bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential control over postganglionic neurons in rat cardiac ganglia by NA and DmnX neurons: anatomical evidence
Authors: Cheng Zixi (Jack) , Zhang Hong , Guo Shang Z. , Wurster Robert , Gozal David
Published: March 2004

In previous single-labeling experiments, we showed that neurons in the nucleus ambiguus (NA) and the dorsal motor nucleus of the vagus (DmnX) project to intrinsic cardiac ganglia. Neurons in these two motor nuclei differ significantly in the size of their projection fields, axon caliber, and endings in cardiac ganglia. These differences in NA and DmnX axon cardiac projections raise the question as to whether they target the same, distinct, or overlapping populations of cardiac principal neurons. To address this issue, we examined vagal terminals in cardiac ganglia and tracer injection sites in the brain stem using two different anterograde tracers {1,1′-dioleyl-3,3,3′,3′-tetramethylindocarbocyanine methanesulfonate and 4-[4-(dihexadecylamino)-styryl]- N-methylpyridinium iodide} and confocal microscopy in male Sprague-Dawley rats. We found that 1) NA and DmnX neurons innervate the same cardiac ganglia, but these axons target separate subpopulations of principal neurons and 2) axons arising from neurons in the NA and DmnX in the contralateral sides of the brain stem enter the cardiac ganglionic plexus through separate bundles and preferentially innervate principal neurons near their entry regions, providing topographic mapping of vagal motor neurons in left and right brain stem vagal nuclei. Because the NA and DmnX project to distinct populations of cardiac principal neurons, we propose that they may play different roles in controlling cardiac function.
Acute arterial hypertension inhibits proximal tubular fluid reabsorption in normotensive rat but not in SHR
Authors: Walstead Christopher , Yip Kay-Pong
Published: March 2004

The effect of acute arterial hypertension on proximal tubular fluid reabsorption was investigated in Sprague-Dawley rats and spontaneously hypertensive rats (SHR) by measuring proximal tubular flow with a nonobstructive optical method. Under control conditions, spontaneous tubular flow was oscillating at 0.02-0.03 Hz in Sprague-Dawley rats. Acute hypertension induced an immediate increase of mean tubular flow (50% increase after 20 min of hypertension) and augmentation of oscillatory amplitude. Acute hypertension did not alter single-nephron blood flow as measured by laser-Doppler velocimetry ( n = 12), suggesting that the increase of tubular flow was due to inhibition of reabsorption but not increase of filtration. By contrast, spontaneous tubular flow was fluctuating aperiodically in SHR. Acute hypertension did not induce a continuous increase of tubular flow or an increase in amplitude of fluctuations ( n = 15). When apical Na+/H+ exchanger activity of proximal tubule was monitored, acute hypertension did not alter the activity in SHR ( n = 8), while similar procedures had been shown to inhibit apical Na+/H+ exchanger activity of proximal tubules by more than 40% in Sprague-Dawley rats. These observations suggest that acute hypertension inhibits proximal tubular fluid reabsorption by inhibiting apical Na+/H+ exchanger activity in Sprague-Dawley rats and that this mechanism is impaired in SHR.
Hydrogen sulfide as an endogenous regulator of vascular smooth muscle tone in trout
Authors: Dombkowski Ryan A. , Russell Michael J. , Olson Kenneth R.
Published: March 2004

Hydrogen sulfide (H2S) is an endogenous vasodilator in mammals, but its presence and function in other vertebrates is unknown. We generated H2S from NaHS and examined the effects on isolated efferent branchial arteries from steelhead (stEBA) or rainbow (rtEBA) trout. H2S concentration was measured colorimetrically (CM) and with ion-selective electrodes (ISE) in rainbow trout plasma. NaHS produced a triphasic response consisting of a relaxation (phase 1), constriction (phase 2), and relaxation (phase 3) in both unstimulated vessels and in stEBA precontracted with carbachol (Carb). Phase 1 and phase 3 in stEBA were decreased and phase 2 increased in unstimulated vessels by K+ATP channel inhibition (glibenclamide), or a cocktail of inhibitors of cyclooxygenase, lipoxygenase, and cytochrome P-450 (indomethacin, esculetin, and clotrimazole). Inhibition of soluble guanylate cyclase with ODQ or NS-2028 inhibited phase 3 in stEBA, although NaHS decreased cGMP production by rtEBA. stEBA phase 2 contractions were partially inhibited by the myosin light chain kinase inhibitor, ML-9, but unaffected by L-type calcium channel inhibition (methoxyverapamil), whereas contraction in rtEBA was partially inhibited by nifedipine or removal of extracellular calcium. Phase 3 relaxations were more pronounced in stEBA precontracted with Carb and norepinephrine (NE) than those contracted by KCl or K2SO4. stEBA phase 2 and phase 3 responses were dose dependent (EC50 = 1.1 ± 1.2 × 10-3 M and 6.7 ± 0.9 × 10-5 M, respectively; n = 7). NaHS was also vasoactive in steelhead bulbus arteriosus, celiacomesenteric arteries, and anterior cardinal veins. Rainbow trout plasma sulfide concentration was 4.0 ± 0.3 × 10-5 M, n = 4 (CM) and 3.8 ± 0.4 × 10-5 M, n = 9 (ISE); similar to phase 3 EC50. Because NaHS has substantial vasoactive effects at physiological plasma concentrations, we propose that its soluble derivative, H2S, is a tonically active endogenous vasoregulator in trout.
Phenotypic modulation of cultured bladder smooth muscle cells and the expression of inducible nitric oxide synthase
Authors: Johansson Rebecka , Persson Katarina
Published: March 2004

