Separate and combined effects of dehydration and thirst sensation on exercise performance in the heat.

Using intravenous infusion, we separated the physiologic consequences of 3% body mass dehydration from the conscious awareness of fluid replacement on time trial (TT) performance in the heat. Eleven trained cyclists performed 90 min of steady-state (50% V ˙ O 2 peak ) cycling followed by a self-paced 20-km TT in a hot-dry (35 °C, 10% relative humidity, wind speed 3.0 m/s) environment while euhydrated-not thirsty (EU-NT), euhydrated-thirsty (EU-T), dehydrated-not thirsty (DH-NT), or dehydrated-thirsty (DH-T). Thirst was manipulated by providing (NT) or withholding (T) ad libitum 35 °C water oral rinse. Distinct hydration states existed, with 0.4 ± 0.5% dehydration following the 20-km TT (EU) compared with 3.2 ± 0.6% in DH (P < 0.001). Greater perceived thirst existed in T (7 ± 2 on a 1-9 scale) than NT (4 ± 2, P < 0.001) after the TT. No significant differences in power output existed during the TT between hydration (EU 202.9 ± 36.5 W vs DH 207.0 ± 35.9 W, P = 0.362) and thirst conditions (NT 203.3 ± 35.6 W vs T 206.6 ± 36.8 W, P = 0.548), nor were there differences in completion time (P = 0.832) or pacing profile (P = 0.690). Within the range of up to 3% body mass loss, neither the physiologic effects from lowered hydration status nor the perception of thirst, separately or combined, affected sustained submaximal exercise performance in the heat for a healthy and fit population.

Detection of Helicobacter pylori by PCR but not culture in water and biofilm samples from drinking water distribution systems in England.

AIMS: To investigate treated water distribution systems in England as a source of Helicobacter pylori. METHODS AND RESULTS: Water and biofilms were obtained from 11 domestic and seven educational properties and from hydrants, reservoirs and water meters supplied by three water utilities. Samples were cultured on nonselective and antibiotic containing media combined with immunomagnetic separation concentration. Viable helicobacters were not detected in any of the 151 samples but Helicobacter-specific PCR assays detected DNA in 26% of samples from domestic properties, schools and hydrants with the highest frequency in biofilms (42%). Direct sequencing of six selected amplicons confirmed >95% sequence homology to H. pylori. CONCLUSIONS: While viable helicobacters were not isolated, evidence was obtained for the presence of Helicobacter DNA, including that of H. pylori. Biofilms on surfaces within water distribution systems may act either as sites for the passive accumulation of helicobacters or as potentially important reservoirs of infection. SIGNIFICANCE AND IMPACT OF THE STUDY: Our findings strengthen evidence that H. pylori may be transmitted through drinking water. However, there is currently no evidence that viable cells can survive the disinfection levels used in UK mains supplies and the health risk from this source remains unclear.

Priming prostate carcinoma cells for increased apoptosis is associated with up-regulation of the caspases.

BACKGROUND: The potential to prime prostatic carcinoma cell lines for apoptosis represents an exciting strategy for the treatment of patients with this disease. The ability and the underlying molecular mechanisms involved in sensitizing both androgen-sensitive and androgen-insensitive cell types to a range of apoptotic-inducing agents are investigated by the authors. METHODS: Primary and secondary cell lines were pretreated with diethyl-maleate (DEM) prior to the induction of apoptosis by Fas antibody (1 microg/mL), cycloheximide (1 microg/mL), etoposide (62.5 microM), and radiation (5 grays). It was demonstrated previously that DEM (50 microM) increases the sensitivity to apoptosis induced by these agents. The effects of DEM on both protein and RNA expression was determined by Western blot analysis and a ribonuclease protection assay, respectively. The effects of DEM on intracellular glutathione (GSH) levels and its intracellular distribution also were assessed. RESULTS: DEM did not affect the expression of the caspases at the transcriptional level but was associated with increased procaspase-3 and caspase-8 protein levels. DEM preincubation restored sensitivity to Fas antibody and radiation-induced apoptosis in cells from the LNCaP-bcl-2 transfectant cell line that, normally, are resistant to these apoptotic stimuli. It is that DEM chemically depletes intracellular thiol levels. Although no depletion in total intracellular thiol GSH was observed at these concentrations of DEM, trafficking of GSH from the nucleus to the cytosol was demonstrated. CONCLUSIONS: Identification of the caspases as a potential target for chemical manipulation may serve as an effective, adjuvant-based approach in the treatment of patients with prostate carcinoma and, in particular, for immunotherapy and radiation-based strategies that rely on the activation of these death-effector proteases.

