A three-compartment apparatus alters the brain concentration of cytokines and neurotrophic factors in cocaine-induced CPP in mice.

Cocaine-induced neuroinflammation plays an important role in the pathophysiology of drug addiction. Evidence suggests that the immune response contributes for memory consolidation related to place preference behavior underlying cocaine administration in mice. Conditioned place preference (CPP) is a protocol extensively used to study the rewarding and/or aversive motivational effects of drug abuse in rodents, reproducing cocaine-seeking behavior in humans. Besides the variety of apparatus used in the CPP protocol, whether different types of apparatus are able to induce the same conditioned behavior response and neurobiological changes remains to be fully explored. We hypothesize that the immune response is involved in the cocaine-induced CPP and that the type of apparatus might influence this response. Herein, two- and three-compartment apparatuses were tested using the behavioral model of CPP. Cocaine-induced CPP was demonstrated in both apparatuses. However, mice injected with cocaine had decreased levels of IL-1ß, IL-6, IL-10, and GDNF in the pre-frontal cortex, and decreased CX3CL1 in the striatum, in the CPP protocol using three compartments compared to controls. While similar levels were seen in the CPP protocol using two compartments. In conclusion, the current study demonstrated that the type of apparatus might influence the investigation of neurobiological mechanisms associated with cocaine-induced CPP. Our data also suggest that the three compartment-apparatus seems to be a more appropriate model to investigate the neuroinflammatory response related to cocaine addiction.

Capybara Oil Improves Hepatic Mitochondrial Dysfunction, Steatosis, and Inflammation in a Murine Model of Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD) is recognized as the most common cause of liver dysfunction worldwide and is commonly associated with obesity. Evidences suggest that NAFLD might be a mitochondrial disease, which contributes to the hepatic steatosis, oxidative stress, cytokine release, and cell death. Capybara oil (CO) is a rich source of polyunsaturated fatty acids (PUFA), which is known to improve inflammation and oxidative stress. In order to determine the effects of CO on NAFLD, C57Bl/6 mice were divided into 3 groups and fed a high-fat diet (HFD) (NAFLD group and NAFLD + CO group) or a control diet (CG group) during 16 weeks. The CO (1.5 g/kg/daily) was administered by gavage during the last 4 weeks of the diet protocol. We evaluated plasma liver enzymes, hepatic steatosis, and cytokine expression in liver as well as hepatocyte ultrastructural morphology and mitochondrial function. CO treatment suppressed hepatic steatosis, attenuated inflammatory response, and decreased plasma alanine aminotransferase (ALT) in mice with NAFLD. CO was also capable of restoring mitochondrial ultrastructure and function as well as balance superoxide dismutase and catalase levels. Our findings indicate that CO treatment has positive effects on NAFLD improving mitochondrial dysfunction, steatosis, acute inflammation, and oxidative stress.

Neurotrophic Factors in Parkinson's Disease: What Have we Learned from Pre-Clinical and Clinical Studies?

BACKGROUND: Parkinson´s Disease (PD) is a chronic, progressive condition, being the second most common neurodegenerative disorder worldwide. The classical features include: bradykinesia, resting tremor, rigidity and festination. These neurological alterations are probably due to the death of dopaminergic neurons in the Substantia Nigra pars compacta and consequent reduction of dopamine input into the striatum. The decrease of dopamine levels may also be involved in the emergence of non-motor symptoms, including cognitive impairment, anxiety and depression symptoms. Neurotrophic Factors (NF) are proteins that modulate neuronal function, development, and survival. It has been reported that NF might exert a protective role in PD. OBJECTIVE: We aim to discuss the emerging evidence from pre-clinical and clinical studies regarding the role of NF in PD as well as their potential as promising therapeutic strategies. METHODS: We carried out an extensive literature search in PubMed central. RESULTS: Pre-clinical studies using NF to treat PD are divergent probably due to several methodological differences, thus precluding any conclusion. Clinical studies findings obtained with the administration of NF in patients with PD were even more disappointed. On the other hand, pre-clinical and clinical studies generally support that physical activity is a low-cost, non-pharmacologic strategy with good results to treat PD. CONCLUSION: The use of NF as a treatment for PD is still a promise not incorporated in clinical practice. Methods to deliver NFs, doses and compounds administered, side effects, population characteristics and duration of disease may probably contribute to the unsuccessful results.

