bluuebunny/crossref_metadata_2025 huggingface dataset:
Ca2+ spark sites in smooth muscle cells are numerous and differ in number of ryanodine receptors, large-conductance K+ channels, and coupling ratio between them
Authors: ZhuGe Ronghua , Fogarty Kevin E. , Baker Stephen P. , McCarron John G. , Tuft Richard A. , Lifshitz Lawrence M. , Walsh John V.
Published: August 2004

Ca2+ sparks are highly localized Ca2+ transients caused by Ca2+ release from sarcoplasmic reticulum through ryanodine receptors (RyR). In smooth muscle, Ca2+ sparks activate nearby large-conductance, Ca2+-sensitive K+ (BK) channels to generate spontaneous transient outward currents (STOC). The properties of individual sites that give rise to Ca2+ sparks have not been examined systematically. We have characterized individual sites in amphibian gastric smooth muscle cells with simultaneous high-speed imaging of Ca2+ sparks using wide-field digital microscopy and patch-clamp recording of STOC in whole cell mode. We used a signal mass approach to measure the total Ca2+ released at a site and to estimate the Ca2+ current flowing through RyR [ ICa(spark)]. The variance between spark sites was significantly greater than the intrasite variance for the following parameters: Ca2+ signal mass, ICa(spark), STOC amplitude, and 5-ms isochronic STOC amplitude. Sites that failed to generate STOC did so consistently, while those at the remaining sites generated STOC without failure, allowing the sites to be divided into STOC-generating and STOC-less sites. We also determined the average number of spark sites, which was 42/cell at a minimum and more likely on the order of at least 400/cell. We conclude that 1) spark sites differ in the number of RyR, BK channels, and coupling ratio of RyR-BK channels, and 2) there are numerous Ca2+ spark-generating sites in smooth muscle cells. The implications of these findings for the organization of the spark microdomain are explored.
Behavior of Ca2+ waves in multicellular preparations from guinea pig ventricle
Authors: Kurebayashi Nagomi , Yamashita Haruyo , Nakazato Yuji , Daida Hiroyuki , Ogawa Yasuo
Published: August 2004

Ca+ waves have been implicated in Ca2+ overload-induced cardiac arrhythmias. To deepen understanding of the behavior of Ca2+ waves in a multicellular system, consecutive two-dimensional Ca2+ images were obtained with a confocal microscope from surface cells of guinea pig ventricular papillary muscles loaded with fluo 3 or rhod 2. In intact muscles, no Ca2+ waves were detected under the resting condition, whereas they were frequently observed during the rest immediately after high-frequency stimulations where cytoplasmic Ca2+ concentration and Ca2+ stored in the sarcoplasmic reticulum (SR) were gradually decreasing. The intervals of Ca2+ waves increased as they occurred later, their amplitudes and velocities remaining unchanged. A SERCA inhibitor reversibly prolonged the wave intervals. In Na+-free/Ca2+-free medium where neither Ca2+ influx nor Na+/Ca2+ exchange took place, recurrent Ca2+ waves emerged at constant intervals in each cell. These results are consistent with the conclusion that the loading level of the SR is critical for induction of Ca2+ waves. Each cell independently exhibited its own regular rhythm of Ca2+ wave with a distinct interval. These waves propagated in either direction along the longitudinal axis within a muscle cell, but seldom beyond the cell boundary. In contrast, in partially damaged muscles that showed spontaneous Ca2+ waves at rest in normal Krebs solution, their propagation often was unidirectional, decreasing in frequency. In these cases, however, Ca2+ waves rarely moved beyond the cellular boundary. The gradient of the cytoplasmic Ca2+ concentration was suggested to be the cause of the one-way propagation.
Transforming growth factor-β1 signaling contributes to development of smooth muscle cells from embryonic stem cells
Authors: Sinha Sanjay , Hoofnagle Mark H. , Kingston Paul A. , McCanna Mary E. , Owens Gary K.
Published: August 2004

Knockout of transforming growth factor (TGF)-β1 or components of its signaling pathway leads to embryonic death in mice due to impaired yolk sac vascular development before significant smooth muscle cell (SMC) maturation occurs. Thus the role of TGF-β1 in SMC development remains unclear. Embryonic stem cell (ESC)-derived embryoid bodies (EBs) recapitulate many of the events of early embryonic development and represent a more physiological context in which to study SMC development than most other in vitro systems. The present studies showed induction of the SMC-selective genes smooth muscle α-actin (SMαA), SM22α, myocardin, smoothelin-B, and smooth muscle myosin heavy chain (SMMHC) within a mouse ESC-EB model system. Significantly, SM2, the SMMHC isoform associated with fully differentiated SMCs, was expressed. Importantly, the results showed that aggregates of SMMHC-expressing cells exhibited visible contractile activity, suggesting that all regulatory pathways essential for development of contractile SMCs were functional in this in vitro model system. Inhibition of endogenous TGF-β with an adenovirus expressing a soluble truncated TGF-β type II receptor attenuated the increase in SMC-selective gene expression in the ESC-EBs, as did an antibody specific for TGF-β1. Of interest, the results of small interfering (si)RNA experiments provided evidence for differential TGF-β-Smad signaling for an early vs. late SMC marker gene in that SMαA promoter activity was dependent on both Smad2 and Smad3 whereas SMMHC activity was Smad2 dependent. These results are the first to provide direct evidence that TGF-β1 signaling through Smad2 and Smad3 plays an important role in the development of SMCs from totipotential ESCs.
Genistein stimulates electrogenic Cl− secretion in mouse jejunum
Authors: Baker Michael J. , Hamilton Kirk L.
Published: August 2004