Cytological transformations associated with parietal cell stimulation: critical steps in the activation cascade.

Cultured rabbit parietal cells were used to evaluate morphological responses to activators and inhibitors of HCl secretion. Immunofluorescence was used to localize the proton pump protein, H, K-ATPase, and the apical membrane-cytoskeletal linker protein, ezrin; fluorescent-labeled phalloidin was used as a marker of F-actin. Treatment of healthy control parietal cells with secretagogues resulted in exaggerated swelling of apical membrane vacuoles, presumably with the accumulation of HCl and water. Thus stimulation-associated swelling of apical vacuoles was blocked by inhibitors that work at various steps in the secretion-activation cascade. When secretion was blocked by agents that prevent the translocation of H,K-ATPase-rich tubulovesicles to apical membrane vacuoles (such as H2-receptor antagonists and protein kinase A inhibitors), the general resting morphology was maintained. ME-3407 (a functional analogue of wortmannin) was unique in preventing H, K-ATPase redistribution and effecting the delocalization of ezrin from apical membrane vacuoles. When secretion was blocked by agents that inhibit the H+ pump or induce H+ backflux, the translocation of H,K-ATPase to apical membrane vacuoles occurred but the large vacuolar swelling associated with HCl and H2O accumulation was greatly diminished. These data support the membrane recycling/recruitment hypothesis of HCl secretion in which H, K-ATPase-rich tubulovesicles are recruited from a cytoplasmic domain to the apical surface, and they are inconsistent with models proposing that the tubulovesicles, regardless of shape, are contiguous with the apical plasma membrane. These studies also demonstrate the utility of the parietal cell culture model in distinguishing a general site of action for various inhibitors and antisecretory agents.

Promoting family-centered care in high risk pregnancy.

Complications during pregnancy heralds a time of increased stress and anxiety for the obstetric patient and her family. Separation from family and home is a common stressor. Incorporating key elements of family-centered care into the policies of the obstetric units that support the health care team to include the family in the plan of care are crucial to family functioning during this difficult time. Assessment of the family and interventions to support family functioning can be structured using the Calgary Family Assessment Model and the Calgary Family Intervention Model. Models that emphasize the family assist the health care team to shift from a plan focused on the medical aspects of care to one that is incorporated to include the woman's family, the base of her support.

Modulation of Na+ current inactivation by stimulation of protein kinase C in cardiac cells.

Modulation of the inward Na+ current (I(Na)) by protein kinase C (PKC) was investigated by intracellular perfusion of a peptide corresponding to the catalytic subunit of PKC (PKCP). The effects of PKC activation independent of membrane-receptor pathways were studied in neonatal rat ventricular myocytes using whole-cell patch-clamp techniques. Perfusion with 2 nmol/L PKCP caused a depolarizing shift in steady state half-inactivation relative to control (-83.2+/-1.3 versus -74.9+/-1.6 mV for control versus PKCP, respectively) without a change in current-voltage relationships or peak I(Na). The development of resting inactivation was slowed by PKCP (tau, 69.1+/-7.6 [control] versus 100.4+/-5.1 ms). Open-channel inactivation, estimated by measuring I(Na) decay from peak current at test voltages between -10 and +30 mV was significantly slowed by PKCP. Recovery from inactivation was more rapid during PKCP perfusion, with a shortening of both the fast (tau(f)) and slow (tau(s)) components of tau (tau(f), 38.5+/-7.0 [control] versus 14.2+/-4.7 ms; tau(s), 163.4+/-47.9 [control] versus 51.3+/-9.2 ms). All of the effects of PKCP on I(Na) were antagonized by the PKC inhibitors chelerythrine chloride or staurosporine or by down-regulation of PKC using phorbol ester preincubation. We conclude that the actions of PKC on the Na+ channel result in slowing the development of inactivation and accelerating reactivation, resulting in less resting inactivation.

