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Activation of the GPCR sphingosine-1-phosphate receptor 1 (S1P1) by sphingosine-1-phosphate (S1P) regulates key physiological processes. S1P1 activation also has been implicated in pathologic processes, including autoimmunity and inflammation; however, the in vivo sites of S1P1 activation under normal and disease conditions are unclear. Here, we describe the development of a mouse model that allows in vivo evaluation of S1P1 activation. These mice, known as S1P1 GFP signaling mice, produce a S1P1 fusion protein containing a transcription factor linked by a protease cleavage site at the C terminus as well as a β-arrestin/protease fusion protein. Activated S1P1 recruits the β-arrestin/protease, resulting in the release of the transcription factor, which stimulates the expression of a GFP reporter gene. Under normal conditions, S1P1 was activated in endothelial cells of lymphoid tissues and in cells in the marginal zone of the spleen, while administration of an S1P1 agonist promoted S1P1 activation in endothelial cells and hepatocytes. In S1P1 GFP signaling mice, LPS-mediated systemic inflammation activated S1P1 in endothelial cells and hepatocytes via hematopoietically derived S1P. These data demonstrate that S1P1 GFP signaling mice can be used to evaluate S1P1 activation and S1P1-active compounds in vivo. Furthermore, this strategy could be potentially applied to any GPCR to identify sites of receptor activation during normal physiology and disease., Introduction Sphingosine-1-phosphate receptor 1 (S1P1), originally named EDG-1, is the founding member of a family of 5 GPCRs with high affinity for the lipid ligand sphingosine-1-phosphate (S1P) (1-3). Activation ofS1P1 [...]

Diabetes remains a major public health issue. According to the American Diabetes Association, 23.5 million, or 10.7% of people in the USA aged 20 years and older, have diabetes. Type-2 diabetes is treated both by controlling the diet and with oral hypoglycemic drugs. However, for many patients, achieving a tight control of glucose is difficult with current regimens. This chapter discusses a relatively low-cost dietary supplement that could be used as an adjuvant therapy for type-2 diabetes. A review of the literature indicates that cysteine-rich whey protein improves glucose metabolism in diabetic animals and type-2 diabetic patients. Similarly, in animal studies, improvement in glucose metabolism is observed after supplementation with L-cysteine, or molecules containing a cysteine moiety. This chapter discusses the biochemical mechanisms by which L-cysteine can upregulate the insulin-dependent signaling cascades of glucose metabolism. Further studies are needed to examine whether clinical interventions using L-cysteine as an adjuvant therapy indeed help to control glycemia and vascular inflammation in the diabetic patient population. Key words: L-cysteine, vascular inflammation, insulin resistance, whey protein, diabetes. Le diabete continue d'etre un enjeu majeur en sante publique. Selon l'American Diabetes Association, 23,5 millions ou 10,7% des personnes dans les E.-U. agees de 20 ans ou plus sont atteintes de diabete. Le diabete de type 2 est traite par la diete et par des medicaments hypoglycemiants oraux. Cependant, plusieurs patients eprouvent des difficultes a controler rigoureusement leur glucose avec les regimes actuels. Ce chapitre traite d' un supplement alimentaire relativement bon marche qui pourrait etre utilise comme adjuvant a la therapie du diabete de type 2. Une revue de la litterature indique que la proteine lactoserique riche en cysteine ameliore le metabolisme du glucose chez des animaux diabetiques et chez les patients atteints de diabete de type 2. De la meme facon, une amelioration du metabolisme du glucose est observee chez l'animal a la suite d'une supplementation en L-cysteine ou en molecules qui contiennent une entite L-cysteine. Ce chapitre discute des mecanismes biochimiques par lesquels la L-cysteine peut reguler a la hausse les cascades de signalisation du metabolisme du glucose dependantes de l'insuline. Des etudes additionnelles sont requises pour determiner si les interventions cliniques qui utilisent la L-cysteine comme adjuvant therapeutique aident effectivement a controler la glycemie et l'inflammation vasculaire chez les patients diabetiques. Mots-cles : L-cysteine, inflammation vasculaire, resistance a l'insuline, proteine lactoserique, diabete. [Traduit par la Redaction], Introduction Diabetes has become an epidemic, and remains a major public health issue worldwide. Intensive blood glucose control dramatically reduces the devastating complications that result from poorly controlled diabetes. However, [...]

The hypoxia-inducible factor (HIF) family of transcription factors directs a coordinated cellular response to hypoxia that includes the transcriptional regulation of a number of metabolic enzymes. Chuvash polycythemia (CP) is an autosomal recessive human disorder in which the regulatory degradation of HIF is impaired, resulting in elevated levels of HIF at normal oxygen tensions. Apart from the polycythemia, CP patients have marked abnormalities of cardiopulmonary function. No studies of integrated metabolic function have been reported. Here we describe the response of these patients to a series of metabolic stresses: exercise of a large muscle mass on a cycle ergometer, exercise of a small muscle mass (calf muscle) which allowed noninvasive in vivo assessments of muscle metabolism using 31p magnetic resonance spectroscopy, and a standard meal tolerance test. During exercise, CP patients had early and marked phosphocreatine depletion and acidosis in skeletal muscle, greater accumulation of lactate in blood, and reduced maximum exercise capacities. Muscle biopsy specimens from CP patients showed elevated levels of transcript for pyruvate dehydrogenase kinase, phosphofructokinase, and muscle pyruvate kinase. In cell culture, a range of experimental manipulations have been used to study the effects of HIF on cellular metabolism. However, these approaches provide no potential to investigate integrated responses at the level of the whole organism. Although CP is relatively subtle disorder, our study now reveals a striking regulatory role for HIF on metabolism during exercise in humans. These findings have significant implications for the development of therapeutic approaches targeting the HIF pathway. exercise | lactate | glycolysis | Chuvash polycythemia | von Hippel-Lindau doi/10.1073/pnas.1002339107

Evidence from macaque monkey tracing studies suggests connectivity-based subdivisions within the precuneus, offering predictions for similar subdivisions in the human. Here we present functional connectivity analyses of this region using resting-state functional MRI data collected from both humans and macaque monkeys. Three distinct patterns of functional connectivity were demonstrated within the precuneus of both species, with each subdivision suggesting a discrete functional role: (i) the anterior precuneus, functionally connected with the superior parietal cortex, paracentral Iobule, and motor cortex, suggesting a sensorimotor region; (ii) the central precuneus, functionally connected to the dorsolateral prefrontal, dorsomedial prefrontal, and multimodal lateral inferior parietal cortex, suggesting a cognitive/associative region; and (iii) the posterior precuneus, displaying functional connectivity with adjacent visual cortical regions. These functional connectivity patterns were differentiated from the more ventral networks associated with the posterior cingulate, which connected with limbic structures such as the medial temporal cortex, dorsal and ventromedial prefrontal regions, posterior lateral inferior parietal regions, and the lateral temporal cortex. Our findings are consistent with predictions from anatomical tracer studies in the monkey, and provide support that resting-state functional connectivity (RSFC) may in part reflect underlying anatomy. These subdivisions within the precuneus suggest that neuroimaging studies will benefit from treating this region as anatomically (and thus functionally) heterogeneous. Furthermore, the consistency between functional connectivity networks in monkeys and humans provides support for RSFC as a viable tool for addressing crossspecies comparisons of functional neuroanatomy. brain connectivity/functional MRI/posteromedial cortex/resting state doi/10.1073/pnas.0905314106

Results of several studies have confirmed the presence of brown adipose tissue among health human adults. The results are also indicative of the fact that activated brown adipose tissue has the potential to contribute substantially to energy expenditure, and thus, may be further researched as a means of treating obesity.

A study was conducted to investigate the importance and role of brown adipose tissue in relation to obesity and body-mass index among adult humans. Results indicated that the tissue played a vital role in adult human metabolism and was also found to be more prevalent among adult women more than adult men.