Phenotypic modulation of smooth muscle is associated with various pathological conditions, including bladder dysfunction. Cytoskeletal dynamics modulate the cell phenotype and were recently shown to be involved in regulation of inducible nitric oxide synthase (iNOS). We tested the hypothesis that the cell differentiation status affects iNOS expression, and that iNOS is preferentially expressed in immature dedifferentiated bladder smooth muscle cells (BSMC). Isolated rat BSMC were put into different stages of differentiation by serum deprivation on laminin-coated plates in the presence of IGF-I and by interaction with Rho signaling and actin polymerization. iNOS and smooth muscle-myosin heavy chain (SM-MHC) protein expression were investigated with Western blot analysis. Our results showed iNOS protein in BSMC exposed to interleukin-1β (2 ng/ml) + TNF-α (50 ng/ml). Growth of BSMC in serum-free medium on laminin in the presence of IGF-I increased SM-MHC expression, whereas cytokine-induced iNOS was inhibited. Disruption of F-actin with latrunculin B (0.5 μM) potentiated iNOS expression and decreased SM-MHC expression. Rho inhibition with C3 (2.5 μg/ml) increased iNOS expression, whereas SM-MHC expression was slightly decreased. Rho-kinase inhibition with Y-27632 (10 μM) mediated a decrease in iNOS and a slight increase in SM-MHC expression. In conclusion, the capacity of BSMC to express iNOS was negatively correlated to differentiation status measured as SM-MHC expression. Actin cytoskeletal dynamics and Rho signaling are involved in regulation of cytokine-induced iNOS expression in BSMC. Phenotypic changes and impairment in actin cytoskeleton formation may potentiate cytokine activation and in turn increase nitric oxide production in the bladder during disease.
Loss of vasomotor responsiveness to the μ-opioid receptor ligand endomorphin-1 in adjuvant monoarthritic rat knee joints
Authors: McDougall Jason J. , Barin A. Kursat , McDougall Chelsea M.
Published: March 2004

Endomorphin-1 is a short-chain neuropeptide with a high affinity for the μ-opioid receptor and has recently been localized in acutely inflamed knee joints where it was found to reduce inflammation. The present study examined the propensity of endomorphin-1 to modulate synovial blood flow in normal and adjuvant-inflamed rat knee joints. Under deep urethane anesthesia, endomorphin-1 was topically applied to exposed normal and 1 wk adjuvant monoarthritic knee joints (0.1 ml bolus; 10-12-10-9mol). Relative changes in articular blood flow were measured by laser Doppler perfusion imaging and vascular resistances in response to the opioid were calculated. In normal knees, endomorphin-1 caused a dose-dependent increase in synovial vascular resistance and this effect was significantly inhibited by the specific μ-opioid receptor antagonist d-Phe-Cys-Tyr-d-Trp-Orn-Thr-Pen-Thr amide (CTOP) ( P < 0.0001, 2-factor ANOVA, n = 5-7). One week after adjuvant inflammation, the hypoaemic effect of endormophin-1 was completely abolished ( P < 0.0001, 2-factor ANOVA, n = 5-7). Immunohistochemical analysis of normal and adjuvant-inflamed joints showed a ninefold increase in endomorphin-1 levels in the monoarthritic knee compared with normal control. Western blotting and immunohistochemistry revealed a moderate number of μ-opioid receptors in normal knees; however, μ-opioid receptors were almost undetectable in arthritic joints. These findings demonstrate that peripheral administration of endomorphin-1 reduces knee joint blood flow and this effect is not sustainable during advanced inflammation. The loss of this hypoaemic response appears to be due to downregulation of μ-opioid receptors as a consequence of endomorphin-1 accumulation within the arthritic joint.
Effect of a perinatal high-salt diet on blood pressure control mechanisms in young Sprague-Dawley rats
Authors: Swenson Steven J. , Speth Robert C. , Porter James P.
Published: March 2004

In the present investigation we sought to determine if a perinatal high-salt treatment affects blood pressure at an early age (30 days), and if so, to determine the mechanisms responsible for the hypertension. Pregnant dams were given an 8% NaCl diet [high-salt (HS) rats] during the final one-third of gestation and throughout the suckling period. After weaning, the pups continued to receive the high-salt diet until testing at age 30 days. Control groups received a normal-salt diet (NS rats). In HS rats, mean arterial pressure (MAP) was significantly increased (110 ± 5 vs. 96 ± 3 mmHg) compared with NS rats. Blockade of brain AT1 receptors with intracerebroventricular losartan decreased MAP in HS but not NS rats. Blockade of α-adrenergic receptors with intravenous phentolamine or ganglionic transmission with intravenous chlorisondamine produced a greater decrease in MAP in HS rats. Baroreflex control of heart rate was assessed using a four-parameter logistics function. The mid-range MAP (p3) was significantly increased in the HS rats. No other baroreflex parameters were affected. Specific binding of 125I-[Sar1,Ile8]ANG II to AT1 receptors was increased in the subfornical organ (SFO) of the HS rats. Expression of AT1a receptor mRNA was greater in both SFO and PVN of the HS rats. These data suggest that even at an early age, Sprague-Dawley rats treated with a perinatal high-salt diet are hypertensive. The elevated blood pressure appears to be caused by increased sympathetic nervous activity, resulting, in part, from increased brain AT1 receptor activation.
Acute inhibition of the hypothalamic paraventricular nucleus decreases renal sympathetic nerve activity and arterial blood pressure in water-deprived rats
Authors: Stocker Sean D. , Keith Kimberly J. , Toney Glenn M.
Published: March 2004