We used the short-circuit current ( Isc) technique to investigate the effects of the isoflavone genistein on the electrogenic Cl− secretion of the mouse jejunum. Genistein stimulated a sustained increase in Isc that was dose dependent. Bumetanide inhibited 76 ± 5% of the genistein-stimulated Isc consistent with activation of Cl− secretion. Genistein failed to stimulate Isc following maximal activation of the cAMP pathway by forskolin. In addition, forskolin had a reduced effect on Isc of the mouse jejunum in the presence of genistein. Glibenclamide, a blocker of CFTR, eliminated the genistein-stimulated increase of Isc and reduced the forskolin-activated Isc. Clotrimazole, a Ca2+-activated K+ channel blocker, failed to reduce the genistein-stimulated Isc. Vanadate, a blocker of tyrosine-dependent phosphatases, reduced the genistein-activated Isc. Tyrphostin A23, a tyrosine kinase inhibitor, reduced basal Isc, after which genistein failed to stimulate Isc. These data suggest that genistein activated a sustained Cl− secretory response of the mouse jejunum and that the effect of genistein was via a tyrosine-dependent phosphorylation pathway.
Kinetic properties of myosin heavy chain isoforms in mouse skeletal muscle: comparison with rat, rabbit, and human and correlation with amino acid sequence
Authors: Andruchov Oleg , Andruchova Olena , Wang Yishu , Galler Stefan
Published: August 2004

Stretch activation kinetics were investigated in skinned mouse skeletal muscle fibers of known myosin heavy chain (MHC) isoform content to assess kinetic properties of different myosin heads while generating force. The time to peak of stretch-induced delayed force increase ( t3) was strongly correlated with MHC isoforms [ t3 given in ms for fiber types containing specified isoforms; means ± SD with n in parentheses: MHCI 680 ± 108 ( 13 ), MHCIIa 110.5 ± 10.7 ( 23 ), MHCIIx(d) 46.2 ± 5.2 ( 20 ), MHCIIb 23.5 ± 3.3 (76)]. This strong correlation suggests different kinetics of force generation of different MHC isoforms in the following order:MHCIIb > MHCIIx(d) > MHCIIa ≫ MHCI. For rat, rabbit, and human skeletal muscles the same type of correlation was found previously. The kinetics decreases slightly with increasing body mass. Available amino acid sequences were aligned to quantify the structural variability of MHC isoforms of different animal species. The variation in t3 showed a correlation with the structural variability of specific actin-binding loops (so-called loop 2 and loop 3) of myosin heads ( r = 0.74). This suggests that alterations of amino acids in these loops contribute to the different kinetics of myosin heads of various MHC isoforms.
Endothelial progenitor cells for postnatal vasculogenesis
Authors: Asahara Takayuki , Kawamoto Atsuhiko
Published: August 2004

In the past decade, researchers have defined committed stem or progenitor cells from various tissues, including bone marrow, peripheral blood, brain, liver, and reproductive organs, in both adult animals and humans. Whereas most cells in adult organs are composed of differentiated cells, which express a variety of specific phenotypic genes adapted to each organ's environment, quiescent stem or progenitor cells are maintained locally or in the systemic circulation and are activated by environmental stimuli for physiological and pathological tissue regeneration. Recently, endothelial progenitor cells (EPCs) were isolated from peripheral blood CD34, Flk-1, or AC133 antigen-positive cells, which are considered to include a hematopoietic stem cell population, and were shown to be incorporated into foci of neovascularization. This finding, that circulating EPCs may home to sites of neovascularization and differentiate into endothelial cells in situ, is consistent with “vasculogenesis,” a critical paradigm for embryonic neovascularization, and suggests that vasculogenesis and angiogenesis may constitute complementary mechanisms for postnatal neovascularization. Previous reports demonstrating therapeutic potential of EPC transplantation in animal models of hindlimb and myocardial ischemia opened the way to the clinical application of cell therapy: the replacement of diseased or degenerating cell populations, tissues, and organs. In this review, we summarize biological features of EPCs and speculate on the utility of EPCs for vascular and general medicine.
Three distinct mechanisms of HCO3− secretion in rat distal colon
Authors: Vidyasagar Sadasivan , Rajendran Vazhaikkurichi M. , Binder Henry J.
Published: August 2004

