bluuebunny/crossref_metadata_2025 huggingface dataset:
Ammonia Response To Constant Exercise: Differences To The Lactate Response
Authors: Ogino Kazuhide , Kinugawa Toru , Osaki Shuichi , Kato Masahiko , Endoh Akihiro , Furuse Yoshiyuki , Uchida Kazuhiko , Shimoyama Masaki , Igawa Osamu , Hisatome Ichiro , Shigemasa Chiaki
Published: March 2003

1. We evaluated the plasma ammonia response to constant exercise at different intensities. Ten healthy male volunteers were asked to perform constant exercise for 15 min at five different intensities: 80, 90, 100, 110 and 120% of their ventilatory threshold (VT). Blood concentrations of lactate, ammonia and hypoxanthine were measured during and after exercise.

2. The concentration of lactate increased continuously during exercise intensities equivalent to 100, 110 and 120% VT. Plasma ammonia began to increase at 6 min exercise and continued increasing during exercise at all five exercise intensities. Plasma hypoxanthine levels also increased continuously during exercise at all exercise intensities; however, they peaked at 5–10 min after exercise. The response of plasma ammonia and hypoxanthine increased with increasing intensities of exercise.

3. While the extent of the increase in lactate levels during exercise at 100, 110 and 120% VT was significantly higher than that at 80% VT, only the increase in ammonia and hypoxanthine levels at 120% VT were significantly higher than those at 80% VT.

4. In conclusion, the plasma ammonia response to constant exercise differed to the lactate and ammonia responses to short‐term exhaustive exercise.
Genetically Targeted Calcium Sensors Enhance The Study Of Organelle Function In Living Cells
Authors: Petrou Steven , Bowser David N , Nicholls Robert A , Panchal Rekha G , Smart Megan L , Reilly Angela M , Williams David A
Published: March 2003

1. Understanding the regulation of calcium (Ca2+), the most common of the mineral ions within the human body, has always been of extreme interest to physiologists. While the importance of Ca2+ in contributing to physiological events through regulation of levels has been significantly established, seldom is consideration given to the intricacies of this ion and its mechanics in producing such diverse physiological responses in different regions of the cell.

2. The present review will summarize new methodologies used in our laboratories for the study of two major intracellular organelles, mitochondria and the nucleus. These techniques are based predominantly on the use of molecular biological approaches to both create and then target protein‐based sensor molecules to specific intracellular locations.

3. The regulation of Ca2+ in the mitochondria and nucleus is of particular interest to us because of the central involvement of these organelles in: (i) cardiac cell responses during ischaemia/reperfusion; and (ii) the control of gene expression, respectively.
Effect Of Fentanyl On Baroreflex Control Of Circumflex Coronary Conductance
Authors: Moore Peter G , Quail Anthony W , Cottee David BF , McIlveen Stephen A , White Saxon W
Published: March 2003

1. Fentanyl, a synthetic μ‐opioid receptor agonist, is the preferred induction and maintenance anaesthetic agent in cardiac surgery.

2. Its actions on myocardial blood flow are poorly understood. There are reports of intra‐operative myocardial ischaemia. Its reported actions on cardiorespiratory control vary widely, but do involve hypertension, bradycardia and peripheral vasoconstriction.

3. Accordingly, the postulate that fentanyl would cause coronary vasoconstriction and myocardial disadvantage was examined in awake dogs with a continuous wave Doppler flow probe mounted on the circumflex coronary artery.

4. Continuous intravenous infusion of fentanyl citrate (550 ng/kg per min) raised plasma concentrations of fentanyl to 3.37 ng/mL in a linear fashion at 20 min. There was a fall in core temperature of 0.7°C and, although no apparent depression of ventilation or fall in arterial or coronary sinus PO2, there was a rise in PCO2 and H+ concentration. Some dogs salivated and panted transiently. Thus, fentanyl may reset temperature regulation in low doses but, at higher doses, is associated with metabolic acidosis.

5. In sinus rhythm, the arterial pressure of the dogs fell slightly, then rose to 115% of resting control. Circumflex flow and conductance rose early, then conductance steadily declined to 83%. Heart rate fell, then rose before returning to pre‐infusion levels. The early circumflex coronary vasodilator effects, but not the later vasoconstrictor effects, were reduced in dogs with paced hearts.

6. In dogs with paced hearts, a dose–effect study using 138, 275, 550 and 1100 ng/kg per min fentanyl suggested that, at low plasma concentrations of 1–2 ng/mL, vasodilatation does occur in both coronary and systemic circulations; however, at higher doses, intense coronary and systemic vasoconstriction supervenes.

7. The dose–response effect of fentanyl on arterial baroreflex control of circumflex conductance was examined during the immediate 8 s circumflex vasodilator response to a step rise in aortic pressure caused by inflation of an intra‐aortic balloon. At low plasma concentrations of fentanyl, baroreflex control of circumflex conductance appears to be enhanced but, with increasing plasma concentrations of fentanyl, appears to be depressed.

8. Therefore, the effects of fentanyl are dose dependent. At low plasma concentrations, left ventricular blood flow and its baroreflex control is enhanced but, at higher concentrations, it is depressed.
Effect Of Plasma Cholesterol On Red Blood Cell Oxygen Transport
Authors: Buchwald Henry , O'Dea Thomas J , Menchaca Hector J , Michalek Van N , Rohde Thomas D
Published: March 2003

1. Oxygen (O2) transfer from the blood to tissues is a function of the red blood cell (RBC) O2 saturation (SO2), the plasma O2 content being negligible. Under conditions of increased tissue O2 demand, the SO2 of arterial blood does not change appreciably (97%); however, the SO2 of mixed venous blood, equal to that of the perfused tissues, can go as low as 20%.