The present study was performed to determine whether sympathetic outflow and arterial blood pressure in water-deprived rats are dependent on the ongoing neuronal activity of the hypothalamic paraventricular nucleus (PVN). Renal sympathetic nerve activity (RSNA), mean arterial blood pressure (MAP), and heart rate were recorded in urethane-α-chloralose-anesthetized rats that were deprived of water but not food for 48 h before experiments. Acute inhibition of the PVN by bilateral microinjection of the GABAA agonist muscimol (100 pmol/side) significantly decreased RSNA in water-deprived rats (-26.7 ± 4.7%, n = 7) but was without effect in control rats (1.3 ± 6.3%, n = 7). Similarly, injection of muscimol produced a greater decrease in MAP in water-deprived rats than in control rats (-46 ± 3 vs. -16 ± 3 mmHg, respectively), although baseline MAP was not different between groups (105 ± 4 vs. 107 ± 4 mmHg, respectively). Neither bilateral microinjection of isotonic saline vehicle (100 nl/side) into the PVN nor muscimol (100 pmol/side) outside the PVN altered RSNA or MAP in either group. In addition, ganglionic blockade with hexamethonium (30 mg/kg iv) significantly decreased MAP in both groups; however, the decrease in MAP was significantly greater in water-deprived rats than in control rats (62 ± 2 vs. 48 ± 2 mmHg, respectively). Collectively, these findings suggest that sympathetic outflow contributes more to the maintenance of blood pressure in the water-deprived rat, and this depends, at least partly, on the ongoing activity of PVN neurons.
High levels of myocardial antioxidant defense in aging nondiabetic normotensive Zucker obese rats
Authors: Conti Marc , Renaud Isabelle M. , Poirier Bruno , Michel Odile , Belair Marie-France , Mandet Chantal , Bruneval Patrick , Myara Isaac , Chevalier Jacques
Published: March 2004

Chronic renal failure often induces left ventricular hypertrophy. We assessed whether the heart is affected in the Zucker obese rat, a model of chronic renal failure associated with obesity, glucose intolerance, and insulin resistance without hypertension or hyperglycemia. After systemic blood pressure measurement, the heart, the aorta, and the kidneys were removed from anesthetized 9- and 13-mo-old Zucker obese and lean control male rats ( n = 33, n = 24, n = 25, and n = 21, respectively). Determination of left ventricular geometry, quantification of myocardium collagen density, and measurement of heart antioxidant enzyme activity were made, as well as aorta and kidney parameters. Mean blood pressure remained at a normal range whatever the age and group considered. Whereas kidney structure and function were severely impaired, no sign of myocardial infarction or inflammatory process was noticed. A moderate left ventricular hypertrophy was observed in 13-mo-old obese rats. While heart malondialdehyde was stable with age and among groups, antioxidant enzyme activity was higher in obese rats. In conclusion, in the absence of hypertensive or hyperglycemic disorders, the heart seems to display a sufficient line of defense against oxidative stress during the development of cardiac hypertrophy.
Electrophysiological properties of the L-type Ca2+current in cardiomyocytes from bluefin tuna and Pacific mackerel
Authors: Shiels H. A. , Blank J. M. , Farrell A. P. , Block B. A.
Published: March 2004

Tunas are capable of exceptionally high maximum metabolic rates; such capability requires rapid delivery of oxygen and metabolic substrate to the tissues. This requirement is met, in part, by exceptionally high maximum cardiac outputs, opening the possibility that myocardial Ca2+delivery is enhanced in myocytes from tuna compared with those from other fish. In this study, we investigated the electrophysiological properties of the cardiac L-type Ca2+channel current ( ICa) to test the hypothesis that Ca2+influx would be larger and have faster kinetics in cardiomyocytes from Pacific bluefin tuna ( Thunnus orientalis) than in those from its sister taxon, the Pacific mackerel ( Scomber japonicus). In accordance with this hypothesis, ICain atrial myocytes from bluefin tuna had significantly greater peak current amplitudes and faster fast inactivation kinetics (-4.4 ± 0.2 pA/pF and 25.9 ± 1.6 ms, respectively) than those from mackerel (-2.7 ± 0.5 pA/pF and 32.3 ± 3.8 ms, respectively). Steady-state activation, inactivation, and recovery from inactivation were also faster in atrial myocytes from tuna than from mackerel. In ventricular myocytes, current amplitude and activation and inactivation rates were similar in both species but elevated compared with those of other teleosts (Vornanen M. Am J Physiol Regul Integr Comp Physiol 272: R1432-R1440, 1997). These results indicate enhanced ICain atrial myocytes from bluefin tuna compared with Pacific mackerel; this enhanced ICamay be associated with elevated cardiac performance, because ICadelivers the majority of Ca2+involved in excitation-contraction coupling in most fish hearts. Similarly, ICais enhanced in the ventricle of both species compared with other teleosts and may play a role in the robust cardiac performance of fishes of the family Scombridae.
HIFs and tumors—causes and consequences
Authors: Höpfl Gisele , Ogunshola Omolara , Gassmann Max
Published: March 2004

For most organisms oxygen is essential for life. When oxygen levels drop below those required to maintain the minimum physiological oxygen requirement of an organism or tissue it is termed hypoxia. To counteract possible deleterious effects of such a state, an immediate molecular response is initiated causing adaptation responses aimed at cell survival. This response is mediated by the hypoxia-inducible factor-1 (HIF-1), which is a heterodimer consisting of an α- and a β-subunit. HIF-1α protein is stabilized under hypoxic conditions and therefore confers selectivity to this response. Hypoxia is characteristic of tumors, mainly because of impaired blood supply resulting from abnormal growth. Over the past few years enormous progress has been made in the attempt to understand how the activation of the physiological response to hypoxia influences neoplastic growth. In this review some aspects of HIF-1 pathway activation in tumors and the consequences for pathophysiology and treatment of neoplasia are discussed.
Lactic and hydrochloric acids induce different patterns of inflammatory response in LPS-stimulated RAW 264.7 cells
Authors: Kellum John A. , Song Mingchen , Li Jinyou
Published: March 2004

