Early-life stress influences ion balance in developing zebrafish (Danio rerio).

As a key endocrine axis involved in responding to stress, the hypothalamic-pituitary-interrenal axis plays dual roles in mobilizing energy and maintaining ionic/osmotic balance in fishes. Although these roles have been examined independently in detail in adult fishes, less attention has been paid to the effects of an endogenous stress response during early life, particularly with respect to its potential effects on ionic/osmotic balance. The present study tested the hypothesis that exposure of zebrafish to stress during early development would alter ion balance later in life. Zebrafish at three developmental stages (4, 7, or 15 days post-fertilization, dpf) were subjected to an air-exposure stressor twice a day for 2 days, causing elevation of whole-body cortisol levels. Individuals stressed early in life exhibited decreased survival and growth, altered cortisol responses to a subsequent air-exposure stressor, and increased whole-body Na+ and Ca2+ concentrations. Changes in whole-body Ca2+ concentrations were accompanied by increased ionocyte abundance at 7 dpf and increased rates of Ca2+ uptake from the environment. Differences in whole-body ion concentrations at 15 and 35 dpf were not accompanied by altered ion uptake rates. Across all ages examined, air-exposure stress experienced at 7 dpf was particularly effective at eliciting phenotypic changes, suggesting a critical window at this age for a stress response to influence development. These findings demonstrate that early-life stress in zebrafish triggers developmental plasticity, with age-dependent effects on both the cortisol stress axis and ion balance.

The effect of seasonality on normal haematological and innate immune parameters of rainbow trout Oncorhynchus mykiss L.

It is well established that seasonality dominates the life history of fish by controlling the timing of physiological events such as reproduction, food intake, locomotor activity and growth performance. Seasonal differences in immune competence and prevalence of disease have been well documented in humans. The aim of this study was to determine if season influences the immune response of rainbow trout (Oncorhynchus mykiss L.). Thus, a 12-month trial was carried out in which plasma lysozyme activity and respiratory burst of head kidney macrophages (two innate immune parameters) and total red and white blood cell counts (two haematological parameters) were monitored at monthly intervals. Since photoperiodic information is thought to be conveyed via melatonin secretion, plasma melatonin levels were also measured at four seasonal points (day and night). A general seasonal influence was observed in the parameters measured in these fish, with the exception of respiratory burst activity of head kidney macrophages, with the parameters highest in summer and lowest in winter for total white blood cell counts and lysozyme activity.

s-IgA response in females following a single bout of moderate intensity exercise in cold and thermoneutral environments.

Previous research has linked exercise under unfavourable environmental conditions to decreased concentration and/or secretion rate of secretory immunoglobulin A (s-IgA). The purpose of this study was to investigate the influence of moderate exercise in a thermoneutral (TN) and a cold (COLD) environment on the concentration of s-IgA and the secretion rate of s-IgA in moderately active females. Sixteen females (23.4 +/- 6.8 yrs, 61.1 +/- 5.9 kg, 1.64 +/- 0.07 m) served as subjects for this study. On separate occasions, one week apart, all subjects ran or walked for 30 minutes at 71 % of individual heart rate reserve on an outdoor (1 degrees C) or an indoor track (24 degrees C). Unstimulated saliva samples were collected for 4-min immediately pre (PR), post (POST), and 30-min post (30POST) exercise. Profile of Mood State (POMS) questionnaires were completed pre and post exercise intervention and Global Mood State (GMS) was calculated. The absolute concentration of s-IgA, and total non-specific protein were measured and the secretion rate of s-IgA and s-IgA:Protein were calculated. Analysis of Variance (ANOVA) revealed that flow rate increased in COLD and decreased in TN such that it was significantly lower (POST) and (30POST) in the TN environment when compared to COLD. There was a significant increase in the secretion rate of s-IgA at 30POST for the COLD trial and a significant decrease at 30POST for the TN trial. No significant differences were found for the concentration of s-IgA, the s-IgA:Protein:ratio, or GMS. Stepwise regression revealed that with all predictors in the model, only the concentration of s-IgA accounted for a significant proportion of the variance in cold temperature, while in the TN environment, both the concentration of s-IgA and the flow rate accounted for significant proportions of the variance. These findings suggest that moderate exercise in cold temperatures does not decrease secretion rate of s-IgA, while exercise in TN temperatures does.