It has been difficult to establish whether we are limited to the heart muscle cells we are born with or if cardiomyocytes are generated also later in life. We have taken advantage of the integration of carbon-14, generated by nuclear bomb tests during the Cold War, into DNA to establish the age of cardiomyocytes in humans. We report that cardiomyocytes renew, with a gradual decrease from 1% turning over annually at the age of 25 to 0.45% at the age of 75. Fewer than 50% of cardiomyocytes are exchanged during a normal life span. The capacity to generate cardiomyocytes in the adult human heart suggests that it may be rational to work toward the development of therapeutic strategies aimed at stimulating this process in cardiac pathologies.

Allometric relationships are important sources of information for many types of anthropological and biological research. The baseline for all allometric relationships is isometry (or geometric similarity), the principal that shape is invariant of size. Here, we formally test for geometric similarity in modern humans, looking at the maximum lengths of four long bones (humerus, radius, femur, and tibia). We use Jolicoeur's multivariate allometry method to examine globally distributed samples of human populations, both collectively and individually. Results indicate that humans are not geometrically similar, although morphological deviations from isometry are small. KEY WORDS allometry; geometric similarity; long bones

The 5-hydroxytryptamine 3 ([5-HT.sub.3]) receptor is expressed widely in the central and peripheral nervous systems, where it mediates or modulates a wide range of physiological processes. The receptor is targeted by drugs administered for nausea and/or emesis and irritable bowel syndrome and has been proposed as a potential drug target in various psychiatric disorders. The [5-HT.sub.3] receptor is a pentameric ligand-gated ion channel and belongs to the Cys-loop receptor family. In contrast to the immense heterogeneity characterizing other Cysloop receptors, native [5-HT.sub.3] receptors historically have been considered a much more homogenous receptor population. However, the recent discovery of additional [5-HT.sub.3] subunits and the dawning realization that central and peripheral [5-HT.sub.3] receptor populations might comprise several subtypes characterized by distinct functional properties has emphasized the complexity of human [5-HT.sub.3] receptor signaling. In this review potential implications of these findings and of the entirely new layer of interindividual diversity introduced to the [5-HT.sub.3] receptor system by genetic variations will be outlined.

Abstract: High-frequency stimulation of peripheral nerve bundles is frequently used in clinical tests and physiologic experiments to study presynaptic and postsynaptic effects. To understand the postsynaptic effects, it is important [...]

Biomedical signal monitoring systems have been rapidly advanced with electronic and information technologies in recent years. However, most of the existing physiological signal monitoring systems can only record the signals without the capability of automatic analysis. In this paper, we proposed a novel brain--computer interface (BCI) system that can acquire and analyze electroencephalogram (EEG) signals in real-time to monitor human physiological as well as cognitive states, and, in turn, provide warning signals to the users when needed. The BCI system consists of a four-channel biosignal acquisition/amplification module, a wireless transmission module, a dual-core signal processing unit, and a host system for display and storage. The embedded dual-core processing system with multitask scheduling capability was proposed to acquire and process the input EEG signals in real time. In addition, the wireless transmission module, which eliminates the inconvenience of wiring, can be switched between radio frequency (RF) and Bluetooth according to the transmission distance. Finally, the real-time EEG-based drowsiness monitoring and warning algorithms were implemented and integrated into the system to close the loop of the BCI system. The practical online testing demonstrates the feasibility of using the proposed system with the ability of real-time processing, automatic analysis, and online warning feedback in real-world operation and living environments. Index Terms--Brain-computer interface (BCI), electroencephalogram (EEG), online, drowsiness detection, wireless.

1/f scaling has been observed throughout human physiology and behavior, but its origins and meaning remain a matter of debate. Some argue that it is a byproduct of ongoing processes in the brain or body and therefore of limited relevance to psychological theory. Others argue that 1/f scaling reflects a fundamental aspect of all physiological and cognitive functions, namely, that they emerge in the balance of independent versus interdependent component activities. In 4 experiments, series of key-press responses were used to test between these 2 alternative explanations. The critical design feature was to take 2 measures of each key-press response: reaction time and key-contact duration. These measures resulted in 2 parallel series of intrinsic fluctuations for each series of key-press responses. Intrinsic fluctuations exhibited 1/f scaling in both reaction times and key-contact durations, yet the 2 measures were uncorrelated with each other and separately perturbable. These and other findings indicate that 1/f scaling is too pervasive to be idiosyncratic and of limited relevance. It is instead argued that 1/f scaling reflects the coordinative, metastable basis of cognitive function. Keywords: 1/f noise and scaling, criticality, metastability, relative coordination, emergent cognitive function

Humans had reached Tasmania by 35 000 years bp and were in residence at the peak of the last ice age. Curiously, the settlements in the coldest period are concentrated [...]

Nedergaard J, Bengtsson T, Cannon B. Unexpected evidence for active brown adipose tissue in adult humans. Am J Physiol Endocrinol Metab 293: E444-E452, 2007. First published May 1, 2007; doi: 10.1152/ajpendo.00691.2006.--The contention that brown adipose tissue is absent in adult man has meant that processes attributed to active brown adipose tissue in experimental animals (mainly rodents), i.e., classical nonshivering thermogenesis, adaptive adrenergic thermogenesis, diet-induced thermogenesis, and antiobesity, should be either absent or attributed to alternative (unknown) mechanisms in man. However, serendipidously, as a consequence of the use of fluorodeoxyglucose positron emission tomography (FDG PET) to trace tumor metastasis, observations that may change that notion have recently been made. These tomography scans have visualized symmetrical areas of increased tracer uptake in the upper parts of the human body; these areas of uptake correspond to brown adipose tissue. We examine here the published observations from a viewpoint of human physiology. The human depots are somewhat differently located from those in rodents, the main depots being found in the supraclavicular and the neck regions with some additional paravertebral, mediastinal, para-aortic, and suprarenal localizations (but no interscapular). Brown adipose tissue activity in man is acutely cold induced and is stimulated via the sympathetic nervous system. The prevalence of active brown adipose tissue in normal adult man can be only indirectly estimated, but it would seem that the prevalence of active brown adipose tissue in the population may be at least in the range of some tens of percent. We conclude that a substantial fraction of adult humans possess active brown adipose tissue that thus has the potential to be of metabolic significance for normal human physiology as well as to become pharmaceutically activated in efforts to combat obesity. fluorodesoxyglucose; positron emission tomography; glucose uptake; nonshivering thermogenesis

It has been shown by Pawar et al. (2007) that the motion artifacts induced by body movement activity (BMA) in a single-lead wearable electrocardiogram (ECG) signal recorder, while monitoring an ambulatory patient, can be detected and removed by using a principal component analysis (PCA)-based classification technique. However, this requires the ECG signal to be temporally segmented so that each segment comprises of artifacts due to a single type of body movement activity. In this paper, we propose a simple, recursively updated PCA-based technique to detect transitions wherever the type of body movement is changed. Index Terms--Body movement activity, motion artifacts, principal component analysis, transition detection, wearable ECG.