HCO3− secretion has long been recognized in the mammalian colon, but it has not been well characterized. Although most studies of colonic HCO3− secretion have revealed evidence of lumen Cl− dependence, suggesting a role for apical membrane Cl−/HCO3− exchange, direct examination of HCO3− secretion in isolated crypt from rat distal colon did not identify Cl−-dependent HCO3− secretion but did reveal cAMP-induced, Cl−-independent HCO3− secretion. Studies were therefore initiated to determine the characteristics of HCO3− secretion in isolated colonic mucosa to identify HCO3− secretion in both surface and crypt cells. HCO3− secretion was measured in rat distal colonic mucosa stripped of muscular and serosal layers by using a pH stat technique. Basal HCO3− secretion (5.6 ± 0.03 μeq·h−1·cm−2) was abolished by removal of either lumen Cl− or bath HCO3−; this Cl−-dependent HCO3− secretion was also inhibited by 100 μM DIDS (0.5 ± 0.03 μeq·h−1·cm−2) but not by 5-nitro-3-(3-phenylpropyl-amino)benzoic acid (NPPB), a Cl− channel blocker. 8-Bromo-cAMP induced Cl−-independent HCO3− secretion (and also inhibited Cl−-dependent HCO3− secretion), which was inhibited by NPPB and by glibenclamide, a CFTR blocker, but not by DIDS. Isobutyrate, a poorly metabolized short-chain fatty acid (SCFA), also induced a Cl−-independent, DIDS-insensitive, saturable HCO3− secretion that was not inhibited by NPPB. Three distinct HCO3− secretory mechanisms were identified: 1) Cl−-dependent secretion associated with apical membrane Cl−/HCO3− exchange, 2) cAMP-induced secretion that was a result of an apical membrane anion channel, and 3) SCFA-dependent secretion associated with an apical membrane SCFA/HCO3− exchange.
Extracellular acidification elicits a chloride current that shares characteristics with ICl(swell)
Authors: Nobles Muriel , Higgins Christopher F. , Sardini Alessandro
Published: August 2004

A Cl− current activated by extracellular acidification, ICl(pHac), has been characterized in various mammalian cell types. Many of the properties of ICl(pHac) are similar to those of the cell swelling-activated Cl− current ICl(swell): ion selectivity (I− > Br− > Cl− > F−), pharmacology [ ICl(pHac) is inhibited by 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS), 1,9-dideoxyforskolin (DDFSK), diphenylamine-2-carboxylic acid (DPC), and niflumic acid], lack of dependence on intra- or extracellular Ca2+, and presence in all cell types tested. ICl(pHac) differs from ICl(swell) in three aspects: 1) its rate of activation and inactivation is very much more rapid, currents reaching a maximum in seconds rather than minutes; 2) it exhibits a slow voltage-dependent activation in contrast to the fast voltage-dependent activation and time- and voltage-dependent inactivation observed for ICl(swell); and 3) it shows a more pronounced outward rectification. Despite these differences, study of the transition between the two currents strongly suggests that ICl(swell) and ICl(pHac) are related and that extracellular acidification reflects a novel stimulus for activating ICl(swell) that, additionally, alters the biophysical properties of the channel.
Lipids: potential regulators of nitric oxide generation
Authors: Chikani Gentle , Zhu Weifei , Smart Eric J.
Published: August 2004

The endothelium is a dynamic organ that secretes several biologically active substances and plays a major functional role in the health of an organism in both physiological and pathological conditions. For instance, the endothelium is involved in control of the exchange of plasma and tissue biomolecules, regulation of vessel tone, inflammation, lipid metabolism, vessel growth and remodeling, and modulation of coagulation and fibrinolysis ( 6 , 12 , 15 ). The endothelium generates nitric oxide, which is a key regulator of vasodilation and plays important roles in preventing, or in some cases promoting, numerous cardiovascular diseases ( 41 ). Several recent studies have examined the interplay between lipids and nitric oxide generation, especially in relation to atherosclerosis ( 14 , 29 , 34 , 36 , 40 ). The endothelium is continuously exposed to circulating lipids in the form of lipoproteins and protein carriers that may have a direct impact on nitric oxide synthesis and function. The purpose of this review is to illustrate some of the recent findings that link lipids (plasma and cellular) to nitric oxide generation (see Fig. 1 ).
First-phase insulin secretion: does it exist in real life? Considerations on shape and function
Authors: Caumo Andrea , Luzi Livio
Published: August 2004

To fulfill its preeminent function of regulating glucose metabolism, insulin secretion must not only be quantitatively appropriate but also have qualitative, dynamic properties that optimize insulin action on target tissues. This review focuses on the importance of the first-phase insulin secretion to glucose metabolism and attempts to illustrate the relationships between the first-phase insulin response to an intravenous glucose challenge and the early insulin response following glucose ingestion. A clear-cut first phase occurs only when the β-cell is exposed to a rapidly changing glucose stimulus, like the one induced by a brisk intravenous glucose administration. In contrast, peripheral insulin concentration following glucose ingestion does not bear any clear sign of biphasic shape. Coupling data from the literature with the results of a β-cell model simulation, a close relationship between the first-phase insulin response to intravenous glucose and the early insulin response to glucose ingestion emerges. It appears that the same ability of the β-cell to produce a pronounced first phase in response to an intravenous glucose challenge can generate a rapidly increasing early phase in response to the blood glucose profile following glucose ingestion. This early insulin response to glucose is enhanced by the concomitant action of incretins and neural responses to nutrient ingestion. Thus, under physiological circumstances, the key feature of the early insulin response seems to be the ability to generate a rapidly increasing insulin profile. This notion is corroborated by recent experimental evidence that the early insulin response, when assessed at the portal level with a frequent sampling, displays a pulsatile nature. Thus, even though the classical first phase does not exist under physiological conditions, the oscillatory behavior identified at the portal level does serve the purpose of rapidly exposing the liver to elevated insulin levels that, also in virtue of their up-and-down pattern, are particularly effective in restraining hepatic glucose production.
Phase of menstrual cycle affects lysine requirement in healthy women
Authors: Kriengsinyos Wantanee , Wykes Linda J. , Goonewardene Laksiri A. , Ball Ronald O. , Pencharz Paul B.
Published: August 2004