Mucosal IgA response to repeated wingate tests in females.

The purpose of this study was to examine the effect of repeated bouts of brief, intermittent, maximal effort exercise on the concentration of secretory IgA (S-IgA) and incidence of upper respiratory tract infection (URTI) in females. Twenty-six non-smoking, active, adult females participated in this study. The exercise intervention consisted of three consecutive, all-out 30-second Wingate leg cycling tests (0.075 kg/kg body mass), which were separated by three minutes of recovery. Unstimulated saliva was collected prior to (PRE) and five minutes after completion (POST) of the exercise testing protocol. IgA, protein and osmolality were measured, and secretory IgA (SIgA), IgA:Protein, and IgA: Osmolality were calculated. In addition, subjects completed standard logs indicating signs and symptoms of upper respiratory tract infection (URTI) for three weeks following the test. Saliva flow rates (478.5 +/- 50; 345.4 +/- 50 microl/min), S-IgA (55.8 +/- 4.7; 35.4 +/- 3.6 microg/min), and IgA: Protein ratio (30.7 +/- 3.0; 17.5 +/- 1.8 microg IgA/mg protein) were significantly lower POST compared to PRE (P < 0.05). No significant differences were observed for raw IgA, protein, osmolality, and IgA: osmolality. The results of this study show that brief, intermittent, maximal effort exercise results in an acute decrease in S-IgA in healthy adult females in less than eight minutes. However, this exercise induced transient decrease did not appear to be associated with an increase in clinical symptoms of URTI in the weeks following the exercise test intervention.

Weight-bearing exercise and markers of bone turnover in female athletes.

Weight-bearing activity provides an osteogenic stimulus, while effects of swimming on bone are unclear. We evaluated bone mineral density (BMD) and markers of bone turnover in female athletes (n = 41, age 20.7 yr) comparing three impact groups, high impact (High, basketball and volleyball, n = 14), medium impact (Med, soccer and track, n = 13), and nonimpact (Non, swimming, n = 7), with sedentary age-matched controls (Con, n = 7). BMD was assessed by dual-energy X-ray absorptiometry at the lumbar spine, femoral neck (FN), Ward's triangle, and trochanter (TR); bone resorption estimated from urinary cross-linked N-telopeptides (NTx); and bone formation determined from serum osteocalcin. Adjusted BMD (g/cm; covariates: body mass index, weight, and calcium and calorie intake) was greater at the FN and TR in the High group (1.27 +/- 0.03 and 1.05 +/- 0.03) than in the Non (1.05 +/- 0.04 and 0.86 +/- 0.04) and Con (1.03 +/- 0.05 and 0.85 +/- 0.05) groups and greater at the TR in the Med group (1.01 +/- 0.03) than in the Non (0.86 +/- 0.04) and Con (0.85 +/- 0.05) groups. Total body BMD was higher in the High group (4.9 +/- 0.12) than in the Med (4.5 +/- 0.12), Non (4.2 +/- 0.14), and Con (4.1 +/- 0.17) groups and greater in the Med group than in the Non and Con groups. Bone formation was lower in the Non group (19.8 +/- 2.6) than in the High (30.6 +/- 3.0) and Med (32.9 +/- 1.9, P < or = 0.05) groups. No differences in a marker of bone resorption (NTx) were noted. This indicates that women who participate in impact sports such as volleyball and basketball had higher BMDs and bone formation values than female swimmers.

