The effect of infrared and X-ray radiation on the production of reactive oxygen species in blood and induction of cytogenetic damage in the bone marrow of mice in vivo

The goal of the present work was to study the effect of infrared light (IRL) at a wavelength of 850 nm modulated by a frequency of 101 Hz with a mode of power 22 mW/cm2 and X-rays with a voltage of 200 kV at a dose rate of 1 Gy/min on the production of reactive oxygen species (ROS) in blood cells using luminol-dependent chemiluminescence, as well as on the induction of a cytogenetic damage in bone marrow cells of mice by the in vivo micronucleus test. The experiments performed have shown: 1) the level of the ROS production in blood of the mice exposed to IRL and X-rays at an adapting dose of 0.1 Gy reaches the peak value after 0.5 h and drops to the ROS level in untreated animals 5 h after either exposure; 2) irradiation of mice with IRL and X-rays at a dose of 0.1 Gy induces adaptive responses both in blood cells and bone marrow cells of mice. These adaptive responses were revealed only 5 h after both exposures, when the level of ROS production decreased to the ROS level in untreated animals; they are equal in magnitude and dynamics and persist up to 2 months.     

Effect of nitric oxide synthase inhibitor on diaphragmatic function after resistive loading

We studied the effect of a nitric oxide synthase inhibitor, Nomega-Nitro-L-arginine-methyl-ester (L-NAME), on in vitro diphragmatic function both at rest (control) or after inspiratory resistive loading (IRL). Sprague-Dawley rats were anesthetized, instrumented, and then the following experimental groups: (1) controls; (2) L-NAME (100 mg/kg/body weight intravenously alone); (3) IRL alone; and (4) L-NAME + IRL. The IRL protocol consisted of applying a variable resistor to the inspiratory limb of a two-way valve at 70% of maximal airway pressure until apnea. After the experiment, the animals were sacrificed and diaphragmatic strips were obtained for activity of constitutive nitric oxide synthase (cNOS) and measurements of in vitro contractile properties: tetanic (Po) and twitch tensions (Pt). cNOS activity was significantly decreased in the L-NAME and L-NAME + IRL groups (P < or = 0.05) as compared with control and IRL groups. L-NAME alone did not affect Po or Pt. However, in both IRL groups, with and without was a significant decrease in Po and Pt. This reduction was comparable in both groups. In summary, our data showed that L-NAME resulted in a significant decrease cNOS activity, but in vitro contractility was impaired.     

Effect of inspiratory resistive loaded breathing and hypoxemia on diaphragmatic function in the piglet.

The combined effects of inspiratory resistive loaded breathing (IRL) and hypoxemia on transdiaphragmatic pressure (Pdi) in nine 1-mo-old Yorkshire piglets were studied. IRL was adjusted to increase spontaneously generated Pdi five to six times above baseline but maintain arterial PCO2 < 70 Torr to prevent hypercapnic depression of diaphragmatic contractility. Measurements of ventilation, blood gases and pH, Pdi, diaphragmatic electromyogram, Pdi during phrenic nerve stimulation, diaphragmatic blood flow, and end-expiratory lung volume were obtained at baseline, after 2 h of IRL, and then after 1 h of hypoxemia (arterial PO2 approximately 40 Torr) combined with IRL. Diaphragmatic muscle samples were obtained after study completion and immediately frozen in liquid nitrogen for determination of tissue ATP, phosphocreatine, lactate, and glycogen levels. Ten 1-mo-old piglets were subjected to IRL alone and served as controls. IRL alone resulted in significant impairment of Pdi generation. The addition of hypoxemia for 1 h did not further compromise Pdi in comparison to control animals who were subjected to IRL alone. Blood flow to both the costal and crural segments of the diaphragm increased significantly during IRL; the addition of the hypoxemic stress resulted in further significant augmentation of blood flow to both segments of the diaphragm. No differences were noted in diaphragmatic muscle tissue ATP, phosphocreatine, or glycogen between control and IRL animals or between control and IRL plus hypoxemia animals. Muscle lactate levels increased significantly in the IRL plus hypoxemia animals only. The data from this study suggest that moderate hypoxemia during resistive-loaded breathing in the piglet does not accentuate diaphragmatic fatigue.     

