Different biochemical strategies of two Neotropical fish to cope with the impairment of nitrogen excretion during air exposure

The exposure of fish to air is normally expected to interfere with the nitrogen excretion process. Hoplias malabaricus and Hoplerythrinus unitaeniatus, two teleost species, display distinct behaviors in response to decreases in natural reservoir water levels, although they may employ similar biochemical strategies. To investigate this point, plasma levels of ammonia, urea, uric acid, and the two urea cycle enzymes, ornithine carbamoyl transferase (OCT) and arginase (ARG), as well as glutamine synthetase (GS) were determined for both species after exposure to air. Plasma ammonia increased gradually during exposure to air, but only H. malabaricus showed increased concentrations of urea. Plasma uric acid remained very low in both fish. Enzymatic activities (mean ± SD, µmol min-1 g protein-1) of H. malabaricus showed significant increases (P<0.05, N = 6) in OCT from 0.84 ± 0.05 to 1.42 ± 0.03, in ARG from 8.07 ± 0.47 to 9.97 ± 0.53 and in GS from 1.15 ± 0.03 to 2.39 ± 0.04. The OCT and ARG enzymes remained constant in H. unitaeniatus (N = 6), but GS increased from 1.49 ± 0.02 to 2.06 ± 0.03. Although these species are very closely related and share the same environment, their biochemical strategies in response to exposure to air or to increased plasma ammonia are different.

Eye color as an indicator of social rank in the fish Nile tilapia

We investigated the association of eye color with the dominant-subordinate relationship in the fish Nile tilapia, Oreochromis niloticus. Eye color pattern was also examined in relation to the intensity of attacks. We paired 20 size-matched fish (intruder: 73.69 ± 11.49 g; resident: 75.42 ± 8.83 g) and evaluated eye color and fights. These fish were isolated in individual aquaria for 10 days and then their eye color was measured 5 min before pairing (basal values). Twenty minutes after pairing, eye color and fights were quantified for 10 min. Clear establishment of social hierarchy was observed in 7 of 10 pairs of fish. Number of attacks ranged from 1 to 168 among pairs. The quartile was calculated for these data and the pairs were then divided into two classes: low-attack (1 to 111 attacks - 2 lower quartiles) or high-attack (112 to 168 attacks - 2 higher quartiles). Dominance decreased the eye-darkening patterns of the fish after pairing, while subordinance increased darkening compared to dominance. Subordinate fish in low-attack confrontations presented a darker eye compared to dominant fish and to the basal condition. We also observed a paler eye pattern in dominants that shared low-attack interactions after pairing compared to the subordinates and within the group. However, we found no differences in the darkening pattern between dominants and subordinates from the high-attack groups. We conclude that eye color is associated with social rank in this species. Moreover, the association between eye color and social rank in the low-attack pairs may function to reduce aggression.

Effect of chronic fish oil supplementation on renal function of normal and cachectic rats

In the present study we determined the effect of chronic diet supplementation with n-3 PUFA on renal function of healthy and cachectic subjects by providing fish oil (1 g/kg body weight) to female rats throughout pregnancy and lactation and then to their offspring post-weaning and examined its effect on renal function parameters during their adulthood. The animals were divided into four groups of 5-10 rats in each group: control, control supplemented with fish oil (P), cachectic Walker 256 tumor-bearing (W), and W supplemented with fish oil (WP). Food intake was significantly lower in the W group compared to control (12.66 ± 4.24 vs 25.30 ± 1.07 g/day). Treatment with fish oil significantly reversed this reduction (22.70 ± 2.94 g/day). Tumor growth rate was markedly reduced in the P group (16.41 ± 2.09 for WP vs 24.06 ± 2.64 g for W). WP group showed a significant increase in mean glomerular filtration rate compared to P and control (1.520 ± 0.214 ml min-1 kg body weight-1; P < 0.05). Tumor-bearing groups had low urine osmolality compared to control rats. The fractional sodium excretion decreased in the W group compared to control (0.43 ± 0.16 vs 2.99 ± 0.87%; P < 0.05), and partially recovered in the WP group (0.90 ± 0.20%). In summary, the chronic supplementation with fish oil used in this study increased the amount of fat in the diet by only 0.1%, but caused remarkable changes in tumor growth rate and cachexia, also showing a renoprotective function.

