Dynamic changes in renal sodium handling during sympathetic stimulation in healthy human males.

The temporal response of changes in renal sodium reabsorption during increased renal sympathetic nerve activity has not been investigated. Central hypovolemia by application of lower-body negative-pressure (LBNP) elicits baroreceptor mediated sympathetic reflexes to maintain arterial blood pressure. We hypothesized, that during 90 min LBNP, the renal sodium retention would increase rapidly, remain increased during intervention, and return to baseline immediately after end of intervention. METHODS: 30 young, healthy, sodium loaded, non-obese males were exposed to -15 mmHg LBNP, -30 mmHg LBNP, -15 mmHg LBNP + renin blockade or time-control (0 mmHg LBNP) for 90 min. Urine was collected every 15 min during 90 min of intervention and 60 min of recovery to identify a possible relation between time of intervention and renal response. RESULTS: All intervention groups exhibited a comparable reduction in distal sodium excretion at the end of the intervention (P = 0.46 between groups; -15 mmHg: -3.1 ± 0.9 %, -30 mmHg: -2.9 ± 0.6 %, -15 mmHg + aslikiren: -1.8 ± 0.6 %). -15 mmHg+Aliskiren resulted in a slower onset, but all groups exhibited a continued reduction in sodium excretion after 1 h of recovery despite return to baseline of renin, aldosterone, diuresis and cardiovascular parameters. CONCLUSION: Sympathetic stimulation for 90 min via LBNP at -30 mmHg LBNP compared to -15 mmHg did not result in a greater response in fractional Na+ excretion, suggesting that additional baroreceptor unloading did not cause further increases in renal sodium reabsorption. Changes in distal Na+ excretion were linear with respect to time (dose) of intervention, but seem to exhibit a saturation-like effect at a level around 4 %. The lack of recovery after 1 h is also a new finding that warrants further investigation.

A method for assessment of the dynamic response of the arterial baroreflex.

AIM: The baroreflex is a key mechanism in cardiovascular regulation, and alterations in baroreceptor function are seen in many diseases, including heart failure, obesity and hypertension. We propose a new method for analysing baroreceptor function from continuous blood pressure (BP) and heart rate (HR) in both health and disease. METHODS: Forty-eight-hour data series of BP and HR were collected with telemetry. Sprague Dawley rats on standard chow (n = 11) served as controls, while rats on a high-fat, high-fructose (HFHC) diet (n = 6) constituted the obese-hypertensive model. A third group of rats underwent autonomic blockade (n = 6). An autoregressive-moving-average with exogenous inputs (ARMAX) model was applied to the data and compared with the α-coefficient. RESULTS: Autonomic blockade caused a significant reduction in the strength of the baroreflex as estimated by ARMAX [ARMAX- baroreflex sensitivity (BRS)] -0.03 ± 0.01 vs. -0.19 ± 0.04 bpm heartbeat-1) . Both methods showed a ~50% reduction in BRS in the obese-hypertensive group compared with control (body weight 531 ± 27 vs. 458 ± 19 g, P < 0.05; mean arterial pressure 119 ± 3 vs. 102 ± 1 mmHg, P < 0.05; ARMAX-BRS -0.08 ± 0.01 vs. -0.15 ± 0.01 bpm heartbeat-1 , P < 0.05; α-coefficient BRS 0.51 ± 0.07 vs. 0.89 ± 0.07 ms mmHg-1 , P < 0.05). The ARMAX method additionally showed the open-loop gain of the baroreflex to be reduced by ~50% in the obese-hypertensive group (-2.3 ± 0.3 vs. -4.1 ± 0.3 bpm, P < 0.05), while the rate constant was similar between groups. CONCLUSION: The ARMAX model represents an efficient method for estimating several aspects of the baroreflex. The open-loop gain of the baroreflex was attenuated in obese-hypertensive rats compared with control, while the time response was similar. The algorithm can be applied to other species including humans.

An epidemiological study of cataracts in wild and farmed lumpfish (Cyclopterus lumpus L.) and the relation to nutrition.