Diaphragmatic force and substrate response to resistive loaded breathing in the piglet

Inspiratory resistive loaded (IRL) breathing results in hypoventilation and diaphragmatic fatigue in the piglet. We studied the effects of 6 h of IRL on ten 1-mo-old piglets. The load was adjusted to increase spontaneously generated transdiaphragmatic pressure five to six times baseline. Six 1-mo-old piglets acted as controls and were identically instrumented but were not subjected to IRL. Measurements of ventilation, blood gases and pH, diaphragmatic electromyogram, force-frequency curve, blood flow, and end-expiratory lung volume were obtained hourly. Diaphragmatic muscle samples were obtained after 6 h for determination of ATP, phosphocreatine, lactate, and glycogen levels. No changes occurred in the control animals. IRL resulted in a significant decrease in ventilation, an increase in diaphragmatic EMG, onset of abdominal expiratory muscle activity, and a fall in end-expiratory lung volume by 1 h. The force-frequency curve adjusted for lung volume change fell by 20% at all frequencies of stimulation at 1 h and by 40% at 6 h. Blood flow to the costal and crural diaphragm increased by 51 and 141%, respectively. No differences were noted in ATP, phosphocreatine, lactate, or glycogen between control and IRL animals. It is concluded that submaximal spontaneous contractions of the piglet diaphragm over a 6-h period cause a substantial decrease in its maximal force-generating capacity that is not related to substrate depletion.     

The basis of asymmetry in IS2 transposition

In the first step of IS2 transposition, the formation of an IS2 minicircle, the roles of the two IS ends differ. Terminal cleavage initiates exclusively at the right inverted repeat (IRR) - the donor end - whereas IRL is always the target. At the resulting minicircle junction, the two abutted ends are separated by a spacer of 1 or 2 basepairs. In this study, we have identified the determinants of donor and target function. The inability of IRL to act as a donor results largely from two sequence differences between IRL and IRR - an extra basepair between the conserved transposase binding sequences and the end of the element, and a change of the terminal dinucleotide from CA-3' to TA-3'. These two changes also impose a characteristic size on the minicircle junction spacer. The only sequences required for the efficient target function of IRL appear to be contained within the segment from position 11-42. Although IRR can function as a target, its shorter length and additional contacts with transposase (positions 1-7) result in minicircles with longer, and inappropriate, spacers. We propose a model for the synaptic complex in which the terminus of IRL makes different contacts with the transposase for the initial and final strand transfer steps. The sequence differences between IRR and IRL, and the behavioural characteristics of IRL that result from them, have probably been selected because they optimize expression of transposase from the minicircle junction promoter, Pjunc.     

Linear skin markings in common bottlenose dolphins (Tursiops truncatus) from the Indian River Lagoon,Florida

A previously undescribed skin abnormality, referred to as “linear skin markings” (LSM), has been identified in free-ranging common bottlenose dolphins (Tursiops truncatus) in the Indian River Lagoon, Florida (IRL). The lesions were identified during photo-identification surveys conducted from 2002 and 2015. LSM presented as distinct, parallel lines running dorso-ventrally on the torso and varied in length and width. The goals of this study were to determine (1) prevalence of the condition in IRL dolphins, (2) age and sex distribution of affected animals, (3) spatial and temporal distribution patterns, (4) duration of the condition, and (5) development of hypotheses regarding the etiology of the condition. Among 1,357 individual dolphins identified during the study period, 96 (7.0%) showed evidence of LSM. Nearly all (98.8%) cases with an established home range occurred in the northern and central regions of the IRL. The majority of cases of known sex were female (85%), of which 100% had given birth to one or more calves. The mean age of animals with LSM when first observed was 7.3 with a range of 1–20 years. The maximum observed duration of LSM was 15 years. Once observed, the condition persisted indefinitely. The etiology of LSM has not been established.     