Subterminal viral DNA nucleotides as specific recognition signals for human immunodeficiency virus type 1 and visna virus integrases under magnesium-dependent conditions.

Many reports describe the characteristics of susceptible viral DNA substrates to various retroviral integrases during in vitro reactions in which manganese serves as the divalent cation cofactor for site-specific nicking. However, manganese is known to alter the specificity of some endonucleases and magnesium may be the divalent cation used during retroviral integration in vivo. To address these concerns, we identified conditions under which the integrases of human immunodeficiency virus type 1 and visna virus were optimally active with magnesium (the first time such activity was shown for visna virus integrase) and used these conditions to test the susceptibility of a series of oligodeoxynucleotide substrates. The data show that two base pairs immediately internal to the conserved CA dinucleotide near the termini of retroviral DNA are selectively recognized by the two integrases and that the final six base pairs of viral DNA contain sufficient sequence information for specific recognition and cleavage by each enzyme. The results validate the importance of the subterminal viral DNA positions even in the presence of magnesium and identify viral DNA positions that functionally interact with integrase. The data obtained under magnesium-dependent conditions, which were obtained with substrates containing single and multiple base-pair substitutions and two different retroviral integrases, are consistent with those previously obtained with manganese. Thus, the large body of manganese-dependent data identifying terminal viral DNA positions that are important in substrate recognition by various integrases likely reflects interactions that are biologically relevant.

Effects of acetaminophen and ibuprofen on renal function in the stressed kidney.

Exercise, salt restriction, and/or dehydration causes transient reductions in renal function that may be buffered by vasodilatory prostaglandins (PGs). Over-the-counter (OTC) analgesics have the potential to alter renal hemodynamics by inhibiting renal PGs. Therefore, we tested the renal effects of the maximal recommended dose of acetaminophen (Acet, 4 g/day) and ibuprofen (Ibu, 1.2 g/day) vs. a placebo (Pl) in humans subjected to progressive renal stresses. After baseline measurements, 12 fit young (25 +/- 1 yr) men and women underwent 3 days of a low (10 meq/day)-sodium diet while taking one of the drugs or Pl (crossover design). Day 4 involved dehydration (-1.6% body wt) followed by 45 min of treadmill exercise (65% maximal O2 uptake) in the heat (36 degreesC). These combined stressors caused dramatic decreases in effective renal plasma flow, glomerular filtration rate (GFR), and sodium excretion. Baseline GFR (range: 118-123 ml/min) decreased to 78 +/- 4, 73 +/- 5, and 82 +/- 5 ml/min postexercise in the Acet, Ibu, and Pl trials, respectively, with a significantly greater decrease in GFR in the Ibu trial (P < 0. 05 vs. Pl). OTC Ibu has small but statistically significant effects on GFR during exercise in a sodium- and volume-depleted state; OTC Acet was associated with no such effects.

Carbon disulfide and N,N-diethyldithiocarbamate generate thiourea cross-links on erythrocyte spectrin in vivo.

CS2, a known neurotoxicant, is used in the viscose production of rayon and is also a decomposition product of N, N-diethyldithiocarbamate, a metabolic product of the drug disulfiram used in alcohol aversion therapy. Previous in vitro investigations have demonstrated the ability of CS2 to cross-link proteins through thiourea, dithiocarbamate ester, and disulfide structures. Although in vivo studies have supported protein cross-linking as both a mechanism of neurotoxicity and a potential biomarker of effect, the chemical structures responsible for CS2-mediated protein cross-linking in vivo have not been elucidated. In the present study, the structure of one type of stable protein cross-link produced on erythrocyte spectrin by CS2 in vivo is determined. Rats were exposed to 50, 500, and 800 ppm CS2 for 13 weeks by inhalation or to 3 mmol/kg N,N-diethyldithiocarbamate administered orally on alternating days for 8 weeks. Erythrocyte spectrin preparations from control and exposed rats were hydrolyzed using 6 N HCl and separated by size-exclusion chromatography. The fraction that coeluted with the synthetic deuterated lysine-lysine thiourea internal standard was derivatized with 3-[4'-[(N,N,N-trimethylamino)ethylene]phenyl] 2-isothiocyanate and analyzed by liquid chromatography tandem mass spectrometry using selected reaction monitoring detection. Lysine-lysine thiourea was detected in spectrin preparations obtained from CS2-treated rats at 500 and 800 ppm and N, N-diethyldithiocarbamate-treated rats, but not from controls. These results establish that CS2-mediated protein cross-linking occurs in vivo through the generation of Lys-Lys thiourea and that diethyldithiocarbamate can, through in vivo release of CS2, produce the same cross-linking structure. This observation supports the utility of cross-linking of peripheral proteins as a specific dosimeter of internal exposure for CS2 and provides a mechanistic explanation to account for the high-molecular-weight neurofilament protein species isolated from rats exposed to CS2 or N, N-diethyldithiocarbamate.