Lysophosphatidylcholine modulates cardiac I(Na) via multiple protein kinase pathways.

Lysophosphatidylcholine (LPC) is a naturally occurring intracellular phospholipid metabolite that has been implicated in arrhythmogenesis during ischemia. LPC has been shown to affect the cardiac Na+ current (I(Na)), but the mechanism of modulation remains undescribed. Recently, low concentrations of LPC have been shown to activate protein kinase C (PKC) independent of the receptor-delineated pathway. The purposes of this study were to describe the effects of intracellularly introduced LPC on I(Na) and to determine if these effects were mediated by kinases. Modulation of I(Na) was studied in ventricular cells with LPC (1 nmol/L to 1 micromol/L) internally applied using whole-cell patch-clamp techniques. Intracellular LPC caused a dose-dependent depolarizing shift of steady state inactivation that was accompanied by a change in slope factor. The development of resting inactivation from closed states was delayed 40%, whereas the recovery from inactivation was significantly accelerated. These results were mimicked by another bioactive lipid, lysophosphatidylethanolamine, or by a peptide analogue of PKC, which is a potent stimulator of endogenous PKC activity. Maximal recruitable current was significantly increased by LPC but not by PKC activation. Some of the effects of LPC on I(Na) could be partially inhibited by the specific PKC inhibitor chelerythrine chloride or by downregulation of PKC with phorbol ester pretreatment. However, genistein, a specific tyrosine kinase inhibitor, completely inhibited all the modulation of I(Na) caused by LPC. These data suggest that LPC modulates I(Na) in cardiac myocytes by a pathway that involves both PKC-dependent and tyrosine kinase- dependent phosphorylation.

Hepatic basolateral sodium-dependent-bile acid transporter expression in two unusual cases of hypercholanemia and in extrahepatic biliary atresia.

The recent cloning of a human sodium-dependent bile acid transporter (NTCP) permits analysis of its expression in human liver disease and investigation of potential primary defects in its expression. NTCP from normal human liver (NHL) was first characterized in detail. Northern blotting of RNA from NHL revealed a 1.8-kb NTCP transcript. Western blotting of crude NHL plasma membranes using a carboxyterminal antipeptide antibody showed that NTCP is a 39-kd polypeptide that is N-glycosylated to a final molecular weight of 56 kd. Indirect immunofluorescent analysis of NHL sections indicated that the NTCP protein is expressed on the basolateral surface of hepatocytes. We hypothesized that the clinical phenotype of a defect in NTCP might be hypercholanemia in the relative absence of liver disease. Accordingly, the coding region of the NTCP gene of two children with this phenotype was sequenced after reverse transcription/polymerase chain reaction (RT/PCR) amplification. No primary defects in the deduced NTCP amino acid sequence were found. Despite the extremely high serum bile salt levels (235 and 126 micromol/L) in these two patients, NTCP messenger RNA (mRNA) and protein expression were quantitatively normal, in contrast to the published observations in a rat model of cholestasis secondary to common bile duct ligation. Hepatic steady-state NTCP mRNA levels in a group of 23 pre- and postportoenterostomy biliary atresia patients were inversely related to total bilirubin, indicating that extrahepatic bile duct obstruction leads to down-regulation of NTCP mRNA levels, similar to that observed in rat common bile duct ligation. Therefore the lack of down-regulation in the two patients with hypercholanemia indicates that elevated serum bile salts are not sufficient to down-regulate NTCP expression, these two patients have abnormal responses to hypercholanemia, or these two patients have a defect in a gene other than NTCP that influences hepatic clearance of bile salts.