The aim of this study was to investigate whether the phases of the menstrual cycle affect lysine requirement in healthy adult females, as determined by the indicator amino acid oxidation (IAAO) method. Five healthy females with regular menstrual cycles were studied at seven graded levels of lysine intake, in random order, with an oral [13C]phenylalanine tracer protocol in both the follicular and luteal phases. A total of 14 studies were conducted for each subject. Breath and plasma samples were collected according to the standard IAAO protocol. Serum 17β-estradiol and progesterone concentrations were measured on each IAAO study day. The rate of release of13CO2from [13C]phenylalanine oxidation (F13CO2) was measured, and a two-phase linear regression crossover model was applied to determine lysine requirement. F13CO2was higher during the luteal phase ( P < 0.001) and was positively associated with serum concentrations of 17β-estradiol and progesterone. The F13CO2data were adjusted for subjects and sex hormones and used to define breakpoints for lysine requirements. The lysine requirement of healthy females in the luteal phase was 37.7 mg·kg−1·day−1and higher ( P = 0.025) than that of females in the follicular phase (35.0 mg·kg−1·day−1). At all lysine intake levels, plasma amino acids were lower and phenylalanine oxidation was higher in the luteal relative to the follicular phase. Therefore, we reason that the higher lysine requirement observed in the luteal phase is probably due to higher amino acid catabolism.
Cell-cell interactions in regulating lung function
Authors: Boitano Scott , Safdar Zeenat , Welsh Donald G. , Bhattacharya Jahar , Koval Michael
Published: August 2004

Tight junction barrier formation and gap junctional communication are two functions directly attributable to cell-cell contact sites. Epithelial and endothelial tight junctions are critical elements of the permeability barrier required to maintain discrete compartments in the lung. On the other hand, gap junctions enable a tissue to act as a cohesive unit by permitting metabolic coupling and enabling the direct transmission of small cytosolic signaling molecules from one cell to another. These components do not act in isolation since other junctional elements, such as adherens junctions, help regulate barrier function and gap junctional communication. Some fundamental elements related to regulation of pulmonary barrier function and gap junctional communication were presented in a Featured Topic session at the 2004 Experimental Biology Conference in Washington, DC, and are reviewed in this summary.
Spectrum of ion channels in alveolar epithelial cells: implications for alveolar fluid balance
Authors: Kemp Paul J. , Kim Kwang-Jin
Published: August 2004

The efficient transition from placental to atmospheric delivery of oxygen at birth is critically dependent on rapid reabsorption of fetal lung fluid. In the perinatal period, this process is driven by active transepithelial sodium transport and is almost exclusively dependent on expression and modulation of the amiloride-sensitive epithelial sodium channel (ENaC). However, later in development, the amiloride sensitivity of the reabsorptive response, which must be sustained to keep the lungs effectively dry, wanes as a function of postnatal age. This Featured Topic (Experimental Biology Meeting, Washington, DC, April, 2004) presented exciting new evidence to demonstrate that, in addition to ENaC, the adult alveolar epithelium expresses a plethora of amiloride-insensitive ion channels, including cystic fibrosis transmembrane conductance regulator, proton channels, voltage-dependent potassium channels, and cyclic nucleotide-gated cation channels. Furthermore, important evidence for selective modulation of ENaC subunits in the lung in response to cardiovascular disease was demonstrated. Finally, it is clear that newly emerging models of human alveolar epithelium in combination with the novel lung slice electrophysiological preparation will ensure that the ascription of function to specific ion channels in the in situ human lung will soon be a real possibility.
Formation and stability ofS-nitrosothiols in RAW 264.7 cells
Authors: Zhang Yanhong , Hogg Neil
Published: August 2004

S-Nitrosothiols have been suggested to be mediators of many nitric oxide-dependent processes, including apoptosis and vascular relaxation. Thiol nitrosation is a poorly understood process in vivo, and the mechanisms by which nitric oxide can be converted into a nitrosating agent have not been established. There is a discrepancy between the suggested biological roles of nitric oxide and its known chemical and physical properties. In this study, we have examined the formation of S-nitrosothiols in lipopolysaccharide-treated RAW 264.7 cells. This treatment generated 17.4 ± 1.0 pmol/mg of protein (means ± SE, n =27) of intracellular S-nitrosothiol that slowly decayed over several hours. S-Nitrosothiol formation depended on the formation of nitric oxide and not on the presence of nitrite. Extracellular thiols were nitrosated by cell-generated nitric oxide. Oxygenated ferrous hemoglobin inhibited the formation of S-nitrosothiol, indicating the nitrosation occurred more slowly than diffusion. We discuss several mechanisms for S-nitrosothiol formation and conclude that the nitrosation propensity of nitric oxide is a freely diffusible element that is not constrained within an individual cell and that both nitric oxide per se and nitric oxide-derived nitrosating agents are able to diffuse across cell membranes. To achieve intracellular localization of the nitrosation reaction, mechanisms must be invoked that do not involve the formation of nitric oxide as an intermediate.
Role of tubular secretion and carbonic anhydrase in vertebrate renal sulfate excretion
Authors: Pelis Ryan M. , Renfro J. Larry
Published: August 2004