Cholecystokinin (CCK), peptide YY (PYY), and ghrelin have been proposed to act as satiety hormones. CCK and PYY are stimulated during meal intake by the presence of nutrients in the small intestine, especially fat, whereas ghrelin is inhibited by eating. The sequence of events (fat intake followed by fat hydrolysis and CCK release) suggests that this process is crucial for triggering the effects. The aim of this study was therefore to investigate whether CCK mediated the effect of intraduodenal (ID) fat on ghrelin secretion and PYY release via CCK-1 receptors. Thirty-six male volunteers were studied in three consecutive, randomized, double-blind, cross-over studies: 1) 12 subjects received an ID fat infusion with or without 120 mg orlistat, an irreversible inhibitor of gastrointestinal lipases, compared with vehicle; 2) 12 subjects received ID long-chain fatty acids (LCF), ID medium-chain fatty acids (MCF), or ID vehicle; and 3) 12 subjects received ID LCF with and without the CCK-1 receptor antagonist dexloxiglumide (Dexlox) or ID vehicle plus intravenous saline (placebo). ID infusions were given for 180 min. The effects of these treatments on ghrelin concentrations and PYY release were quantified. Plasma hormone concentrations were measured in regular intervals by specific RIA systems. We found the following results. 1) ID fat induced a significant inhibition in ghrelin levels (P < 0.01) and a significant increase in PYY concentrations (P < 0.004). Inhibition of fat hydrolysis by orlistat abolished both effects. 2) LCF significantly inhibited ghrelin levels (P < 0.02) and stimulated PYY release (P < 0.008), whereas MCF were ineffective compared with controls. 3) Dexlox administration abolished the effect of LCF on ghrelin and on PYY. ID fat or LCF significantly stimulated plasma CCK (P < 0.006 and P < 0.004) compared with saline. MCF did not stimulate plasma CCK release. In summary, fat hydrolysis is essential to induce effects on ghrelin and PYY through the generation of LCF, whereas MCF are ineffective. Furthermore, LCF stimulated plasma CCK release, suggesting that peripheral CCK is the mediator of these actions. The CCK-1 receptor antagonist Dexlox abolished the effect of ID LCF, on both ghrelin and PYY. Generation of LCF through hydrolysis of fat is a critical step for fat-induced inhibition of ghrelin and stimulation of PYY in humans; the signal is mediated via CCK release and CCK-1 receptors. peptide YY; cholecystokinin

The ability of modified low-density lipoptoteins (LDLs) to induce both proliferation and death of endothelial cells is considered to be a mechanism of early atherosclerosis development. We previously showed that carbamylated LDL (cLDL) induces human coronary artery endothelial cell (HCAEC) death in vitro. This effect is similar to the atherogenic action of oxidized LDL (oxLDL) that induces the proliferation and death of endothelial cells. The present study was designed to analyze a potential proliferative effect of cLDL and whether proliferation caused by modified LDLs is related to cell death. Cultured HCAECs were exposed to different concentrations of modified LDL or native LDL for varying periods of time. Cell proliferation measured by bromodeoxyuridine incorporation and S-phase analysis was dosedependently increased in the presence of cLDL (6.25-200 [micro]g/ml). The proliferation induced by cLDL or oxLDL was associated with cell death and increased phosphorylation of extracellular signal-regulated kinase (ERK) and c-Jun N[H.sub.2]- terminal kinase (JNK). Inhibition of cLDL- or oxLDL-induced proliferation by aphidicolin (1 [micro]g/ml) was protective against both short-term cell death measured by lactate dehydrogenase release into the medium and long-term cell viability visualized by cell multiplication. Inhibition of ERK phosphorylation led to a significant decrease of DNA synthesis and cell rescue from injury by modified LDLs, while inhibition of JNK phosphorylation had an only partial rescue effect without involvement in cell proliferation. These data are the first evidence that endothelial cell death induced by cLDL or oxLDL is mediated by cell proliferation through the mitogen-activated protein kinase pathway. carbamylation; carbamylated low-density lipoprotein; oxidized low- density lipoprotein; mitogen-activated protein kinase; atherosclerosis

Insulin increases plasma nonesterified fatty acid (NEFA) clearance in humans, but whether this is independent of change in plasma NEFA appearance is currently unknown. Nine nondiabetic men (age: 28 [+ or -] 3 yr, body mass index: 27.2 [+ or -] 1.7 kg/[m.sup.2]) underwent euglycemic clamps to maintain low (LINS) vs. high (HINS) physiological insulin levels for 6 h. An intravenous infusion of hepafin + Intralipid (HI) was performed during 4 of the 6 h of the clamps (in the last 4 h at LINS and in the first 4 h at HINS), whereas saline infusion (SAL) was administered in the remaining 2 h to modulate plasma NEFA levels independently of plasma insulin levels. Four experimental conditions were obtained in each individual: LINS with saline (LINS/SAL) and with HI infusion (LINS/HI) and HINS with saline (HINS/SAL) and with HI infusion (HINS/HI). Plasma palmitate appearance during HINS/SAL was lower than during the three other experimental conditions (P < 0.05). In contrast, plasma linoleate appearance, as expected, was increased by HI independently of insulin level (P < 0.02). Plasma palmitate clearance during HINS/SAL was higher than LINS/SAL and LINS/HI (P < 0.008), and this increase was blunted during HINS/HI. We observed a linear decrease in plasma palmitate clearance with increasing plasma NEFA appearance independent of insulin levels. Plasma NEFA levels increased exponentially with increase in plasma NEFA appearance. We conclude that insulin stimulates plasma NEFA clearance by reducing the endogenous appearance rate of NEFA. The relationship between plasma NEFA level and appearance rate is nonlinear. human; insulin action; lipid metabolism; fatty acid metabolism

The research work on the genetic variants, which interact with environmental stimuli to alter aspects of human physical performance and related intermediate phenotypes, is presented.

A stereo-radiographic 3D reconstruction technique is used to investigate the external and internal body geometry of European adults in order to understand human anthropometry. The analysis has resulted in a unique geometrical database enabling improvement of numerical models of the human body for crash test simulation and has offered various possibilities in the anthropometry field.

The influence of the degrees of freedom of the body on postural stability in sitting postures is investigated. The results have showed that a model, taking into account only the degrees of freedom of the lumbar and thoracic spine and pelvis, looks like a better predictor of postural sway than the total number of degrees of freedom of the body.

Temporal and geographic variability in adult body size can be a useful indicator of a population's adaptation. The southern African Cape supported foraging populations exclusively until some pastoralism is seen, ca. 2000 BP. This paper describes and interprets body-size patterns among foragers, as deduced from maximum femoral lengths and femoral head diameters, using 127 individually dated adult skeletons from the western (70) and southern (57) regions of the Cape (64 male, 60 female, 3 sex undetermined). Estimated statures are comparable to historic Khoesan samples, but show lower values and greater variance during the fifth/ fourth millennium before the present among both sexes and both biomes. Variation in femoral length does not correlate with diet protein, as reflected in stable isotope ([[delta].sup.13]C and [[delta].sup.15]N) values. Positive correlations between femoral head diameter and isotopic indicators suggest greater body mass with more reliance on marine protein. A decline in femoral length begins at around 4000 BP, a time when archaeologists suggest that population growth led to the incorporation of lower-ranked food resources, and to reduced mobility. A clearly identifiable linear recovery, beginning at around 3000 BP, greatly predates the earliest evidence of pastoralism on the Cape. Apparent problems of food sufficiency were addressed and solved, within a hunting and gathering economy. KEY WORDS Later Stone Age; hunter-gatherers; Khoesan; bioarchaeology; stature; stable isotopes; paleodiet

Butyrate, a short-chain fatty acid, is the major energy fuel for the colonocytes. We have previously reported that monocarboxylate transporter isoform 1 (MCT1) mediates uptake of butyrate by human colonic Caco-2 cells. To better understand the mechanisms of MCT1 expression and regulation in the human intestine, we examined the activity and regulation of MCT1 promoter in Caco-2 cells. The transcription initiation site in the MCT1 promoter was identified as a guanine nucleotide 281 bp upstream from the translation initiation site and is surrounded by a guanine-cytosine-rich area. The promoter was found to be highly active when transfected into Caco-2 cells, and its activity decreased with deletions at its 5'-end. Gel mobility shift experiments showed binding of the transcription factors upstream stimulatory factor (USF)1 and 2 to the site -114 to -119 of the MCT1 promoter. With the use of site-directed mutagenesis and promoter activity in Caco-2 cells, the USF proteins appeared to have a repressor role on the MCT1 promoter, which was further confirmed by cotransfecting expression vectors encoding USF1 and 2 in Caco-2 cells and determining endogenous MCT1 expression in USE2 overexpressed cells. The two potential SP1 binding sites found in the same region of the promoter were found not to be involved in its regulation. short-chain fatty acids: transcriptional regulation; cis elements; mutagenesis; upstream stimulatory factors

