bluuebunny/crossref_metadata_2025 huggingface dataset:
Treating childhood acute myeloid leukaemia with the AML‐BFM‐83 protocol: experience in a developing country
Authors: Chan L. L. , Abdel‐Latif M. E. , Ariffin W. A. , Ariffin H. , Lin H. P.
Published: October 2004

Treatment for childhood acute myeloid leukaemia (AML) consists of remission induction chemotherapy followed by postremission chemotherapy with or without bone marrow transplantation. The AML Berlin–Frankfurt–Munster (BFM)‐83 protocol with induction–consolidation–maintenance chemotherapy for 2 years has been reported to result in a 6‐year event‐free survival (EFS) and event‐free interval (EFI) of 49% and 61% respectively. A total of 174 Malaysian children were treated with this protocol between 1985 and 1999. The 5‐year EFS and EFI was 30.7% and 48.0% respectively. The overall mortality from sepsis was 24%, which needs urgent address. The 5‐year EFS for patients treated before 1993 and after 1993 was 18.6% and 41.3%, respectively (P = 0.04), while the EFI was 32% and 60.6% respectively (P = 0.034). The improvement seen after 1993 was related to a reduction in induction deaths for that period and probably reflected increased capability and familiarity to cope with the demands of the AML‐BFM‐83 protocol and accompanying complications in the treatment of AML.
Drink‐Spiking and sexual assault: Innovative approaches to a difficult problem
Authors: Costello Mayet , Blyth Caroline
Published: September 2004

The Sutherland Drink‐Spiking Project is a collaborative interagency project initiated by the Southern Sydney Sexual Assault Service. Focusing on selected intervention as well as primary and secondary prevention with drink‐spiking and sexual assault in southern Sydney, this project is distinctive due to its primary focus on perpetrators potential perpetrators, and their peers. Beyond this, the project also provided an opportunity for social workers to meaningfully engage with and apply innovative project management practice. This is notable since reform initiatives in public sector agencies in New South Wales are encouraging the use of project management practice and social workers need to engage with these ‘new directions’ or risk becoming marginalised. This paper explores The Sutherland Drink‐Spiking Project and demonstrates how applying a project management methodology benefited social work practice.
International social work practicum in India
Authors: Pawar Manohar , Hanna Georgina , Sheridan Rachael
Published: September 2004

The present study discusses the process of organising social work placements in India for two Australian students and presents the students' and the field education coordinator's reflections on practicum experiences in the school and hospital setting. Several steps need to be taken to plan and implement international placements. Undertaking a practicum in another unknown culture is as challenging as it is rewarding. Cross‐cultural comparative dimensions naturally emerge in our thinking, feeling and doing, and help us appreciate what we have in our own country. Learning occurs both within and outside the practicum. Positive mental attitude, psychological make up, an open mind, planning and preparation do help. However, overseas placements have their own quota of surprises. Any student aspiring to undertake an international practicum should be prepared to face some unexpected situations as they unfold. Overall, effective learning did occur and horizons did expand.
Interaction equations for multiaxial fatigue assessment of welded structures
Authors: BÄCKSTRÖM M. , MARQUIS G.
Published: September 2004

Multiaxial fatigue data from 233 welded test specimens taken from eight different studies have been evaluated based on three published interaction equations for normal and shear stress. The interaction equations were obtained from SFS 2378, Eurocode 3 and International Institute of Welding (IIW) recommendations. Fatigue classes for normal and shear stress were obtained directly from the design guidance documents. Additionally, mean fatigue strengths were determined by regression analysis of bending only and torsion only data for different specimen types. In some cases, the S–N slopes assumed by the different standards were not appropriate for the test data. Specimens that showed significantly different cracking locations or cracking mode between bending and torsion were not easily correlated by the interaction equations. Interaction equations work best in cases where both the normal stress and the shear stress tend to produce crack initiation and growth in the same location and in the same direction. The use of a damage summation of 0.5 for non‐proportional loading as recommended by IIW was consistent with experimental observations for tube‐to‐plate specimens. Other codes used a damage sum of unity.
Psychiatric medication‐induced obesity: treatment options
Authors: Schwartz T. L. , Nihalani N. , Virk S. , Jindal S. , Chilton M.
Published: September 2004

A majority of psychiatric medications are known to generate weight gain and ultimately obesity in some patients. The authors undertook a comprehensive literature review in order to provide a better understanding of novel treatment options in regards to alleviating weight gained by use of antidepressants, antipsychotics, and mood stabilizers. There are no agents for management of this weight gain approved by the Food and Drug Administration (FDA), and existing studies on options are mainly uncontrolled, small‐scale projects with limited power to produce coherent conclusions. There is a clear need for larger studies on existing options, and future psychotropics without these side‐effects are currently in the pipeline.
Direct effects of sex steroid hormones on adipose tissues and obesity
Authors: Mayes J. S. , Watson G. H.
Published: September 2004