The renal proximal tubule of vertebrates performs an essential role in controlling plasma SO42−concentration ([SO42−]). Although net tubular SO42−reabsorption is the predominate control process in terrestrial vertebrates, a facilitated secretory flux is also present. In contrast, marine teleosts obtain excess SO42−from drinking, and increased plasma [SO42−] is prevented predominately through net tubular secretion. Tubular SO42−secretion is accomplished by at least two electroneutral anion exchange processes in series. Movement of SO42−into the cell across the basolateral membrane is pH dependent, suggesting SO42−/OH−exchange. Luminal HCO3−and Cl−can facilitate SO42−movement out of the cell across the brush-border membrane. The molecular identities of the anion exchangers are unknown but are probably homologues of SO42−transporters in the mammalian SLC26 gene family. In all species tested, glucocorticoids increase renal SO42−excretion. Whereas glucocorticoids downregulate SO42−reabsorptive mechanisms in terrestrial vertebrates, they may also stimulate a mediated secretory flux. In the marine teleost, cortisol increases the level of SO42−/HCO3−exchange at the brush-border membrane, tubular carbonic anhydrase (CA) activity, CAII protein, and a proportion of tubular SO42−secretion that is CA dependent. CA activity is required for about one-half of this net SO42−secretion but is also required for about one-half of the net reabsorption in bird proximal epithelium. A CA-SO42−/anion exchanger metabolon arrangement is proposed that may speed both the secretory and reabsorptive processes.
Biochemistry of exercise-induced metabolic acidosis
Authors: Robergs Robert A. , Ghiasvand Farzenah , Parker Daryl
Published: August 2004

The development of acidosis during intense exercise has traditionally been explained by the increased production of lactic acid, causing the release of a proton and the formation of the acid salt sodium lactate. On the basis of this explanation, if the rate of lactate production is high enough, the cellular proton buffering capacity can be exceeded, resulting in a decrease in cellular pH. These biochemical events have been termed lactic acidosis. The lactic acidosis of exercise has been a classic explanation of the biochemistry of acidosis for more than 80 years. This belief has led to the interpretation that lactate production causes acidosis and, in turn, that increased lactate production is one of the several causes of muscle fatigue during intense exercise. This review presents clear evidence that there is no biochemical support for lactate production causing acidosis. Lactate production retards, not causes, acidosis. Similarly, there is a wealth of research evidence to show that acidosis is caused by reactions other than lactate production. Every time ATP is broken down to ADP and Pi, a proton is released. When the ATP demand of muscle contraction is met by mitochondrial respiration, there is no proton accumulation in the cell, as protons are used by the mitochondria for oxidative phosphorylation and to maintain the proton gradient in the intermembranous space. It is only when the exercise intensity increases beyond steady state that there is a need for greater reliance on ATP regeneration from glycolysis and the phosphagen system. The ATP that is supplied from these nonmitochondrial sources and is eventually used to fuel muscle contraction increases proton release and causes the acidosis of intense exercise. Lactate production increases under these cellular conditions to prevent pyruvate accumulation and supply the NAD+needed for phase 2 of glycolysis. Thus increased lactate production coincides with cellular acidosis and remains a good indirect marker for cell metabolic conditions that induce metabolic acidosis. If muscle did not produce lactate, acidosis and muscle fatigue would occur more quickly and exercise performance would be severely impaired.
Production of arginine by the kidney is impaired in a model of sepsis: early events following LPS
Authors: Lortie Mark J. , Satriano Joseph , Gabbai Francis B. , Thareau Sonia , Khang Ser , Deng Aihua , Pizzo Donald P. , Thomson Scott C. , Blantz Roland C. , Munger Karen A.
Published: August 2004

Lipopolysaccharide (LPS) is used experimentally to elicit the innate physiological responses observed in human sepsis. We have previously shown that LPS causes depletion of plasma arginine before inducible nitric oxide synthase (iNOS) activity, indicating that changes in arginine uptake and/or production rather than enhanced consumption are responsible. Because the kidney is the primary source of circulating arginine and renal failure is a hallmark of septicemia, we determined the time course of changes in arginine metabolism and kidney function relative to iNOS expression. LPS given intravenously to anesthetized rats caused a decrease in mean arterial blood pressure after 120 min that coincided with increased plasma nitric oxide end products (NOx) and iNOS expression in lung and liver. Interestingly, impairment of renal function preceded iNOS activity by 30–60 min and occurred in tandem with decreased renal arginine production. The baseline rate of renal arginine production was ∼60 μmol·h−1·kg−1, corresponding to an apparent plasma half-life of ∼20 min, and decreased by one-half within 60 min of LPS. Calculations based on the systemic production and clearance show that normally only 5% of kidney arginine output is destined to become nitric oxide and that <25% of LPS-impaired renal production was converted to NOx in the first 4 h. In addition, we provide novel observations indicating that the kidney appears refractory to iNOS induction by LPS because no discernible enhancement of renal NOx production occurred within 4 h, and iNOS expression in the kidney was muted compared with that in liver or lung. These studies demonstrate that the major factor responsible for the rapid decrease in extracellular arginine content following LPS is impaired production by the kidney, a phenomenon that appears linked to reduced renal perfusion.
Effects of forebrain circumventricular organ ablation on drinking or salt appetite after sodium depletion or hypernatremia
Authors: Fitts Douglas A. , Freece Julia A. , Van Bebber Julie E. , Zierath Dannielle K. , Bassett John E.
Published: August 2004

