bluuebunny/crossref_metadata_2025 huggingface dataset:
Circumventricular Organs: Gateways to the Brain Subcellular Mechanisms Of Area Postrema Activation
Authors: Hay Meredith
Published: March 2003

1. The present paper reviews some of the recent studies regarding the cellular mechanims regulating area postrema (AP) neuronal activity.

2. The calcium currents involved in activation and regulation of AP neuronal activity most likely belong to the Q‐ and R‐family of calcium channels.

3. Angiotensin II increases AP activity by facilitating inward calcium currents and inhibiting calcium‐activated potassium currents.

4. The heterogeneity observed regarding responsiveness to glutamate and 5‐hydroxytryptamine and peptides may reflect multiple populations of AP neurons, each with specific roles in the integration of afferent and peptidergic neurotransmission.
Circumventricular Organs: Gateways to the Brain Role Of Circumventricular Organs In Pro‐Inflammatory Cytokine‐Induced Activation Of The Hypothalamic– Pituitary–Adrenal Axis
Authors: Buller Kathryn M
Published: March 2003

1. The past 15 years has seen the emergence of a new field of neuroscience research based primarily on how the immune system and the central nervous system can interact. A notable example of this interaction occurs when peripheral inflammation, infection or tissue injury activates the hypothalamic– pituitary–adrenal axis (HPA).

2. During such assaults, immune cells release the pro‐ inflammatory cytokines interleukin (IL)‐1, IL‐6 and tumour necrosis factor‐α into the general circulation.

3. These cytokines are believed to act as mediators for HPA axis activation. However, physical limitations of cytokines impede their movement across the blood–brain barrier and, consequently, it has been unclear as to precisely how and where IL‐1β signals cross into the brain to trigger HPA axis activation.

4. Evidence from recent anatomical and functional studies suggests two neuronal networks may be involved in triggering HPA axis activity in response to circulating cytokines. These are catecholamine cells of the medulla oblongata and the circumventricular organs (CVO).

5. The present paper examines the role of CVO in generating HPA axis responses to pro‐inflammatory cytokines and culminates with a proposed model based on cytokine signalling primarily involving the area postrema and catecholamine cells in the ventrolateral and dorsal medulla.
Rigor Force Responses Of Permeabilized Fibres From Fast And Slow Skeletal Muscles Of Aged Rats
Authors: Plant David R , Lynch Gordon S
Published: March 2003

1. Ageing is generally associated with a decline in skeletal muscle mass and strength and a slowing of muscle contraction, factors that impact upon the quality of life for the elderly. The mechanisms underlying this age‐related muscle weakness have not been fully resolved. The purpose of the present study was to determine whether the decrease in muscle force as a consequence of age could be attributed partly to a decrease in the number of cross‐bridges participating during contraction.

2. Given that the rigor force is proportional to the approximate total number of interacting sites between the actin and myosin filaments, we tested the null hypothesis that the rigor force of permeabilized muscle fibres from young and old rats would not be different.

3. Permeabilized fibres from the extensor digitorum longus (fast‐twitch; EDL) and soleus (predominantly slow‐twitch) muscles of young (6 months of age) and old (27 months of age) male F344 rats were activated in Ca2+‐buffered solutions to determine force–pCa characteristics (where pCa = –log10[Ca2+]) and then in solutions lacking ATP and Ca2+ to determine rigor force levels.

4. The rigor forces for EDL and soleus muscle fibres were not different between young and old rats, indicating that the approximate total number of cross‐bridges that can be formed between filaments did not decline with age. We conclude that the age‐related decrease in force output is more likely attributed to a decrease in the force per cross‐bridge and/or decreases in the efficiency of excitation–contraction coupling.
Metalloendopeptidases EC 3.4.24.15 and EC 3.4.24.16 And Bradykinin B2 Receptors Do Not Play Important Roles In Renal Wrap Hypertension In Rabbits
Authors: Norman M Ursula , Lew Rebecca A , Smith A Ian , Denton Kate M , Evans Roger G
Published: March 2003

1. The aim of the present study was to determine the effects of the metalloendopeptidase (EP) 24.15 and 24.16 inhibitor N‐[1‐(R,S)‐carboxy‐3‐phenylpropyl]‐Ala‐Aib‐Tyr‐p‐aminobenzoate (JA‐2) on haemodynamics and renal function in conscious rabbits with two‐kidney, two‐wrapped hypertension. We have also examined the role of endogenous bradykinin in the maintenance phase of this form of renovascular hypertension and whether inhibition of bradykinin degradation contributes to any potential effects of JA‐2.

2. In two preliminary operations, rabbits were equipped with transit‐time ultrasound flow probes for measuring cardiac output (CO) and renal blood flow (RBF) and had both kidneys wrapped in cellophane. Starting 4 weeks after the last operation, rabbits underwent four studies (3–5 days apart), during which they were treated with combinations of the bradykinin B2 receptor antagonist icatibant or its vehicle (1 mL/kg bodyweight 0.9% w/v NaCl) and JA‐2 or its vehicle (1 mL/kg of a 5% w/v 2‐hydroxypropyl‐β‐cyclodextrin, 2.5% v/v dimethylsulphoxide solution). Renal function was monitored using standard renal clearance methods.

3. Icatibant (10 μg/kg) had no significant effects on systemic haemodynamic variables (mean arterial pressure, heart rate or CO), renal haemodynamic variables (RBF or glomerular filtration rate), urine flow or sodium excretion. At 5 mg/kg plus 3 mg/kg per h, JA‐2 also did not affect any of these variables, either after icatibant vehicle treatment or after icatibant treatment.