We aimed to determine whether there were differences in the extent and time course of skeletal muscle myofibrillar protein synthesis (MPS) and muscle collagen protein synthesis (CPS) in human skeletal muscle in an 8.5-h period after bouts of maximal muscle shortening (SC; average peak torque = 225 [+ or -] 7 N*m, means [+ or -] SE) or lengthening contractions (LC; average peak torque = 299 [ + or -] 18 N*m) with equivalent work performed in each mode. Eight healthy young men (21.9 [+ or -] 0.6 yr, body mass index 24.9 [+ or -] 1.3 kg/[m.sup.2]) performed 6 sets of 10 maximal unilateral LC of the knee extensors on an isokinetic dynamometer. With the contralateral leg, they then performed 6 sets of maximal unilateral SC with work matched to the total work performed during LC (10.9 [+ or -] 0.7 vs. 10.9 [+ or -] 0.8 kJ, P = 0.83). After exercise, the participants consumed small intermittent meals to provide 0.1 g*[kg.sup.-1] * [h.sup.-1] of protein and carbohydrate. Prior exercise elevated MPS above rest in both conditions, but there was a more rapid rise after LC (P < 0.01). The increases (P < 0.001) in CPS above rest were identical for both SC and LC and likely represent a remodeling of the myofibrillar basement membrane. Therefore, a more rapid rise in MPS after maximal LC could translate into greater protein accretion and muscle hypertrophy during chronic resistance training utilizing maximal LC. eccentric; concentric; resistance exercise; z-band streaming

Myostatin inhibits myoblast proliferation and differentiation in developing muscle. Mounting evidence suggests that myostatin also plays a limiting role in growth/ repair/regeneration of differentiated adult muscle by inhibiting satellite cell activation. We tested the hypothesis that myostatin mRNA expression would decrease after resistance loading (RL) with a blunted response in older (O) females (F) who have shown minimal hypertrophy [vs. males (M)] after long-term RL. As myostatin is thought to modulate cell cycle activity, we also studied the response of gene transcripts key to stimulation (cyclin B1 and D1) and inhibition ([p21.sup.cip] and [p27.sup.kip]) of the cell cycle, along with the muscle-specific load-sensitive mitogen mechano-growth factor (MGF). Twenty young (Y; 20-35 yr, 10 YF, 10 YM) and 18 O (60-75 yr, 9 OF, 90M) consented to vastus lateralis biopsy before and 24 h after a bout of RL (3 sets x 8-12 repetitions to volitional fatigue of squat, leg press, knee extension). Gene expression levels were determined by relative RT-PCR with 18S as an internal standard and analyzed by age x gender x load repeated-measures ANOVA. A load effect was found for tour transcripts (P < 0.005) including myostatin, cyclin D1, [p27.sup.kip], and MGF as mRNA levels decreased for myostatin (-44%) and [p27.sup.kip] (-16%) and increased for cyclin D1 (34%) and MGF (49%). For myostatin, age x load and gender x load interactions (P < 0.05) were driven by a lack of change in OF, while marked declines were noted in YM (-56%), YF (-48%), and OM (-40%). Higher cyclin D1 levels in OF led to a main age effect (36%, O > Y) and an age x gender interaction (66%, OF > YF vs. 10%, OM > YM: P < 0.05). An age x gender x load interaction (P < 0.05) for cyclin D1 resulted from a 48% increase in OF. Post hoc testing within groups revealed a significant increase in MGF after RL in YM only (91%, P < 0.05). Higher levels of cyclin B1 in O (27%, O > Y) led to a main age effect (P < 0.05). An age x load interaction for cyclin B1 (P < 0.05) was driven by a 26% increase in Y with no change in O after RL. No age or gender differences, or load-mediated changes, were detected in levels of [p21.sup.cip] mRNA expression. These data clearly demonstrate that RL downregulates myostatin expression and alters genes key to cell cycle progression. However, failure to reduce myostatin expression may play a role in limiting RL-induced hypertrophy in OF. sarcopenia; mechano-growth factor; [p27.sup.kip]; aging; resistance exercise

Cellular and molecular regulation of renal biotin uptake in humans is not well defined. The contribution of the human [Na.sup.+]-dependent multivitamin transporter (hSMVT) to carrier-mediated biotin uptake by human proximal tubular epithelial cells is not clear. The aim of this study was to address these issues, with the human-derived proximal tubular epithelial HK-2 cells used as a model. First, we characterized the mechanism of biotin uptake by these cells and obtained evidence for involvement of an [Na.sup.+]-, temperature-, and energy-dependent carrier-mediated uptake system. This system was inhibited by the biotin structural analog desthiobiotin, pantothenic acid, and lipoate. These findings suggest involvement of the hSMVT system in the uptake process. This was confirmed by demonstrating that the hSMVT system is expressed in HK-2 cells at the protein and mRNA levels and by selective silencing of the hSMVT gene with the use of gene-specific small interfering RNAs, which led to specific and significant inhibition of carrier-mediated biotin uptake. Of the two recently cloned promoters of the hSMVT gene, promoter 1 was more active than promoter 2 in these cells. Pretreatment of HK-2 cells with modulators of PKC- and [Ca.sup.2+]/ calmodulin-mediated pathways (but not those that modulate PKA-, protein tyrosine kinase-, or nitric oxide-mediated pathways) led to significant alterations in biotin uptake. Maintaining the HK-2 cells in a biotin-deficient growth medium led to a marked upregulation in biotin transport, which was associated with an increase in hSMVT protein and RNA levels and an increase in activity of the hSMVT promoters. These results demonstrate that biotin uptake by human renal epithelial cells occurs via the hSMVT system and that the process is regulated by intracellular PKC- and [Ca.sup.2+]/calmodulin-mediated pathways. The uptake process appears to be adaptively regulated by extracellular biotin level, which involves transcriptional regulatory mechanism(s). transport regulation; renal biotin uptake; human sodium-dependent multivitamin transporter; human kidney epithelial cells

Proximal tubule cells extract dicarboxylates from filtrate and blood, using cotransporters located in the brush border [sodium dicarboxylate cotransporter (NaDC-1)] and basolateral cell membrane (NaDC-3). We expressed the human NaDC-3 (hNaDC-3) in Xenopus laevis oocytes and characterized it by the two-electrode voltage-clamp technique. At -60 mV, succinate (4 carbons) and glutarate (5 carbons) generated inward currents due to translocation of three sodium ions and one divalent dicarboxylate, whereas oxalate (2 carbons) and malonate (3 carbons) did not. The cis-dicarboxylate maleate produced currents smaller in magnitude, whereas the trans-dicarboxylate fumarate generated currents similar to succinate. The substituted succinate derivatives, malate, 2,2- and 2,3-dimethylsuccinate, and 2,3-dimercaptosuccinate elicited inward currents, whereas aspartate and guanidinosuccinate showed hardly detectable currents. The C-5 dicarboxylates glutarate and [alpha]-ketoglutarate produced larger currents than succinate; glutamate and folate failed to cause inward currents. Kinetic analysis revealed, at -60 mV, [K.sub.0.5] values of 25 [+ or -] 12 [micro]M for succinate and 45 [+ or -] 13 [micro]M for [alpha]-ketoglutarate, values close to the plasma concentration of these compounds. For both compounds, the [K.sub.0.5] was independent of voltage, whereas the maximal current increased with hyperpolarization. As opposed to the rat and flounder orthologs, hNaDC-3 was hardly inhibited by lithium concentrations up to 5 mM. In the absence of sodium, however, lithium can mediate succinate-dependent currents. The narrow substrate specificity prevents interaction of drugs with dicarboxylate-like structure with hNaDC-3 and ensures sufficient support of the proximal tubule cells with [alpha]-ketoglutarate for anion secretion via organic anion transporter 1 or 3. [alpha]-ketoglutarate; folate; kidney; tricarboxylic acid cycle intermediates; lithium sensitivity; succinate