In many previous studies, one or the other forebrain circumventricular organ, the subfornical organ (SFO) or organum vasculosum laminae terminalis (OVLT), was lesioned to test whether it was critical for the behavioral or physiological responses to sodium depletion and hypernatremia. These studies conflict in their conclusions. The present study was designed to create discrete lesions of both the SFO and OVLT in the same animals and to compare these with rats having a lesion of only the SFO or OVLT. Both the OVLT-lesioned group and the combined SFO + OVLT-lesioned group drank significantly more water and saline on a daily basis than Controls or SFO-lesioned rats. In both sodium depletion and hypertonic saline testing, rats with SFO lesions displayed transient deficits in salt appetite or thirst responses, whereas the rats with single OVLT lesions did not. In the sodium depletion test, but not in the hypernatremia test, rats with lesions of both the SFO and OVLT exhibited the largest deficit. The data support the hypothesis that a combined lesion eliminates redundancy and is more effective than a single lesion in sodium depletion tests. The interpretation of the OVLT lesion-only data may have been complicated by a tendency to drink more fluid on a daily basis, because some of those animals drank copious water in addition to saline even very early during the salt appetite test.
Membrane lipids and sodium pumps of cattle and crocodiles: an experimental test of the membrane pacemaker theory of metabolism
Authors: Wu B. J. , Hulbert A. J. , Storlien L. H. , Else P. L.
Published: August 2004

The influence of membrane lipid composition on the molecular activity of a major membrane protein (the sodium pump) was examined as a test of the membrane pacemaker theory of metabolism. Microsomal membranes from the kidneys of cattle (Bos taurus) and crocodiles (Crocodylus porosus) were found to possess similar sodium pump concentrations, but cattle membranes showed a four- to fivefold higher enzyme (Na+-K+-ATPase) activity when measured at 37°C. The molecular activity of the sodium pumps (ATP/min) from both species was fully recoverable when delipidated pumps were reconstituted with membrane from the original source (same species). The results of experiments involving species membrane crossovers showed cattle sodium pump molecular activity to progressively decrease from 3,245 to 1,953 ( P < 0.005) to 1,031 ( P < 0.003) ATP/min when subjected to two cycles of delipidation and reconstitution with crocodile membrane as a lipid source. In contrast, the molecular activity of crocodile sodium pumps progressively increased from 729 to 908 ( P < 0.01) to 1,476 ( P = 0.01) ATP/min when subjected to two cycles of delipidation and reconstitution with cattle membrane as a lipid source. The lipid composition of the two membrane preparations showed similar levels of saturated (∼31–34%) and monounsaturated (∼23–25%) fatty acids. Cattle membrane had fourfold more n-3 polyunsaturated fatty acids (11.2 vs. 2.9%) but had a reduced n-6 polyunsaturate content (29 vs. 43%). The results support the membrane pacemaker theory of metabolism and suggest membrane lipids and their polyunsaturates play a significant role in determining the molecular activity of the sodium pump.
On morphometric measurement of oxygen diffusing capacity in middle ear gas exchange
Authors: Kanick Stephen Chad , Doyle William J. , Ghadiali Samir N. , Federspiel William J.
Published: August 2004

An accurate mathematical model of transmucosal gas exchange is prerequisite to understanding middle ear (ME) physiology. Current models require experimentally measured gas species time constants for all extant conditions as input parameters. However, studies on pulmonary gas exchange have shown that a morphometric model that incorporates more fundamental physiochemical and anatomic parameters accurately simulates transport from which the species time constants can be derived for all extant conditions. Here, we implemented a variant of that model for ME gas exchange that requires the measurement of diffusional length (τ) for the ME mucosa. That measure contributes to the mucosal diffusing capacity and reflects the resistance to gas flow between air space and capillary. Two methods for measuring τ have been proposed: linear distance between the air-mucosal boundary and capillary and the harmonic mean of all contributing pathway lengths. Oxygen diffusing capacity was calculated for different ME mucosal geometries by using the two τ measures, and the results were compared with those predicted by a detailed, two-dimensional finite element analysis. Predictive accuracy was improved by incorporating the harmonic τ measure, which captures important information regarding variations in capillary shape and distribution. However, compared with the oxygen diffusing capacity derived from the finite element analysis, both measures yielded nonlinear, positively biased estimates. The morphometric techniques underestimate diffusion length by failing to account for the curvilinear gas flow pathways predicted by the finite element model.
Synaptic Integration in Rat Frontal Cortex Shaped by Network Activity
Authors: Léger Jean-François , Stern Edward A. , Aertsen Ad , Heck Detlef
Published: August 2004

Neocortical neurons in vivo are embedded in networks with intensive ongoing activity. How this network activity affects the neurons’ integrative properties and what function this may imply at the network level remain largely unknown. Most of our knowledge regarding synaptic communication and integration is based on recordings in vitro, where network activity is strongly diminished or even absent. Here, we present results from two complementary series of experiments based on intracellular in vivo recordings in anesthetized rat frontal cortex. Specifically, we measured 1) the relationship between the excursions of a neuron’s membrane potential and the spiking activity in the surrounding network and 2) how the summation of several inputs to a single neuron changes with the different levels of its membrane potential excursions and the associated states of network activity. The combination of these measurements enables us to assess how the level of network activity influences synaptic integration. We present direct evidence that integration of synaptic inputs in frontal cortex is linear, independent of the level of network activity. However, during periods of high network activity, the neurons’ response to synaptic input is markedly reduced in both amplitude and duration. This results in a drastic shortening of its window for temporal integration, requiring more precise coordination of presynaptic spike discharges to reliably drive the neuron to spike under conditions of high network activity. We conclude that ongoing activity, as present in the active brain, emphasizes the need for neuronal cooperation at the network level, and cannot be ignored in the exploration of cortical function.
Hair-Cell Versus Afferent Adaptation in the Semicircular Canals
Authors: Rabbitt R. D. , Boyle R. , Holstein G. R. , Highstein S. M.
Published: August 2004