Chronic hormone replacement therapy alters thermoregulatory and vasomotor function in postmenopausal women.

This investigation examined effects of chronic (>/=2 yr) hormone replacement therapy (HRT), both estrogen replacement therapy (ERT) and estrogen plus progesterone therapy (E+P), on core temperature and skin blood flow responses of postmenopausal women. Twenty-five postmenopausal women [9 not on HRT (NO), 8 on ERT, 8 on E+P] exercised on a cycle ergometer for 1 h at an ambient temperature of 36 degrees C. Cutaneous vascular conductance (CVC) was monitored by laser-Doppler flowmetry, and forearm vascular conductance (FVC) was measured by using venous occlusion plethysmography. Iontophoresis of bretylium tosylate was performed before exercise to block local vasoconstrictor (VC) activity at one skin site on the forearm. Rectal temperature (Tre) was approximately 0.5 degrees C lower for the ERT group (P < 0.01) compared with E+P and NO groups at rest and throughout exercise. FVC: mean body temperature (Tb) and CVC: Tb curves were shifted approximately 0.5 degrees C leftward for the ERT group (P < 0.0001). Baseline CVC was significantly higher in the ERT group (P < 0.05), but there was no interaction between bretylium treatment and groups once exercise was initiated. These results suggest that 1) chronic ERT likely acts centrally to decrease Tre, 2) ERT lowers the Tre at which heat-loss effector mechanisms are initiated, primarily by actions on active cutaneous vasodilation, and 3) addition of exogenous progestins in HRT effectively blocks these effects.

Decreased active vasodilator sensitivity in aged skin.

Older men and women respond to local and reflex-mediated heat stress with an attenuated increase in cutaneous vascular conductance (CVC). This study was performed to test the hypothesis that an augmented or sustained noradrenergic vasoconstriction (VC) may play a role in this age-related difference. Fifteen young (22 +/- 1 yr) and 15 older (66 +/- 1 yr) men exercised at 50% peak oxygen uptake in a 36 degrees C environment. Skin perfusion was monitored at two sites on the right forearm by laser-Doppler flowmetry: one site pretreated with bretylium tosylate (BT) to block the local release of norepinephrine and thus VC and an adjacent control site. Blockade of reflex VC was verified during whole body cooling using a water-perfused suit. CVC (perfusion divided by mean arterial pressure) at each site was reported as a percentage of the maximal CVC (%CVCmax) induced at the end of each experiment by prolonged local heating at 42 degrees C. Neither age nor BT affected the %CVCmax (75-86%) attained at high core temperatures. During the early rise phase of CVC, the %CVCmax-change in esophageal temperature (delta T(es)) curve was shifted to the right in the older men (effective delta T(es) associated with 50% CVC response for young, 0.22 +/- 0.04 and 0.39 +/- 0.04 degrees C and for older, 0.73 +/- 0.04 and 0.85 +/- 0.04 degrees C at control and BT sites, respectively). BT had no interactive effect on this age difference, suggesting a lack of involvement of the VC system in the attenuated CVC response of individuals over the age of 60 yr. Additionally, increases in skin vascular conductance were quantitatively compared by measuring increases in total forearm vascular conductance (FVC, restricted to the forearm skin under these conditions). After the initial approximately 0.2 degrees C increase in T(es), FVC was 40-50% lower in the older men (P < 0.01) for the remainder of the exercise. Decreased active vasodilator sensitivity to increasing core temperature, coupled with structural limitations to vasodilation, appears to limit the cutaneous vascular response to exertional heat stress in older subjects.