2. Tissue O2 availability is limited by the exposure time to a RBC, which decreases under conditions of maximum stress (< 1 s). If the O2 unloading time was to increase significantly, because of a decrease in the RBC diffusion constant or an increase in the RBC membrane thickness, the RBC O2 unloading time would exceed tissue (e.g. cardiac) transit time and O2 transfer would be impaired.

3. Cholesterol constitutes the non‐polar, hydrophobic lipid of the enveloping layer of the RBC membrane. As the cholesterol content of the RBC increases, the fluidity of the membrane decreases and the lipid shell stiffens.

4. Early studies demonstrated that high blood cholesterol concentrations were associated with reduced blood O2 transport; in essence, the haemoglobin dissociation curve was shifted to the left.

5. Current investigations have shown that the cholesterol RBC membrane barrier to O2 diffusion delayed O2 entry into the RBC during saturation and delayed O2 release from the RBC during desaturation. In an analysis of 93 patients divided by their cholesterol concentration into five groups, the percentage change in blood O2 diffusion was inversely proportional to the cholesterol concentration.

6. The RBC membrane cholesterol is in equilibrium with the plasma cholesterol concentration. It stands to reason that as the plasma cholesterol increases, the RBC membrane becomes impaired and O2 transport is reduced.

7. The implications of this new perspective on O2 transport include the ability to increase tissue oxygenation by lowering plasma cholesterol.
Experimental Biology 2000 Symposium on Differential Control of Sympathetic Outflow NEUROANATOMICAL SPECIFICITY OF THE CIRCUITS CONTROLLING SYMPATHETIC OUTFLOW TO DIFFERENT TARGETS
Authors: Sved Alan F , Cano Georgina , Card J Patrick
Published: March 2003

1. Despite the emerging framework that central neural pathways controlling the activity of the sympathetic nervous system are capable of producing highly selective responses, the specific neural pathways governing different sympathetic outflows are poorly understood.

2. Anatomical studies suggest that five brain areas, namely the rostral ventrolateral medulla, the rostral ventromedial medulla, the caudal raphe nuclei, the region containing the A5 noradrenergic neurons and the paraventricular hypothalamic nucleus, provide dominant supraspinal innervation of sympathetic preganglionic neurons.

3. The anatomical parcellation of different functions within and among these cell groups is uncertain. However, recent studies using transynaptic retrograde labelling of neural pathways connected to various sympathetic targets suggest that the circuits controlling these different targets may be partially distinct. Similarly, anatomical studies relying on stimulus‐evoked expression of immediate early genes, such as c‐fos, suggest that different sympathetic responses may be controlled by distinct neural circuits.

4. Thus, although many similarities exist in the anatomical circuits innervating different sympathetic targets, possibly supporting the orchestration of global sympathetic responses, differences are also discernible.
Proceedings of the Australian Physiological and Pharmacological Society Symposium: The Hypothalamus LEPTIN AND THE REGULATION OF FOOD INTAKE AND THE NEUROENDOCRINE AXIS IN SHEEP
Authors: Clarke Iain J , Henry Belinda , Iqbal Javed , Goding James W
Published: March 2003

1. Leptin is secreted by fat and acts on the brain.

2. Central infusion of leptin reduces food intake but does not alter endocrine secretions in normally fed sheep.

3. Leptin treatment can correct for altered hormonal secretion in fasted animals.

4. Alterations in bodyweight (leptin status) affect the expression of a number of genes in the hypothalamus that are involved in the regulation of food intake and neuroendocrine function.

5. Leptin receptors are found in both the hypothalamus and pituitary and direct action of leptin can be demonstrated on the somatotrophs in the pituitary.
Dynamic Aspects Of Platelet Adhesion Under Flow
Authors: Dopheide Sacha M , Yap Cindy L , Jackson Shaun P
Published: March 2003

1. Cell–cell and cell–matrix adhesive interactions are critical for a wide range of physiological processes, including embryogenesis, inflammation, immunity and haemostasis.

2. The ability of circulating blood cells, such as platelets and leucocytes, to adhere to sites of vascular injury is complicated by the presence of blood flow, which imposes hydrodynamic forces on adhesion contacts.

3. To overcome this problem, platelets and leucocytes have evolved specific adhesion receptors with unique biomechanical properties that enable these cells to adhere to the vessel wall under flow conditions.

4. Platelet adhesion in the normal circulation appears to be a multiple‐step process involving an initial reversible interaction between the platelet adhesion receptor glycoprotein Ib‐IX‐V and the vascular adhesion protein von Willebrand factor. Once tethered to the vessel wall, platelets form irreversible adhesion contacts through the binding of one or more platelet integrins to specific subendothelial matrix proteins.

5. There is now a wealth of evidence demonstrating that these receptors not only mediate platelet adhesion, but also transduce signals leading to platelet activation.

6. In the present review, we will briefly discuss the current understanding of the specific roles of individual platelet receptors in supporting the haemostatic function of platelets and discuss mechanisms by which these receptors induce platelet activation.
Prophylactic Effects Of Pilocarpine Hydrochloride On Xerostomia Models Induced By X‐Ray Irradiation In Rats
Authors: Asari Tetsuya , Komatsu Yoshimitsu , Misawa Keiko , Hara Kiyoto , Akahane Masuo
Published: March 2003

1. In the present study, we investigated the prophylactic effects of pilocarpine hydrochloride on xerostomia models induced by either single (15 Gy) or repeated (8.6 Gy ×3 days) X‐ray irradiation in rats. Pilocarpine hydrochloride was administered orally 90 min before each irradiation session. Then, 7 days later, salivary volume, amylase activity and protein concentration in the saliva secreted from the right parotid gland were measured before and after a subsequent administration of pilocarpine hydrochloride (intraduodenal).