4. Our data do not support major roles for endogenous bradykinin or bradykinin degradation by EP 24.15/24.16 in the control of systemic and renal haemodynamics or renal excretory function in two‐kidney, two‐wrapped hypertension in rabbits.
Inflammation As A Key Event In The Development Of Neointima Following Vascular Balloon Injury
Authors: Wainwright Cl , Miller Am , Wadsworth Rm
Published: March 2003

1. The present review discusses the current evidence to implicate leucocytes as key players in the development of neointima in arteries that have been subjected to balloon angioplasty injury.

2. There is substantial clinical evidence that leucocytes are activated after angioplasty, as determined by increased plasma levels of both leucocyte granulation products and soluble leucocyte and endothelial cell adhesion molecules.

3. Experimental evidence to implicate leucocytes in neointimal formation comes from studies that demonstrate leucocyte accumulation occurs within the vascular wall soon after injury and that induction of leukopenia prevents neointimal formation.

4. The evidence implicating specific adhesion molecules and cytokines in the key events leading to neointimal formation is discussed.
Blood Pressure Lowering In Diabetes: A Brief Review Of The Current Evidence And Description Of A New Trial
Authors: Patel Anushka , Chalmers John , Neal Bruce , Chapman Neil , Girgis Seham , MacMahon Stephen
Published: March 2003

1. Diabetes is a major global public health problem. The prevalence of this disease is predicted to increase sharply in the coming decades, particularly in less‐developed regions of the world.

2. Most premature morbidity and mortality associated with diabetes relates to markedly increased risks of major cardiovascular diseases, such as myocardial infarction and stroke (macrovascular events), as well as microvascular complications, such as nephropathy and retinopathy.

3. Hypertension is a prevalent and important risk factor for vascular events in these patients. However, observational data demonstrate a continuous relationship between blood pressure and risk of vascular events, suggesting that even those individuals considered normotensive may benefit from blood pressure lowering.

4. Trials of blood pressure lowering among mostly hypertensive individuals with diabetes have demonstrated benefit of intervention on macrovascular and microvascular outcomes. Recent data may suggest additional effects of angiotensin‐ converting enzyme inhibitors independent of blood pressure lowering.

5. Issues where data are lacking with respect to blood pressure lowering in diabetes include the effects of blood pressure lowering among non‐hypertensive individuals and the effects of blood pressure lowering regimens based on different classes of drug.

6. Data expected to address some of these issues are being collected. These include a prospective meta‐analysis of blood pressure‐lowering trials with large numbers of patients with diabetes. A new large‐scale randomised trial, ADVANCE (Action in Diabetes and Vascular Disease), is also described.
Effect Of Vasodilatory β‐Adrenoceptor Blockers On Cardiovascular Haemodynamics In Anaesthetized Rats
Authors: Takahashi Hakuo , Masaki Hiroya , Komiyama Yutaka , Masuda Midori , Nishimura Masato
Published: March 2003

1. The effect of vasodilatory β‐adrenoceptor blockers on regional blood flow in the major organs of anaesthetized rats was investigated using radioactive microspheres. The administration of propranolol and saline was used as a control.

2. Intravenous injections of carvedilol (2 mg/rat), celiprolol (20 mg/rat) and bopindolol (1 mg/rat) equally decreased systemic blood pressure (SBP) by approximately 20 mmHg, whereas propranolol (1 mg/rat) decreased SBP only slightly but not significantly.

3. Heart rate was significantly decreased by carvedilol, celiprolol, bopindolol and propranolol.

4. Coronary blood flow was markedly increased by carvedilol, but not by the other three drugs.

5. Cardiac output tended to decrease following the administration of all four drugs.

6. Total peripheral vascular resistance was not significantly affected by carvedilol, celiprolol and bopindolol, but was markedly increased following propranolol.

7. Renal blood flow was markedly increased by carvedilol.

8. Blood flow in the brown fat was markedly increased by carvedilol and bopindolol, but not by celiprolol and propranolol.

9. These findings indicate that the newer vasodilatory beta‐blockers, such as carvedilol and bopindolol, have a beneficial effect on the regional circulation in contrast with the classical beta‐blocker propranolol.

10. The regional haemodynamic effects observed in the present study following intravenous injection of the beta‐blockers may help explain the clinical experience that vasodilatory beta‐blockers increase insulin sensitivity and decrease mortality in patients with congestive heart failure.
Responses To Microinjections Of Endomorphin And Nociceptin Into The Medullary Cardiovascular Areas
Authors: Sapru HN , Chitravanshi VC
Published: March 2003

1. Cardiovascular effects of microinjections of nociceptin and endomorphin‐2 into the following medullary areas were studied in urethane‐anaesthetized rats: chemoreceptor projection site (CPS), intermediate portion of the nucleus tractus solitarius (I‐NTS), caudal ventrolateral medullary depressor area (CVLM) and rostral ventrolateral medullary pressor area (RVLM).

2. Microinjections of nociceptin or endomorphin‐2 (0.6 mmol/L each) into the CPS and RVLM elicited depressor and bradycardic responses, whereas similar injections into the I‐NTS and CVLM elicited pressor and tachycardic responses.