This article describes the possible role of the endothelial cell-surface coat, containing proteoglycans (PGs) with connected glycosaminoglycans (GAGs), in maintaining glomerular permselectivity. Primary human glomerular endothelial cells (HGEC) in culture were treated with the nephrosis-inducing agent puromycin aminonucleoside (PAN). Analysis was made by TaqMan real-time PCR, Western blot analysis, and by metabolic labeling with [[sup.35]S]sulfate. The HGECs express several PGs: syndecan, versican, glypican, perlecan, decorin, and biglycan, which may contribute to the glomerular charge barrier. PAN treatment downregulated both the protein expression (by 25%) and the mRNA expression (by 37 [+ or -] 6%, P < 0.001, n = 8) of versican compared with control. Transferases important for chondroitin and heparan sulfate biosynthesis were also significantly downregulated by PAN, resulting in less sulfate groups, shorter GAG chains, and reduced PG net-negative charge. Moreover, analysis of the cell media after PAN treatment revealed a reduced content of [[sup.35]S]sulfate-labeled PGs (40% of control). We conclude that PAN may cause proteinuria by affecting the endothelial cell-surface layer and not only by disrupting the foot process arrangement of the podocytes. Thus the endothelium may be a more important component of the glomerular barrier than hitherto acknowledged. glomerular basal membrane; glycosaminoglycan; heparan sulfate; chondroitin sulfate; glycocalyx; nephrotic syndrome

Cerebral endothelial cells in the rat, pig, and, most recently, human have been shown to express several types of receptors specific for glutamate. High levels of glutamate disrupt the cerebral endothelial barrier via activation of N-methyl-D-aspartate (NMDA) receptors. We have previously suggested that this glutamate-induced barrier dysfunction was oxidant dependent. Here, we provide evidence that human cerebral endothelial cells respond to glutamate by generating an intracellular oxidant stress via NMDA receptor activation. Cerebral endothelial cells loaded with the oxidant-sensitive probe dihydrorhodamine were used to measure intracellular reactive oxygen species (ROS) formation in response to glutamate receptor agonists, antagonists, and second message blockers. Glutamate (1 mM) significantly increased ROS formation compared with sham controls (30 min). This ROS response was significantly reduced by 1) MK-801, a noncompetitive NMDA receptor antagonist; 2) 8-(N,N-diethylamino)-n-octyl-3,4,5- trimethoxybenzoate, an intracellular [Ca.sup.2+] antagonist; 3) La[Cl.sub.3], an extracellular [Ca.sup.2+] channel blocker; 4) diphenyleiodonium, a hemeferryl- containing protein inhibitor; 5) itraconazole, a cytochrome P-450 3A4 inhibitor; and 6) cyclosporine A, which prevents mitochondrial membrane pore transition required for mitochondrial-dependent ROS generation. Our results suggest that the cerebral endothelial barrier dysfunction seen in response to glutamate is [Ca.sup.2+] dependent and may require several intracellular signaling events mediated by oxidants derived from reduced nicotinamide adenine dinucleotide oxidase, cytochrome P-450, and the mitochondria. reactive oxygen species; mitochondria; reduced nicotinamide adenine dinucleotide oxidase; arachidonic acid; human; brain

Although in physiological conditions RR interval and systolic arterial pressure (SAP) are likely to interact in a closed loop, the traditional cross-spectral analysis cannot distinguish feedback (FB) from feedforward (FF) influences. In this study, a causal approach was applied for calculating the coherence from SAP to RR ([K.sub.s-r]) and from RR to SAP ([K.sub.r-s]) and the gain and phase of the baroreflex transfer function. The method was applied, compared with the noncausal one, to RR and SAP series taken from 15 healthy young subjects in the supine position and after passive head-up tilt. For the low frequency (0.04-0.15 Hz) spectral component, the enhanced FF coupling ([K.sub.r-s] = 0.59 [+ or -] 0.21, significant in 14 subjects) and the blunted FB coupling ([K.sub.s-r] = 0.17 [+ or -] 0.17, significant in 4 subjects) found at rest indicated the prevalence of nonbaroreflex mechanisms. The tilt maneuver recovered FB influences ([K.sub.s-r] = 0.47 [+ or -] 0.16, significant in 14 subjects), which were stronger than FF interactions ([K.sub.s-r] = 0.34 [+ or -] 0.19, significant in 9 subjects). At the respiratory frequency, the RR-SAP regulation was balanced at rest ([K.sub.s-r] = 0.30 [+ or -] 0.18 and [K.sub.r-s] = 0.29 [+ or -] 0.20, significant in 11 and 8 subjects) and shifted toward FB mechanisms after tilt ([K.sub.s-r] = 0.35 [+ or -] 0.19 and [K.sub.r-s] = 0.19 [+ or -] 0.11, significant in 14 and 8 subjects). The causal baroreflex gain estimates were always lower than the corresponding noncausal values and decreased significantly from rest to tilt in both frequency bands. The tilt-induced increase of the phase lag from SAP to RR suggested a shift from vagal to sympathetic modulation. Thus the importance of nonbaroreflex interactions pointed out the necessity of accounting for causality in the cross-spectral analysis of the interactions between cardiovascular variables in healthy humans. cross-spectral analysis; coherence and transfer function; cardiovascular regulation; feedback and feedforward mechanisms; nonbaroreflex interactions

Previous work indicates that sympathetic nerves participate in the vascular responses to direct cooling of the skin in humans. We evaluated this hypothesis further in a four-part series by measuring changes in cutaneous vascular conductance (CVC) from forearm skin locally cooled from 34 to 29[degrees]C for 30 min. In part 1. bretylium tosylate reversed the initial vasoconstriction (-14 [+ or -] 6.6% control CVC, first 5 rain) to one of vasodilation (+ 19.7 [+ or -] 7.7%) but did not affect the response at 30 rain (-30.6 [+ or -] 9% control, -38.9 [+ or -] 6.9% bretylium: both P < 0.05, P > 0.05 between treatments). In part 2, yohimbine and propranolol (YP) also reversed the initial vasoconstriction (14.3 [+ or -] 4.2% control) to vasodilation (+ 26.3 [+ or -] 12.1% YP), without a significant effect on the 30-rain response (-26.7 [+ or -] 6.1% YP, -43.2 [+ or -] 6.5% control: both P < 0.05, P > 0.05 between sites). In part 3, the NPY Y1 receptor antagonist BIBP 3226 had no significant effect on either phase of vasoconstriction (P > 0.05 between sites both times). In part 4, sensory nerve blockade by anesthetic cream (Em1a) also reversed the initial vasoconstriction (-20.1 [+ or -] 6.4% control) to one of vasodilation (+213.4 [+ or -] 87.0% Em1a), whereas the final levels did not differ significantly (37.7 [+ or -] 10.1% control, -37.2 [+ or -] 8.7% Em1a: both P < 0.05, P > 0.05 between treatments). These results indicate that local cooling causes cold-sensitive afferents to activate sympathetic nerves to release norepinephrine, leading to a local cutaneous vasoconstriction that masks a nonneurogenic vasodilation. Later, a vasoconstriction develops with or without functional sensory or sympathetic nerves. human: peripheral circulation: local control of blood flow: skin circulation: microdialysis; iontophoresis: neuropeptide Y; norepinephrine: axon reflex