2. In irradiated no‐pretreatment rats, irradiation induced a significant reduction in both spontaneous and pilocarpine hydrochloride‐stimulated secretion (both total salivary volume and flow rate), regardless of the protocol used for X‐ray exposure. In irradiated, pilocarpine hydrochloride‐pretreated rats, salivary secretion was increased after stimulation by pilocarpine hydrochloride (intraduodenal) to a degree that depended on the pretreatment dose of pilocarpine hydrochloride (p.o.) in both xerostomia models.

3. There were no differences in amylase or protein concentrations between irradiated rats pretreated with pilocarpine hydrochloride and irradiated no‐pretreatment control rats.

4. A decrease in the weight of the parotid gland was observed in rats exposed to either the single dose or repeated irradiation protocols. Changes in the submandibular gland were less marked than those in the parotid gland. These changes in gland weight were not affected by pilocarpine hydrochloride pretreatment.

5. The responsiveness of the parotid gland to subsequent stimulation with pilocarpine hydrochloride was apparently preserved in both xerostomia models by pretreatment with pilocarpine hydrochloride, which itself increased salivary secretion. This suggests that pilocarpine hydrochloride may exert functional protective effects against xerostomia that occurs following irradiation therapy through a stimulation of salivary secretion.
Ongoing Nicotinic And Non‐Nicotinic Inputs To Inhibitory Neurons In The Mouse Colon
Authors: Powell Andrew K , Fida Rosa , Bywater Robert AR
Published: March 2003

1. Intracellular microelectrodes were used to record spontaneous and evoked inhibitory junction potentials (IJP) from the circular muscle layer of the mid‐distal region of the mouse isolated colon in the presence of nifedipine (1 μmol/L) and hyoscine (1 μmol/L).

2. The length of the tissue preparation (> 1 cm) or the presence of the mucosa had no effect on the frequency of spontaneous IJP.

3. Hexamethonium (500 μmol/L) reduced the frequency of spontaneous IJP to approximately 70% of the control frequency, whereas D‐tubocurarine (280 μmol/L) reduced the frequency to approximately 17% of control. Apamin (250 nmol/L) abolished all spontaneous IJP activity.

4. The greater inhibition of spontaneous IJP in the presence of D‐tubocurarine compared with hexamethonium is discussed as a possible ‘apamin‐like’ effect.

5. Although electrically evoked IJP (single pulse at 15 V, 0.6 msec) were not significantly affected by hexamethonium, D‐tubocurarine and apamin reduced the amplitude of evoked IJP to approximately 65 and 50% of control, respectively.

6. These results suggest that the properties of spontaneous IJP cannot be inferred by a study of evoked IJP alone.
Participation Of Atp In Cell Volume Regulation In The Endothelium After Hypotonic Stress
Authors: Shinozuka Kazumasa , Tanaka Naoko , Kawasaki Kumiko , Mizuno Hideya , Kubota Yoko , Nakamura Kazuki , Hashimoto Michio , Kunitomo Masaru
Published: March 2003

1. The role of ATP in the regulatory volume decrease (RVD) after hypotonic cell swelling was examined in cultured endothelial cells isolated from the rat caudal artery.

2. Hypotonic stress increased [Ca2+]i in addition to increasing the overflow of ATP and cell volume. The hypotonicity induced increase in [Ca2+]i was prevented by pyridoxalphosphate‐6‐azophenyl‐1‐2′,4′‐disulphonic acid (PPADS; a P2 purinoceptor antagonist), U‐73122 (a phospholipase C inhibitor) and thapsigargin (a Ca2+ pump inhibitor). However, the hypotonicity induced increase in cell volume was potentiated by PPADS, U‐73122 and thapsigargin.

3. Similar changes were observed in cells treated with 2‐methylthioATP, a P2Y purinoceptor agonist, but not by α,β‐methylene ATP, a P2X purinoceptor agonist. Thus, it appears that the responses observed following hypotonic stress are mediated by activation of P2Y purinoceptors.

4. On the basis of these findings, it is suggested that ATP, which is released by hypotonicity, may participate in the RVD as a substantial regulator or initiator via P2 purinoceptor‐induced increases in [Ca2+]i.
Taurine Inhibits Development Of Atherosclerotic Lesions In apolipoprotein E‐Deficient Mice‡
Authors: Kondo Yukiko , Toda Yoshihisa , Kitajima Hideaki , Oda Hiroaki , Nagate Takatoshi , Kameo Kazuya , Murakami Shigeru
Published: March 2003

1. The effects of taurine on the development of atherosclerotic lesions were investigated in apolipoprotein (apo) E‐deficient mice, an animal model with severe hypercholesterolaemia and extensive atherosclerosis. These mice were fed a normal laboratory chow containing 2% taurine for 12 weeks.

2. Serum total cholesterol was significantly elevated after 12 weeks treatment with taurine. This elevation was due to increases in very low‐density lipoprotein‐ and low‐density lipoprotein–cholesterol.

3. Despite such effects on serum lipoproteins, analysis using en face oil red O staining revealed that taurine reduced the area of arterial lipid accumulation by 28%, as measured quantitatively as an index of atherosclerosis. Histological examination also demonstrated a decrease in the size of aortic lesions in taurine‐treated mice.

4. Serum levels of thiobarbituric acid reactive substances (TBARS) in apoE‐deficent mice were higher than in normolipidaemic C57BL/6J mice. Serum TBARS levels were significantly decreased by 12 weeks treatment of apoE‐deficient mice with taurine.