3. The mechanism of cardiovascular responses to microinjections of these opioid peptides into different medullary areas involved in cardiovascular function can be postulated as follows: the direct effect of nociceptin and endomorphin‐2 on neurons is usually inhibitory. Because the activation of CPS and RVLM by microinjections of L‐glutamate results in pressor and tachycardic responses, inhibition
of these areas by nociceptin and endomorphin‐2 elicits depressor and bradycardic
responses. Similarly, activation of neurons in the I‐NTS and CVLM by microinjections
of L‐glutamate elicits depressor and bradycardic responses. Therefore, inhibition of these areas by microinjections of these opioid peptides elicits an increase in blood pressure and heart rate.
Effects Of Combined Pioglitazone And Metformin On Diabetes And Obesity In Wistar Fatty Rats
Authors: Suzuki Masami , Odaka Hiroyuki , Suzuki Noriko , Sugiyama Yasuo , Ikeda Hitoshi
Published: March 2003

1. The effects of combined treatment with pioglitazone·HCl and metformin on diabetes and obesity were investigated in Wistar fatty rats, which are hyperglycaemic and hypertriglyceridaemic and have higher plasma levels of total ketone bodies than lean rats.

2. Plasma glucose was significantly decreased when pioglitazone·HCl or metformin was administered alone and combined treatment accentuated this decrease. The administration of pioglitazone·HCl, but not metformin, also decreased plasma levels of triglyceride and total ketone bodies.

3. The glycogen content of skeletal muscle was not increased by pioglitazone·HCl or metformin alone, but was increased by combined treatment (P = 0.003, ANOVA).

4. Pioglitazone·HCl produced increased food intake and bodyweight in hyperphagic Wistar fatty rats; however, concurrent administration of metformin significantly ameliorated these pioglitazone·HCl‐induced increases.

5. These results indicate that combined treatment with pioglitazone·HCl and metformin induces a marked hypoglycaemic effect accompanied by a reduction in plasma levels of total ketone bodies and prevention of excessive bodyweight gain in Wistar fatty rats. These favourable effects suggest that the combination would be beneficial in treating patients with type 2 diabetes.
Altered Expression Of Vascular Natriuretic Peptide Receptors In Experimental Hypertensive Rats
Authors: Lee JongUn , Kim Sunmi , Jung Mehye , Oh YoonWha , Kim Soo Wan
Published: March 2003

1. The aim of the present study was to determine whether the regulation of vascular natriuretic peptide receptors (NPR) is related to the local renin–angiotensin system (RAS).

2. Male Sprague‐Dawley rats were made two‐kidney, one‐clip (2K1C) and deoxycorticosterone acetate (DOCA)‐salt hypertensive to activate and inhibit the RAS, respectively. Another model of hypertension was induced by treatment with an inhibitor of nitric oxide synthesis, namely NG‐nitro‐L‐arginine methyl ester
(L‐NAME).

3. The mRNA expression of NPR‐A, NPR‐C, angiotensin‐ converting enzyme (ACE) and angiotensin AT1 receptors was determined in the thoracic aorta by semiquantitative reverse transcription–polymerase chain reaction. The particulate guanylyl cyclase activity stimulated by atrial natriuretic peptide (ANP) was also determined in the membrane fraction of the thoracic aorta.

4. The plasma concentrations of ANP were increased significantly in the three models of hypertension. Plasma renin activity was increased in 2K1C hypertension, decreased in DOCA‐salt hypertension and not significantly altered in L‐NAME hypertension.

5. The mRNA expression of NPR‐A and NPR‐C was decreased, whereas that of ACE and AT1 receptors was increased in 2K1C and L‐NAME hypertension.
The mRNA expression of NPR‐A and NPR‐C was increased, whereas that of ACE and AT1 receptors was decreased in DOCA‐salt hypertension.

6. The particulate guanylyl cyclase activity was decreased in 2K1C and L‐NAME hypertension and increased in DOCA‐salt hypertension.

7. The vascular expression of NPR may be reciprocally regulated by local RAS activity.
Decrease In Catechol‐O‐Methyltransferase Activity In The Liver Of Streptozotocin‐Induced Diabetic Rats
Authors: Wang JP , Liu IM , Tzeng TF , Cheng JT
Published: March 2003

1. The present study compared the activity of catechol‐ O‐methyltransferase (COMT) in the liver and plasma of streptozotocin (STZ)‐induced diabetic rats with that in normal rats. The activity of COMT was estimated by the metabolism of noradrenaline to metanephrine (MN), both measured by high‐performance liquid chromatography with electrochemical detection.

2. Formation of MN was decreased in the liver of STZ‐ diabetic rats compared with normal rats. The amount of MN was also decreased in plasma obtained from STZ‐diabetic rats. A Michaelis–Menten plot showed a reduction in the maximum velocity and an increase in the Km for COMT in liver samples from STZ‐diabetic rats.

3. The role of hyperglycaemia in the lowering of COMT activity was then investigated using phlorizin or insulin at doses sufficient to normalize plasma glucose from STZ‐diabetic rats. Both insulin and phlorizin treatment of STZ‐diabetic rats for 4 days restored the activity of COMT to that seen in normal rats. Thus, correction of hyperglycaemia in STZ‐diabetic rats can reverse the decrease in COMT activity. The activity of COMT is lower in STZ‐diabetic rats than in normal rats mainly due to the higher plasma glucose.
Effects Of A 1 Day Fast On Biohumoral Variables Associated With Human Circadian Rhythmicity
Authors: Uezono Keiko , Kawasaki Terukazu , Itoh Kazue , Cugini Pietro
Published: March 2003

1. The aim of the present study was to investigate the effects of a short (1 day) fast by testing biohumoral variables associated with the human circadian rhythm.

2. Fifteen clinically healthy male volunteers (32 ± 8 years old) participated in the study. Subjects were fed a control diet for 7 days. The last day was a control day and the following 8th day was the fasting day. Each subject was asked to collect urine seven times over a 24 h period. Chemical and hormonal variables were measured in each fractionated urine specimen. The time‐ qualified urinary excretion rates were biometrically analysed using conventional and chronobiological methods.