Twelve groups of farmed lumpfish and one of wild lumpfish were screened for cataract and sampled for fish muscle tissue, whole heart and both eye lenses to investigate possible relations between cataract and tissue free amino acid concentrations. Cataract prevalence ranged from 20% to 100%, with the highest average score of 7.3 (max 8) and incidences of severe cataract (>5) in all groups. Cataract could not be explained by suboptimal histidine concentrations in the feed. Neither muscle nor cardiac tissues had concentrations of free histidine compounds. The lumpfish lens contained N-acetylhistidine (NAH), of which low concentrations were strongly related to cataract severity. However, no correlation between lens NAH and cataract severity was found in the present sample set. Wild lumpfish had higher levels compared to farmed lumpfish, suggesting that the farmed lumpfish may have been deficient in histidine or have a higher utilization of NAH due to osmotic problems. Thus, cataract in farmed lumpfish may be related to primary or secondary disturbed nutrient metabolism or malnutrition, shown by the high levels of specific amino acids in different tissues, which may cause osmotic imbalance and cataract development. This nutritional or environmental-related welfare problem deserves further research.

Short-term effects of liraglutide on kidney function and vasoactive hormones in type 2 diabetes: a randomized clinical trial.

AIMS: To investigate the effects of a single dose of 1.2 mg liraglutide, a once-daily glucagon-like peptide-1 (GLP-1) receptor agonist, on key renal variables in patients with type 2 diabetes. METHODS: The study was a placebo-controlled, double-blind, crossover trial in 11 male patients with type 2 diabetes. Measurements included (51) Cr-EDTA plasma clearance estimated glomerular filtration rate (GFR) and MRI-based renal blood flow (RBF), tissue perfusion and oxygenation. RESULTS: Liraglutide had no effect on GFR [95% confidence interval (CI) -6.8 to 3.6 ml/min/1.73 m(2) ] or on RBF (95% CI -39 to 30 ml/min) and did not change local renal blood perfusion or oxygenation. The fractional excretion of lithium increased by 14% (p = 0.01) and sodium clearance tended to increase (p = 0.06). Liraglutide increased diastolic and systolic blood pressure (3 and 6 mm Hg) and heart rate (2 beats per min; all p < 0.05). Angiotensin II (ANG II) concentration decreased by 21% (p = 0.02), but there were no effects on other renin-angiotensin system components, atrial natriuretic peptides (ANPs), methanephrines or excretion of catecholamines. CONCLUSIONS: Short-term liraglutide treatment did not affect renal haemodynamics but decreased the proximal tubular sodium reabsorption. Blood pressure increased with short-term as opposed to long-term treatment. Catecholamine levels were unchanged and the results did not support a GLP-1-ANP axis. ANG II levels decreased, which may contribute to renal protection by GLP-1 receptor agonists.

Enhanced alveolar fluid clearance following 72 h of continuous isoproterenol infusion in rats.

AIM: We wished to determine the effect of continuous ß-receptor stimulation on alveolar fluid clearance and to elucidate the mechanisms behind this effect. METHODS: Alveolar fluid clearance was measured in anaesthetized rats pretreated for 72 h with the ß-agonist isoproterenol (200 µg kg(-1) h(-1) sc) or vehicle. Alveolar fluid clearance in artificially ventilated rats was determined over 1 h by infusion of isotonic Ringer solution containing (125) I-albumin into the lungs. Additionally, alveolar fluid clearance was determined when amiloride or l-cis-diltiazem was added to the solution to block ENaC and cyclic nucleotide-gated (CNG) channels respectively. RESULTS: Isoproterenol treatment induced a 42% increase in alveolar fluid clearance (18.9 ± 1.4%) vs. vehicle (13.3 ± 3.3%). Addition of amiloride resulted in a net decrease of 8% in both groups, while l-cis-diltiazem caused a net decrease of 12% in isoproterenol-treated animals, but only 5% in vehicle-treated animals. Western blotting showed that isoproterenol treatment increased the abundance of the α-ENaC and ß-ENaC subunits (223 ± 51% and 274 ± 55% of vehicle, respectively) but we saw no changes in protein level of the γ-EnaC subunit. CONCLUSION: Continuous ß-adrenoceptor stimulation with isoproterenol enhances alveolar fluid clearance through alternative pathways involving l-cis-diltiazem-sensitive channels.