5. Thus, taurine prevents the formation of atherosclerotic lesions, independently of serum cholesterol levels, and the results suggest that the anti‐oxidative effects of tauirne are related to its anti‐atherosclerotic actions.
Nociceptin/Orphanin Fq Modulates The Cardiovascular, But Not Renal, Responses To Stress In Spontaneously Hypertensive Rats
Authors: Kapusta Daniel R , Dayan Lisa A , Kenigs Velga A
Published: March 2003

1. The central administration of the endogenous opioid‐like peptide nociceptin/orphanin FQ (N/OFQ) produces marked cardiovascular depressor and renal sympathoinhibitory responses in conscious animals. These findings are evidence that central N/OFQ may modulate the cardiovascular and renal responses to acute environmental stress.

2. The changes in cardiovascular and renal function produced by intracerebroventricular (i.c.v.) N/OFQ were measured in conscious spontaneously hypertensive rats (SHR) under basal conditions and during the acute environmental stimulus of air jet stress.

3. In SHR, central N/OFQ produced profound hypotensive, bradycardic, renal sympathoinhibitory (delayed) and water‐diuretic effects by a pathway that does not involve activation of central α2‐adrenoceptors or classical opioid receptors.

4. Intracerebroventricular injection of N/OFQ prevented the pressor response and blunted the tachycardia to air jet stress. A similar renal sympathoexcitatory and antinatriuretic response was observed in conscious SHR during air stress, before and after i.c.v. N/OFQ.

5. These findings are evidence that, in conscious SHR, i.c.v. N/OFQ selectively inhibited the neural responses to air jet stress by attenuating sympathetic outflow to the heart and, potentially, vasculature, but not to the kidneys. Central endogenous N/OFQ systems may be activated and contribute to regional changes in sympathetic outflow during acute stress.
Central Mechanisms Underlying Short‐ And Long‐Term Regulation Of The Cardiovascular System
Authors: Dampney RAL , Coleman MJ , Fontes MAP , Hirooka Y , Horiuchi J , Li Y‐W , Polson JW , Potts PD , Tagawa T
Published: March 2003

1. Sympathetic vasomotor nerves play a major role in determining the level of arterial blood pressure and the distribution of cardiac output. The present review will discuss briefly the central regulatory mechanisms that control the sympathetic outflow to the cardiovascular system in the short and long term.

2. In the short term, the sympathetic vasomotor outflow is regulated by: (i) homeostatic feedback mechanisms, such as the baroreceptor or chemoreceptor reflexes; or (ii) feed‐forward mechanisms that evoke cardiovascular changes as part of more complex behavioural responses.

3. The essential central pathways that subserve the baroreceptor reflex and, to a lesser extent, other cardiovascular reflexes, have been identified by studies in both anaesthetized and conscious animals. A critical component of these pathways is a group of neurons in the rostral ventrolateral medulla that project directly to the spinal sympathetic outflow and that receive inputs from both peripheral receptors and higher centres in the brain.

4. Much less is known about the central pathways subserving feed‐forward or ‘central command’ responses, such as the cardiovascular changes that occur during exercise or that are evoked by a threatening or alerting stimulus. However, recent evidence indicates that the dorsomedial hypothalamic nucleus is a critical component of the pathways mediating the cardiovascular response to an acute alerting stimulus.

5. Long‐term sustained changes in sympathetic vasomotor activity occur under both physiological conditions (e.g. a change in salt intake) and pathophysiological conditions (e.g. heart failure). There is evidence that the paraventricular nucleus in the hypothalamus is a critical component of the pathways mediating these changes.

6. Understanding the central mechanisms involved in the long‐term regulation of sympathetic activity and blood pressure is a major challenge for the future. As a working hypothesis, a model is presented of the postulated central mechanisms that result in sustained changes in sympathetic vasomotor activity that are evoked by different types of chronic stimulation.
Chronic Hypoxia Preconditioning Increases Survival In Rats Suffering From Heatstroke
Authors: Wen Hsiao‐Chuan , Lee Chin‐Cheng , Lee Wen‐Chuan , Huang Kuo‐Sheng , Lin Mao‐Tsun
Published: March 2003

1. In the present study, we assessed the protective effects of chronic hypoxia preconditioning against heatstroke‐induced injury in urethane‐anaesthetized rats. Heatstroke was induced by exposing the animals to an ambient temperature of 42°C. The time at which both the mean arterial pressure (MAP) and local cerebral blood flow (CBF) in the striatum began to decrease from peak levels was taken as the onset of heatstroke. Control rats were exposed to a temperature of 24°C.

2. Mean arterial pressure, CBF, blood pH, PaO2, PaCO2 and survival time (the interval between onset of heatstroke and cardiac arrest) after heat stress were all lower than in control rats (in which ‘survival time’ was defined as > 360 min). However, blood lactate concentrations were greater in rats exposed to heat. Rats placed at high altitude (HA), when exposed to the same heat stress (42°C) survived much longer (113 ± 26 min; n = 8) than rats maintained at sea level (SL; 20 ± 2 min; n = 8).

3. After the onset of heatstroke, blood pH and lactate concentrations were found to be significantly higher and lower, respectively, in HA rats than in SL rats.

4. Western blot assay revealed that chronic hypoxia preconditioning induced heat shock protein (HSP) 72 expression in both the kidneys and lungs.

5. Thus, it appears that the observed benefit of chronic hypoxia preconditioning is related to attenuation of tissue acidification and elevations of HSP72 expression in both kidneys and lungs during heatstroke.
Involvement Of α‐Calcitonin Gene‐Related Peptide In Heat Stress‐Induced Delayed Preconditioning In Rat Hearts
Authors: Peng Jun , Lu Rong , Xiao Liang , Deng Han‐Wu , Li Yuan‐Jian
Published: March 2003

1. Previous studies have shown that hyperthermia is capable of activating capsaicin‐sensitive sensory nerves and stimulating the release of neurotransmitters from their peripheral terminals. Calcitonin gene‐related peptide (CGRP) has recently been found to participate in delayed cardioprotection in rat isolated hearts.