3. During fasting, significant incremental changes were detected in the urinary excretion rates of potassium, aldosterone, 17‐hydroxycorticosteroids and adrenaline and significant decremental changes were detected in the excretion rates of sodium, chloride, creatinine, urea nitrogen, uric acid, 17‐ketosteroids, noradrenaline and dopamine. The circadian rhythmicity of the variables was well preserved and remained almost stable throughout the fasting phase.

4. Fasting affected the mean oscillatory levels and oscillatory amplitudes of variables, suggesting that nutrients may have played roles as tonic and phasic modulators on the mechanisms that physiologically regulate ircadian rhythmicity.
Effects Of Blockade Of α‐ and β‐Adrenoceptors And Neuropeptide Y1 Receptors, As Well As Brachial Plexus Blockade, On Endothelium‐Dependent Vasodilation In The Human Forearm
Authors: Johansson Kristina , Eriksson Mats , Wahlqvist Inger , Mühlen Bengt von zur , Lind Lars
Published: March 2003

1. The aim of the present study was to investigate the effects of α‐adrenoceptor
blockade (phentolamine), β‐adrenoceptor blockade (propranolol), neuropeptide
Y1 receptor blockade and neurogenic blockade (brachial plexus) on endothelium‐dependent vasodilation (EDV) in the human forearm.

2. Forty‐four young healthy volunteers underwent forearm blood flow (FBF) measurements, using venous occlusion plethysmography, during local intra‐arterial infusions of methacholine (MCh; inducing EDV) and sodium nitroprusside (SNP; inducing endothelium‐independent vasodilation (EIDV)). These measurements were undertaken at baseline and were repeated with either concomitant local intra‐arterial infusion of phentolamine (n = 8),
propranolol (n = 7) or saline (n = 6) in the forearm, neuropeptide
Y1 receptor blockade (n = 12) given i.v. or during axillary plexus
blockade (n = 11).

3. Both α‐adrenoceptor blockade and neurogenic blockade induced an
upward shift in the dose–response curve for both EDV and EIDV. β‐Adrenoceptor
blockade did not change resting FBF or EIDV, but induced a significant decrease in
EDV (P = 0.015). Neuropeptide Y1 receptor blocker induced no significant changes in resting FBF, EDV and EIDV and neither did saline. No changes in blood pressure or heart rate were induced by any of the blockades.

4. Whereas β‐adrenoceptor blockade impaired EDV, α‐adrenoceptor blockade and neurogenic blockade caused a general vasodilation that was not endothelium dependent. Neuropeptide Y does not seem to influence blood flow in the resting forearm.
Functional Analysis Of Histamine Receptor Subtypes Involved In Endothelium‐Mediated Relaxation Of The Human Uterine Artery
Authors: Spitaler Michaela M , Hammer Astrid , Malli Roland , Graier Wolfgang F
Published: March 2003

1. This work was designed to introduce human uterine arteries as a new model for cardiovascular research. Advantages of this model include considerable availability of tissue because of the appearance of uterus myomatosus in post‐menopausal women who undergo surgery and the chance to work on dysfunctional and healthy vessels.

2. Histamine evoked relaxation of the uterine artery that was prevented by removal of the endothelium or by the presence of NG‐nitro‐L‐arginine.

3. Receptor antagonists for histamine H1 (mepyramine) and H2 (ranitidine) receptors increased the EC50 of histamine by 112‐ and 67‐fold, respectively.

4. Remarkably, isolated uterine arteries could be stored in incubators for 5 days without any change in contractility to phenylephrine and endothelium‐dependent relaxation to acetylcholine and histamine.

5. Endothelial cells could be isolated and cultured in high purity, as demonstrated by histochemical staining of factor VIII, low CD45‐RO for macrophages and no smooth muscle α‐actin. In addition, cultured human uterine artery endothelial cells could be used for single cell Ca2+ measurements.

6. In agreement with our findings in the intact vessel, histamine‐initiated elevation of the intracellular free Ca2+ concentration was reduced in the presence of mepyramine and ranitidine by 59 and 55%, respectively.

7. These data indicate that, in the human uterine artery, H1 and H2 receptors are involved in histamine‐induced endothelium‐dependent relaxation that is mediated by nitric oxide.

8. In addition, this vessel can be stored for possible virus‐mediated gene expression for 5 days without any loss of reagibility.

9. Finally, endothelial cells can be isolated and cultured from the human uterine artery and maintain their reactivity to histamine in culture.
Problems In The Use Of Herbal And Natural Substances, With A Specific Example Concerning The Cardiovascular System
Authors: Pang Peter KT , Benishin Christina , Lewanczuk Richard , Shan Jacqueline
Published: March 2003

1. There has been increasing awareness and use of natural preparations for health purposes by consumers.

2. However, recent studies have repeatedly shown that many natural products marketed as nutraceuticals or health food do not deliver the health benefit as claimed and are inconsistent from batch to batch.

3. The present paper describes the scientific rationale of such inconsistency and uses an antihypertensive preparation as an example to demonstrate the significant value of natural products if developed scientifically and properly.
Intratracheal Administration Of Glucocorticoids Using Surfactant As A Vehicle
Authors: Nimmo Alan J , Carstairs Jill R , Patole Sanjay K , Whitehall John , Davidson Kate , Vink Robert
Published: March 2003

1. Glucocorticoids are an effective treatment in the amelioration of chronic lung disease in neonates. However, systemic administration of glucocorticoids to neonates is associated with significant side‐effects that preclude them as an early intervention to prevent onset of the condition. Conversely, local intratracheal administration of glucocorticoids may prevent inflammatory insult to the lungs without the development of systemic side‐effects. We therefore investigated whether local intratracheal delivery of corticosteroids could be facilitated using surfactant as a vehicle.