The time course and extent of adaptation in semicircular canal hair cells was compared to adaptation in primary afferent neurons for physiological stimuli in vivo to study the origins of the neural code transmitted to the brain. The oyster toadfish, Opsanus tau, was used as the experimental model. Afferent firing-rate adaptation followed a double-exponential time course in response to step cupula displacements. The dominant adaptation time constant varied considerably among afferent fibers and spanned six orders of magnitude for the population (∼1 ms to >1,000 s). For sinusoidal stimuli (0.1–20 Hz), the rapidly adapting afferents exhibited a 90° phase lead and frequency-dependent gain, whereas slowly adapting afferents exhibited a flat gain and no phase lead. Hair-cell voltage and current modulations were similar to the slowly adapting afferents and exhibited a relatively flat gain with very little phase lead over the physiological bandwidth and dynamic range tested. Semicircular canal microphonics also showed responses consistent with the slowly adapting subset of afferents and with hair cells. The relatively broad diversity of afferent adaptation time constants and frequency-dependent discharge modulations relative to hair-cell voltage implicate a subsequent site of adaptation that plays a major role in further shaping the temporal characteristics of semicircular canal afferent neural signals.
Carboxyl Tail Region of the Kv2.2 Subunit Mediates Novel Developmental Regulation of Channel Density
Authors: Blaine Judith T. , Taylor Alison D. , Ribera Angeles B.
Published: August 2004

Molecular mechanisms underlying the acquisition of stable electrical phenotypes in developing neurons remain poorly defined. As Xenopus embryonic spinal neurons mature, they initially exhibit dramatic changes in excitability due to a threefold increase in voltage-gated potassium current ( IKv) density. Later when mature neurons begin synapse formation, IKvdensity remains stable. Elevation of Kv1.1 and Kv2.1 RNA levels indicates that excess transcript levels of these Kv genes can increase current density in both young and mature neurons. In contrast, Kv2.2 overexpression increases IKvdensity in young but not mature neurons despite the presence of protein translated from injected RNA at this stage. Because protein domains can determine biophysical as well as subcellular localization properties of channel subunits, we tested whether a region of the Kv2.2 subunit regulated functional expression in mature neurons. We focused on the large cytoplasmic carboxy tail, a region that differs most between Kv2.2 and the structurally related Kv2.1 subunit. Chimeric Kv2 subunits were generated in which different regions of the large cytoplasmic carboxyl tail were exchanged between Kv2.1 and Kv2.2 subunits. All chimeric Kv2 subunits induced voltage-gated potassium currents when expressed heterologously in oocytes. In vivo chimeric subunits increased IKvdensity in young neurons on overexpression in the developing embryo. In contrast, in mature neurons, only those chimeras lacking a domain in the proximal carboxy terminus, proxC, increased IKvdensity when overexpressed. Thus the proxC domain mediates developmental and subunit-specific regulation of IKvand identifies a novel function for protein domains.
Transforming Tonic Firing Into a Rhythmic Output in theAplysiaFeeding System: Presynaptic Inhibition of a Command-Like Neuron by a CPG Element
Authors: Hurwitz Itay , Susswein Abraham J. , Weiss Klaudiusz R.
Published: August 2004

Tonic stimuli can elicit rhythmic responses. The neural circuit underlying Aplysia californica consummatory feeding was used to examine how a maintained stimulus elicits repetitive, rhythmic movements. The command-like cerebral-buccal interneuron 2 (CBI-2) is excited by tonic food stimuli but initiates rhythmic consummatory responses by exciting only protraction-phase neurons, which then excite retraction-phase neurons after a delay. CBI-2 is inhibited during retraction, generally preventing it from exciting protraction-phase neurons during retraction. We have found that depolarizing CBI-2 during retraction overcomes the inhibition and causes CBI-2 to fire, potentially leading CBI-2 to excite protraction-phase neurons during retraction. However, CBI-2 synaptic outputs to protraction-phase neurons were blocked during retraction, thereby preventing excitation during retraction. The block was caused by presynaptic inhibition of CBI-2 by a key buccal ganglion retraction-phase interneuron, B64, which also causes postsynaptic inhibition of protraction-phase neurons. Pre- and postsynaptic inhibition could be separated. First, only presynaptic inhibition affected facilitation of excitatory postsynaptic potentials (EPSPs) from CBI-2 to its followers. Second, a newly identified neuron, B54, produced postsynaptic inhibition similar to that of B64 but did not cause presynaptic inhibition. Third, in some target neurons B64 produced only presynaptic but not postsynaptic inhibition. Blocking CBI-2 transmitter release in the buccal ganglia during retraction functions to prevent CBI-2 from driving protraction-phase neurons during retraction and regulates the facilitation of the CBI-2 induced EPSPs in protraction-phase neurons.
Electrophysiological Responses to Light of Neurons in the Eye and Statocyst of Lymnaea stagnalis
Authors: Sakakibara Manabu , Aritaka Tomoyo , Iizuka Akira , Suzuki Hiroyuki , Horikoshi Tetsuro , Lukowiak Ken
Published: August 2004