Detection and characterisation of SCCmec remnants in multiresistant methicillin-susceptible Staphylococcus aureus causing a clonal outbreak in a Swedish county.

The purpose of this study was to investigate if multiresistant methicillin-susceptible Staphylococcus aureus (MR-MSSA) causing a clonal outbreak in Östergötland County, Sweden, were derived from methicillin-resistant S. aureus (MRSA) by carrying remnants of SCCmec, and, if so, to characterise this element. A total of 54 MSSA isolates with concomitant resistance to erythromycin, clindamycin and tobramycin from 49 patients (91% clonally related, spa type t002) were investigated with the BD GeneOhm MRSA assay and real-time polymerase chain reaction (PCR) targeting the SCCmec integration site/SCCmec right extremity junction. DNA sequencing of one isolate representing the MR-MSSA outbreak clone was performed by massive parallel 454 pyrosequencing. All isolates that were part of the clonal outbreak carried SCCmec remnants. The DNA sequencing revealed the carriage of a pseudo-SCC element 12 kb in size, with a genomic organisation identical to an SCCmec type ΙΙ element, except for a 41-kb gap. This study demonstrates the presence of a pseudo-SCC element resembling SCCmec type II among MR-MSSA, suggesting possible derivation from MRSA. The presence of SCCmec remnants should always be considered when SCCmec typing is used for MRSA detection, and may not be suitable in locations with a high prevalence of MR-MSSA, since this might give a high number of false-positive results.

Effects of renal denervation on the NKCC2 cotransporter in the thick ascending limb of the loop of Henle in rats with congestive heart failure.

AIM: Congestive heart failure (CHF) is associated with increased renal sympathetic nerve activity and renal sodium retention. Rats with CHF display increased expression of the Na-K-2Cl cotransporter (NKCC2) in the renal medullary thick ascending limb of the loop of Henle (mTAL), and arginine vasopressin (AVP)-stimulated cAMP formation in mTAL segments is increased in rats with CHF. The aim of the study was to investigate the role of RSNA on cAMP formation and NKCC2 expression in mTAL in rats with CHF. METHODS: Congestive heart failure was induced in male Wistar rats by ligation of the left anterior descending coronary artery. Bilateral surgical renal denervation (DNX) was performed 3 weeks later. Two weeks after DNX, mTAL segments were isolated and stimulated with AVP. RESULTS: Congestive heart failure rats displayed increased mTAL NKCC2 expression (2.5 ± 0.5 vs. 1 ± 0.2 in Sham rats), which was abolished by DNX. Bilateral denervation decreased basal cAMP levels in unstimulated tubules from CHF rats (CHF: 12.56 ± 7.73 fmol µg(-1) protein vs. DNX-CHF: 7.94 ± 4.33; P < 0.05), as well as from Sham rats (SHAM: 4.70 ± 1.38 vs. DNX-SHAM: 2.36 ± 1.52; P < 0.05). mTAL segments from DNX-CHF and DNX-Sham rats showed decreased AVP (10(-6) M)-stimulated cAMP formation, compared with CHF (CHF: 11.92 ± 4.89 fmol µg(-1) protein vs. DNX-CHF: 4.68 ± 2.47; P < 0.05) and Sham (SHAM: 10.78 ± 5.59 vs. DNX-SHAM: 4.89 ± 2.62; P < 0.05). CONCLUSION: These results indicate that the renal sympathetic nerves have an effect on NKCC2 expression in the mTAL and might have an effect on cAMP formation in the TAL in CHF.

Rapid MRSA test in exposed persons: costs and savings in hospitals.