2. Addition of dexamethasone to either diluted or commercial artificial surfactant, Survanta (Abbott Industries, Sydney, NSW, Australia), did not alter the surface properties of the surfactant.

3. After intratracheal instillation to rats, radiolabelled dexamethasone in Survanta was well distributed throughout all four lobes of the lungs. A concentration gradient of the steroid was observed between the root and the peripheral sections of all lobes.

4. Our results suggest that surfactant is an effective vehicle for intratracheal delivery of glucocorticoids. Moreover, we propose that prophylactic intratracheal administration of glucocorticoids administered shortly after birth may prevent inflammatory insult to the lungs and thereby reduce the likelihood of chronic lung disease developing.
Clinical trials and tribulations
Authors: McInnes Gordon T
Published: March 2003

1. Pharmacologists should be involved in all stages of drug development. Often neglected is the final step, the clinical trials and other studies that determine clinical utility. The present article illustrates how pharmacoepidemiology can facilitate evaluation of the clinical potential of different drugs used to treat hypertension.

2. The evidence base for the drug treatment of hypertension is very strong. Large‐scale outcome trials, largely based on diuretics, indicate that stroke events are prevented to the extent expected from blood pressure reduction, but there appears to be a shortfall in the prevention of coronary heart disease events.

3. On theoretical grounds, newer agents may be expected to have benefits in coronary heart disease prevention beyond blood pressure reduction. Recent trials with angiotensin‐converting enzyme inhibitors and calcium channel blockers suggest no advantage over conventional drugs, but shortcomings in these studies mean that each is uninformative.

4. Observational studies based on pharmacoepidemiological principles offer an alternative approach to evaluating outcomes in treated hypertensives.

5. Evidence from the Glasgow Blood Pressure Clinic database suggest that there are outcome differences between antihypertensive agents. Angiotensin‐converting enzyme inhibitor treatment is associated with a mortality advantage, whereas calcium channel blocker therapy is associated with a poorer prognosis. Preliminary findings from a primary care database support these observations.

6. Long‐term follow up of a well‐documented high‐risk clinical population may allow detection of outcome differences not apparent in relatively short‐term clinical trials.

7. Appropriate interpretation of observational data necessitates an understanding of the strengths and limitations of observational data. Clinical pharmacologists have a critical role in design and evaluation of pharmacoepidemiology studies.
Effects of glutathione S‐transferase inhibitors on nitroglycerin action in pig isolated coronary arteries
Authors: Matsuzaki Toshihiro , Sakanashi Mayuko , Nakasone Junko , Noguchi Katsuhiko , Miyagi Kanako , Sakanashi Makiko , Kukita Ichiro , Aniya Yoko , Sakanashi Matao
Published: March 2003

1. The present study was designed to clarify the role of glutathione S‐transferase (GST) in the vasorelaxation response and development of tolerance to nitroglycerin (GTN) using GST inhibitors.

2. In pig isolated coronary arteries, GST activity was significantly changed to 77 and 82, or 69% of the control level (100%) following treatment with bromosulphophthalein (BSP; 10−3 and 10−4 mol/L) or ethacrynic acid (ETA; 10−4 mol/L), both GST inhibitors, respectively, but not following treatment with 10−3 and 10−4 mol/L GTN (GST activity 97 and 98% of control, respectively).

3. In KCl‐contracted coronary artery strips pre‐incubated with 10−5 and 10−4 mol/L GTN, 10−4 and 10−3 mol/L BSP or 10−4 mol/L ETA, concentration‐dependent relaxations produced by GTN were significantly decreased compared with control.

4. 8‐Bromo cGMP (8‐Br‐cGMP), a membrane‐permeable cGMP analogue, produced concentration‐dependent relaxations in GTN‐pretreated arterial strips that were identical to control responses. However, there was weak but significant decrease in concentration‐dependent relaxations in response to 8‐Br‐cGMP in BSP‐ and ETA‐pretreated arteries.

5. The cGMP content in coronary arteries was significantly increased with GTN, GTN + BSP or GTN + ETA to similar high levels compared with control.

6. The results of the present study show that BSP and ETA decrease GTN‐ and 8‐Br‐cGMP‐induced vasorelaxation, but have no effect on the GTN‐induced increase in cGMP content in coronary arteries, suggesting a possibility that the GST inhibitors may have depressant actions on GTN‐ and 8‐Br‐cGMP‐induced vasorelaxation through direct inhibition of the vasorelaxation of vascular smooth muscle themselves, in addition to having inhibitory effects GST activity.
Responses to endothelium‐derived factors and their interaction in mesenteric arteries from Wistar‐Kyoto and stroke‐prone spontaneously hypertensive rats
Authors: Sekiguchi Fumiko , Nakahira Tomohiro , Kawata Kyoko , Sunano Satoru
Published: March 2003

1. Responses to endothelium‐derived nitric oxide (EDNO), indomethacin‐sensitive endothelium‐derived contracting factor (EDCF) and hyperpolarization by endothelium‐derived hyperpolarizing factor (EDHF) and the interaction among these factors in mesenteric arteries from 16‐week‐old Wistar Kyoto (WKY) rats and age‐matched stroke‐prone spontaneously hypertensive rats (SHRSP) were studied, observing the time‐course of the response to 10−5 mol/L acetylcholine (ACh).

2. The effects of EDNO, EDCF and EDHF were blocked by Nω‐nitro‐l‐arginine (10−4 mol/L), indomethacin (10−5 mol/L) and a combination of apamin (5 × 10−6 mol/L) and charybdotoxin (10−7 mol/L), respectively.