Nitric oxide production in the ventilatory muscles in response to acute resistive loading

The effect of muscle activation on muscle nitric oxide (NO) production remains controversial. Whereas NO release increases in in vitro activated muscles and in vivo limb muscles, diaphragmatic NO synthase (NOS) activity declines after 3 h of inspiratory resistive loading (IRL). We tested in this study the hypotheses that acute IRL decreases diaphragmatic NO derivatives levels and reduces protein expression of neuronal (nNOS), endothelial (eNOS), and inducible (iNOS) NO synthases, as well as 3-nitrotyrosine formation. Anesthetized, tracheostomized, spontaneously breathing adult rats were subjected to IRL (50% of the maximum inspiratory pressure) for 1, 3, or 6 h. Quietly breathing rats served as controls. After 3 h of IRL, muscle eNOS and nNOS protein levels rose by 80 and 60% of control values, respectively. Whereas eNOS expression did not change any further, nNOS expression reached 550% of control values after 6 h of IRL. Strong iNOS protein expression was detected in the diaphragms after 6 h of IRL. Total NO derivatives levels in the diaphragm declined during IRL as a result of reduction in nitrate, nitrite, and nitrosothiols. Diaphragmatic protein tyrosine nitration decreased in response to IRL, and this reduction was mainly due to reduced tyrosine nitration of enolase and aldolase. We conclude that diaphragmatic NO derivatives levels decline in response to IRL and that the rise in diaphragmatic NOS protein expression may be a compensatory response designed to counterbalance the decline in NOS activity.     

Structural organization of the termini of the L and S components of the genome of pseudorabies virus.

The sequences of several hundred nucleotides around the junctions between the L and S components in concatemeric DNA and in mature virion DNA were ascertained. The two ends of the mature genome (which are joined in concatemeric DNA) show no sequence homology. Several directly repeated elements are present near both ends of the genome. Furthermore, the last 82 nucleotides at the left end of the L component (and of the genome) are repeated in inverted form (inverted repeat within the L component [IRL]) approximately 350 to 600 nucleotides downstream (depending on the virus isolate) bracketing the UL2 component. A comparison between the sequences at the right and left ends of the L component of the genome showed patchy homology, probably representing a vestigial inverted repeat bracketing the L component (IRL). Furthermore, less than 5% of the genomes have an L component that is in the orientation opposite to that of most of the viral genomes, indicating that the vestigial IRL that brackets the UL sequence may be sufficient to mediate inversion of the L component in some of the genomes. On the other hand, the UL2 component, which is bracketed by a perfect IRL, does not invert to a greater extent than does the L component (if it inverts at all). Analysis of the nucleotide sequence at the concatemeric junction of three different pseudorabies virus isolates showed almost complete sequence conservation. The sequence and organization of the repeated elements in the different isolates were almost identical, despite their different histories and origins. The high degree of conservation of these repeated elements implies that they may fulfill an essential function in the life cycle of the virus.     

Environmental drivers of habitat use by common bottlenose dolphins (Tursiops truncatus) in the Indian River Lagoon,Florida, USA

Anthropogenic impacts in estuarine systems can influence marine mammal habitat use, population dynamics, fitness, and mortality events. The objective was to examine habitat use among the resident common bottlenose dolphin (Tursiops truncatus) population inhabiting the Indian River Lagoon, Florida, and the influences of variation in environmental factors and prey availability in 2003–2015. We utilized photo-identification surveys, stratified random samples of prey, and environmental variables collected monthly. Kernel density estimation was used to determine the magnitude-per-unit area of dolphins across the IRL by wet and dry seasons each year, the values were used as a response variable in classification and regression tree analyses with fish community and environmental variables as predictors. Spatial patterns in dolphin density in the IRL were associated with salinity and dissolved oxygen levels, which are in part associated with freshwater discharges of nutrient and algae laden waters from the region's storm water management system. These findings isolate locations of concern for management of dolphin habitat, and anthropogenic drivers of dolphin distributions requiring further research.     

