mechanisms of learning and memory in the mammalian brain. We have previously shown that adult hyperthyroid rats showed a delay in the acquisition of a place learning task and attenuated long-term potentiation. However, changes in LTD in hyperthyroidism remain unclear. Rats were administered 0.2 mg/kg/day of L-thyroxine for 21 days starting at postnatal day 40 to induce hyperthyroidism. LTD was induced in the dentate gyrus using low-frequency stimulation (LFS) of the perforant pathway. The mRNA expressions of p38 mitogen-activated protein kinase (p38-MAPK) and protein phosphatase 1 (PP1) were evaluated using a quantitative reverse transcriptase PCR. In control rats, a standard LFS protocol induced a slight depression of the population spike (PS) amplitude during the induction phase of LTD (76±13% vs. baseline), but a small potentiation of the PS amplitude was observed in the early (107±18%) and late (111±20%) phases of LTD. Interestingly, in the hyperthyroid rats, the same LFS protocol induced a reliable LTD in the dentate gyrus of the hippocampus as evidenced by a marked depression in the PS amplitude during the induction (54±6% vs. baseline) and the early phases (56±8%) of LTD. Elevated mRNA levels of p38-MAPK and PP1 were observed in the hippocampus of the LFS-treated hyperthyroid rats compared with the hippocampus of the vehicle-treated hyperthyroid rats. No significant change in p38-MAPK or PP1 mRNA expression was observed in the euthyroid rats. The present study shows that a standard LFS protocol can induce a durable depression of synaptic strength and an upregulation of PP1 and p38-MAPK mRNA in hyperthyroid rats. We conclude that hyperthyroidism can induce molecular changes associated with degeneration of the hippocampus. The relationship between the levels of thyroid hormone and dementia requires further investigation.

Natriuretic Peptides as Cardiovascular Safety Biomarkers in Rats: Comparison With Blood Pressure, Heart Rate, and Heart Weight.

Cardiovascular (CV) toxicity is an important cause of failure during drug development. Blood-based biomarkers can be used to detect CV toxicity during preclinical development and prioritize compounds at lower risk of causing such toxicities. Evidence of myocardial degeneration can be detected by measuring concentrations of biomarkers such as cardiac troponin I and creatine kinase in blood; however, detection of functional changes in the CV system, such as blood pressure, generally requires studies in animals with surgically implanted pressure transducers. This is a significant limitation because sustained changes in blood pressure are often accompanied by changes in heart rate and together can lead to cardiac hypertrophy and myocardial degeneration in animals, and major adverse cardiovascular events (MACE) in humans. Increased concentrations of NPs in blood correlate with higher risk of cardiac mortality, all-cause mortality, and MACE in humans. Their utility as biomarkers of CV function and toxicity in rodents was investigated by exploring the relationships between plasma concentrations of NTproANP and NTproBNP, blood pressure, heart rate, and heart weight in Sprague Dawley rats administered compounds that caused hypotension or hypertension, including nifedipine, fluprostenol, minoxidil, L-NAME, L-thyroxine, or sunitinib for 1-2 weeks. Changes in NTproANP and/or NTproBNP concentrations were inversely correlated with changes in blood pressure. NTproANP and NTproBNP concentrations were inconsistently correlated with relative heart weights. In addition, increased heart rate was associated with increased heart weights. These studies support the use of natriuretic peptides and heart rate to detect changes in blood pressure and cardiac hypertrophy in short-duration rat studies.