OBJECTIVE: To study a rapid Xpert polymerase chain reaction (PCR) method in detecting methicillin-resistant Staphylococcus aureus (MRSA) in patients and healthcare workers (HCW) exposed to MRSA, and to estimate savings associated to isolation or work restriction. METHODS: A test set of four double (one for the growth and one for the rapid test) pre-wet swabs from the nose, throat, hands/wrists and perineum was studied by a growth method and by the Xpert MRSA test. RESULTS: The total correspondence between the growth and the rapid test was 92.8%. The overall sensitivity, specificity, positive and negative predictive values were for the Xpert MRSA test: 87%, 99.6%, 68.5% and 99.9%, and for the growth test: 76%, 100%, 100%, and 99.8%, assuming a prevalence of MRSA of 0.01%. Among the MRSA positive persons, the Xpert and growth tests detected MRSA in 44.6% and 40% of nose samples, respectively, 38.2% and 45.5% throat samples, 30.8% and 11.5% hands/wrists samples, 44% and 38% perineum samples, and in 81.8% and 77.3% wound samples, respectively. By combining four anatomical sites, the detection rate increased to 87.5% by both methods. The cost for each Xpert and growth test was euro50 and euro6.25, respectively. The rapid test would save at least euro925 per exposed HCW and euro550 per patient that were MRSA negative. CONCLUSION: The MRSA Xpert test is easy to perform, has a high negative predictive value, and may be used to control healthcare workers and patients exposed to MRSA. Sampling from multiple anatomical locations is recommended. Still, more then 10% of MRSA positive cases may not be found.

AP214, an analogue of alpha-melanocyte-stimulating hormone, ameliorates sepsis-induced acute kidney injury and mortality.

Sepsis remains a serious problem in critically ill patients with the mortality increasing to over half when there is attendant acute kidney injury. alpha-Melanocyte-stimulating hormone is a potent anti-inflammatory cytokine that inhibits many forms of inflammation including that with acute kidney injury. We tested whether a new alpha-melanocyte-stimulating hormone analogue (AP214), which has increased binding affinity to melanocortin receptors, improves sepsis-induced kidney injury and mortality using a cecal ligation and puncture mouse model. In the lethal cecal ligation-puncture model of sepsis, severe hypotension and bradycardia resulted and AP214 attenuated acute kidney injury of the lethal model with a bell-shaped dose-response curve. An optimum AP214 dose reduced acute kidney injury even when it was administered 6 h after surgery and it significantly improved blood pressure and heart rate. AP214 reduced serum TNF-alpha and IL-10 levels with a bell-shaped dose-response curve. Additionally; NF-kappaB activation in the kidney and spleen, and splenocyte apoptosis were decreased by the treatment. AP214 significantly improved survival in both lethal and sublethal models. We have shown that AP214 improves hemodynamic failure, acute kidney injury, mortality and splenocyte apoptosis attenuating pro- and anti-inflammatory actions due to sepsis.

Losartan decreases vasopressin-mediated cAMP accumulation in the thick ascending limb of the loop of Henle in rats with congestive heart failure.

INTRODUCTION: Vasopressin (AVP) stimulates sodium reabsorption and Na,K,2Cl-cotransporter (NKCC2) protein level in the thick ascending limb (TAL) of Henle's loop in rats. Rats with congestive heart failure (CHF) have increased protein level of NKCC2, which can be normalized by angiotensin II receptor type-1 (AT(1)) blockade with losartan. AIM: In this study, we investigated whether CHF rats displayed changes in AVP stimulated cAMP formation in the TAL and examined the role of AT(1) receptor blockade on this system. METHOD: CHF was induced by ligation of the left anterior descending coronary artery (LAD). SHAM-operated rats were used as controls. Half of the rats were treated with losartan (10 mg kg day(-1) i.p.). RESULTS: CHF rats were characterized by increased left ventricular end diastolic pressure. Measurement of cAMP in isolated outer medullary TAL showed that both basal and AVP (10(-6) m) stimulated cAMP levels were significantly increased in CHF rats (25.52 +/- 4.49 pmol cAMP microg(-1) protein, P < 0.05) compared to Sham rats (8.13 +/- 1.14 pmol cAMP microg(-1) protein), P < 0.05). Losartan significantly reduced the basal level of cAMP in CHF rats (CHF: 12.56 +/- 1.93 fmol microg(-1) protein vs. Los-CHF: 7.49 +/- 1.08, P < 0.05), but not in Sham rats (SHAM: 4.66 +/- 0.59 vs. Los-SHAM: 4.75 +/- 0.71). AVP-mediated cAMP accumulation was absent in both groups treated with losartan (Los-SHAM: 4.75 +/- 0.71 and Los-CHF: 7.49 +/- 1.08). CONCLUSION: The results indicate that the increased NKCC2 protein level in the mTAL from CHF rats is associated with increased cAMP accumulation in this segment. Furthermore, the finding that AT(1) receptor blockade prevents AVP-mediated cAMP accumulation in both SHAM and CHF rats suggests an interaction between angiotensin II and AVP in regulation of mTAL Na reabsorption.