Metabolic acidosis frequently complicates sepsis and septic shock and may be deleterious to cellular function. Different types of metabolic acidosis (e.g., hyperchloremic and lactic acidosis) have been associated with different effects on the immune response, but direct comparative studies are lacking. Murine macrophage-like RAW 264.7 cells were cultured in complete medium with lactic acid or HCl to adjust the pH between 6.5 and 7.4 and then stimulated with LPS ( Escherichia coli 0111:B4; 10 ng/ml). Nitric oxide (NO), IL-6, and IL-10 levels were measured in the supernatants. RNA was extracted from the cell pellets, and RT-PCR was performed to amplify corresponding mediators. Gel shift assay was also performed to assess NF-κB DNA binding. Increasing concentrations of acid caused increasing acidification of the media. Trypan blue exclusion and lactate dehydrogenase release demonstrated that acidification did not reduce cell viability. HCl significantly increased LPS-induced NO release and NF-κB DNA binding at pH 7.0 but not at pH 6.5. IL-6 and IL-10 expression (RNA and protein) were reduced with HCl-induced acidification, but IL-10 was reduced much more than IL-6 at low pH. By contrast, lactic acid significantly decreased LPS-induced NO, IL-6, and IL-10 expression in a dose-dependent manner. Lactic acid also inhibited LPS-induced NF-κB DNA binding. Two common forms of metabolic acidosis (hyperchloremic and lactic acidosis) are associated with dramatically different patterns of immune response in LPS-stimulated RAW 264.7 cells. HCl is essentially proinflammatory as assessed by NO release, IL-6-to-IL-10 ratios, and NF-κB DNA binding. By contrast, lactic acidosis is anti-inflammatory.
A paracrine signaling role for serotonin in rat taste buds: expression and localization of serotonin receptor subtypes
Authors: Kaya Namik , Shen Tiansheng , Lu Shao-gang , Zhao Fang-li , Herness Scott
Published: March 2004

Recent advances in peripheral taste physiology now suggest that the classic linear view of information processing within the taste bud is inadequate and that paracrine processing, although undemonstrated, may be an essential feature of peripheral gustatory transduction. Taste receptor cells (TRCs) express multiple neurotransmitters of unknown function that could potentially participate in a paracrine role. Serotonin is expressed in a subset of TRCs with afferent synapses; additionally, TRCs respond physiologically to serotonin. This study explored the expression and cellular localization of serotonin receptor subtypes in TRCs as a possible route of paracrine communication. RT-PCR was performed on RNA extracted from rat posterior taste buds with 14 primer sets representing 5-HT1 through 5-HT7 receptor subtype families. Data suggest that 5-HT1A and 5-HT3 receptors are expressed in taste buds. Immunocytochemistry with a 5-HT1A-specific antibody demonstrated that subsets of TRCs were immunopositive for 5-HT1A. With the use of double-labeling, serotonin- and 5-HT1A-immunopositive cells were observed exclusively in nonoverlapping populations. On the other hand, 5-HT3-immunopositive taste receptor cells were not observed. This observation, combined with other data, suggests 5-HT3 is expressed in postsynaptic neural elements within the bud. We hypothesize that 5-HT release from TRCs activates postsynaptic 5-HT3 receptors on afferent nerve fibers and, via a paracrine route, inhibits neighboring TRCs via 5-HT1A receptors. The role of the 5-HT1A-expressing TRC within the taste bud remains to be explored.
Lethality during continuous anthrax lethal toxin infusion is associated with circulatory shock but not inflammatory cytokine or nitric oxide release in rats
Authors: Cui Xizhong , Moayeri Mahtab , Li Yan , Li Xuemei , Haley Michael , Fitz Yvonne , Correa-Araujo Rosaly , Banks Steven M. , Leppla Stephen H. , Eichacker Peter Q.
Published: March 2004

Although circulatory shock related to lethal toxin (LeTx) may play a primary role in lethality due to Bacillus anthracis infection, its mechanisms are unclear. We investigated whether LeTx-induced shock is associated with inflammatory cytokine and nitric oxide (NO) release. Sprague-Dawley rats with central venous and arterial catheters received 24-h infusions of LeTx (lethal factor 100 μg/kg; protective antigen 200 μg/kg) that produced death beginning at 9 h and a 7-day mortality rate of 53%. By 9 h, mean arterial blood pressure, heart rate, pH, and base excess were decreased and lactate and hemoglobin levels were increased in LeTx nonsurvivors compared with LeTx survivors and controls (diluent only) ( P ≤ 0.05 for each comparing the 3 groups). Despite these changes, arterial oxygen and circulating leukocytes and platelets were not decreased and TNF-α, IL-1β, IL-6, and IL-10 levels were not increased comparing either LeTx nonsurvivors or survivors to controls. Nitrate/nitrite levels and tissue histology also did not differ comparing LeTx animals and controls. In additional experiments, although 24-h infusions of LeTx and Esherichia coli LPS produced similar mortality rates (54 and 56%, respectively) and times to death (13.2 ± 0.8 vs. 11.0 ± 1.7 h, respectively) compared with controls, only LPS reduced circulating leukocytes, platelets, and IL-2 levels and increased TNF-α, IL-1α and -1β, IL-6, IL-10, interferon-γ, granulocyte macrophage-colony stimulating factor, RANTES, migratory inhibitory protein-1α, -2, and -3, and monocyte chemotactic protein-1, as well as nitrate/nitrite levels (all P ≤ 0.05 for the effects of LPS). Thus, in contrast to LPS, excessive inflammatory cytokine and NO release does not appear to contribute to the circulatory shock and lethality occurring with LeTx in this rat model. Although therapies to modulate these host mediators may be applicable for shock caused by LPS or other bacterial toxins, they may not with LeTx.
Spinal control of erection by glutamate in rats
Authors: Rampin Olivier , Monnerie Régine , Jérôme Nathalie , McKenna Kevin , Maurin Yves
Published: March 2004