3. The response to EDNO observed in the absence of EDCF and EDHF was not different between preparations from WKY rats and SHRSP. The response to EDCF observed in the absence of EDNO and EDHF was slightly greater in preparations from SHRSP. The response to EDHF in the absence of EDNO and EDCF was much greater in preparations from WKY rats.

4. Endothelium‐derived contracting factor attenuated the relaxation in response to EDNO, the attenuation being greater in preparations from SHRSP. Relaxation in response to EDNO was blocked by EDHF in preparations from WKY rats, but not in preparations from SHRSP.

5. The response to EDCF was augmented by both EDNO and EDHF. The augmentation was greater in preparations from SHRSP.

6. The response to EDHF was attenuated by EDNO in preparations from WKY rats, but not in preparations from SHRSP. The response to EDHF was attenuated by EDCF in preparations from both WKY rats and SHRSP, the attenuation being greater in preparations from SHRSP.

7. These results suggest that there are interactions among these factors in terms of their release or the response to ACh in mesenteric arteries that differ between preparations from WKY rats and SHRSP. In addition, involvement of factors other than these three factors, which also differs between preparations from WKY rats and SHRSP, is suggested.
Primary mesenchyme cell‐ring pattern formation in 2D‐embryos of the sea urchin
Authors: Katow Hideki , Nakajima Yoko , Uemura Isao
Published: March 2003

Primary mesenchyme cell (PMC) migration during PMC‐ring pattern formation was analyzed using computer‐assisted time‐lapse video microscopy in spread embryos (2D‐embryo) of the sea urchin, Mespilia globulus, and a computer simulation. The PMC formed a near normal ring pattern in the 2D‐embryos, which were shown to be an excellent model for the examination of cell behavior in vivo by time‐lapse computer analysis. The average migration distance of the ventro‐lateral PMC aggregate‐forming cells (AFC) and that of the dorso‐ventral PMC cable‐forming cells (CFC) showed no significant difference. All PMC took a rather straightforward migration path to their destinations with little lag time after ingression. This in vivo cell behavior fitted well to a computer simulation with a non‐diffusable chemotaxis factor in the cyber‐cell migration field. This simulation suggests that PMC recognize their destination from a very early moment of cell migration from the vegetal plate, and implicates that a chemoattractive region is necessary for making the PMC migration pattern. The left‐ and right‐lateral AFC and dorso and ventral CFC were each derived from an unequally divided one‐quarter segment of the vegetal plate. This suggests that AFC and CFC have a distinctive ancestor in the vegetal plate, and the PMC are a heterogeneous population at least in terms of their destination in the PMC‐ring pattern.
Mutations with sensory ray defect unmask cuticular glycoprotein antigens in Caenorhabditis elegans male tail
Authors: Ko Frankie C. F. , Chow King L.
Published: March 2003

Caenorhabditis elegans male tail has nine pairs of bilaterally symmetric ray processes extended into a cuticular fan. The formation of these structures involves both cell lineage differentiation and cellular morphogenesis. Nine mutations were examined, all of which presented an amorphous ray phenotype. Glycoconjugates carrying an N‐acetylglucosamine (GlcNAc) epitope were detected at a high level in their male bursa. It was shown that these antigens are not responsible for the morphological defects. It was further demonstrated that these ram and mab gene products represent critical components for male tail cuticle organization. Mutations of them abolish the integrity of the male bursal cuticle and unmask the underlying GlcNAc epitope.
Heteroplasmic conjugates formed by the fusion of starfish oocyte pairs with a 12 minute time difference in the maturation phase
Authors: Yoneda Mitsuki
Published: March 2003

Two starfish oocytes with a 12 min time difference in the maturation phase were fused together with electric pulses to make a heteroplasmic conjugate. The starfish used were Asterina pectinifera. The emergence of the first meiotic spindle and the extrusion of the polar bodies in the conjugate were timed. Under polarization microscopy two meiotic spindles emerged with a time difference of 10–11 min, which is close to the time difference in the maturation phase between the original oocytes before fusion. In contrast, subsequent formation of the first two polar bodies occurred successively with a short time lag of 1–3 min between them. Times for the formation of both polar bodies were midway between the anticipated times for polar body formation in respective non‐fused control oocytes. Thus, in one nucleus the meiotic division was delayed, while in another nucleus it was accelerated, in a single heteroplasmic conjugate. These two sets of observations indicate the presence of a certain control system that regulates progression of the cell cycle at a point during the period from the entry into metaphase through to late anaphase of meiosis I in starfish oocytes. This type of cell cycle control in starfish oocytes is obviously distinct from the currently accepted view of the cell cycle control by the spindle assembly checkpoint that monitors unattached kinetochores of mitotic chromosomes.
Relationship between ATP level and respiratory rate in sea urchin embryos
Authors: Fujiwara Akiko , Kamata Yasuyuki , Asami Koichi , Yasumasu Ikuo
Published: March 2003