Biphasic changes of epithelial sodium channel abundance and trafficking in common bile duct ligation-induced liver cirrhosis.

We hypothesize that dysregulation of the epithelial sodium channel (ENaC) may be responsible for the increased sodium retention in liver cirrhosis. Liver cirrhosis was induced by common bile duct ligation (CBDL). We examined the abundance of ENaC subunits and type 2 isoform of 11beta-hydroxysteroid dehydrogenase (11betaHSD2) in the kidney by immunoblotting and immunohistochemistry at 6 or 8 weeks after operation. At 6 weeks, cirrhotic rats had developed ascites and displayed a positive sodium balance. The urinary sodium excretion and fractional excretion of sodium were decreased, while plasma aldosterone was unchanged. The abundance of ENaC subunits was not changed in the cortex and outer stripe of the outer medulla (OSOM). In contrast, immunoperoxidase microscopy revealed an increased apical targeting of alpha-, beta- and gammaENaC in late distal convoluted tubule, connecting tubule and collecting duct. Moreover, 11betaHSD2 abundance was decreased in the cortex/OSOM and inner stripe of the outer medulla. At 8 weeks, urinary sodium excretion and fractional excretion of sodium were not changed, while the plasma aldosterone level was decreased. The expression of ENaC subunits was decreased in the cortex/OSOM. Immunoperoxidase microscopy confirmed decreased expression of ENaC subunits, whereas subcellular localization was not changed. These results suggest that increased apical targeting of ENaC subunits and diminished abundance of 11betaHSD2 may contribute to promote sodium retention in the sodium-retaining stage of liver cirrhosis (at 6 weeks). The subsequent decreased expression and reduced targeting of ENaC subunits may play a role in promoting sodium excretion in the later stage of liver cirrhosis (at 8 weeks).

Magnesium depletion enhances cisplatin-induced nephrotoxicity.

PURPOSE: Nephrotoxicity and magnesium (Mg)-depletion are well-known side effects to cisplatin (CP) treatment. The purpose of this present study was to investigate the role of Mg on CP induced changes in renal function. CP induced renal dysfunction was achieved by treatment with CP or vehicle (2.5 mg/kg) once weekly for 3 weeks. Since the CP-induced renal damage, including tubular reabsorption defects, is most prominent within the outer medulla (OM), changes in the expression pattern of OM aquaporins and sodium transporters including the Na,K-ATPase (alpha-subunit), type III Na,H-exchanger (NHE3), aquaporin 1 (AQP1) and 2 (AQP2) and the Na,K,2Cl-cotransporter (NKCC2) were investigated by semi-quantitative Western blotting. EXPERIMENTAL DESIGN: Rats had access to either a diet with standard Mg or to a Mg-depleted diet. Cisplatin was administered to female Wistar rats once a week for 3 weeks according to four regimens: (1) Cisplatin 2.5 mg/kg body weight i.p., to rats on a diet with standard Mg, (2) Cisplatin 2.5 mg/kg body weight i.p., to rats on a diet with low Mg, (3) Isotonic NaCl 2.5 ml/kg body weight i.p., to rats on a diet with standard Mg, (4) Isotonic NaCl 2.5 ml/kg body weight i.p., to rats on a diet with low Mg. RESULTS: CP had no effect on plasma creatinine or urea in rats with standard Mg intake, but the expression of all five transporters was significantly reduced when compared to vehicle treated rats on standard Mg-intake. Vehicle treated rats on low Mg-intake had a significant reduction in the expression of Na,K-ATPase, NHE3 and NKCC2, but unchanged expression levels of AQP1 or AQP2 when compared to standard treated controls. Forty percent of the CP-treated rats on low Mg-intake died during the experiment and the remaining animals had marked increased plasma creatinine and urea. Furthermore, the Western blot analysis revealed an almost complete disappearance of all four transporters, suggesting a dramatic synergistic effect of CP and Mg-depletion on renal function including the expression pattern of outer medullary sodium transporters and aquaporins. CONCLUSIONS: This study indicates a substantial additive effect of Mg-depletion on cisplatin induced renal toxicity as evidenced by significant changes in plasma creatinine and urea, renal failure induced mortality and loss of renal transporters. This should give cause for concern since the nephrotoxicity observed during cisplatin treatment might be substantiated by the known Mg-loss associated with cisplatin treatment especially in patients suffering from intense gastro-intestinal side effects.