The lumbosacral spinal network controlling penile erection is activated by information from peripheral and supraspinal origins. We tested the hypothesis that glutamate, released by sensory afferents from the genitals, activates this proerectile network. In anesthetized intact and T8 spinalized (i.e., freed from supraspinal inhibition) male rats, the parameters of electrical stimulation of the dorsal penile nerve (DPN) that elicited intracavernous pressure (ICP) rises were determined. In T8 spinalized rats, DPN stimulations were applied in the presence of d(-)-2-amino-5-phosphonopentanoic acid (d-AP5), a competitive NMDA receptor antagonist, or of 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulphonamide (NBQX), an AMPA-kainate receptor antagonist, injected intrathecally at the lumbosacral level. Both antagonists, alone or in combination, dose dependently decreased the ICP rise and increased its latency. In conscious rats, reflexive erections were depressed by d-AP5 and NBQX, as revealed by an increased latency of the first erection and by decreases of the number of rats displaying erections, of the number of erection clusters and of the number of erections per cluster. In anesthetized rats, the combined administration of the glutamatergic agonists NMDA and AMPA elicited ICP rises in the absence of DPN stimulation. In contrast, both agonists moderately decreased the ICP rise elicited by DPN stimulation but did not affect its latency. These results support our hypothesis that glutamate, released on stimulation of the genitals and acting at AMPA and NMDA receptors, is a potent activator of the spinal proerectile network.
Chronic exercise lowers the defended body weight gain and adiposity in diet-induced obese rats
Authors: Levin Barry E. , Dunn-Meynell Ambrose A.
Published: March 2004

The effects of running wheel exercise and caloric restriction on the regulation of body weight, adiposity, and hypothalamic neuropeptide expression were compared in diet-induced obese male rats over 6 wk. Compared with sedentary controls, exercising rats had reduced body weight gain (24%), visceral (4 fat pads; 36%) and carcass (leptin; 35%) adiposity but not insulin levels. Hypothalamic arcuate nucleus (ARC) proopiomelanocortin (POMC) mRNA expression was 25% lower, but ARC neuropeptide Y (NPY), agouti-related peptide, dorsomedial nucleus (DMN) NPY, and paraventricular nucleus (PVN) corticotropin-releasing hormone (CRH) expression was comparable to controls. Sedentary rats calorically restricted to 85% of control body weight reduced their visceral adiposity (24%), leptin (64%), and insulin (21%) levels. ARC NPY (23%) and DMN NPY (60%) were increased, while ARC POMC (40%) and PVN CRH (14%) were decreased. Calorically restricted exercising rats ran half as much as ad libitum-fed exercising rats and had less visceral obesity than comparably restricted sedentary rats. When sedentary restricted rats were refed after 4 wk, they increased intake and regained the weight gain and adiposity of sedentary controls. While refed exercising rats and sedentary rats ate comparable amounts, refed exercising rats regained weight and adiposity only to the level of ad libitum-fed exercising rats. Thus exercise lowers the defended level of weight gain and adiposity without a compensatory increase in intake and with a very different profile of hypothalamic neuropeptide expression from calorically restricted rats. This may be due to exercise-related factors other than plasma insulin and leptin.
No effect of dietary calcium on body weight of lean and obese mice and rats
Authors: Zhang Qinmin , Tordoff Michael G.
Published: March 2004

Recent epidemiological and animal studies have led to the hypothesis that low dietary calcium intakes contribute to obesity. Here, we evaluated whether calcium influenced the body weight of normal-weight and obese rodents. All experiments involved female C57BL/6J mice or Sprague-Dawley rats fed normal- or high-energy-density diets (3.8 or 4.7 kcal/g). Calcium intake was manipulated by allowing mice to drink sweetened 30 mM CaCl2 solution or feeding mice and rats diets differing in calcium content (0.2%, 0.6%, or 1.8% Ca2+). Blood samples were taken from rats to confirm that the diets had their intended effects on metabolism. There were no effects of the calcium manipulations on energy intake, body weight, or carcass fat content and no simple relation between calciotropic hormones and body weight. One experiment found a significant decrease in body weight gain of lean and obese rats fed the 1.8% Ca2+ diet, but we suspect that this was due to forced consumption of the unpalatable diet, reducing growth. These studies provide little support for the hypothesis that dietary calcium contributes to the etiology or maintenance of obesity.
Thermal injury impairs cardiac protein synthesis and is associated with alterations in translation initiation
Authors: Lang Charles H. , Frost Robert A. , Vary Thomas C.
Published: March 2004

The purpose of the present study was to determine whether burn injury decreases myocardial protein synthesis and potential contributing mechanisms for this impairment. To address this aim, thermal injury was produced by a 40% total body surface area full-thickness scald burn in anesthetized rats, and the animals were studied 24 h later. Burn decreased the in vivo-determined rate of myocardial protein synthesis and translation efficiency by 25% but did not alter the protein synthetic rate in skeletal muscle. To identify potential mechanisms responsible for regulating mRNA translation in cardiac muscle, we examined several eukaryotic initiation factors (eIFs) and elongation factors (eEFs). Burn failed to alter eIF2B activity or the total amount or phosphorylation status of either eIF2α or eIF2Bϵ in heart. In contrast, hearts from burned rats demonstrated 1) an increased binding of the translational repressor 4E-BP1 with eIF4E, 2) a decreased amount of eIF4E associated with eIF4G, and 3) a decreased amount of the hyperphosphorylated γ-form of 4E-BP1. These changes in eIF4E availability were not seen in gastrocnemius muscle where burn injury did not decrease protein synthesis. Furthermore, constitutive phosphorylation of mTOR, S6K1, the ribosomal protein S6, and eIF4G were also decreased in hearts from burned rats. Burn did not appear to adversely affect elongation because there was no significant difference in the myocardial content of eEF1α or eEF2 or the phosphorylation state of eEF2. The above-mentioned burn-induced changes in mRNA translation were associated with an impairment of in vitro myocardial performance. Finally, 24 h postburn, the cardiac mRNA content of IL-1β, IL-6, and high-mobility group protein B1 (but not TNF-α) was increased. In summary, these data suggest that thermal injury specifically decreases cardiac protein synthesis in part by decreasing mRNA translation efficiency resulting from an impairment in translation initiation associated with alterations in eIF4E availability and S6K1 activity.
Kinetics and regulation of a Ca2+-activated Cl-conductance in mouse renal inner medullary collecting duct cells
Authors: Boese S. H. , Aziz O. , Simmons N. L. , Gray M. A.
Published: March 2004