Sex steroid hormones are involved in the metabolism, accumulation and distribution of adipose tissues. It is now known that oestrogen receptor, progesterone receptor and androgen receptor exist in adipose tissues, so their actions could be direct. Sex steroid hormones carry out their function in adipose tissues by both genomic and nongenomic mechanisms. In the genomic mechanism, the sex steroid hormone binds to its receptor and the steroid–receptor complex regulates the transcription of given genes. Leptin and lipoprotein lipase are two key proteins in adipose tissues that are regulated by transcriptional control with sex steroid hormones. In the nongenomic mechanism, the sex steroid hormone binds to its receptor in the plasma membrane, and second messengers are formed. This involves both the cAMP cascade and the phosphoinositide cascade. Activation of the cAMP cascade by sex steroid hormones would activate hormone‐sensitive lipase leading to lipolysis in adipose tissues. In the phosphoinositide cascade, diacylglycerol and inositol 1,4,5‐trisphosphate are formed as second messengers ultimately causing the activation of protein kinase C. Their activation appears to be involved in the control of preadipocyte proliferation and differentiation. In the presence of sex steroid hormones, a normal distribution of body fat exists, but with a decrease in sex steroid hormones, as occurs with ageing or gonadectomy, there is a tendency to increase central obesity, a major risk for cardiovascular disease, type 2 diabetes and certain cancers. Because sex steroid hormones regulate the amount and distribution of adipose tissues, they or adipose tissue‐specific selective receptor modulators might be used to ameliorate obesity. In fact, hormone replacement therapy in postmenopausal women and testosterone replacement therapy in older men appear to reduce the degree of central obesity. However, these therapies have numerous side effects limiting their use, and selective receptor modulators of sex steroid hormones are needed that are more specific for adipose tissues with fewer side effects.
Substrate metabolism, nutrient balance and obesity development in children and adolescents: a target for intervention?
Authors: Sun M. , Schutz Y. , Maffeis C.
Published: September 2004

Obesity results from the organism's inability to maintain energy balance over a long term. Childhood obesity and its related factors and pathological consequences tend to persist into adulthood. A cluster of factors, including high energy density in the diet (high fat intake), low energy expenditure, and disturbed substrate oxidation, favour the increase in fat mass. Oxidation of three major macronutrients and their roles in the regulation of energy balance, particularly in children and adolescents, are discussed. Total glucose oxidation is not different between obese and lean children; exogenous glucose utilization is higher whereas endogenous glucose utilization is lower in obese compared with lean children. Carbohydrate composition of the diet determines carbohydrate oxidation regardless of fat content of the diet. Both exogenous and endogenous fat oxidation are higher in obese than in lean subjects. The influence of high fat intake on accumulation of fat mass is operative rather over a long term. Several future directions are addressed, such that a combination of increased physical activity and modification in diet composition, in terms of energy density and glycemic index, is recommended for children and adolescents.
Leerstellen in Der Deutschen Gedenkkultur: Die Streitschriften Von W.G. Sebald und Klaus Briegleb
Authors: Hahn Hans‐Joachim
Published: September 2004

Der nachstehende Aufsatz analysiert zwei breit diskutierte Veröffentlichungen von W.G. Sebald und Klaus Briegleb zum Gegenstand deutscher Erinnerungspolitik hinsichtlich des Nationalsozialismus und der Zerstörung deutscher Städte während der alliierten Bombenangriffe. Obwohl Sebald die Weise untersucht, in der die alliierten Luftangriffe in der deutschen Literatur dargestellt und nicht dargestellt wurden, während Briegleb an einigen Texten von Mitgliedern der Gruppe 47 deren Beitrag zum Verhältnis von Juden und Nichtjuden in Deutschland nach 1945 reflektiert, beschäftigen sich beide Autoren mit offensichtlichen Lücken und Leerstellen der deutschen Nachkriegskultur. Insbesondere wird dabei gezeigt, auf welche Weise beide Veröffentlichungen zur unbeendeten Debatte um das komplexe Verhältnis von deutscher Täterschaft und Opfererfahrung beitragen. Zum anderen wird eine Antwort auf die Frage versucht, warum den Texten innerhalb des deutschen Feuilletons eine beinahe antagonistische Rezeption zuteil wurde.
Border Looking: The Cold War Visuality of the Sudeten German Expellees and its Afterlife
Authors: Komska Yuliya
Published: September 2004