The effect of spironolactone on experimental periodontitis in rats.

BACKGROUND: Elevated levels of tumour necrosis factor (TNF) have been found in patients with adult periodontitis. Animal studies have shown that TNF plays an important role in the pathogenesis of periodontitis. New findings suggest that the aldosterone-inhibitor spironolactone possesses an anti-TNF effect. The purpose of the study was to determine the anti-TNF effect of spironolactone in an endotoxic shock rat model and to disclose the effect of oral administration of spironolactone on the development of experimental periodontitis in rats. METHODS: The study was divided in two parts. Part 1: oral administration of spironolactone (100 mg/kg) followed by intravenous lipopolysaccharide (1 mg/kg) infusion 45 min later. Blood samples were taken before and 90 min after lipopolysaccharide infusion to determine the TNF levels in spironolactone treated and non-treated rats. Part 2: oral administration of spironolactone [100 mg/(kg day)] starting 2 days prior to induction of experimental periodontitis established by peridental ligatures. Morphometrical and radiographical registrations of alveolar bone destruction were carried out to determine the effect of spironolactone on the progression of experimental periodontitis. RESULTS: In part 1 the endotoxic shock model showed a significant reduction in TNF levels in the spironolactone-treated group compared to the non-treated group, suggesting that spironolactone acts as a TNF inhibitor. In part 2 spironolactone-treated rats did not demonstrate significantly less alveolar bone destruction compared to non-treated rats. CONCLUSIONS: The insignificant effect of spironolactone treatment could be explained by the fast metabolism of spironolactone and that spironolactone does not completely inhibit TNF production in rats. Moreover, many other cytokines and mediators involved in alveolar bone destruction may account for the lacking response to spironolactone.

Complete genomic sequences of astroviruses from sheep and turkey: comparison with related viruses.

The genomes of astroviruses infecting sheep and turkey were sequenced. Detailed analyses of these sequences were performed, including comparison with the other complete astrovirus sequences available as well as with other RNA virus sequences, with focus on the non-structural proteins and RNA sequences. Earlier postulated functional astrovirus RNA motifs and protein domains could in most cases be recognised in the sheep and turkey astrovirus sequences. In addition, analyses of the available astrovirus sequences revealed: two protein regions with the potential for forming coiled coils, differences in the postulated transmembrane region, a similarity between the putative astrovirus nuclear localisation signal and calicivirus genome-linked proteins, and a stretch of a highly conserved RNA sequence with a possible role in the astrovirus capsid gene expression. The present analyses contribute to the deciphering of pertinent functions of the astrovirus genomes.

Collecting duct function in liver cirrhotic rats with early sodium retention.