In sea urchin embryos, the rate of respiration, as a result of electron transport through the mitochondrial respiratory chain, was enhanced after hatching without any change in the intrinsic capacity of electron transport in mitochondria. The increase in respiratory rate after hatching was accompanied by an evident decrease in intracellular adenosine triphosphate (ATP) concentration without any change in intracellular levels of adenosine diphosphate (ADP) and adenosine monophosphate (AMP). Adenosine triphosphate is proposed to fortify acceptor control of respiration at high concentrations and to reduce the respiratory rate even in the presence of ADP, the acceptor. The relationships between the respiratory rate and intracellular ATP concentration in embryos were the same as those in mitochondria isolated from embryos, obtained in the presence of ADP at the same concentration as in the embryos. Probably, the respiratory rate is enhanced after hatching because of the decrease in the level of ATP. In embryos kept in a medium containing adenosine, intracellular ATP concentration increased especially after hatching, without any change in the ADP level, and the respiratory rate after hatching was made as low as the rate expected, based on the relationships obtained on isolated mitochondria. The respiratory rate in embryos probably depends on intracellular ATP concentration, irrespective of the developmental stage in early development.
Beneficial effect of nicotinamide on the proportion of insulin cells in developing chick pancreas
Authors: Mngomezulu W. T. , Kramer Beverley
Published: March 2003

Previous studies have suggested that nicotinamide increases the number of insulin cells both in vivo and in vitro. However, the question remains as to whether there is in fact an increase and whether this increase is caused by the proliferation of progenitor cells, or by replication of existing insulin cells. In order to investigate this, the endodermal component of dorsal pancreatic buds of 5‐day‐old chick embryos was cultured on Matrigel in a serum‐free medium (Ham’s F12‐ITS) to which nicotinamide, at a concentration of 5 and 10 m
M, respectively, was added. Control explants were cultured in Ham’s F12‐ITS medium without nicotinamide. After 7 days in culture the buds were incubated with bromodeoxyuridine (BrdU) and then processed for immunocytochemistry. Localization of insulin, BrdU and glucagon was carried out on adjacent serial sections. The proportion of insulin cells was 6.76, 11.32 and 16.86% in control, 5 and 10 m
M nicotinamide‐treated explants, respectively. Hence adding nicotinamide to the culture medium induced a 1.7‐ and 2.5‐fold increase in the proportion of insulin cells when compared to the controls. These proportions were significantly different from that of control explants (P < 0.05). However, a very small number of insulin cells were found to be proliferating, suggesting that the increase in the proportion of insulin cells had resulted from stimulation of progenitor cells and not proliferation of existing insulin cells.
Effects of rat Axin domains on axis formation in Xenopus embryos
Authors: Fukui Akimasa , Kishida Shosei , Kikuchi Akira , Asashima Makoto
Published: March 2003

Wnt signaling plays an important role in axis formation in early vertebrate development. Axin is one Wnt signaling regulator that inhibits this pathway. The effects of the injection of mRNA of several rat Axin (rAxin) mutants on axis formation in Xenopus embryos were examined. It was found that rAxin mutants containing only a regulation of G‐protein signaling (RGS) domain fragment or with deletion of the RGS domain induced axis formation. Because the RGS domain is a major adenomatous polyposis coli gene product (APC)‐binding domain, APC association with glycogen synthase kinase 3β (GSK3β) on the Axin molecule may be important in inhibition of axis formation. The ventralizing activities of wild‐type rAxin and a mutant in which the Dishevelled and Axin (DIX) domain was deleted (ΔDIX mutant) were examined. Histological examination and gene expression revealed that the ventralizing activity of the ΔDIX mutant was weaker than that of wild‐type rAxin. This finding suggests that the C‐terminus of rAxin contributes to the inhibition of Wnt signaling in Xenopus embryos. Furthermore, an rAxin mutant that contained both the RGS and GSK3β‐binding domains affected both the dorsal and ventral sides of blastomeres, mediated ectodermal fate and induced expansion of notochord and/or endoderm, but did not induce axis formation.
A novel role of differentiation‐inducing factor‐1 in Dictyostelium development, assessed by the restoration of a developmental defect in a mutant lacking mitogen‐activated protein kinase ERK2
Authors: Kuwayama Hidekazu , Oyama Masakazu , Kubohara Yuzuru , Maeda Mineko
Published: March 2003

It has been previously reported that the differentiating wild‐type cells of Dictyostelium discoideum secrete a diffusible factor or factors that are able to rescue the developmental defect in the mutant lacking extracellular signal‐regulated kinase 2 (ERK2), encoded by the gene erkB. In the present study, it is demonstrated that differentiation‐inducing factor‐1 (DIF‐1) for stalk cells can mimic the role of the factor(s) and the mechanism of the action of DIF‐1 in the erkB null mutant is also discussed. The mutant usually never forms multicellular aggregates, because of its defect in cyclic adenosine monophosphate (cAMP) signaling. In the presence of 100 n
M DIF‐1, however, the mutant cells formed tiny slugs, which eventually developed into small fruiting bodies. In contrast, DIF‐1 never rescued the developmental arrest of other Dictyostelium mutants lacking adenylyl cyclase A (ACA), cAMP receptors cAR1 and cAR3, heterotrimeric G‐protein, the cytosolic regulator of ACA, or the catalytic subunit of cAMP‐dependent protein kinase (PKA‐C). Most importantly, it was found that DIF‐1 did not affect the cellular cAMP level, but rather elevated the transcriptional level of pka during the development of erkB null cells. These results suggest that DIF‐1 may rescue the developmental defect in erkB null cells via the increase in PKA activity, thus giving the first conclusive evidence that DIF‐1 plays a crucial role in the early events of Dictyostelium development as well as in prestalk and stalk cell induction.
Rapid patterning in 2‐D cultures of Dictyostelium cells and its relationship to zonal differentiation
Authors: Hirano Takashi , Sawai Satoshi , Sawada Yasuji , Maeda Yasuo
Published: March 2003