Cosmid-derived markers anchoring the bovine genetic map to the physical map.

The mapping strategy for the bovine genome described in this paper uses large insert clones as a tool for physical mapping and as a source of highly polymorphic microsatellites for genetic typing, and was one objective of the BovMap Project funded by the European Union (UE). Eight-three cosmid and phage clones were characterized and used to physically anchor the linkage groups defining all the bovine autosomes and the X Chromosome (Chr). By combining physical and genetic mapping, clones described in this paper have led to the identification of the linkage groups corresponding to Chr 9, 12, 16, and 25. In addition, anchored loci from this study were used to orient the linkage groups corresponding to Chr 3, 7, 8, 9, 13, 16, 18, 19, and 28 as identified in previously published maps. Comparison of the estimated size of the physical and linkage maps suggests that the genetic length of the bovine genome may be around 4000 cM.

Monitoring hemodialysis changes with bioimpedance. What do we really measure?

Changes in electrical resistance (delta R), water (delta H2O), and blood variables (Na+, K+, Cl-, protein [Pr], osmolality [Osmo], hemoglobin [Hb], and hematocrit [Hct]) were monitored during hemodialysis in 20 subjects. Average delta H2O was 1.69 +/- 0.91 liters. Blood, R, and circumferences (C) were measured for the upper limb (UL), lower limb (LL), and total body (TB) before, midway, and at the end of dialysis. Segmental volumes (V) and specific resistivity (rho) were computed. delta Hct and delta Pr were stronger predictors of delta H2O than delta R. Forward stepwise regression analysis using delta R at 5, 50, and 548 kHz, delta Pr, and delta Hct showed that delta Hct and delta Pr alone accounted for 77% of the variance in delta TBW. Adding delta R increased this at most to 81%. Both the UL and LL had significant increases in rho. The volumetric change of the UL was nonsignificant, whereas that of the LL accounted for over one half of the delta H2O. Single frequency bioimpedance cannot be used to monitor dialytic delta H2O unless one accounts for fluid composition changes. Monitoring bioimpedance changes over only a single body segment is also questioned.

Sequence analysis of a full-length cDNA for the murine pro alpha 2(I) collagen chain: comparison of the derived primary structure with human pro alpha 2(I) collagen.

Comparison of the nucleotide sequence and primary structure of murine and human pro alpha 2(I) collagen indicates a high degree of homology: 87% at the nucleotide level and 87% at the amino acid level, with the greatest degree of variability in the amino- and carboxy-pro-peptide domains. The homology is greatest in the triple helical domain, repeating [Gly-X-Y]338, exhibiting 90% homology at the amino acid level, with only X and Y position residue substitutions. The X and Y residues show 86% homology between murine and human pro alpha 2(I) collagen triple helices, with no truly nonconservative substitutions.

Maximizing communication skills in graduate and postgraduate health-care education through medical writing.