Stimulation of endothelin B receptor modulates the inflammatory activation of rat astrocytes

Inside the brain tissue, endothelins play numerous important biological roles. One of the targets, astrocytes, predominantly display endothelin receptor subtype B (ET(B)). On cultured primary rat astroglial cells, we analyzed the effect of IRL1620, a selective ET(B) receptor agonist, on the production of nitric oxide (NO) and the synthesis of interleukin (IL)-6 and tumor necrosis factor (TNF)-alpha. We performed these experiments in the presence or absence of interferon-gamma (IFN-gamma) and/or lipopolysaccharide (LPS). IRL1620 decreases NO production under basal conditions and after IFN-gamma stimulation. However, during LPS-induced NO production, IRL1620 enhances this release. The basal IL-6 secretion and especially the LPS-induced synthesis are enhanced by the IRL1620 stimulation. The LPS-dependent TNF-alpha production is increased by the ET(B) stimulation. The IRL1620-induced decrease of basal NO production is not dependent on Ca2+ entry or on phospholipase C (PLC) activation, as shown by the use of LaCl3 and U73122, respectively. In the presence of LPS, the IRL1620 potentiation of NO production is inhibited by LaCl3 and U73122. The IRL1620-induced increase of IL-6 is dependent on PLC activation. These results suggest that endothelins can have dual effects depending on the costimulatory factors present. Endothelins thus have important immunomodulatory functions in the brain.     

Effect of naloxone on spectral shifts of the diaphragm EMG during inspiratory loading

Shifts in the power spectrum of the diaphragm EMG to lower frequencies may occur in the presence of fatiguing inspiratory flow-resistive loads (IRL). However, such a shift of the centroid frequency (fc) could follow a reduction in central output through a differential reduction in end-inspiratory high-frequency power (HFP). In unanesthetized goats, we tested the hypothesis that activation of the endogenous opioid system by IRL would differentially reduce central respiratory output, causing a reduction in fc. IRL was imposed for 180 min after which naloxone (0.1 mg/kg, NLX) was given. fc was computed from the power spectral density estimated by the Welch method. IRL reduced fc from 148.0 +/- 9.8 (SE) Hz at base line to 141.1 +/- 8.9 Hz or to 95.5 +/- 1.3% of base line by 180 min (both P less than 0.05). NLX increased fc to 148.9 +/- 9.9 Hz or to 100.6 +/- 1.1% of base line (both P less than 0.05). The decline in fc during IRL was found to be the result of a reduction in HFP, predominantly toward the end of inspiration. The reversibility of this fc shift with NLX suggests a central mechanism consequent to elaboration of endogenous opioids and not a peripheral (muscular) event consequent to muscle fatigue.     

Differences in endothelin receptor types in the vasculature of Bothrops jararaca (Viperidae) and Oxyrhopus guibei (Colubridae) snakes

Endothelins (ETs) are vasoactive peptides evolutionary well conserved that exert their effects through two specific receptors (ET(A) and ET(B)) widely distributed in all vertebrates. In snakes, the presence and function of endothelins and their receptors are still scarcely described. We have recently demonstrated the presence of ET(A) and ET(B2) receptors in the snake Bothrops jararaca (Bj). In the present work we showed that distinctively from Bj, the vascular contraction induced by endothelin in Oxyrhopus guibei (Og) snake is mediated only by ET(A) receptors. Selective ET(B) agonists (SRTX-c and IRL(1620)) and antagonists (IRL(1038) and BQ(788)) were ineffective in Og preparations of isolated aorta. We also showed that ET-1 response on Og arterial blood pressure was monophasic hypertensive as opposed to biphasic (hypotension followed by hypertension) in Bj. Furthermore, we characterized the relaxing properties of endothelin receptor ET(B1) in pre-contracted aorta preparations. We showed that IRL(1620) induced relaxation of pre-contracted Bj aorta but was ineffective in relaxing Og preparations. IRL(1620) relaxing effect on Bj aorta was abolished by l-NAME, indicating involvement of NO release, and was reduced by selective ET(B) antagonists. Our findings suggest that Og snake has a more primitive spectrum of ET receptors (only ET(A) receptor) than Bj (presence of ET(A), ET(B1) and ET(B2) receptors).     