Liver cirrhosis is a chronic disease associated with sodium retention due to increased tubular sodium reabsorption. However, the exact tubular site of increased sodium reabsorption in uncertain. We have recently demonstrated selective hypertrophy of the inner stripe of the outer medulla (ISOM) in rats with liver cirrhosis induced by common bile duct ligation (CBL). The present study was designed in order to measure Na-K-ATPase activity in the two major tubular segments located in the ISOM: the thick ascending limb of henles (MTAL) and the collecting ducts (OMCD) in CBL rats. Sham-operated rats were used as controls. In addition, the natriuretic response to amiloride (0.2 mg kg(-1) h(-1) i.v) was examined in conscious, chronically instrumented rats during conditions where amiloride-induced volume losses were replaced continuously using a servo-controlled i.v. volume replacement system. For 4-5 weeks after CBL, cirrhotic rats showed sodium retention relative to control rats without any sign of ascites. Plasma levels of sodium and aldosterone were normal, but plasma vasopressin was increased. Effective renal plasma flow was significantly increased, whereas glomerular filtration rate (GFR) and renal lithium handling were normal. The CBL rats showed a blunted natriuretic response to amiloride (DeltaFE(Na): 1.17 +/- 0.15% vs. 1.65 +/- 0.13%; P < 0.05). In rats with CBL, Na-K-ATPase activity per mm tubular length was decreased in the OMCD and unchanged in the TAL segment. These results suggest that increased tubular sodium reabsorption in liver cirrhotic rats with early sodium retention is localized in segments proximal to the collecting ducts.

Role of sodium depletion in acute antidiuretic effect of bendroflumethiazide in rats with nephrogenic diabetes insipidus.

The mechanisms underlying the acute antidiuretic response to bendroflumethiazide (BFTZ; 0.25 mg/h for 3 h) in rats with nephrogenic diabetes insipidus (NDI) was investigated. NDI was induced in conscious chronically instrumented female Wistar rats either by chronic lithium administration (40-60 mmol Li/kg of diet for 4 weeks) or by acute infusion of V2 antagonist OPC-31260 (0.2 mg/h). Renal clearance experiments were performed in conscious rats instrumented with permanent catheters. During experiments total body water content was held constant by i.v. replacement of urine production (V) with 150 mM glucose. One group in addition received i.v. replacement of urinary sodium losses. In both models of NDI, BFTZ-induced antidiuresis was associated with a decrease in the delivery of tubular fluid to the distal nephron, as measured by lithium clearance (C(Li)). Both the antidiuresis and the decrease in C(Li) could be prevented by sodium replacement. BFTZ did not affect distal water handling as measured by V/C(Li). BFTZ did not induce antidiuresis in normal rats with water diuresis. It is concluded that in rats with NDI, thiazide-induced antidiuresis can be entirely explained by a fall in distal delivery of tubular fluid related to sodium depletion. This contrasts the response in rats with central diabetes insipidus, where thiazides in addition increase distal water reabsorption.

Molecular epidemiology of calicivirus infections in Norway.

The national reference laboratory for calicivirus diagnostics monitors the epidemiology of calicivirus infections in Norway. During winter 1998-1999, 406 fecal samples were received from patients with suspected calicivirus infection. Of these, 76 (19%) were calicivirus positive by a nested reverse transcription-polymerase chain reaction. A number of alternative PCR designs were employed to disclose false negatives, but none were found. One half of the PCR positive samples were sequenced in order to investigate whether various cases represented the same outbreak, and to what extent a single or multiple subtypes were responsible for the morbidity during this season. The sequence data revealed that the majority of cases represented a genotype related to the Lordsdale strain, whereas the remaining cases seemed more sporadic. Most often, samples from particular outbreaks were highly homogeneous. However, in a few cases, samples connected with the same outbreak proved to contain epidemiologically independent strains.

Yeast COQ4 encodes a mitochondrial protein required for coenzyme Q synthesis.