Using the whole cell patch-clamp technique, a Ca2+-activated Cl-conductance (CaCC) was transiently activated by extracellular ATP (100 μM) in primary cultures of mouse inner medullary collecting duct (IMCD) cells and in the mouse IMCD-K2 cell line. ATP also transiently increased intracellular Ca2+concentration ([Ca2+]i) from ∼100 nM to peak values of ∼750 nM in mIMCD-K2 cells, with a time course similar to the ATP-induced activation and decay of the CaCC. Removal of extracellular Ca2+had no major effect on the peak Cl-conductance or the increase in [Ca2+]iinduced by ATP, suggesting that Ca2+released from intracellular stores directly activates the CaCC. In mIMCD-K2 cells, a rectifying time- and voltage-dependent current was observed when [Ca2+]iwas fixed via the patch pipette to between 100 and 500 nM. Maximal activation occurred at ∼1 μM [Ca2+]i, with currents losing any kinetics and displaying a linear current-voltage relationship. From Ca2+-dose-response curves, an EC50value of ∼650 nM at -80 mV was obtained, suggesting that under physiological conditions the CaCC would be near fully activated by mucosal nucleotides. Noise analysis of whole cell currents in mIMCD-K2 cells suggests a single-channel conductance of 6–8 pS and a density of ∼5,000 channels/cell. In conclusion, the CaCC in mouse IMCD cells is a low-conductance, nucleotide-sensitive Cl-channel, whose activity is tightly coupled to changes in [Ca2+]iover the normal physiological range.
Smad expression during kidney development
Authors: Vrljicak P. , Myburgh D. , Ryan A. K. , van Rooijen M. A. , Mummery C. L. , Gupta I. R.
Published: March 2004

Signaling by the transforming growth factor (TGF)-β superfamily is important during kidney development. Here, we describe the spatial and temporal expression patterns of the Smads, the transcription factors that translate TGF-β signals into gene expression. RT-PCR data and in situ hybridization analysis showed that the receptor-regulated (R) Smads (Smad1, -2, -3, -5, and -8), the common partner Smad (Smad4), and the inhibitory (I) Smads (Smad6 and -7) were all expressed during mouse kidney development from embryonic day 12 until the end of nephrogenesis at postnatal day 15. Each Smad had a distinct spatial distribution. All were expressed by mesenchymal cells in the nephrogenic zone and were downregulated once these cells began to epithelialize. The common partner Smad, Smad4, was present in uninduced mesenchymal cells and at ureteric bud tips. The bone morphogenetic-responsive R-Smads, Smad1, -5, and -8, were mainly expressed in the nephrogenic zone, whereas the TGF-β- responsive R-Smads were predominantly noted in the medullary interstitium. Expression of the I-Smad Smad7 was also seen in mesenchymal cells in the interstitium. Based on the observed patterns of expression, we speculate that individual or combinations of Smads may play specific roles in cell-fate determination during kidney development.
Angiotensin II blockade prevents hyperglycemia-induced activation of JAK and STAT proteins in diabetic rat kidney glomeruli
Authors: Banes Amy K. , Shaw Séan , Jenkins John , Redd Heather , Amiri Farhad , Pollock David M. , Marrero Mario B.
Published: March 2004

Clinical and animal studies show that treatment with angiotensin-converting enzyme (ACE) inhibitors or ANG II-receptor antagonists slows progression of nephropathy in diabetes, indicating ANG II plays an important role in its development. We previously reported that hyperglycemia augments both ANG II-induced growth and activation of Janus kinase (JAK)2 and signal transducers and activators of transcription (STAT) proteins in cultured rat mesangial cells. Furthermore, we demonstrated that the tyrosine kinase enzyme JAK2 plays a key role in both ANG II- and hyperglycemia-induced growth in these cells. We hypothesized that the ACE inhibitor captopril and the ANG II-receptor antagonist candesartan would hinder hyperglycemic-induced activation of JAK and STAT proteins in rat glomeruli, demonstrating that ANG II plays an important role in the activation of these proteins in vivo. Adult male Sprague-Dawley rats were given either streptozotocin (STZ; 60 mg/kg iv) or vehicle, and glomeruli were isolated 2 wk later. Activation of JAK and STAT proteins was evaluated by Western blot analysis for specific tyrosine phosphorylation. Groups of rats were given captopril (75–85 mg·kg-1·day-1), candesartan (10 mg· kg-1·day-1), or the JAK2 inhibitor AG-490 (5 mg·kg-1·day-1) for the study's duration. STZ stimulated glomerular phosphorylation of JAK2, STAT1, STAT3, and STAT5. Phosphorylation was reduced in rats treated with captopril, candesartan, and AG-490. Furthermore, both candesartan and AG-490 inhibited STZ-induced increases in urinary protein excretion. In conclusion, our studies demonstrate that hyperglycemia induces activation of JAK2 and the STATs in vivo via an ANG II-dependent mechanism and that these proteins may be involved in the early kidney damage associated with diabetes.
Gene expression alterations during HGF-induced dedifferentiation of a renal tubular epithelial cell line (MDCK) using a novel canine DNA microarray
Authors: Balkovetz Daniel F. , Gerrard Edward R. , Li Shixiong , Johnson David , Lee James , Tobias John W. , Rogers Katherine K. , Snyder Richard W. , Lipschutz Joshua H.
Published: March 2004