Effect of intracellular and extracellular acidosis on sodium current in ventricular myocytes.

Conduction slowing is an essential element in the generation of ischemic ventricular arrhythmias and is determined in part by the inward Na+ current (INa). Because intracellular acidosis is an early consequence of ischemia, we hypothesized that lowering intracellular pH (pHi) would reduce or kinetically modulate INa and thus affect cardiac conduction. To test this hypothesis, the whole cell patch-clamp method was used to measure INa in neonatal rat ventricular myocytes exposed to varying extracellular pH (pHo 6.4-7.4), while perfusing the cells with acidic solutions (pHi 6.2-7.2). With simultaneous acidification of pHo and pHi there was a progressive increase in time to peak current, a 31% decrease in peak INa (298 +/- 18 to 206 +/- 16 pA/pF), and a complex slowing of inactivation kinetics. At the most extreme levels of acidification, there was a 5-mV hyperpolarizing shift in steady-state inactivation and a 6-mV depolarizing shift in activation. Independent changes of pHo and pHi indicate that the reduction of peak INa is a function of pHo. However, steady-state inactivation is modulated by pHi. The time course of activation and inactivation appears to depend on both pHo and pHi. We conclude that both intracellular and extracellular acidosis are significant but distinct modulators of INa amplitude and kinetics in cardiac myocytes.

Lactic acidosis transiently increases metabolic rate of turtle myocytes.

We measured O2 consumption as an estimate of metabolic rate in isolated calcium-tolerant ventricular myocytes of turtles (Chrysemys picta belli) at control pH 7.8 and in the same solution brought to pH 7.4 and 7.0 with additions of lactic acid. Our aim was to test the hypothesis that lactic acidosis caused metabolic depression by initiating downregulation of Na+ channels, and thus Na(+)-K(+)-ATPase (Na+ pump) activity, which we would measure as a decrease in O2 consumption. Myocyte O2 consumption was measured in reptilian N-tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid-buffered Ringer solution and in nomially Na(+)- and Ca(2+)-free solution, thus estimating the Na+ pump component of metabolic rate. Lowering extracellular pH from 7.8 to 7.0 resulted in a significant increase in metabolic rate of cells in Ringer solution but not those in Na(+)- and Ca(2+)-free solution. This result was unchanged by the addition of 2 mM Ca2+ to Na(+)-free cell suspensions, indicating that the difference was due to the presence of Na+. Addition of 100 microM amiloride to cells in Ringer solution at pH 7.0 abolished the increase in O2 consumption, suggesting that the apparent increase in Na(+)-K(+)-ATPase activity was secondary to Na(+)-H+ exchange. Intracellular pH was measured using 5,5-dimethyl[14C]oxazolidine-2,4-dione. Cells treated with amiloride and those in Na(+)- and Ca(2+)-free solution did not regulate intracellular pH following acidosis and maintained basal metabolic rate. These data suggest that the Na(+)-H+ exchanger is an important contributor to intracellular pH regulation in the myocyte but increases Na+ pump activity and metabolic rate immediately following acidosis.

The role of personality in the management of partial dentures.

A sample of fifty-three partially dentate subjects who had had partial dentures provided in the last 3 years completed a personality questionnaire (Cattell's 16 PFQ). At the same time they completed a clinical questionnaire and were examined by two of the authors. The purpose of this was to assess the patients' satisfaction with their partial dentures. The results of the personality questionnaire showed that this group of partial denture wearers differed from the general population on only two factors. They were less intelligent and more self-sufficient. Interestingly they differed from a similar group of complete denture wearers on a further three factors. The partial denture group were more self-assured, self-sufficient and more relaxed. On the basis of the clinical questionnaire and examination, five partially dentate patients were found to be dissatisfied. Their personality scores were significantly more unstable than the satisfied group of partial denture wearers. Although based on a relatively small group, the results do suggest that personality may have a marked effect on the patients' adaption to partial dentures. When compared to a similar group of edentulous patients, marked differences were found, suggesting that the loss of the natural teeth in some individuals affects behaviour.