2. The purpose of the present study was to explore whether the delayed cardioprotection by heat stress in vivo involves the expression and release of CGRP.

3. Sprague‐Dawley rats were pretreated with whole‐body hyperthermia (rectal 42°C) for 15 min, 24 h before the experiments and then the left main coronary artery of rat hearts was subjected to a 45 min occlusion followed by 3 h reperfusion. The degree of myocardial injury was evaluated by measurement of infarct size and plasma creatine kinase (CK) activity. The plasma levels of CGRP and expression of CGRP (α
and β isoforms) mRNA in lumbar dorsal root ganglia at 4, 8, 16 or 24 h after heat stress treatment were measured.

4. Pretreatment with hyperthermia significantly reduced infarct size and CK release. Heat stress also significantly increased plasma concentrations of CGRP and the expression of α‐CGRP mRNA, but not β‐CGRP mRNA. The effect of heat stress was completely abolished by pretreatment with capsaicin (50 mg/kg, s.c.), which selectively depletes transmitters in capsaicin‐sensitive sensory nerves.

5. In summary, the results suggest that the delayed cardioprotection by heat stress involves the synthesis and release of CGRP and that the protection is mainly mediated by the α‐CGRP isoform.
Effects Of Endotoxin‐Induced Shock On Withdrawal Contractions In The Guinea‐Pig Isolated Ileum
Authors: Fatehi M , Tanomand A , Fatehi‐Hassanabad Z
Published: March 2003

1. Taking into consideration the effect of septic shock on releasing various mediators, the present study was undertaken in an attempt to elucidate the effect of endotoxin on naloxone‐induced withdrawal contractions in the guinea‐pig isolated ileum.

2. To induce withdrawal contractions, preparations removed either from saline‐ or endotoxin (2.5 mg/kg in 0.4 mL, i.p.)‐treated animals were incubated for 5 min with morphine (40 μmol/L) and naloxone (50 μmol/L) was then applied.

3. In tissues removed from endotoxin‐treated animals, a significant reduction in withdrawal contractions was observed. In control preparations, indomethacin (1 μmol/L
for 15 min) significantly reduced naloxone‐induced contractions, whereas NG‐nitro‐L‐arginine
methyl ester (L‐NAME; 10 μmol/L for 15 min) had no effect.
However, indomethacin pretreatment of tissues removed from endotoxin‐treated animals
did not modify the withdrawal contractions, whereas L‐NAME pretreatment enhanced the amplitude of the withdrawal‐induced contractions.

4. These results suggest that attenuation of the withdrawal contractions in guinea‐pig isolated ileum induced by endotoxin‐pretreatment may be due, in part, to the activation of an L‐arginine–nitric oxide pathway.
Insulin‐Enhanced Liposome‐Mediated Gene Transfer Into A Gastric Carcinoma Cell Line
Authors: Ohmiya Naoki , Emi Nobuhiko , Niwa Yasumasa , Goto Hidemi , Hayakawa Tetsuo
Published: March 2003

1. Liposome‐mediated transfection is useful due to no DNA constraints, lower immunogenicity and easy preparation. However, it has the disadvantage of low transfection efficiency. We aimed to test whether lipofection efficiency could be enhanced in gastrointestinal cell lines by the growth‐promoting effect of insulin.

2. To assess the effect of insulin on lipofection efficiency and the cell cycle, expression of green fluorescent protein (GFP) and DNA distribution in gastric (MKN1), colonic (HT29) and pancreatic (BxPC3) carcinoma cell lines was analysed using flow cytometry.

3. The percentage of positive cells with GFP was significantly higher in MKN1 cells in culture medium with 5 mg/mL insulin than without insulin, whereas the percentage was the same in HT29 and BxPC3 cells with insulin as without insulin. The percentage of S phase fraction MKN1 cells with insulin was greater than without insulin, whereas the percentage of S phase fractions of HT29 and BxPC3 cells was the same with or without insulin. Lipofection efficiency correlated with the percentage of S phase fraction.

4. Insulin has the potential to enhance efficiency of lipofection into a sensitive cell line by increasing cellular proliferation.
Effects of cytokines on contractile and dilator responses of airway smooth muscle
Authors: Shore Stephanie A , Moore Paul E
Published: March 2003

1. Increased bronchoconstrictor responses to contractile agonists and decreased dilator responses to β‐adrenoceptor agonists are characteristics of human asthma. One explanation for these features of asthma is that cytokines released in the asthmatic airway have direct effects on airway smooth muscle cells that alter their phenotype.

2. The present review summarizes data indicating that inflammatory cytokines, such as interleukin (IL)‐1β and tumour necrosis factor‐α, T helper (h) 1 cytokines, such as interferon‐γ, and Th2 cytokines, such as IL‐13 and IL‐5, have the capacity to enhance contractile responses and/or decrease relaxant responses of airway smooth muscle. These effects are observed in smooth muscle from human airways and airway smooth muscle of other species.

3. Understanding the mechanistic basis for the effects of these cytokines may prove to be an important step in improving the efficacy of β‐adrenoceptor agonists for the treatment of asthma.
Heat stress increases endothelium‐dependent relaxations and prevents reperfusion‐induced endothelial dysfunction
Authors: Richard Vincent , Laude Karine , Artigues Cecile , Kaeffer Nathalie , Henry Jean‐Paul , Thuillez Christian
Published: March 2003

1. Heat stress has been widely used to stimulate the expression of stress proteins and is associated with various cardiovascular changes, including anti‐ischaemic effects. However, the effect of heat stress on endothelial function is less clear.

2. Heat stress was induced in anaesthetized rats by increasing body temperature to 42°C for 15 min. Twenty‐four hours later, segments of rat aorta and mesenteric and coronary arteries were mounted in organ chambers.