Hepatocyte growth factor (HGF) elicits a broad spectrum of biological activities, including epithelial cell dedifferentiation. One of the most widely used and best-studied polarized epithelial cell lines is the Madin-Darby canine kidney (MDCK) cell line. Here, we describe and validate the early response of polarized monolayers of MDCK cells stimulated with recombinant HGF using a novel canine DNA microarray designed to query 12,473 gene sequences. In our survey, eight genes previously implicated in the HGF signaling pathway were differentially regulated, demonstrating that the system was responsive to HGF. Also identified were 117 genes not previously known to be involved in the HGF pathway. The results were confirmed by real-time PCR or Western blot analysis for 38 genes. Of particular interest were the large number of differentially regulated genes encoding small GTPases, proteins involved in endoplasmic reticulum translation, proteins involved in the cytoskeleton, the extracellular matrix, and the hematopoietic and prostaglandin systems.
Preservation of complex I function during hypoxia-reoxygenation-induced mitochondrial injury in proximal tubules
Authors: Feldkamp Thorsten , Kribben Andreas , Roeser Nancy F. , Senter Ruth A. , Kemner Sarah , Venkatachalam Manjeri A. , Nissim Itzhak , Weinberg Joel M.
Published: March 2004

Inhibition of complex I has been considered to be an important contributor to mitochondrial dysfunction in tissues subjected to ischemia-reperfusion. We have investigated the role of complex I in a severe energetic deficit that develops in kidney proximal tubules subjected to hypoxia-reoxygenation and is strongly ameliorated by supplementation with specific citric acid cycle metabolites, including succinate and the combination of α-ketoglutarate plus malate. NADH: ubiquinone reductase activity in the tubules was decreased by only 26% during 60-min hypoxia and did not change further during 60-min reoxygenation. During titration of complex I activity with rotenone, progressive reduction of NAD+to NADH was detected at >20% complex I inhibition, but substantial decreases in ATP levels and mitochondrial membrane potential did not occur until >70% inhibition. NAD+was reduced to NADH during hypoxia, but the NADH formed was fully reoxidized during reoxygenation, consistent with the conclusion that complex I function was not limiting for recovery. Extensive degradation of cytosolic and mitochondrial NAD(H) pools occurred during either hypoxia or severe electron transport inhibition by rotenone, with patterns of metabolite accumulation consistent with catabolism by both NAD+glycohydrolase and pyrophosphatase. This degradation was strongly blocked by α-ketoglutarate plus malate. The data demonstrate surprisingly little sensitivity of these cells to inhibition of complex I and high levels of resistance to development of complex I dysfunction during hypoxia-reoxygenation and indicate that events upstream of complex I are important for the energetic deficit. The work provides new insight into fundamental aspects of mitochondrial pathophysiology in proximal tubules during acute renal failure.
A vibrator prevents streaming during close-arterial infusion into the kidney
Authors: Hamza Shereen M. , Kaufman Susan
Published: March 2004

Close-arterial infusion of test substances allows one to study the responses of a selected vascular bed without inducing confounding systemic effects. Unfortunately, laminar flow patterns within the artery cause streaming of the injected factor, so that distribution within the target organ is not homogeneous. We describe a reliable method of overcoming these problems. Specifically, we attach a vibrator (i-Vibe egg) to the syringe containing the test substance. We showed that, without vibration, infusion of a solution of Evans blue (0.5% wt/vol) results in uneven distribution of the dye in the kidney. Vibration of the syringe during infusion allows for uniform coloration of the kidney surface. There is also functional improvement of drug distribution during vibration. Renal blood flow was measured during intrarenal infusion of phenylephrine (150 μl, 0.05–0.5 μg). Vibration caused a significant leftward shift in the dose-response curve, i.e., the phenylephrine-induced reduction in renal blood flow was enhanced by vibration. This cheap, simple method for ensuring adequate mixing of intra-arterially infused substances will facilitate not only the study of renal function in the rat but also infusion of test and therapeutic substances into other organs.
Differential regulation of PHEX expression in bone and parathyroid gland by chronic renal insufficiency and 1,25-dihydroxyvitamin D3
Authors: Brewer Angela J. , Canaff Lucie , Hendy Geoffrey N. , Tenenhouse Harriet S.
Published: March 2004

Mutations in the PHEX gene are responsible for X-linked hypophosphatemia, a renal phosphate-wasting disorder associated with defective skeletal mineralization. PHEX is predominantly expressed in bones and teeth and in the parathyroid gland of patients with chronic renal failure and tertiary hyperparathyroidism. The purpose of the present study was to examine the effects of renal insufficiency and 1,25-dihydroxyvitamin D3[1,25(OH)2D3] on the regulation of PHEX expression in rat tibia and parathyroid gland. In rats fed a high-phosphate (Pi) diet, ⅚ nephrectomy elicited a significant increase in the serum parathyroid hormone (PTH) concentration that was associated with a significant increase in the abundance of PHEX mRNA and protein in the tibia and a significant increase in PHEX mRNA in the parathyroid gland. In contrast, 1,25(OH)2D3administration to intact rats fed a control diet elicited a significant decrease in the serum PTH concentration that was accompanied by a significant decrease in PHEX mRNA and protein abundance in the tibia and a significant decrease in PHEX mRNA in the parathyroid gland. In addition, the increases in serum PTH levels and PHEX mRNA in the tibia and parathyroid gland in ⅚ nephrectomized rats fed a high-Pidiet were blunted by 1,25(OH)2D3. Serum PTH concentration was positively and significantly correlated with tibial PHEX mRNA and protein abundance. In summary, we demonstrate that PHEX expression in the tibia and parathyroid gland is increased by chronic renal insufficiency and decreased by 1,25(OH)2D3administration and suggest that PTH status may play an important role in mediating these changes in PHEX expression.
p38 MAP kinase mediates mechanically induced COX-2 and PG EP4receptor expression in podocytes: implications for the actin cytoskeleton
Authors: Martineau Louis C. , McVeigh Lyne I. , Jasmin Bernard J. , Kennedy Chris R. J.
Published: March 2004