We aimed to investigate the interaction between the arterial baroreflex and muscle metaboreflex [as reflected by alterations in the dynamic responses shown by leg blood flow (LBF: by the ultrasound Doppler method), leg vascular conductance (LVC), mean arterial blood pressure (MAP), and heart rate (HR)] in humans. In 12 healthy subjects (10 men and 2 women), who performed sustained 1-min handgrip exercise at 50% maximal voluntary contraction followed immediately by an imposed postexercise muscle ischemia (PEMI), 5-S periods of neck pressure (NP: 50 mmHg) or neck suction (NS: -60 mmHg) were used to evaluate carotid baroreflex function both at rest (Con) and during PEMI. First, the decreases in LVC and LBF and the augmentation of MAP elicited by NP were all greater during PEMI than in Con ([DELTA]LVC, -1.2 [+ or -] 0.2 vs. -1.9 [+ or -] 0.2 ml x [min.sup.-1] x mm[Hg.sup.-1]; [DELTA]LBF, 97.3 [+ or -] 11.2 vs. -177.0 [+ or -] 21.8 ml/min; [DELTA]MAP, 6.7 [+ or -] 1.2 vs. 11.5 [+ or -] 1.4 mmHg, Con vs. PEMI: each P < 0.05). Second, in Con, NS significantly increased both LVC and LBF (ALVC, 0.9 [+ or -] 0.2 ml x [min.sup.-1] x mm[Hg.sup.-1]; [DELTA]LBF, 46.6 [+ or -] 9.8 ml/min; significant change from baseline: each P < 0.057, and, whereas during PEMI no significant increases in LVC and LBF occurred during NS itself ([DELTA]LVC, 0.2 [+ or -] 0.1 ml x [min.sup.-1] x mm[Hg.sup.-1]; [DELTA]LBF, 10.8 [+ or -] 9.6 ml/min; each P > 0.05), a decrease was evident in each parameters at 5 s after the cessation of NS. Third, during PEMI. the decrease in MAP elicited by NS was smaller ([DELTA]MAP. -8.4 [+ or -] 1.0 vs. 5.8 [+ or -] 0.4 mmHg, Con vs. PEMI; P < 0.057, and it recovered to its initial level more quickly after NS (vs. Con). Finally, however, the HR responses to NS and NP were not different between PEMI and Con. These results suggest that during muscle metaboreflex activation in humans, the arterial baroreflex dynamic effect on peripheral vascular conductance is modulated, as exemplified by 1) an augmentation of the NP-induced LVC decrease, and 2) a loss of the NS-induced LVC increase. skeletal muscle metaboreflex: carotid baroreflex; exercise

Pulmonary emphysema results from an excessive degradation of lung parenchyma associated with a failure of alveolar repair. Secretion by pulmonary fibroblasts of hepatocyte growth factor (HGF) and keratinocyte growth factor (KGF) is crucial to an effective epithelial repair after lung injury. We hypothesized that abnormal HGF or KGF secretion by pulmonary fibroblasts could play a role in the development of emphysema. We measured in vitro production of HGF and KGF by human fibroblasts cultured from emphysematous and normal lung samples. HGF and KGF production was quantified at basal state and after stimulation. Intracellular content of HGF was lower in emphysema (1.52 pg/[micro]g, range of 0.15-7.40 pg/[micro]g) than in control fibroblasts (14.16 pg/[micro]g, range of 2.50-47.62 pg/[micro]g; P = 0.047). HGF production by emphysema fibroblasts (19.3 pg/[micro]g protein, range of 10.4-39.2 pg/[micro]g) was lower than that of controls at baseline (57.5 pg/[micro]g, range of 20.4-116 pg/[micro]g; P = 0.019) and after stimulation with interleukin-1[beta] or prostaglandin [E.sub.2]. Neither retinoic acids (alltrans and 9-cis) nor N-acetylcysteine could reverse this abnormality. KGF production by emphysema fibroblasts (5.3 pg/[micro]g, range of 2.2-9.3 pg/[micro]g) was similar to that of controls at baseline (2.6 pg/[micro]g, range of 1-6.1 pg/[micro]g; P = 0.14) but could not be stimulated with interleukin-1[beta]. A decreased secretion of HGF by pulmonary fibroblasts could contribute to the insufficient alveolar repair in pulmonary emphysema. pneumocytes; fibroblast growth factor-7; alveolar repair; retinoids

In human muscle the role of myoglobin (Mb) and its relationship to factors such as muscle perfusion and metabolic capacity are not well understood. We utilized nuclear magnetic resonance (NMR) to simultaneously study the Mb concentration ([Mb]), perfusion, and metabolic characteristics in calf muscles of athletes trained long term for either sprint or endurance running after plantar flexion exercise and cuff ischemia. The acquisitions for [sup.1]H assessment of Mb desaturation and concentration, arterial spin labeling measurement of muscle perfusion, and [sup.31]p spectroscopy to monitor high-energy phosphate metabolites were interleaved in a 4-T magnet. The endurance-trained runners had a significantly elevated [Mb] (0.28 [+ or -] 0.06 vs. 0.20 [+ or -] 0.03 mmol/kg). The time constant of creatine rephosphorylation ([tau]PCr), an indicator of oxidative capacity, was both shorter in the endurance-trained group (34 [+ or -] 6 vs. 64 [+ or -] 20 s) and negatively correlated with [Mb] across all subjects (r = 0.58). The time to reach maximal perfusion after cuff release was also both shorter in the endurance-trained group (306 [+ or -] 74 vs. 560 [+ or -] 240 s) and negatively correlated with [Mb] (r = 0.56). Finally, Mb reoxygenation rate tended to be higher in the endurance-trained group and was positively correlated with [tau]PCr (r = 0.75). In summary, these NMR data reveal that [Mb] is increased in human muscle with a high oxidative capacity and a highly responsive vasculature, and the rate at which Mb resaturates is well correlated with the rephosphorylation rate of Cr, each of which support a teleological role for Mb in [O.sub.2] transport within highly oxidative human skeletal muscle. postexercise hyperemia; skeletal muscle; perfusion; oxygenation; phosphocreatine

Byline: A. D. Nozdrachev (1), O. N. Mikhailova (2), E. L. Polyakov (3), M. S. Rudas (4) Author Affiliation: (1) St. Petersburg State University, St. Petersburg, 199034, Russia (2) Institute of Gerontology and Bioregulation, Russian Academy of Medical Sciences, St. Petersburg, Russia (3) Pavlov Institute of Physiology, St. Petersburg, Russia (4) Institute of the Human Brain, Russian Academy of Sciences, St. Petersburg, 197022, Russia Article History: Registration Date: 26/12/2004

We investigated the effects of nitric oxide (NO) on activity of the inwardly rectifying [K.sup.+] channel in cultured human proximal tubule cells, using the cell-attached mode of the patch-clamp technique. An inhibitor of NO synthases, [N.sup.[omega]]-nitro-L-arginine methyl ester (L-NAME; 100 [micro]M), reduced channel activity, which was restored by an NO donor, sodium nitroprusside (SNP; 10 [micro]M) or 8-bromo-cGMP (8-BrcGMP; 100 [micro]M). However, SNP failed to activate the channel in the presence of an inhibitor of soluble guanylate cyclase, 1H-[1,2,4]oxadiazon[4,3-a]quinoxalin-1-one (10 [micro]M). Similarly, the SNP effect was abolished by a protein kinase G (PKG)-specific inhibitor, KT-5823 (1 [micro]M), but not by a protein kinase A-specific inhibitor, KT-5720 (500 nM). Another NO donor, S-nitroso-N-acetyl-1D, L-penicillamine (10 [micro]M), mimicked the SNP-induced channel activation. In contrast to the stimulatory effect of SNP at a low dose (10 [micro]M), a higher dose of SNP (1 mM) reduced channel activity, which was not restored by 8-BrcGMP. Recordings of membrane potential with the slow whole cell configuration demonstrated that L-NAME (100 [micro]M) and the high dose of SNP (1 mM) depolarized the cell by 10.1 [+ or -] 2.6 and 9.2 [+ or -] 1.0 mV, respectively, whereas the low dose of SNP (10 [micro]M) hyperpolarized it by 7.1 [+ or -] 0.7 mV. These results suggested that the endogenous NO would contribute to the maintenance of basal activity of this [K.sup.+] channel and hence the potential formation via a cGMP/PKG-dependent mechanism, whereas a high dose of NO impaired channel activity independent of cGMP/PKG-mediated processes. patch-clamp; human kidney; guanosine 3',5'-cyclic monophosphate; sodium nitroprusside; protein kinase G