3. Heat stress markedly increased relaxation to acetylcholine (ACh) in all three blood vessels studied, without affecting the response to the nitric oxide (NO) donor sydnonimine‐1.

4. Heat stress also increased aortic relaxation to histamine and the calcium ionophore A23187.

5. In the aorta, an inhibitor of NO synthesis abolished the response to ACh in both control and heat stressed‐rings, whereas a cyclo‐oxygenase inhibitor had no effect.

6. Heat stress also prevented completely the impaired response to ACh in coronary arteries isolated from rats subjected to myocardial ischaemia and reperfusion.

7. Thus, heat stress increases the stimulated release of NO the rat aorta and mesenteric and coronary arteries and prevents reperfusion‐induced injury at the level of the coronary endothelium.
Reduced [125I]‐Bolton Hunter substance P binding (NK1 receptors) in the basal forebrain nuclei of aged rats
Authors: Geraghty Dominic P , Maguire Christine M
Published: March 2003

1. Quantitative autoradiography and homogenate radioligand binding of [125I]‐Bolton Hunter substance P ([125I]‐BHSP) were used to compare brain NK1 receptors in young (2 months) and aged (18–20 months) rats.

2. The autoradiographic distribution and density of [125I]‐BHSP binding sites was similar in all cortical regions of young and aged rats. In contrast, the density of [125I]‐BHSP binding sites was significantly (P < 0.05) lower in the basal forebrain nuclei (intermediate part of the lateral septal nuclei, medial septal nucleus and horizontal and vertical nuclei of the diagonal band) of aged rats. In all other brain regions examined, binding densities were almost identical in young and aged rats.

3. Because a population of NK1 receptors ([125I]‐BHSP binding sites) in the basal forebrain nuclei is associated with cholinergic neurons, the decrease in NK1 receptors in aged rats may reflect degeneration of cholinergic neurons and contribute to the motor and cognitive deficiencies that occur with ageing.
Increased Renal Vascular Resistance In Zinc‐Deficient Rats: Role Of Nitric Oxide and Superoxide
Authors: Kurihara Nobutaka , Yanagisawa Hiroyuki , Sato Masamichi , Tien Chang‐Kuen , Wada Osamu
Published: March 2003

1. Zinc deficiency (ZD) induces many kinds of pathological states. However, the effects of ZD on haemodynamics remain unclear. In the present study, we measured mean blood pressure (BP) and renal blood flow (RBF) under anaesthesia and calculated renal vascular resistance (RVR) from these parameters in rats maintained on a ZD diet (0.5 p.p.m. zinc) for 4 weeks.

2. Zinc deficiency did not change mean BP, but significantly reduced RBF and increased RVR (each P < 0.01). In addition, these effects of ZD were reversible.

3. Because Cu/Zn superoxide dismutase (SOD) is a zinc‐containing enzyme and superoxide is a potent scavenger of nitric oxide (NO), a vasodilator, we hypothesized that one of the mechanisms by which ZD increases RVR is by decreasing NO bioavailability by the enhanced formation of superoxide due to low Cu/Zn SOD activity. To test this hypothesis, we observed the roles of NO and superoxide in the mechanism, after having confirmed the low activity of Cu/Zn SOD in the kidneys of ZD rats.

4. Administration of the SOD mimetic tempol (5 mg/kg per min) decreased RVR to a significantly greater extent in ZD rats compared with control, suggesting that superoxide was responsible for the mechanism. Low doses of the NO donor sodium nitroprusside (SNP; 2.0 µg/kg per min, continuous) decreased RVR to a significantly smaller extent in ZD rats compared with control, whereas a high dose of SNP (0.75 mg/kg, bolus) decreased RVR to a significantly greater extent in ZD rats compared with control, suggesting that the mechanism includes an inhibition of NO activity in ZD, which is most likely to be a scavenging of NO by the activated superoxide.

5. In summary, ZD may increase RVR. The mechanism probably includes changes in NO and superoxide activities.
CLINICAL SIGNIFICANCE OF MEASUREMENT OF RESTING ENERGY EXPENDITURE IN CHILDHOOD
Authors: Amagai Teruyoshi , Mouri Takeshi , Kirii Kyoko , Hori Tetsuo , Kaneko Michio , Ohkawa Haruo
Published: March 2003

1. Biliary atresia (BA), as a common disease in Japan, and cystic fibrosis (CF), as an extremely uncommon disease in Japan, were selected to assess the clinical significance of measurement of energy expenditure (EE).

2. Energy expenditure was significantly higher in children with BA than in normal children.

3. Measurement of EE in BA lead to clues to resolving its mechanism by novel assessment of interleukin‐6 and leptin.

4. Energy expenditure in children with CF is also higher, but this has been addressed by nutritional intervention with additional calories.

5. Individualization of EE measurement is necessary in the analysis of pathological mechanisms and nutritional management of patients with both common and uncommon diseases.
Accuracy of flow and sensors of indirect calorimetry for neonates and infants: Using mass spectrometry and a pneumotachogram
Authors: Amagai Teruyoshi , Mouri Takeshi , Ohkawa Haruo , Nishi Isao
Published: March 2003

1. The accuracy of measurements of flow rate and concentrations of O2 and CO2 in expiratory gas by indirect calorimetry (IC) using paediatric and adult circuits were assessed by mass spectrometry and a pneumotachogram, which have been proven as the most reliable instruments for these purposes.

2. In the paediatric circuit, all measurements were demonstrated to be reliable: error rates for flow rate, O2 concentration and CO2 concentration were +3.13, +2.66 and −6.63%, respectively. All were within the 10%, which is acceptable as a biological error.