After their expulsion from Czechoslovakia in 1945–6, the Sudeten Germans continued to revisit the (West) German‐Czech(oslovak) border, a dividing line now about to undergo institutional dissolution within the expanded European Union. Until 1987, their different groups contributed to building what became a chain of pilgrimage sites located along the Iron Curtain and visited well into the present. Closely related to the so‐called ‘Blick in die Heimat’, the environment of these borderland shrines has produced a complex visual regime that I have dubbed ‘border looking’. Within this system, as I demonstrate, the Sudeten German visuality oscillates not only between a re‐appropriating revisionist gaze and a cursory furtive, fragmented glimpse acknowledging the irrevocability of loss, but also between the ‘gaze into the homeland’ and the ‘image of the homeland’. My examination of this condition is intended to be both a contribution to a cultural history of the Cold War and a study of the expellee subjectivity. It allows us to raise questions about the effectiveness of memory inscribed on a geographical margin at a juncture when initiatives advocating a centre to commemorate the expulsion of Germans (‘Zentrum gegen Vertreibungen’) are being countered by pleas for de‐centred, regional remembrance.

Hier kannst du die Heimat,

die Heimat seh’n!1
Flüchtlings‐ und Vertriebenenschicksale im Werk Arno Schmidts
Authors: Menke Timm
Published: September 2004

Als einer der wenigen Zeitzeugen hat Arno Schmidt in seinen Werken die Thematik der Leiden der deutschen Zivilbevölkerung am Ende des Zweiten Weltkriegs aufgegriffen. Von seinem Erstlingswerk in der unmittelbaren Nachkriegszeit, der Erzählung Leviathan (publiziert 1949), bis hin zum Roman Kaff auch Mare Crisium von 1960 hat er sich immer wieder mit dem Schicksal von Vertriebenen und Flüchtlingen auseinandergesetzt. Seine Arbeiten erfahren angesichts der gegenwärtigen Aufarbeitungswelle der Thematik durch die Enkel jener Nachkriegsgeneration eine neue Aktualität und Anerkennung, die ihnen bei ihrer Erstveröffentlichung weitgehend versagt blieben. Schmidts literaturhistorisches Verdienst besteht darin, das Leiden der Vertriebenen nie isoliert zu schildern, sondern es stets in den geschichtlichen Kontext des vom deutschen Boden ausgegangenen Weltkriegs und Völkermords zu stellen.
Erinnerung, Fiktion und Geschichte. Über Die Transformation Des Erlebten Ins Kulturelle Gedächtnis: Walser – Wilkomirski – Grass
Authors: Preusser Heinz‐Peter
Published: September 2004

In alle Erinnerungen mischen sich konstruktive Anteile; das gilt für die schriftlich abgefassten Autobiographien in besonderem Maße. Zuweilen wird hier deutlich die Grenze zur fiktionalen Narration überschritten. Aber gerade der Fall Wilkomirski hat gezeigt, wo die ontologische Differenz gewahrt werden muss zwischen Erinnerungen als Wahrheitsaussagen und reinen Produkten künstlerischer Phantasie. Umgekehrt hat Walser diesen Anspruch gar nicht erhoben und ist dennoch für seinen Roman Ein springender Brunnen dafür haftbar gemacht worden, kein adäquates Bild der Vergangenheit geliefert und Auschwitz verschwiegen zu haben. Schließlich hat Grass mit seiner Novelle Im Krebsgang Geschichte mit den Mitteln der Fiktion ins Bewusstsein rücken wollen; das Vergessene sollte wieder Gestalt erhalten und Allgemeingut werden. Alle drei Autoren bieten Extrempunkte im Prozess der Transformation des Erlebten ins kulturelle Gedächtnis, in denen insbesondere das Verhältnis von Individuellem und Allgemeinem unterschiedlich gewichtet wird. Und sie unternehmen ihre Versuche zu einem historisch neuralgischen Zeitpunkt, in dem die Gesellschaft abzuklären sucht, wie der Holocaust, die NS‐Zeit und die Geschichte des Zweiten Weltkrieges – im Sinne eines Archivierens – erinnert werden sollen.
Delirium in acute stroke: a preliminary study of the role of anticholinergic medications
Authors: Caeiro L. , Ferro J. M. , Claro M. I. , Coelho J. , Albuquerque R. , Figueira M. L.
Published: October 2004