The myogenic response, the inherent ability of blood vessels to rapidly respond to changes in transmural pressure, is involved in local blood flow autoregulation. Animal studies suggest that aging impairs the myogenic response. The purpose of this study was to compare the effects of changes in transmural pressure on mean blood velocity (MBV, cm/s) in young and older subjects. Twelve younger men and women (25 [+ or -] 1 yr) were gender and body composition matched to twelve older men and women (65 [+ or -] 1 yr). A specially designed tank raised or lowered forearm pressure by 50 mmHg within 0.2 s. Brachial artery MBV was measured directly above the site of forearm pressure change using Doppler methods. In response to increasing transmural pressure (i.e., release of +50 mmHg), older subjects compared with younger subjects had significantly lower peak MBV ([DELTA] 12.43 [+ or -] 1.16 vs. [DELTA] 17.97 [+ or -] 2.01 cm/s; P < 0.05), reduced rates in the dynamic fall of MBV after peak values were achieved (vasoconstriction) (-1.88 [+ or -] 0.17 vs. -2.90 [+ or -] 0.28 cm x [s.sup.-1] x [s.sup.-1]; P < 0.05), and lower MBV values with sustained suction. In response to decreasing transmural pressure (i.e., change to +50 mmHg), there was a significantly greater increase in MBV (A peak flow from trough 7.71 [+ or -] 1.32 vs. 4.38 [+ or -] 0.71 cm/s; P < 0.05) and a trend toward a greater rate of rise in MBV (vasodilation; 1.61 [+ or -] 0.29 vs. 0.96 [+ or -] 0.21 cm x [s.sup.-1] x [s.sup.-1]; P = 0.08) in the older subjects. Older subjects compared with the younger subjects exhibited decreased dynamic vasoconstriction, enhanced steady-state constriction, as well as evidence for enhanced dynamic vasodilation responses to sustained alterations in forearm transmural pressure. regional blood flow; blood pressure; circulation

The presence of myofibroblasts arranged parallel to the longitudinal axes of anchoring villi of the placenta has previously been described. Furthermore, it has been suggested that intraplacental blood volume, and hence fetal-maternal oxygen-nutrient exchange, may in part be regulated through the longitudinal contraction of anchoring villi. We demonstrate here that anchoring villi have the ability to contract and relax longitudinally. Anchoring villi from normal term human placentae were dissected and suspended from force-displacement transducers to determine their longitudinal contractility in response to potassium chloride (KC1), [N.sup.[omega]]-nitro-L-arginine methyl ester (L-NAME) and the nitric oxide donors sodium nitroprusside (SNP) and glyceryl trinitrate (GTN). Treatment with both KCl and L-NAME resulted in up to a 62% and 74% increase, respectively, in longitudinal contraction over resting tone. In contrast, both SNP and GTN caused a dose-dependent relaxation of precontracted villi. Immunohistochemistry of longitudinal sections of villi confirmed the presence of [alpha]-actin-containing cells in the extravascular space. Histological staining with hemotoxylin and eosin confirm that the tissue used in these experiments were anchoring villi. These findings suggest that the contraction of anchoring villi may be an important mechanism whereby the placenta may regulate intraplacental volume. placenta; blood flow; smooth muscle

Above a certain level of cerebral activation the brain increases its uptake of glucose more than that of [O.sub.2], i.e., the cerebral metabolic ratio of [O.sub.2]/(glucose + 1/2 lactate) decreases. This study quantified such surplus brain uptake of carbohydrate relative to [O.sub.2] in eight healthy males who performed exhaustive exercise. The arterial-venous differences over the brain for [O.sub.2], glucose, and lactate were integrated to calculate the surplus cerebral uptake of glucose equivalents. To evaluate whether the amount of glucose equivalents depends on the time to exhaustion, exercise was also performed with [[beta].sub.1]-adrenergic blockade by metoprolol. Exhaustive exercise (24.8 [+ or -] 6.1 min; mean [+ or -] SE) decreased the cerebral metabolic ratio from a resting value of 5.6 [+ or -] 0.2 to 3.0 [+ or -] 0.4 (P < 0.05) and led to a surplus uptake of glucose equivalents of 9 [+ or -] 2 mmol. [[beta].sub.1]-blockade reduced the time to exhaustion (15.8 [+ or -] 1.7 min; P < 0.05), whereas the cerebral metabolic ratio decreased to an equally low level (3.2 [+ or -] 0.3) and the surplus uptake of glucose equivalents was not significantly different (7 [+ or -] 1 mmol; P = 0.08). A time-dependent cerebral surplus uptake of carbohydrate was not substantiated and, consequently, exhaustive exercise involves a brain surplus carbohydrate uptake of a magnitude comparable with its glycogen content. carbon dioxide; central fatigue; glucose; glycogen; lactate

Previous studies have provided evidence of a non-noradrenergic contributor to reflex cutaneous vasoconstriction in humans but did not identify the transmitter responsible. To test whether neuropeptide Y (NPY) has a role, in two series of experiments we slowly reduced whole body skin temperature ([T.sub.SK]) from 34.5 to 31.7[degrees]C. In protocol 1, Ringer solution and the NPY receptor antagonist BIBP-3226 alone were delivered intradermally via microdialysis. In protocol 2, yohimbine plus propranolol (Yoh + Pro), Yoh + Pro in combination with BIBP-3226, and Ringer solution were delivered to antagonize locally the vasomotor effects of NPY and norepinephrine. Blood flow was measured by laser Doppler flowmetry (LDF). Mean arterial blood pressure (MAP) was monitored at the finger (Finapres). In protocol 1, cutaneous vascular conductance (CVC) fell by 45%, to 55.1 [+ or -] 5.6% of baseline at control sites (P < 0.05). At BIBP-3226-treated sites, CVC fell by 34.1% to 65.9 [+ or -] 5.0% (P < 0.05; P < 0.05 between sites). In protocol 2, during body cooling, CVC at control sites fell by 32.6%, to 67.4 [+ or -] 4.3% of baseline; at sites treated with Yoh + Pro, CVC fell by 18.7%, to 81.3 [+ or -] 4.4% of baseline (P < 0.05 vs. baseline; P < 0.05 vs. control) and did not fall significantly at sites treated with BIBP-3226 + Yoh + Pro (P > 0.05; P < 0.05 vs. other sites). After cooling, exogenous norepinephrine induced vasoconstriction at control sites (P < 0.05) but not at sites treated with Yoh + Pro + BIBP-3226 (P > 0.05). These results indicate that NPY participates in sympathetically mediated cutaneous vasoconstriction in humans during whole body cooling. sympathetic nervous system; cotransmitters; cold stress; skin blood flow

Activation of the chemokine receptor CXCR3 by its cognate ligands induces several differentiated cellular responses important to the growth and migration of a variety of hematopoietic and structural cells. In the human respiratory tract, human airway epithelial cells (HAEC) release the CXCR3 ligands Mig/CXCL9, IP-10/CXCL10, and I-TAC/CXCL11. Simultaneous expression of CXCR3 by HAEC would have important implications for the processes of airway inflammation and repair. Accordingly, in the present study we sought to determine whether HAEC also express the classic CXCR3 chemokine receptor CXCR3-A and its splice variant CXCR3-B and hence may respond in autocrine fashion to its ligands. We found that cultured HAEC (16-HBE and tracheocytes) constitutively expressed CXCR3 mRNA and protein. CXCR3 rnRNA levels assessed by expression array were -35% of [beta]-actin expression. In contrast, CCR3, CCR4, CCRS, CCR8, and CX3CR1 were G protein-coupled receptors; chemotaxis; inflammation; lung; chronic obstructive pulmonary disease; CXC receptor 3