Avaliação do quadro clínico e perfil bioquímico de bovinos durante indução e tratamento de hipocalcemia / Assessment of clinical and biochemical profile of cattle during induction and treatment of hypocalcemia

O presente trabalho objetivou estudar o quadro sintomatológico, algumas variáveis bioquímicas e a resposta ao tratamento com cálcio de bovinos com hipocalcemia induzida experimentalmente. Foram utilizadas 12 novilhas distribuídas nos grupos controle (n = 5) e tratado (n = 7). Foi infundida solução de EDTA a 5% até o animal apresentar sinais clínicos de hipocalcemia, quando então era iniciado o tratamento com solução contendo cálcio, fósforo, magnésio e glicose, na dose de 1 mL/kg/PV, em 30 minutos, enquanto que o grupo controle recebia apenas solução fisiológica na mesma dose. Exame clínico e coleta de amostras sanguíneas foram realizados nos tempos T0 (basal), T1 (Fase I, caracterizada por tremores musculares), T2 (ao final da infusão com EDTA), T3 (ao final do tratamento) e T4 (24 horas após o término do experimento). Todas as novilhas mostraram diminuição temporária da concentração de cálcio total e livre, fósforo, e apresentaram quadro clássico de hipocalcemia. A taquicardia, a hipofonese e a atonia ruminal desapareceram no decorrer do tratamento, sendo observado aumento no cálcio livre e total e fósforo. O medicamento usado no tratamento dos animais foi eficaz na recuperação do quadro clínico de hipocalcemia dentro de 30 minutos, promovendo retorno das principais variáveis do perfil bioquímico aos valores basais

The present work aims to study the clinical picture, biochemical profile and treatment response in cattle with induced hypocalcaemia. Were utilized 12 heifers randomly distributed in treated (n = 7) and control (n = 5) groups. The induction model was carried on by continuous EDTA infusion into jugular vein until the animals present clinical signs of hypocalcaemia. After that, the treated group received a calcium (Ca) solution enriched with phosphorus, magnesium and glucose with a dose of 1 mL/kg/BW in 30 minutes, meanwhile, the control group was treated with the same dose of physiologic solution. Clinical examination were performed and blood samples were obtained in times T0 (basal time), T1 (beginning of hypocalcaemia); T2 (end of EDTA infusion); T3 (end of treatment) and T4 (24 hours after the induction). All the heifers present temporary blood calcium and phosphorus reduction and demonstrated classical clinical picture of hypocalcaemia. The treated group present full clinical recovery and blood calcium and phosphorus increase. Most evident clinical signs were increasing heart beat, hypophonesis and rumenal atony. Those symptoms were reversed after calcium treatment. The solution used for treatment was efficient on clinical recovery within thirty minutes, promoting the return to basal levels of the most of biochemical's variables

Hepatogenous photosensitization associated with liver copper accumulation in buffalos.

Four 1.5-year-old, male, Murrah buffalos were maintained during eight months without direct solar exposure during a study of copper toxicosis. Four days after solar exposure, all buffalos presented clinical manifestations consistent with acute photosensitization, including anorexia, apathy, loss of body weight, and generalized cutaneous lesions. Gross lesions were characterized by severe erythema, localized edema, fissures, tissue necrosis, gangrene and crust formation with serous exudation. Liver copper concentration was evaluated, and cutaneous biopsies were taken when clinical signs were evident. The liver copper concentration before solar exposure was increased in all animals. Histopathologic examination of the skin revealed hepatogenous photosensitization characterized by parakeratotic hyperkeratosis, acantholysis, degeneration of squamous epithelial cells, epidermal necrosis with atrophy of sweat glands, and multifocal superficial and deep dermal edema. These findings suggest that asymptomatic accumulation of copper within the liver might have induced hepatic insufficiency thereby resulting in secondary photosensitization when these buffalos were exposed to sunlight.

Clinical observations of cattle and buffalos with experimentally induced chronic copper poisoning.

The susceptibility of cattle and buffalos to chronic copper poisoning (CCP) was compared by using cattle (n=10) and buffalo (n=10) steers distributed into two copper supplemented (n=6) and two control (n=4) groups. Supplemented animals received 2 mg copper (Cu)/kg body weight daily for one week, with an additional 2 mg weekly until the end of the experiment (day 105). Three liver biopsies (day 0, 45, and 105) were obtained for mineral analyses; clinical examinations and blood samples were obtained every 15days. Three supplemented cattle and two buffalos with typical manifestations of CCP died. There were no differences in the frequency of mortality between cattle and buffalos; hepatic copper concentration was higher in cattle than buffalos. These findings suggest that buffalos and cattle might be equally susceptible to CCP. However, buffalos accumulate less liver copper than cattle and have a lower threshold of hepatic Cu accumulation, which leads to clinical manifestation of CCP.