A dynamic cytoskeleton allows podocytes to withstand significant mechanical stress on elevation of intraglomerular capillary pressure (Pgc). However, vasoactive hormones, such as prostaglandin E2(PGE2), may challenge the integrity of the actin cytoskeleton, alter podocyte morphology, and compromise glomerular permeability. PGE2synthesis correlates with the onset of proteinuria and increased Pgcfollowing reduced nephron mass. We investigated the interplay among mechanical stress, cyclooxygenase (COX), E-prostanoid (EP) receptor expression, and the actin cytoskeleton, using an in vitro model of cell stretch. Immortalized mouse podocytes grown on flexible silicone membranes were cyclically stretched (5% elongation, 0.5 Hz) for 2 h. EP4and COX-2 mRNA increased three- and sevenfold above nonstretched controls, whereas EP1and COX-1 levels were unchanged. Six hours of stretch resulted in a threefold increase in PGE2-stimulated cAMP accumulation, a measure of EP4receptor function, and an increase in COX-2 protein. The stretch-induced effects on COX-2/EP4expression and EP4-induced cAMP production were attributable to p38 MAP kinase, as blockade of this pathway, but not of ERK or JNK, abrogated the response. These stretch-induced changes in expression were transcriptionally dependent as they were actinomycin D sensitive. Finally, we investigated the influence of enhanced EP4signaling on the actin cytoskeleton. Addition of PGE2resulted in actin filament depolymerization observable only in stretched cells. Our results indicate that key components of the eicosanoid pathway are upregulated by mechanically stimulated p38 MAP kinase in podocytes. Enhanced EP4receptor signaling may undermine podocyte cytoskeletal dynamics and thereby compromise filtration barrier function under conditions of increased Pgc.
Empirical Relationships for Frequency Content Parameters of Earthquake Ground Motions
Authors: Rathje Ellen M. , Faraj Fadi , Russell Stephanie , Bray Jonathan D.
Published: March 2004

The frequency content of an earthquake ground motion is important because it affects the dynamic response of earth and structural systems. Four scalar parameters that characterize the frequency content of strong ground motions are (1) the mean period (Tm), (2) the average spectral period (Tavg), (3) the smoothed spectral predominant period (To), and (4) the predominant spectral period (Tp). Tm and Tavg distinguish the low frequency content of ground motions, while To is affected most by the high frequency content. Tp does not adequately describe the frequency content of a strong ground motion and is not recommended. Empirical relationships are developed that predict three parameters (Tm, Tavg, and To) as a function of earthquake magnitude, site-to-source distance, site conditions, and rupture directivity. The relationships are developed from a large strong-motion database that includes recorded motions from the recent earthquakes in Turkey and Taiwan. The new relationships update those previously developed by the authors and others. The results indicate that three site classes, which distinguish between rock, shallow soil, and deep soil, provide a better prediction of the frequency content parameters and smaller standard error terms than conventional “rock” and “soil” site classes. Forward directivity significantly increases the frequency content parameters, particularly Tm and To, at distances less than 20 km. Each of the frequency content parameters can be predicted with reasonable accuracy, but Tm is the preferred because it best distinguishes the frequency content of strong ground motions.
Evaluation of Modal and FEMA Pushover Analyses: SAC Buildings
Authors: Goel Rakesh K. , Chopra Anil K.
Published: March 2004

This paper comprehensively evaluates the Modal Pushover Analysis (MPA) procedure against the “exact” nonlinear response history analysis (RHA) and investigates the accuracy of seismic demands determined by pushover analysis using FEMA-356 force distributions; the MPA procedure in this paper contains several improvements over the original version presented in Chopra and Goel (2002). Seismic demands are computed for six buildings, each analyzed for 20 ground motions. It is demonstrated that with increasing number of “modes” included, the height-wise distribution of story drifts and plastic rotations estimated by MPA becomes generally similar to trends noted from nonlinear RHA. The additional bias and dispersion introduced by neglecting “modal” coupling and P-Δ effects due to gravity loads in MPA procedure is small unless the building is deformed far into the inelastic range with significant degradation in lateral capacity. A comparison of the seismic demands computed by FEMA-356 NSP and nonlinear RHA showed that FEMA-356 lateral force distributions lead to gross underestimation of story drifts and completely fail to identify plastic rotations in upper stories compared to the values from the nonlinear RHA. The “Uniform” force distribution in FEMA-356 NSP seems unnecessary because it grossly overestimates drifts and plastic rotations in lower stories and grossly underestimates them in upper stories. The MPA procedure resulted in estimates of demand that were much better than from FEMA force distributions over a wide range of responses—from essentially elastic response of Boston buildings to strongly inelastic response of Los Angeles buildings. However, pushover analysis procedures cannot be expected to provide satisfactory estimates of seismic demands for buildings deforming far into the inelastic range with significant degradation of the lateral capacity; for such cases, nonlinear RHA becomes necessary.