Routine post-weaning handling of rats prevents isolation rearing-induced deficit in prepulse inhibition

Rats reared under isolation conditions from weaning present a number of behavioral changes compared to animals reared under social conditions (group housing). These changes include deficits in prepulse inhibition (PPI) of the startle reflex to a loud sound. PPI refers to the reduction of the magnitude of the startle reflex when a relatively weak stimulus (the prepulse) precedes by an appropriate time interval the intense startle-elicing stimulus (the pulse). PPI is useful for studying sensorimotor integration. The present study evaluated the effect of handling on the impairment of PPI induced by isolation-rearing. Male Wistar rats (N = 11-15/group) were housed in groups (5 per cage and handled three times a week) or isolated (housed individually) since weaning (21 days) for 10 weeks when they reach approximately 150 g. The isolated rats were divided into "minimally handled" animals (handled once a week for cleaning purposes only) or "handled" animals (handled three times a week). This handling consisted of grasping the rat by the tail and moving it to a clean cage (approximately 5 s). A statistically significant reduction (52%) in the PPI test was found only in the isolated group with minimal handling while no difference was seen between grouped animals and isolated handled animals. These results indicate that isolation rearing causes disruption in the PPI at adult age, which serves as an index of attention deficit. This change in the sensory processing of information induced by post-weaning isolation can be prevented by handling during the development of the animal.

Losses of immunoreactive parvalbumin amacrine and immunoreactive alphaprotein kinase C bipolar cells caused by methylmercury chloride intoxication in the retina of the tropical fish Hoplias malabaricus

To quantify the effects of methylmercury (MeHg) on amacrine and on ON-bipolar cells in the retina, experiments were performed in MeHg-exposed groups of adult trahiras (Hoplias malabaricus) at two dose levels (2 and 6 µg/g, ip). The retinas of test and control groups were processed by mouse anti-parvalbumin and rabbit anti-alphaprotein kinase C (alphaPKC) immunocytochemistry. Morphology and soma location in the inner nuclear layer were used to identify immunoreactive parvalbumin (PV-IR) and alphaPKC (alphaPKC-IR) in wholemount preparations. Cell density, topography and isodensity maps were estimated using confocal images. PV-IR was detected in amacrine cells in the inner nuclear layer and in displaced amacrine cells from the ganglion cell layer, and alphaPKC-IR was detected in ON-bipolar cells. The MeHg-treated group (6 µg/g) showed significant reduction of the ON-bipolar alphaPKC-IR cell density (mean density = 1306 ± 393 cells/mm²) compared to control (1886 ± 892 cells/mm²; P < 0.001). The mean densities found for amacrine PV-IR cells in MeHg-treated retinas were 1040 ± 56 cells/mm² (2 µg/g) and 845 ± 82 cells/mm² (6 µg/g), also lower than control (1312 ± 31 cells/mm²; P < 0.05), differently from the data observed in displaced PV-IR amacrine cells. These results show that MeHg changed the PV-IR amacrine cell density in a dose-dependent way, and reduced the density of alphaKC-IR bipolar cells at the dose of 6 µg/g. Further studies are needed to identify the physiological impact of these findings on visual function.

Agonistic-like responses from the torus semicircularis dorsalis elicited by GABA A blockade in the weakly electric fish Gymnotus carapo

Findings by our group have shown that the dorsolateral telencephalon of Gymnotus carapo sends efferents to the mesencephalic torus semicircularis dorsalis (TSd) and that presumably this connection is involved in the changes in electric organ discharge (EOD) and in skeletomotor responses observed following microinjections of GABA A antagonist bicuculline into this telencephalic region. Other studies have implicated the TSd or its mammalian homologue, the inferior colliculus, in defensive responses. In the present study, we explore the possible involvement of the TSd and of the GABA-ergic system in the modulation of the electric and skeletomotor displays. For this purpose, different doses of bicuculline (0.98, 0.49, 0.245, and 0.015 mM) and muscimol (15.35 mM) were microinjected (0.1 µL) in the TSd of the awake G. carapo. Microinjection of bicuculline induced dose-dependent interruptions of EOD and increased skeletomotor activity resembling defense displays. The effects of the two highest doses showed maximum values at 5 min (4.3 ± 2.7 and 3.8 ± 2.0 Hz, P < 0.05) and persisted until 10 min (11 ± 5.7 and 8.7 ± 5.2 Hz, P < 0.05). Microinjections of muscimol were ineffective. During the interruptions of EOD, the novelty response (increased frequency in response to sensory novelties) induced by an electric stimulus delivered by a pair of electrodes placed in the water of the experimental cuvette was reduced or abolished. These data suggest that the GABA-ergic mechanisms of the TSd inhibit the neural substrate of the defense reaction at this midbrain level.