A reversible radioligand specific for the ETB receptor: [125I]Tyr13-Suc-[Glu9,Ala11,15]-endothelin-1(8- 21), [125I]IRL 1620.

Suc-[Glu9,Ala11,15]-endothelin(ET)-1(8-21), IRL 1620, is a linear ET-analog specific for the ET-isopeptide-nonselective ETB receptor. The radio-iodinated analog, [125I]IRL 1620, showed a single class of saturable binding to the ETB receptors in porcine lung membranes with a Kd of 18 pM and a Bmax of 930 fmol/mg protein, which are almost comparable to the values obtained with [125I]ET-3 (6 pM and 900 fmol/mg protein). In competitive binding assays with [125I]IRL 1620, unlabeled ET-1, ET-3, IRL 1620 and [monoiodo-Tyr13]-IRL 1620 showed almost identical displacement curves with Ki of 8 to 16 pM. However, [125I]IRL 1620 was dissociated from the binding sites by addition of an excess amount (100 nM) of any of these unlabeled peptides, each with the same t1/2 of 100 min. This was in marked contrast to [125I]ET-3 which was hardly dissociated from the binding sites.     

Nutrient Addition Differentially Affects Ecological Processes of <Emphasis Type="Italic">Avicennia germinans</Emphasis> in Nitrogen versus Phosphorus Limited Mangrove Ecosystems

Nutrient over-enrichment is a major threat to marine environments, but system-specific attributes of coastal ecosystems may result in differences in their sensitivity and susceptibility to eutrophication. We used fertilization experiments in nitrogen (N)- and phosphorus (P)-limited mangrove forests to test the hypothesis that alleviating different kinds of nutrient limitation may have different effects on ecosystem structure and function in natural systems. We compared a broad range of ecological processes to determine if these systems have different thresholds where shifts might occur in nutrient limitation. Growth responses indicated N limitation in Avicennia germinans (black mangrove) forests in the Indian River Lagoon (IRL), Florida, and P limitation at Twin Cays, Belize. When nutrient deficiency was relieved, A. germinans grew out of its stunted form by increasing wood relative to leaf biomass and shoot length relative to lateral growth. At the P-limited site, P enrichment (+P) increased specific leaf area, N resorption, and P uptake, but had no effect on P resorption. At the N-limited site, +N increased both N and P resorption, but did not alter biomass allocation. Herbivory was greater at the P-limited site and was unaffected by +P, whereas +N led to increased herbivory at the N-limited site. The responses to nutrient enrichment depended on the ecological process and limiting nutrient and suggested that N- versus P-limited mangroves do have different thresholds. +P had a greater effect on more ecological processes at Twin Cays than did +N at the IRL, which indicated that the P-limited site was more sensitive to nutrient loading. Because of this sensitivity, eutrophication is more likely to cause a shift in nutrient limitation at P-limited Twin Cays than N-limited IRL.     

Respiratory load compensation in chronic airway obstruction

To assess respiratory neuromuscular function and load compensating ability in patients with chronic airway obstruction (CAO), we studied eight stable patients with irreversible airway obstruction during hyperoxic CO2 rebreathing with and without a 17 cmH2O X l-1 X s flow-resistive inspiratory load (IRL). Minute ventilation (VE), transdiaphragmatic pressure (Pdi), and diaphragmatic electromyogram (EMGdi) were monitored. Pdi and EMGdi were obtained via a single gastroesophageal catheter with EMGdi being quantitated as the average rate of rise of inspiratory (moving average) activity. Based on the effects of IRL on the Pdi response to CO2 [delta Pdi/delta arterial CO2 tension (PaCO2)] and the change in Pdi for a given change in EMGdi (delta Pdi/delta EMGdi) during rebreathing, two groups could be clearly identified. Four patients (group A) were able to increase delta Pdi/delta PaCO2 and delta Pdi/delta EMGdi, whereas in the other four (group B) the IRL responses decreased. All group B patients were hyperinflated having significantly greater functional residual capacity (FRC) and residual volume than group A. In addition the IRL induced percent change in delta Pdi/delta PaCO2, and delta VE/delta PaCO2 was negatively correlated with lung volume so that in the hyperinflated group B the higher the FRC the greater was the decrease in Pdi response due to IRL. In both groups the greater the FRC the greater was the decrease in the ventilatory response to loading. Patients with CAO, even with severe airways obstruction, can effect load compensation by increasing diaphragmatic force output, but the presence of increased lung volume with the associated shortened diaphragm prevents such load compensation.     