The COQ4 gene coding for a component of the coenzyme Q biosynthetic pathway in the yeast Saccharomyces cerevisiae was cloned by a functional complementation of a Q-deficient mutant strain. Yeast coq4 mutant strains harboring the COQ4 gene on either single- or multicopy plasmids acquired the ability to grow on media containing a nonfermentable carbon source, synthesize Q(6), and respire. COQ4 encodes a polypeptide containing 335 amino acids with a calculated molecular mass of 38.6 kDa. By Western blot analysis with a specific antiserum, Coq4p was shown to peripherally associate with the matrix face of the mitochondrial inner membrane. The putative mitochondrial-targeting sequence present at the amino-terminus of the polypeptide efficiently imported it to mitochondria in a membrane-potential-dependent manner. Steady-state levels of COQ4 mRNA were increased during growth on glycerol-containing medium, in accordance with a function in Q biosynthesis. The function of Coq4p is unknown, although its presence is required to maintain a steady-state level of Coq7p, another component of the Q biosynthetic pathway. The results presented here, along with those available from literature, are discussed in light of the recently proposed existence of a multisubunit complex functioning in Q biosynthesis (A. Y. Hsu, T. Q. Do, P. T. Lee, and C. F. Clarke, 2000, Biochim. Biophys. Acta 1484, 287-297).

A defect in coenzyme Q biosynthesis is responsible for the respiratory deficiency in Saccharomyces cerevisiae abc1 mutants.

Ubiquinone (coenzyme Q or Q) is an essential component of the mitochondrial respiratory chain in eukaryotic cells. There are eight complementation groups of Q-deficient Saccharomyces cerevisiae mutants designated coq1-coq8. Here we report that COQ8 is ABC1 (for Activity of bc(1) complex), which was originally isolated as a multicopy suppressor of a cytochrome b mRNA translation defect (Bousquet, I., Dujardin, G., and Slonimski, P. P. (1991) EMBO J. 10, 2023-2031). Previous studies of abc1 mutants suggested that the mitochondrial respiratory complexes were thermosensitive and function inefficiently. Although initial characterization of the abc1 mutants revealed characteristics of Q-deficient mutants, levels of Q were reported to be similar to wild type. The suggested function of Abc1p was that it acts as a chaperone-like protein essential for the proper conformation and functioning of the bc(1) and its neighboring complexes (Brasseur, G., Tron, P., Dujardin, G., Slonimski, P. P. (1997) Eur. J. Biochem. 246, 103-111). Studies presented here indicate that abc1/coq8 null mutants are defective in Q biosynthesis and accumulate 3-hexaprenyl-4-hydroxybenzoic acid as the predominant intermediate. As observed in other yeast coq mutants, supplementation of growth media with Q(6) rescues the abc1/coq8 null mutants for growth on nonfermentable carbon sources. Such supplementation also partially restores succinate-cytochrome c reductase activity in the abc1/coq8 null mutants. Abc1/Coq8p localizes to the mitochondria, and is proteolytically processed upon import. The findings presented here indicate that the previously reported thermosensitivity of the respiratory complexes of abc1/coq8 mutants results from the lack of Q and a general deficiency in respiration, rather than a specific phenotype due to dysfunction of the Abc1 polypeptide. These results indicate that ABC1/COQ8 is essential for Q-biosynthesis and that the critical defect of abc1/coq8 mutants is a lack of Q.

Brain angiotensin receptors and sympathoadrenal regulation during insulin-induced hypoglycemia.

Simultaneous blockade of systemic AT1 and AT2 receptors or converting enzyme inhibition (CEI) attenuates the hypoglycemia-induced reflex increase of epinephrine (Epi). To examine the role of brain AT1 and AT2 receptors in the reflex regulation of Epi release, we measured catecholamines, hemodynamics, and renin during insulin-induced hypoglycemia in conscious rats pretreated intracerebroventricularly with losartan, PD-123319, losartan and PD-123319, or vehicle. Epi and norepinephrine (NE) increased 60-and 3-fold, respectively. However, the gain of the reflex increase in plasma Epi (Deltaplasma Epi/Deltaplasma glucose) and the overall Epi and NE responses were similar in all groups. The ensuing blood pressure response was similar between groups, but the corresponding bradycardia was augmented after PD-123319 (P < 0.05 vs. vehicle) or combined losartan and PD-123319 (P < 0.01 vs. vehicle). The findings indicate 1) brain angiotensin receptors are not essential for the reflex regulation of Epi release during hypoglycemia and 2) the gain of baroreceptor-mediated bradycardia is increased by blockade of brain AT2 receptors in this model.