Lymnaea can be classically conditioned by pairing photic stimulation with a rotational stimulus. The electrophysiological properties of the Lymnaea photoreceptors and statocyst neurons are incompletely known. There are 2 types of ocular photoreceptors and 3 types of statocyst “hair cells.” Type A photoreceptors had a response latency from 200 to 400 ms, with a graded depolarizing response having maximum action spectra at 480–500 nm, corresponding to the βmax of rhodopsin. Additionally they extend their axons in the direction of the other type of photoreceptor neuron, the type T cell. These neurons have a 2-component response to light: a response reversibly reduced in Ca2+-free saline, and a component persisting in Ca2+-free saline. Type T cells send processes into the cerebral ganglion and terminate close to the ending of the statocyst hair cells. Hair cells send their terminal branches to the cerebral ganglia close to the terminations of the type T cells. Caudal hair cells respond to a light flash with a depolarization, whereas the rostral cells respond with a hyperpolarization. The response latency in all hair cells was dependent on the stimulus intensity; the brightest light tested had a latency of 200 ms. The photo-induced response was abolished in Ca2+-free saline, whereas it was still present in high Ca2+–high Mg2+ saline, consistent with the hypothesis that the connection between the photoreceptors and hair cells is monosynaptic. Thus the sensory information necessary for forming an association between photic and rotational stimuli converges on the statocyst neurons.
Short-Term Sound Temporal Envelope Characteristics Determine Multisecond Time Patterns of Activity in Human Auditory Cortex as Shown by fMRI
Authors: Harms Michael P. , Guinan John J. , Sigalovsky Irina S. , Melcher Jennifer R.
Published: August 2004

Functional magnetic resonance imaging (fMRI) of human auditory cortex has demonstrated a striking range of temporal waveshapes in responses to sound. Prolonged (30 s) low-rate (2/s) noise burst trains elicit “sustained” responses, whereas high-rate (35/s) trains elicit “phasic” responses with peaks just after train onset and offset. As a step toward understanding the significance of these responses for auditory processing, the present fMRI study sought to resolve exactly which features of sound determine cortical response waveshape. The results indicate that sound temporal envelope characteristics, but not sound level or bandwidth, strongly influence response waveshapes, and thus the underlying time patterns of neural activity. The results show that sensitivity to sound temporal envelope holds in both primary and nonprimary cortical areas, but nonprimary areas show more pronounced phasic responses for some types of stimuli (higher-rate trains, continuous noise), indicating more prominent neural activity at sound onset and offset. It has been hypothesized that the neural activity underlying the onset and offset peaks reflects the beginning and end of auditory perceptual events. The present data support this idea because sound temporal envelope, the sound characteristic that most strongly influences whether fMRI responses are phasic, also strongly influences whether successive stimuli (e.g., the bursts of a train) are perceptually grouped into a single auditory event. Thus fMRI waveshape may provide a window onto neural activity patterns that reflect the segmentation of our auditory environment into distinct, meaningful events.
Comparison of Oculomotor Neuronal Activity in Paralaminar and Mediodorsal Thalamus in the Rhesus Monkey
Authors: Tanibuchi Ikuo , Goldman-Rakic Patricia S.
Published: August 2004

We previously reported that neurons in the mediodorsal thalamic nucleus (MD) are topographically organized and express spatial and nonspatial coding properties similar to those of the prefrontal areas with which they are connected. In the course of mapping the dorsal thalamus, we also studied neurons in a subset of thalamic nuclei (the caudal part of the ventral lateral nucleus (VLc), the oral part of the ventral posterior lateral nucleus (VPLo), the parvocellular part of the ventral anterior nucleus (VApc)) lateral to the MD and just across the internal medullary lamina. We compared these “paralaminar” neurons to MD neurons by having monkeys perform the same spatial and nonspatial cognitive tasks as those used to investigate the MD; these included two saccadic tasks—one requiring delayed and the other immediate responses—and one picture fixation task. Of the paralaminar thalamic neurons modulated by the saccadic tasks, a majority had saccade-related activity, and this was nearly always spatially tuned. Also, for about half of these neurons, the saccade-related activity occurred exclusively during the delayed-response task. No neurons with event-related activity in the saccadic tasks were preferentially modulated by specific picture stimuli, although other neurons were. All of these results were similar to what we had found for MD neurons. However, in contrast to the high proportion of presaccadic responses observed in the MD, the majority of saccade-related neurons in paralaminar thalamus exhibited mid- or postsaccadic activity, i.e., that started during or after the saccade. Our findings suggest that neurons in the paralaminar thalamus may be possible conduits of oculomotor feedback signals, especially during memory-guided saccades.
Thoughts on Bone Biomechanics
Authors: Oxnard Charles E.
Published: August 2004

Simple simulations of stresses and strains in several types of localised anatomical features have been modelled using basic biomechanical thinking, experimental stress analysis employing photo-elastic methods and theoretical stress and strain analysis using a finite elements approach where the computational program utilises fast LaGrangian analysis of continua. The studies are providing results showing that, for given load proportions, the particular forms of some bones and bony architectural features seem related to stresses and strains that are relatively evenly distributed across surfaces (and therefore mechanically optimal). The results particularly show how apparently opposite (paradoxical) situations can readily occur within anatomical systems. They also provide strong biological information relevant to the assessment of such features of bone architecture in phylogenetic investigations. They even give information relevant to clinical problems.