Purification and properties of a coagulant thrombin-like enzyme from the venom of Bothrops leucurus.

A thrombin-like enzyme from Bothrops leucurus venom, named leucurobin (leuc), was purified by gel filtration, affinity and ion exchange chromatographies. Physicochemical studies indicated that the purified enzyme is a 35 kDa monomeric glycoprotein on SDS-PAGE under reducing conditions, which decreased to 29 kDa after deglycosylation with N-glycosidase F (PNGase F). The amino acid sequence of leuc was determined by automated sequencing of the intact native protein and peptides produced by digestion of the S-pyridyl-ethylated protein with trypsin. The protein sequence exhibits significant similarities with other serine proteases reported from snake venoms, and contains two potential sites of N-linked glycosylation. The proteinase split off fibrinopeptide A (FPA) rapidly from human fibrinogen; however, only negligible traces of fibrinopeptide B (FPB) were observed. In addition, the enzyme released the N-terminal peptide (Mr=4572) containing the first 42 residues from the Bbeta-chain. Leuc could neither activate factor XIII nor release kinins from heat-treated bovine plasma. Its specific clotting activity was equivalent to 198 NIH thrombin U/mg on human fibrinogen. Kinetic properties of leuc were determined using representative chromogenic substrates. The enzyme evoked the gyroxin syndrome when injected into the tail veins of mice at levels of 0.143 microg/g mouse. The inhibitory effects of PMSF and benzamidine on the amidolytic activity suggest that leuc is a serine proteinase, and inhibition by beta-mercaptoethanol revealed the important role of the disulfide bonds in the stabilization of the native structure. Antibothropic serum, SBTI and EDTA had little or no effect on its amidolytic activity. However, the clotting effect of the enzyme was strongly inhibited by antibothropic serum. A Dixon plot showed that the hydrolysis of Bz-L-Arg-pNA by leuc was competitively inhibited by benzamidine (Ki=1.61+/-0.25 mM).

Coagulant thrombin-like enzymes from the venoms of Brazilian and Peruvian bushmaster (Lachesis muta muta) snakes.

Two isoforms of a thrombin-like enzyme designated TLE-B and TLE-P were purified from the venoms of Lachesis muta muta (bushmaster) snakes captured in two different geographical localities, Manaus (Brazil) and Pucallpa (Perú). TLE-B and TLE-P showed Mr values of 44000 and 43000 under reducing conditions on SDS-PAGE, which decreased to 27000 after deglycosylation with N-glycosidase F (PNGase F). The purified proteinases split off fibrinopeptide A rapidly from human fibrinogen and fibrinopeptide B more slowly. In addition, both enzymes released the N-terminal peptide (Mr=4572) containing the first 42 residues from the Bbeta-chain. Their specific clotting activities were equivalent to 1000 and 900 NIH thrombin units/mg on human fibrinogen and 526 and 606 NIH thrombin units/mg on bovine fibrinogen for TLE-B and TLE-P, respectively. Kinetic properties of these enzymes were determined using representative chromogenic substrates. Tryptic peptide mapping of the two native enzymes suggested a large degree of structural similarity. Purified rabbit IgG against TLE-B reacted with both enzymes forming a continuous precipitin line on immunodiffusion. Furthermore, Western blot and indirect ELISA were used to compare the antigenic cross-reactivity for both enzymes as well as the venoms of L. muta muta and Bothrops snakes. Incubation of human alpha2-macroglobulin (alpha2-M) with each enzyme at molar ratios of 1:1, 1:2 and 1:4 enzyme:inhibitor resulted in retarding their clotting activities by approximately 12 times, whereas their amidolytic activities were not affected. However, the Mr 180000 subunits of alpha2-M were not cleaved by these enzymes, suggesting that alpha2-M inhibits TLEs by steric hindrance. Similarly, inhibitions of their clotting activities were obtained using high concentrations of rabbit IgG (40 microg, corresponding to molar ratio enzyme:inhibitor of 1:2) against TLE-B.