Rapid patterning has been observed in confined 2‐D cultures of Dictyostelium discoideum Ax‐2 cells as an outer dark zone and a inner light zone. The width of outer zone was usually ~100 μm, irrespective of the size of cell masses under atmospheric conditions. The width of the outer zone, however, changed depending on external O2 concentrations and reached up to 250 μm at 100% O2. A clear regional difference in tetramethyl rhodamine methyl ester (TMRM) staining was noticed between the outer zone and the inner zone: the inner zone was more strongly stained with TMRM than the outer zone, which faced the air. Using inhibitors of oxidative phosphorylation (dinitrophenol (DNP) or NaN3) and a specific inhibitor of CN‐resistant respiration (benzohydroxamic acid (BHAM)), it has been demonstrated that the outer zone is basically formed by the O2 threshold for oxidative phosphorylation, while the inner cells mainly perform cyanide‐resistant respiration. When cells around the early mound stage (just before prestalk and prespore differentiation) were cultured as 2‐D cell masses, ecmA‐expressing cells (pstA cells), ecmB‐expressing cells (pstB cells) and D19‐expressing cells (prespore; psp cells), arose in a position‐dependent manner in the outer zone. In the inner zone, cell motility seemed to be markedly impaired and neither prestalk nor prespore differentiation occurred. In addition, once‐differentiated prespore cells were found to dedifferentiate rapidly in the inner zone. The reason for dedifferentiation as well as for failure of cells to differentiate in the inner zone is discussed with reference to O2 radicals.
In vitro pancreas formation from Xenopus ectoderm treated with activin and retinoic acid
Authors: Moriya Naomi , Komazaki Shinji , Takahashi Shuji , Yokota Chika , Asashima Makoto
Published: March 2003

In the present study, isolated presumptive ectoderm from Xenopus blastula was treated with activin and retinoic acid to induce differentiation into pancreas. The presumptive ectoderm region of the blastula consists of undifferentiated cells and is fated to become epidermis and neural tissue in normal development. When the region is isolated and cultured in vitro, it develops into atypical epidermis. Isolated presumptive ectoderm was treated with activin and retinoic acid. The ectoderm frequently differentiated into pancreas‐like structures accompanied by an intestinal epithelium‐like structure. Sections of the explants viewed using light and electron microscopy showed some cells clustered and forming an acinus‐like structure, including secretory granules. The pancreas‐specific molecular markers insulin and XlHbox8 were also expressed in the treated explants. The pancreatic hormones, insulin and glucagon, were detected in the explants using immunohistochemistry. Therefore, sequential treatment with activin and retinoic acid can induce presumptive ectoderm to differentiate into a morphological and functional pancreas in vitro. When ectoderm was immediately treated with retinoic acid after treatment with activin, well‐differentiated pronephric tubules were seen in a few of the differentiated pancreases. Treatment with retinoic acid 3–5 h after activin treatment induced frequent pancreatic differentiation. When the time lag was longer than 15 h, the explants developed into axial mesoderm and pharynx. The present study provides an effective system for analyzing pancreas differentiation in vertebrate development.
Involvement of Rel/NF‐κB in regulation of ascidian notochord formation
Authors: Shimada Masumi , Satoh Noriyuki , Yokosawa Hideyoshi
Published: March 2003

The Rel/NF‐κB family is known to be involved in a wide variety of biological processes, including morphogenesis. In the present study, two protochordate cDNA clones encoding Rel/NF‐κB proteins, named As‐rel1 and As‐rel2, were isolated from a fertilized egg cDNA library of the ascidian Halocynthia roretzi. The As‐rel1 protein is a typical Rel/NF‐κB family member, containing a Rel homology domain, a nuclear localization sequence and a C‐terminal putative transcription activation domain, while the As‐rel2 protein is a novel Rel/NF‐κB family member that lacks a nuclear localization sequence and the C‐terminal domain. Northern blot analyses showed that both transcripts were maternally expressed and that their expression changed during development of H. roretzi embryos. Although injection of the As‐rel2 mRNA into H. roretzi fertilized eggs had little effect on embryonic development, injection of the As‐rel1 mRNA interfered greatly with notochord formation, resulting in a shortened tail with a reduced number of notochord cells. In contrast, embryos co‐injected with As‐rel1 and As‐rel2 mRNA developed normally, indicating that the As‐rel2 protein rescued the defect in notochord formation induced by the injection of As‐rel1 mRNA alone. These results strongly suggest that the As‐rel1 protein functions as a suppressor in ascidian notochord formation, while the As‐rel2 protein has an antagonistic effect on the action of the As‐rel1 protein.
XCL‐2 is a novel m‐type calpain and disrupts morphogenetic movements during embryogenesis in Xenopus laevis
Authors: Cao Ying , Zhao Hui , Grunz Horst
Published: March 2003

We identified a novel cDNA, XCL‐2, encoding an m‐type calpain, a calcium‐dependent intracellular protease. This protein has all characteristic structures and active sites of canonical calpains. Zygotic transcription of the gene was first detected at stage 10. It is expressed exclusively in the ventral circumblastoporal collar and the mesoderm‐free zone at the most anterior tip of neural fold in late gastrulae and neurulae. In later stages, expression is only found in cement gland and proctodeum. It is also expressed in a tissue‐specific manner. In adult tissues, various levels of expression were detected in brain, eye, heart, intestine, kidney, lung, stomach and testis, but not in liver, muscle, nerve, ovary, skin and spleen. Overexpression of wild‐type XCL‐2 suggests that this gene is involved in gastrulation movement and convergent extension during gastrulation and neurulation. Overexpression of a dominant‐negative mutant caused a phenotype morphologically similar to, but histologically different from, that caused by overexpression of wild‐type XCL‐2. The mutant phenotype can be rescued by injection of wild‐type XCL‐2. These data suggest that XCL‐2 plays an important role in convergent extension movements during embryogenesis in Xenopus laevis.