Effects of mercury intoxication on the response of horizontal cells of the retina of thraira fish (Hoplias malabaricus)

Methyl mercury (MeHg) is highly neurotoxic, affecting visual function in addition to other central nervous system functions. The effect of mercury intoxication on the amplitude of horizontal cell responses to light was studied in the retina of the fish Hoplias malabaricus. Intracellular responses were recorded from horizontal cells of fish previously intoxicated with MeHg by intraperitoneal injection (IP group) or by trophic exposure (T group). Only one retina per fish was used. The doses of MeHg chloride administered to the IP group were 0.01, 0.05, 0.1, 1.0, 2.0, and 6.0 mg/kg. The amplitudes of the horizontal cell responses were lower than control in individuals exposed to 0.01 (N = 4 retinas), 0.05 (N = 2 retinas) and 0.1 mg/kg (N = 1 retina), whereas no responses were recorded in the 1.0, 2.0, and 6.0 mg/kg groups. T group individuals were fed young specimens of Astyanax sp previously injected with MeHg corresponding to 0.75 (N = 1 retina), 0.075 (N = 8 retinas) or 0.0075 (N = 4 retinas) mg/kg fish body weight. After 14 doses, one every 5 days, the amplitude of the horizontal cell response was higher than control in individuals exposed to 0.075 and 0.0075 mg/kg, and lower in individuals exposed to 0.75 mg/kg. We conclude that intoxication with MeHg affects the electrophysiological response of the horizontal cells in the retina, either reducing or increasing its amplitude compared to control, and that these effects are related to the dose and/or to the mode of administration.

Incidence, risk factors and prognostic factors of acute renal failure in patients admitted to an intensive care unit

The objective of the present study was to assess the incidence, risk factors and outcome of patients who develop acute renal failure (ARF) in intensive care units. In this prospective observational study, 221 patients with a 48-h minimum stay, 18-year-old minimum age and absence of overt acute or chronic renal failure were included. Exclusion criteria were organ donors and renal transplantation patients. ARF was defined as a creatinine level above 1.5 mg/dL. Statistics were performed using Pearsons' chi2 test, Student t-test, and Wilcoxon test. Multivariate analysis was run using all variables with P < 0.1 in the univariate analysis. ARF developed in 19.0% of the patients, with 76.19% resulting in death. Main risk factors (univariate analysis) were: higher intra-operative hydration and bleeding, higher death risk by APACHE II score, logist organ dysfunction system on the first day, mechanical ventilation, shock due to systemic inflammatory response syndrome (SIRS)/sepsis, noradrenaline use, and plasma creatinine and urea levels on admission. Heart rate on admission (OR = 1.023 (1.002-1.044)), male gender (OR = 4.275 (1.340-13642)), shock due to SIRS/sepsis (OR = 8.590 (2.710-27.229)), higher intra-operative hydration (OR = 1.002 (1.000-1004)), and plasma urea on admission (OR = 1.012 (0.980-1044)) remained significant (multivariate analysis). The mortality risk factors (univariate analysis) were shock due to SIRS/sepsis, mechanical ventilation, blood stream infection, potassium and bicarbonate levels. Only potassium levels remained significant (P = 0.037). In conclusion, ARF has a high incidence, morbidity and mortality when it occurs in intensive care unit. There is a very close association with hemodynamic status and multiple organ dysfunction.

Stress responses of the fish Nile tilapia subjected to electroshock and social stressors

Plasma cortisol and glucose levels were measured in 36 adult Nile tilapia males, Oreochromis niloticus (standard length, mean ± SD, 14.38 ± 1.31 cm), subjected to electroshock and social stressors. Pre-stressor levels were determined 5 days after the adjustment of the fish to the experimental aquaria (1 fish/aquarium). Five days later, the effects of stressors on both cortisol and glucose levels were assessed. The following stressors were imposed for 60 min: pairing with a larger resident animal (social stressor), or a gentle electroshock (AC, 20 V, 15 mA, 100 Hz for 1 min every 4 min). Each stressor was tested in two independent groups, one in which stress was quantified immediately after the end of the 60-min stressor imposition (T60) and the other in which stress was quantified 30 min later (T90). Pre-stressor values for cortisol and glucose were not statistically different between groups. Plasma cortisol levels increased significantly and were of similar magnitude for both electroshock and the social stressor (mean ± SD for basal and final samples were: electroshock T60 = 65.47 ± 15.3, 177.0 ± 30.3; T90 = 54.8 ± 16.0, 196.2 ± 57.8; social stress T60 = 47.1 ± 9.0, 187.6 ± 61.7; T90 = 41.6 ± 8.1, 112.3 ± 26.8, respectively). Plasma glucose levels increased significantly for electroshock at both time points (T60 and T90), but only at T90 for the social stressor. Initial and final mean (± SD) values are: electroshock T60 = 52.5 ± 9.2, 115.0 ± 15.7; T90 = 35.5 ± 1.1, 146.3 ± 13.3; social stress T60 = 54.8 ± 8.8, 84.4 ± 15.0; T90 = 34.5 ± 5.6, 116.3 ± 13.6, respectively. Therefore, electroshock induced an increase in glucose more rapidly than did the social stressor. Furthermore, a significant positive correlation between cortisol and glucose was detected only at T90 for the social stressor. These results indicate that a fish species responds differently to different stressors, thus suggesting specificity of fish stress response to a stressor.