In preparation for birth, lung epithelia must switch from net fluid secretion, required for lung development, to net absorption, which prepares the lungs for postnatal gas exchange. The apical membrane amiloride-sensitive epithelial Na channel (ENaC) is the rate-limiting step for [Na.sup.+] and fluid absorption. Expression of [alpha]-ENaC mRNA has been detected in human lung as early as the embryonic stage of development. However, humans express multiple transcripts for [alpha]-ENaC, containing differing 5'-untranslated regions (UTR) with unknown effects on protein translation, and different ontogenies for individual transcripts could provide a novel mechanism for developmental regulation of ENaC function. To assess the relative expression of the two most abundant [alpha]-ENaC transcripts ([alpha]-ENaC1 and [alpha]-ENaC2) during lung development, we performed nonradioactive in situ hybridization using probes specific to the alternative 5'-UTRs. Both transcripts were expressed throughout intrauterine lung development (8 to 40 wk gestation), and expression was localized to the surface epithelial cells of the conductive and respiratory airways in both ciliated cells and nonciliated Clara cells. [alpha]-ENaC mRNA expression was also identified in the serous cells of the submucosal glands surrounding the proximal airways. In the mature prenatal lung, subsets of alveolar type II (ATII) cells expressed one or both of the [alpha]-ENaC transcripts. Our observations demonstrate that a developmentally regulated switch between [alpha]-ENaC 5'-UTR variants is not the trigger by which the developing human lung becomes a fluid-absorbing organ at birth, that individual ATII cells express neither, one, or both of the [alpha]-ENaC transcripts, and that the overall expression is linked to epithelial cell differentiation and lung maturation. fluid absorption; lung maturation; postnatal gas exchange

Transforming growth factor-[beta]1 (TGF-[beta]1) belongs to a family of multifunctional cytokines that regulate a variety of biological processes, including cell differentiation, proliferation, and apoptosis. The effects of TGF-[beta]1 are cell context and cell cycle specific and may be signaled through several pathways. We examined the effect of TGF-[beta]1 on apoptosis of primary human central airway epithelial cells and cell lines. TGF-[beta]1 protected human airway epithelial cells from apoptosis induced by either activation of the Fas death receptor (CD95) or by corticosteroids. This protective effect was blocked by inhibition of the Smad pathway via overexpression of inhibitory Smad7. The protective effect is associated with an increase in the cyclin-dependent kinase inhibitor p21 and was blocked by the overexpression of key gatekeeper cyclins for the [G.sub.1]/S interface, cyclins D1 and E. Blockade of the Smad pathway by overexpression of the inhibitory Smad7 permitted demonstration of a TGF-[beta]-mediated proapoptotic pathway. This proapoptotic effect was blocked by inhibition of the p38 MAPK kinase signaling with the inhibitor SB-203580 and was associated with an increase in p38 activity as measured by a kinase assay. Here we demonstrate dual signaling pathways involving TGF-[beta]1, an antiapoptotic pathway mediated by the Smad pathway involving p21, and an apoptosis-permissive pathway mediated in part by p38 MAPK. cyclin-dependent kinase; cyclin-dependent kinase inhibitor; extracellular regulated kinase; transforming growth factor-[beta] receptor; airway epithelium; apoptosis

The widespread presence of the Na-K-2Cl (NKCC) cotransporter protein suggests that chronic administration of inhibitors may result in adverse effects. Inhibition of the NKCC cotransporter by loop diuretics is felt to underlie the diuretic and the pulmonary smooth muscle relaxant effects of this drug class. However, the fundamental regulation of salt and water movement by this cotransporter suggests that it may also mediate cell volume changes occurring during cell cycle progression. Thus we hypothesized that NKCC cotransporter inhibition by loop diuretics would decrease cellular proliferation. Normal human bronchial smooth muscle cells (BSMC) showed a significant concentration-dependent decrease in cell counts after 7 days of exposure to both bumetanide (n = 5-10) and furosemide (n = 6-16) compared with controls. Proliferation was similarly inhibited in normal human lung fibroblasts (n = 5-9). To determine whether this was due to loss of cells, we performed apoptosis assays on BSMC. Both annexin V-propidinm iodide staining (n = 5-10) and single cell gel electrophoresis assays (n = 4) were negative for necrosis and apoptosis in BSMC exposed to 10 [micro]M bumetanide. Subsequent analysis of the cell cycle by flow cytometry showed that bumetanide-exposed BSMC were delayed in [G.sub.1] phase compared with controls (n = 4-8). This is the first evidence for loop diuretic inhibition of airway smooth muscle cell proliferation. NKCC cotransporter inhibition impeded [G.sub.1]-S phase transition without facilitating cell death. Thus although inhibition by loop diuretics relaxes airway smooth muscle, the NKCC cotransporter may have a more important role in cell proliferation regulation. furosemide; bumetanide

Intestinal mucosal cells and invading leukocytes produce inappropriate levels of cytokines and chemokines in human colitis. However, smooth muscle cells of the airway and vasculature also synthesize cytokines and chemokines. To determine whether human colonic myocytes can synthesize proinflammatory mediators, strips of circular smooth muscle and smooth muscle cells were isolated from human colon. Myocytes and muscle strips were stimulated with 10 ng/ml of IL-1[beta], TNF-[alpha], and IFN-[gamma], respectively. Expression of mRNA for IL-1[beta], IL-6, IL-8, and cyclooxygenase-2 (COX-2) was induced within 2 h and continued to increase for 8-12 h. Regulated on activation, normal T cell-expressed and -secreted (RANTES) mRNA expression was slower, appearing at 8 h and increasing linearly through 20 h. Expression of all five mRNAs was inhibited by 0.1 [micro]M MG-132, a proteosome inhibitor that blocks NF-[kappa]B activation. Expression of IL-1[beta], IL-6, IL-8, and COX-2 mRNA was reduced by 30 [micro]M PP1, an Src family tyrosine kinase inhibitor, and by 25 [micro]M SB-203580, a p38 MAPK inhibitor. MAPK/extracellular regulated kinase-1 inhibitor PD-98059 (25 [micro]M) was much less effective. In conclusion, human colonic smooth muscle cells can synthesize and secrete interleukins (IL-1[beta] and IL-6) and chemokines (IL-8 and RANTES) and upregulate expression of COX-2. Regulation of cytokine, chemokine, and COX-2 mRNA depends on multiple signaling pathways, including Src-family kinases, extracellular regulated kinase, p38 MAPKs, and NF-[kappa]B. SB-203580 was a consistent, efficacious inhibitor of inflammatory gene expression, suggesting an important role of p38 MAPK in synthetic functions of human colonic smooth muscle. cyclooxygenase; cytokine synthesis; mitogen-activated protein kinase-activated protein kinase; nuclear factor-[kappa]B; p38 mitogen-activated protein kinase

Because of attentional limitations, the human visual system can process for awareness and response only a fraction of the input received. Lesion and functional imaging studies have identified frontal, temporal, and parietal areas as playing a major role in the attentional control of visual processing, but very little is known about how these areas interact to form a dynamic attentional network. We hypothesized that the network communicates by means of neural phase synchronization, and we used magnetoencephalography to study transient long-range interarea phase coupling in a well studied attentionally taxing dual-target task (attentional blink). Our results reveal that communication within the fronto-parieto-temporal attentional network proceeds via transient long-range phase synchronization in the beta band. Changes in synchronization reflect changes in the attentional demands of the task and are directly related to behavioral performance. Thus, we show how attentional limitations arise from the way in which the subsystems of the attentional network interact.

This comparison of morphological and neutral genetic variation in 10 human populations was designed to test a neutral hypothesis of cranial evolution in living and recent humans and to explain deviations from neutrality where detected. Overall, among-population differences in extant Homo sapiens cranial morphology are proportional to among-population differences in neutral molecular characteristics. For most of the populations studied, cranial morphology varies among regions in a manner consistent with neutral expectations. Removal of the effects of shared population history and structure by using the partial Mantel's test, however, does not remove the correlation between some aspects of cranial morphology and a measure of coldness of climate. The excess differentiation is most apparent in those population comparisons that involve a Siberian population living in an extremely cold environment. This finding suggests the action of natural selection, associated with regional variation in temperature, leading to among-population differentiation in excess of neutral expectations for some cranial dimensions. Those dimensions reflect the breadth of the skull, cranial vault size and shape, and aspects of nasal morphology. Although morphology for most of the world appears to vary among populations in accordance with neutral expectations in the context of population structure and history, morphology of the Siberian population appears to have undergone adaptation by natural selection.