3. However, in the adult circuit of IC, all measurements were reliable except for measurements of CO2 concentration: error rates for flow rate, O2 concentration and CO2 concentration were +2.82, +1.64 and +11.42%, respectively.

4. A fluctuation phenomenon of expiratory gas concentration was observed only in IC. Mass spectrometry did not show this phenomenon. This phenomenon seems to be derived from the IC machine itself.
Role of Pax‐5 in the regulation of a mid‐hindbrain organizer’s activity
Authors: Funahashi Jun‐ichi , Okafuji Tatsuya , Ohuchi Hideyo , Noji Sumihare , Tanaka Hideaki , Nakamura Harukazu
Published: March 2003

The mes‐metencephalic boundary (isthmus) has been suggested to act as an organizer in the development of the optic tectum. Pax‐5 was cloned as a candidate for regulator of the organizing center. Isthmus‐specific expression of Pax‐5 and analogy with the genetic cascade in Drosophila suggest that Pax‐5 may be at a higher hierarchical position in the gene regulation cascade of tectum development. To examine this possibility, a gain‐of‐function experiment on Pax‐5 was carried out. In ovo electroporation on E2 chick brain with the eucaryotic expression vector that encodes chick Pax‐5 cDNA was used. Not only was a considerable amount of Pax‐5 expressed ectopically in the transfected brain, but irregular bulging of the neuroepithelium was induced in the diencephalon and mesencephalon. At Pax‐5 misexpressing sites, uptake of BrdU was increased. Histological examination of E7 transfected brain revealed that Pax‐5 caused transdifferentiation of diencephalon into the tectum‐like structure. In the bulges of the E7 mesencephalon, differentiation of laminar structure was repressed when compared to the normal side. In transfected embryos, En‐2, Wnt‐1 and Fgf8 were up‐regulated ectopically, and Otx2 was down‐regulated in the diencephalon to mesencephalon. Moreover, Ephrin‐A2, which is expressed specifically in the tectum with a gradient highest at the caudal end, is suggested to be involved in pathfinding of the retinal fibers, and was induced in the bulges. When the mouse Fgf8 expression vector was electroporated, Pax‐5 and chick Fgf8 were also induced ectopically. These results suggest that Pax‐5, together with Fgf8, hold a higher position in the genetic hierarchy of the isthmus organizing center and regulate its activity.
FGF10 can induce Fgf8 expression concomitantly with En1 and R‐fng expression in chick limb ectoderm, independent of its dorsoventral specification
Authors: Ohuchi Hideyo , Nakagawa Takashi , Itoh Nobuyuki , Noji Sumihare
Published: March 2003

The limb bud has a thickened epithelium at the dorsal–ventral boundary, the apical ectodermal ridge (AER), which sustains limb outgrowth and patterning. A secreted molecule fibroblast growth factor (FGF)10 is involved in inducing Fgf8 expression in the prospective AER and mutual interaction between mesenchymal FGF10 and FGF8 in the AER is essential for limb outgrowth. A secreted factor Wnt7a and a homeobox protein Lmx1 are involved in the dorsal patterning of the limb, whereas a homeobox protein Engrailed 1 (En1) is involved in the dorsal–ventral patterning as well as AER formation. Radical fringe (R‐fng), a vertebrate homolog of Drosophila fringe was also found to elaborate AER formation in chicks. However, little is known about the molecular interactions between these factors during AER formation. The present study clarified the relationship between FGF10, Wnt7a, Lmx1, R‐fng and En1 during limb development using a foil‐barrier insertion experiment. It was found that a foil‐barrier inserted into the chick prospective wing mesenchyme lateral to the mesonephric duct blocks AER induction. This experiment was expanded by implanting Fgf10‐expressing cells lateral to the barrier and examined whether FGF10 could rescue the expression of the limb‐patterning genes reported in AER formation. It was found that FGF10 is sufficient to induce Fgf8 expression in the ectoderm of the foil‐inserted limb bud, concomitantly with R‐fng and En1 expression. However, FGF10 could not rescue the expression of the dorsal marker genes, Wnt7a or Lmx1. Thus, it is suggested that epithelial factors of En1 and R‐fng can induce Fgf8 expression in the limb ectoderm in cooperation with a mesenchymal factor FGF10. Some factor(s) other than FGF10, possibly from the paraxial structures medial to the limb mesoderm, is responsible for the initial dorsal–ventral specification of the limb bud.
Expression of helix–loop–helix type negative regulators of differentiation during limb regeneration in urodeles and anurans
Authors: Shimizu‐Nishikawa Keiko , Tazawa Ichiro , Uchiyama Kohji , Yoshizato Katsutoshi
Published: March 2003

The urodele is capable of regenerating its limb by forming a blastema even in the adult. By contrast, the anuran, which is phylogenetically close to the urodele, loses this ability during metamorphosis and forms blastema‐like tissues that develop only into a spike‐like structure in the adult. In order to compare the molecular mechanism of the formation and maintenance of the blastema between the urodele and anuran, the genes encoding helix–loop–helix (HLH) type negative regulators of differentiation were characterized for both the Japanese newt, Cynops pyrrhogaster, and African clawed frog, Xenopus laevis. Cynops homologs of Id2, Id3, and HES1 and Xenopus Id2 were identified. To learn the roles of these genes in regeneration, their expression was examined. The expression of Id2 and Id3 was low in unamputated limbs, but was up‐regulated in blastemas of both adult newt and Xenopus. Interestingly, transcripts of the two Id genes showed specific localizations in the blastema and the expression patterns were very similar in both species through the early to medium bud stage. Id2 was expressed predominantly in the blastemal epidermis, and Id3 was expressed equally in the blastemal epidermis and mesenchyme including cells in precartilage condensations. HES1 expression was up‐regulated in the newt blastemal epidermis. It was thought that the up‐regulation of these genes in the epidermis was related to the proliferation of the cells and that increased expression of these genes in the mesenchyme was related to the undifferentiated state of the blastemal cells. These results and considerations strongly suggested that the state of differentiation is similar in the early to medium bud blastema of both urodeles and anurans. The expression of Id3 remained high through to the digits stage in newts. In contrast, its expression in Xenopus decreased in spike‐like regenerates, which correspond to palette‐digits stage of newt regenerates. From these results, it was suggested that the blastema redifferentiates earlier in the frog than in the newt, and therefore the timing of redifferentiation of the cartilage is crucial for complete regeneration.
Sulfated O‐linked glycans of the vitelline coat as ligands in gamete interaction in the ascidian, Halocynthia roretzi
Authors: Baginski Tomasz , Hirohashi Noritaka , Hoshi Motonori
Published: March 2003