Fish hemoglobins

Vertebrate hemoglobin, contained in erythrocytes, is a globular protein with a quaternary structure composed of 4 globin chains (2 alpha and 2 beta) and a prosthetic group named heme bound to each one. Having myoglobin as an ancestor, hemoglobin acquired the capacity to respond to chemical stimuli that modulate its function according to tissue requirements for oxygen. Fish are generally submitted to spatial and temporal O2 variations and have developed anatomical, physiological and biochemical strategies to adapt to the changing environmental gas availability. Structurally, most fish hemoglobins are tetrameric; however, those from some species such as lamprey and hagfish dissociate, being monomeric when oxygenated and oligomeric when deoxygenated. Fish blood frequently possesses several hemoglobins; the primary origin of this finding lies in the polymorphism that occurs in the globin loci, an aspect that may occasionally confer advantages to its carriers or even be a harmless evolutionary remnant. On the other hand, the functional properties exhibit different behaviors, ranging from a total absence of responses to allosteric regulation to drastic ones, such as the Root effect.

Comparison of I-FISH and G-banding for the detection of chromosomal abnormalities during the evolution of myelodysplastic syndrome

Myelodysplastic syndrome (MDS) patients with a normal karyotype constitute a heterogeneous group from a biological standpoint and their outcome is often unpredictable. Interphase fluorescence in situ hybridization (I-FISH) studies could increase the rate of detection of abnormalities, but previous reports in the literature have been contradictory. We performed I-FISH and conventional karyotyping (G-banding) on 50 MDS patients at diagnosis, after 6 and 12 months or at any time if a transformation to acute myeloid leukemia (AML) was detected. Applying a probe-panel targeting the centromere of chromosomes 7 and 8, 5q31, 5p15.2 and 7q31, we observed one case with 5q deletion not identified by G-banding. I-FISH at 6 and 12 months confirmed the karyotype results. Eight cases transformed to AML during follow-up, but no hidden clone was detected by I-FISH in any of them. The inclusion of I-FISH during follow-up of MDS resulted in a small improvement in abnormality detection when compared with conventional G-banding.

Fish oil ingestion reduces the number of aberrant crypt foci and adenoma in 1,2-dimethylhydrazine-induced colon cancer in rats

We determined the effect of fish oil (FO) ingestion on colonic carcinogenesis in rats. Male Wistar rats received 4 subcutaneous injections (40 mg/kg body weight each) of 1,2-dimethylhydrazine (DMH) at 3-day intervals and were fed a diet containing 18% by weight FO (N = 10) or soybean oil (SO, N = 10) for 36 weeks. At sacrifice, the colon was removed, aberrant crypt foci were counted and the fatty acid profile was determined. Intestinal tumors were removed and classified as adenoma or carcinoma. Liver and feces were collected and analyzed for fatty acid profile. FO reduced the mean (± SEM) number of aberrant crypt foci compared to SO (113.55 ± 6.97 vs 214.60 ± 18.61; P < 0.05) and the incidence of adenoma (FO: 20% vs SO: 100%), but carcinoma occurred equally in FO and SO rats (2 animals per group). The polyunsaturated fatty acid (PUFA) profile of the colon was affected by diet (P < 0.05): total ω-3 (FO: 8.18 ± 0.97 vs SO: 1.71 ± 0.54%) and total ω-6 (FO: 3.83 ± 0.59 vs SO: 10.43 ± 1.28%). The same occurred in the liver (P < 0.05): total ω-3 (FO: 34.41 ± 2.6 vs SO: 6.46 ± 0.59%) and total ω-6 (FO: 8.73 ± 1.37 vs SO: 42.12 ± 2.33%). The PUFA profile of the feces and liver polyamine levels did not differ between groups (P > 0.05). In conclusion, our findings indicate that chronic FO ingestion protected against the DMH-induced preneoplastic colon lesions and adenoma development, but not against carcinoma in rats.