Graduate and postgraduate health-care professional training and postdoctoral fellowship programs that deny trainees opportunities to practice both oral and written communication skills produce an incompletely trained health-care provider unable to compete for faculty positions at university hospitals and affiliated staffs. Therefore, it is imperative that program directors make medical writing a prerequisite to successful completion of postgraduate training programs. To make trainees as well as administrators and faculty aware of the importance of oral and written communication skills, a variety of oral abilities needed for presenting medical findings prior to publication are detailed. The use of 2 x 2 slides to support a presentation as well as transparencies, movies, and videotapes are considered. The poster session/scientific exhibit, now becoming more visible because of increasing attendance at professional meetings, is also explained. Written communication abilities are discussed. Consideration is given to the writing of professional manuscripts for publication in a refereed journal. Other types of written communication include case reports, clinicopathological conferences, letters to the editor, book reviews, books, and book contributions. The opportunity to learn needed skills must be offered in the postgraduate health-care curriculum. Mandatory medical writing will maximize the marketability of black health-care professionals for faculty staff placement. Moreover, the establishment of a "track record" early in a professional career will increase the likelihood that black health providers are awarded grants for research.

Production of alloantisera against class II bovine lymphocyte antigens (BoLA) by cross-immunization between class I matched cattle.

This paper describes the production of alloantisera directed against bovine major histocompatibility complex (MHC) (BoLA) class II antigens in animals whose MHC phenotypes had been defined by one dimensional isoelectric focusing. Animals of closely matched BoLA class I types were selected by serology and subsequently typed for class I and class II by 1D-IEF of immunoprecipitated antigens. Those with similar class I type by both methods, but differing at the class II locus, were chosen for reciprocal immunization. Cross-immunization was by two skin implantations 6 weeks apart. The resulting antisera showed low titre after the first immunization and elevated titre 3 weeks after the second immunization. The sera reacted strongly with cells expressing specific BoLA class II antigens. The pattern of reactivity correlated well with IEF class II typing on a panel of animals representing all of the class II IEF types present in the Friesian population.

An auditory Stroop effect for pitch, loudness, and time.

Increased reaction time to pitch, loudness, or duration of an auditorily presented word whose meaning is in conflict with the response label (i.e., high/low, loud/soft, or fast/slow) demonstrates an auditory Stroop effect. Fifteen normal adults participated in an auditory Stroop test. Analysis of reaction time data supports the existence of an auditory Stroop effect for pitch and loudness, but not duration. The interaction between psychoacoustic and semantic attributes of speech stimuli is discussed.

Relationship between the bovine major histocompatibility complex (BoLA), erythrocyte markers and susceptibility to mastitis in Icelandic cattle.

Milk and blood samples were obtained from three Icelandic dairy herds. The herds were monitored regularly for mastitis incidence. Cell counts, adenosine triphosphate (ATP) and antitrypsin levels of the milk samples were recorded. In addition, red cell and BoLA typing were performed on the blood. Although cell counts and ATP levels showed significant associations with mastitis, antitrypsin levels did not. Red blood cell antigens N'2 and S1 and the lymphocyte antigen detected by the monoclonal antibody M7 were associated with low cell counts, whilst BoLA w6 and w6.1 were associated with high cell counts. BoLA w6.2 and w11 showed significant association with high antitrypsin levels. Only ED116 showed a significant association with mastitis.

Analysis of BoLA w6. Evidence for multiple subgroups.

BoLA w6 is one of 16 specificities agreed upon at the 2nd International BoLA Workshop and many laboratories have produced sera reacting with this specificity. This paper presents evidence for at least four sub-groups in w6. Three Edinburgh sera showing identical reaction patterns in the 1st and 2nd BoLA workshops have been studied by absorption. One serum contained a single antibody population reacting with an epitope common to all w6 positive cells. The other two contained antibodies against the same common epitope, antibodies to epitopes on four sub-groups and an antibody reacting only with one subgroup w6.1. Another Edinburgh serum contained two populations of antibodies. One reacting with the common epitope and one with an epitope on two or possibly three subgroups. Immunisation between w6 positive individuals produced antisera to two subgroups without producing antibodies to common epitopes. At least two additional subgroups are likely to exist. These results indicate the presence of specificities unique to individual subtypes coupled with epitopes common to some and all w6 subgroups.