In the ascidian Halocynthia roretzi, sperm–egg binding is probably mediated through the interaction between α‐L‐fucosidase present on the sperm surface and anionic saccharide chains of the egg vitelline coat. To characterize biologically active glycans, total glycans were chemically released from the glycopeptide fraction of the vitelline coat. The fraction of uncharged glycans and two fractions of negatively charged glycans were separated by diethylaminoethyl‐anion exchange chromatography. In a competitive inhibition assay of fertilization, both anionic fractions showed inhibitory activity, with more anionic glycans being most potent, while uncharged glycans were biologically inactive. Chemical desulfation combined with a competitive inhibition assay of fertilization and ion analysis determined that sulfate groups were responsible for anionic character and crucial for biological activity. Monosaccharide analysis of anionic fractions showed a high content of N‐acetylgalactosamine, galactose, xylose and the presence of arabinose, mannose, N‐acetylglucosamine, glucose and rhamnose. Glycans were O‐linked and galactose and xylose residues were detected at reducing termini. Linkage analysis suggested that 1,4‐linked xylose, 1,3‐linked galactose and N‐acetylgalactosamine residues, substituted to different degrees by sulfate groups on the C‐3 and C‐4 carbons, respectively, constituted the core structures of anionic glycans.
Interactions between cytoskeletal components during myoplasm rearrangement in ascidian eggs
Authors: Chiba Shota , Miki Yuko , Ashida Kazunori , Wada Michiko R. , Tanaka Kimio J. , Shibata Yumiko , Nakamori Rinna , Nishikata Takahito
Published: March 2003

Ooplasmic segregation in ascidian eggs consists of two phases of cytoplasmic movement, the first phase is mediated by the microfilament system and the second is mediated by the microtubule system. Recently, two novel proteins, p58 and myoplasmin‐C1, which are localized to the myoplasm, were suggested to have important roles in muscle differentiation. In order to analyze the molecular mechanisms underlying ooplasmic segregation, the interactions between actin, tubulin, p58 and myoplasmin‐C1 were examined. During the first segregation, microtubule meshwork in the unfertilized egg disappeared. At the second segregation, a novel structure of the microtubules that extended from the sperm aster and localized in the cortical region of the myoplasm was found. Moreover, uniform distribution of the cortical actin filament was observed at the second segregation. During the course of myoplasm rearrangement, p58 and myoplasmin‐C1 are colocalized and can form a molecular complex in vitro. This complex of p58 and myoplasmin‐C1 is a good candidate for a cytoskeletal component of the myoplasm, and is likely to be involved in the correct distribution of cytoplasmic determinants.
Gene transfer into chicken embryos by retrovirus vectors
Authors: Iba Hideo
Published: March 2003

While chickens have many properties that are advantageous for embryological studies, their genetic analysis has been restricted. However, by using retrovirus vector systems in combination with classical techniques of experimental developmental biology, it has recently become possible to analyze the function of genes involved in the development of this organism. Avian retrovirus vectors are unique in that they can be divided into two categories: replication‐competent and replication‐defective (replication‐incompetent). By choosing the vectors correctly, there are many experimental applications of these vectors such as induction of constitutive (or regulated) gene expression in a restricted region of tissues, organs and embryos; cell lineage analysis; and formation of concentration gradients of morphogens in micromass cultures. In this paper, several retrovirus vectors available for the chicken will be introduced and their applications in developmental biology will be reviewed.
Xoom is required for epibolic movement of animal ectodermal cells in Xenopus laevis gastrulation
Authors: Hasegawa Kouichi , Kinoshita Tsutomu
Published: March 2003

Gastrulation is the most dynamic cell movement and initiates the body plan in amphibian development. In contrast to numerous molecular studies on mesodermal induction, the driving force of gastrulation is as yet poorly understood. A novel transmembrane protein, Xoom, was previously reported, which is required for Xenopus gastrulation. In the present study, the role of Xoom during Xenopus gastrulation was further examined in detail. Overexpression and misexpression of Xoom induced overproduction of Xoom protein, but not a changed phenotype. However, Xoom antisense ribonucleic acid (RNA) injection reduced the Xoom protein and caused gastrulation defects without any influence on the involution and translation levels of mesodermal marker genes. Normal migrating activity of dorsal mesodermal cells was recognized in the antisense RNA‐injected explant. Morphological examination using artificial exogastrulation showed that convergent extension of mesodermal cells occurred normally, but the ectodermal cell layer significantly shrank in the antisense RNA‐injected embryo. Comparison of cell shape among various experimental conditions showed that inhibition of cell spreading occurs specifically in the outer ectodermal layer of the antisense RNA‐injected embryo. Cytochemical examination indicated disorganization of F‐actin in the ectodermal cells of the antisense RNA‐injected embryo. These results suggest that Xoom plays an important role in the epibolic movement of ectodermal cells through some regulation of actin filament organization.