Control of respiration in fish, amphibians and reptiles

Fish and amphibians utilise a suction/force pump to ventilate gills or lungs, with the respiratory muscles innervated by cranial nerves, while reptiles have a thoracic, aspiratory pump innervated by spinal nerves. However, fish can recruit a hypobranchial pump for active jaw occlusion during hypoxia, using feeding muscles innervated by anterior spinal nerves. This same pump is used to ventilate the air-breathing organ in air-breathing fishes. Some reptiles retain a buccal force pump for use during hypoxia or exercise. All vertebrates have respiratory rhythm generators (RRG) located in the brainstem. In cyclostomes and possibly jawed fishes, this may comprise elements of the trigeminal nucleus, though in the latter group RRG neurons have been located in the reticular formation. In air-breathing fishes and amphibians, there may be separate RRG for gill and lung ventilation. There is some evidence for multiple RRG in reptiles. Both amphibians and reptiles show episodic breathing patterns that may be centrally generated, though they do respond to changes in oxygen supply. Fish and larval amphibians have chemoreceptors sensitive to oxygen partial pressure located on the gills. Hypoxia induces increased ventilation and a reflex bradycardia and may trigger aquatic surface respiration or air-breathing, though these latter activities also respond to behavioural cues. Adult amphibians and reptiles have peripheral chemoreceptors located on the carotid arteries and central chemoreceptors sensitive to blood carbon dioxide levels. Lung perfusion may be regulated by cardiac shunting and lung ventilation stimulates lung stretch receptors.

Autonomic control of cardiorespiratory interactions in fish, amphibians and reptiles

Control of the heart rate and cardiorespiratory interactions (CRI) is predominantly parasympathetic in all jawed vertebrates, with the sympathetic nervous system having some influence in tetrapods. Respiratory sinus arrhythmia (RSA) has been described as a solely mammalian phenomenon but respiration-related beat-to-beat control of the heart has been described in fish and reptiles. Though they are both important, the relative roles of feed-forward central control and peripheral reflexes in generating CRI vary between groups of fishes and probably between other vertebrates. CRI may relate to two locations for the vagal preganglionic neurons (VPN) and in particular cardiac VPN in the brainstem. This has been described in representatives from all vertebrate groups, though the proportion in each location is variable. Air-breathing fishes, amphibians and reptiles breathe discontinuously and the onset of a bout of breathing is characteristically accompanied by an immediate increase in heart rate plus, in the latter two groups, a left-right shunting of blood through the pulmonary circuit. Both the increase in heart rate and opening of a sphincter on the pulmonary artery are due to withdrawal of vagal tone. An increase in heart rate following a meal in snakes is related to withdrawal of vagal tone plus a non-adrenergic-non-cholinergic effect that may be due to humoral factors released by the gut. Histamine is one candidate for this role.

Vancomycin-resistant enterococcus outbreak in a pediatric intensive care unit: report of successful interventions for control and prevention

The objective of this study is to retrospectively report the results of interventions for controlling a vancomycin-resistant enterococcus (VRE) outbreak in a tertiary-care pediatric intensive care unit (PICU) of a University Hospital. After identification of the outbreak, interventions were made at the following levels: patient care, microbiological surveillance, and medical and nursing staff training. Data were collected from computer-based databases and from the electronic prescription system. Vancomycin use progressively increased after March 2008, peaking in August 2009. Five cases of VRE infection were identified, with 3 deaths. After the interventions, we noted a significant reduction in vancomycin prescription and use (75% reduction), and the last case of VRE infection was identified 4 months later. The survivors remained colonized until hospital discharge. After interventions there was a transient increase in PICU length-of-stay and mortality. Since then, the use of vancomycin has remained relatively constant and strict, no other cases of VRE infection or colonization have been identified and length-of-stay and mortality returned to baseline. In conclusion, we showed that a bundle intervention aiming at a strict control of vancomycin use and full compliance with the Hospital Infection Control Practices Advisory Committee guidelines, along with contact precautions and hand-hygiene promotion, can be effective in reducing vancomycin use and the emergence and spread of vancomycin-resistant bacteria in a tertiary-care PICU.