Investigation of growth phases for bottlenose dolphins using a Bayesian modeling approach

The Gompertz function is the most commonly used growth function for cetacean studies. However, this function cannot represent multiple phases of growth. In this study, we present a Bayesian framework fitting parameters of a triple-logistic growth function to describe multiple phases of growth for bottlenose dolphins ( Tursiops truncatus ), simultaneously fitting and comparing all growth parameters between South Carolina (SC), Mississippi Sound (MSS), and Indian River Lagoon (IRL) cohorts. The fitted functions indicated a preliminary early, rapid growth phase, followed by a second phase of slower growth, and then a moderate growth spurt later in life. Growth parameters between geographic cohorts did not show obvious differences, although asymptotic length for SC dolphins was lower than MSS and IRL dolphins and significantly lower between females from SC and the IRL. Growth rate velocities between the sexes showed females exceed males initially (<1 yr), followed by males gaining an advantage around the ages of 3–4 yr until the age of around 15 yr when growth rates for both sexes approached zero (asymptotic length). This study demonstrates age-related changes in growth rates between bottlenose dolphin sexes and evidence of at least some differences ( i.e. , asymptotic length) across geographic cohorts.     

Effects of hyperoxia and allergic airway inflammation on cough reflex intensity in guinea pigs

Toxic influence of high oxygen concentration on pulmonary function and structures has been known for many years. However, the influence of high oxygen concentration breathing on defensive respiratory reflexes is still not clear. In our previous experiments, we found an inhibitory effect of 100 % oxygen breathing on cough reflex intensity in healthy guinea pigs. The present study was designed to detect the effects of hyperoxia on cough reflex in guinea pigs with allergic airway inflammation. In the first phase of our experiment, the animals were sensitized with ovalbumin. Thirty-two sensitized animals were used in two separate experiments according to oxygen concentration breathing: 100 % or 50 % oxygen for 60 h continuously. In each experiment, one group of animals was exposed to hyperoxia, another to ambient air. The cough reflex was induced both by aerosol of citric acid before sensitization, then in sensitized animals at 24 h and 60 h of exposition to oxygen/air in awake animals, and by mechanical stimulation of airway mucosa in anesthetized animals just after the end of the experiment. In contrast to 50 % oxygen, 100 % oxygen breathing leads to significant decrease in chemically induced cough in guinea pigs with allergic inflammation. No significant changes were present in cough induced by mechanical stimulation of airways.     

Vagal afferents, diaphragm fatigue, and inspiratory resistance in anesthetized dogs

This study tests three hypotheses regarding mechanisms that produce rapid shallow breathing during a severe inspiratory resistive load (IRL): 1) an intact vagal afferent pathway is necessary; 2) diaphragm fatigue contributes to tachypnea; and 3) hypoxia may alter the pattern of respiration. We imposed a severe IRL on pentobarbital sodium-anesthetized dogs, followed by bilateral vagotomy, then by supplemental O2. IRL alone produced rapid shallow breathing associated with hypercapnia and hypoxia. After the vagotomy, the breathing pattern became slow and deep, restoring arterial PCO2 but not arterial PO2 toward the control values. Relief of hypoxia had no effect, and at no time was there any evidence of fatigue of the diaphragm as measured by the response to phrenic nerve stimulation. We conclude that an intact afferent vagal pathway is necessary for the tachypnea resulting from a severe IRL, neither hypoxia nor diaphragm fatigue played a role, and, although we cannot rule out stimulation of vagal afferents, the simplest explanation for the increased frequency in our experiments is increased respiratory drive due to hypercapnia.