The pathogenesis of delirium in acute stroke is incompletely understood. The use of medications with anticholinergic (ACH) activity is associated with an increased frequency of delirium. We hypothesized that the intake of medications with ACH activity is associated with delirium in acute stroke patients. Delirium was assessed using the DSM‐IV‐TR criteria and the Delirium Rating Scale, in a sample of consecutive patients with an acute (≤4 days) cerebral infarct or intracerebral haemorrhage (ICH). We performed a gender and age matched case–control study. Twenty‐two delirious stroke patients (cases) and 52 non‐delirious patients (controls) were compared concerning the intake of ACH medications (i) before stroke, (ii) during hospitalization but before the assessment. The variables associated with delirium on bivariate analysis were entered in a stepwise logistic regression analysis. The final regression model (Nagelkerke R2 = 0.65) retained non‐neuroleptics ACH medication during hospitalization (OR = 24.4; 95% CI = 2.18–250), medical complications (OR = 20.8; 95% CI = 3.46–125), ACH medication taken before stroke (OR = 17.5; 95% CI = 1.00–333.3) and ICH (OR = 16.9; 95% CI = 2.73–100) as independent predictors of delirium. This preliminary result indicates that drugs with subtle ACH activity play a role in the pathogeneses of delirium in acute stroke. Medication with ACH activity should be avoided in acute stroke patients.
The Interaction of nucleotides with the tolbutamide block of cloned atp‐sensitive k+ channel currents expressed in xenopus oocytes: a reinterpretation
Authors: Gribble Fiona M. , Tucker Stephen J. , Ashcroft Frances M.
Published: September 2004

We have examined the mechanism by which nucleotides modulate the tolbutamide block of the β‐cell ATP‐sensitive K+ channel (KATP channel), using wild‐type and mutant KATP channels heterologously expressed in Xenopus oocytes. This channel is composed of sulphonylurea receptor (SUR1) and pore‐forming (Kir6.2) subunits.

The dose–response relation for tolbutamide block of wild‐type KATP currents in the absence of nucleotide showed both a high‐affinity (Ki=2.0 μm) and a low‐affinity (Ki=1.8 mm) site.

The dose–response relation for tolbutamide block of Kir6.2ΔC36 (a truncated form of Kir6.2 which is expressed independently of SUR1) was best fitted with a single, low‐affinity site (Ki=1.7 mm). This indicates that the high‐affinity site resides on SUR1, whereas the low‐affinity site is located on Kir6.2.

ADP (100 μM) had a dual effect on wild‐type KATP currents: the nucleotide enhanced the current in the presence of Mg2+, but was inhibitory in the absence of Mg2+. Kir6.2ΔC36 currents were blocked by 100 μM ADP in the presence of Mg2+.

For wild‐type KATP currents, the blocking effect of 0.5 mm tolbutamide appeared greater in the presence of 100 μm MgADP (84 ± 2%) than in its absence (59 ± 4%). When SUR1 was mutated to abolish MgADP activation of KATP currents (K719A or K1384M), there was no difference in the extent of tolbutamide inhibition in the presence or absence of MgADP.

The ki for tolbutamide interaction with either the high‐ or low‐affinity site was unaffected by 100 μM MgADP, for both wild‐type and K719A‐K1384M currents.

MgGDP (100μM) enhanced wild‐type KATP currents and was without effect on K719A– K1384M currents. It did not affect the ki for tolbutamide block at either the high‐ or low‐affinity site.

Our results indicate that interaction of tolbutamide with the high‐affinity site (on SUR1) abolishes the stimulatory action of MgADP. This unmasks the inhibitory effect of ADP and leads to an apparent increase in channel inhibition. Under physiological conditions, abolition of MgADP activation is likely to constitute the principal mechanism by which tolbutamide inhibits the KATP channel.
Effect of hydroxylamine on photon‐like events during dark adaptation in toad rod photoreceptors
Authors: Leibrock C.S. , Lamb T.D.
Published: September 2004

The suction pipette technique was used to investigate the recovery of toad rod photoreceptors following small bleaches of 0.2‐3% of the rhodopsin.

The reduction in sensitivity and the increase in noise elicited by bleaches were measured, and from these measurements the underlying rate of occurrence of photon‐like events was calculated as a function of time after the bleach.

Exposure to hydroxylamine solution was used to hasten the decomposition of the meta‐rhodopsin photoproducts. The outer segment was exposed to 110 mmhydroxylamine in a low‐Ca2+Ringer solution for a period of 10–50 s beginning 10–17 min after the bleaching exposure.

By the time of the hydroxylamine exposure, the flash sensitivity and response kinetics had returned almost to normal, and were not significantly altered by the exposure.

Following hydroxylamine exposure, the rate of spontaneous photon‐like events in the rods declined rapidly to near pre‐bleach levels.

We conclude that hydroxylamine reduces the rate of occurrence of photon‐like events induced by a bleach, and we postulate that this reduction results from the removal of metarhodopsin (most likely metarhodopsin II) from the outer segment.

Our results are consistent with a model in which photon‐like events result from reversal of the reactions (phosphorylation and capping by arrestin) that lead to inactivation of the activated form of rhodopsin, Rh*.
Ca2+ and Na+ permeability of high‐threshold Ca2+ channels and their volt age‐dependent block by Mg2+ ions in chick sensory neurones
Authors: Carbone E. , Lux H. D. , Carabelli V. , Aicardi G. , Zucker H.
Published: September 2004

The Mg2+ block of Na+ and Ca2+ currents through high‐voltage activated (HVA; L‐ and N‐type) Ca2+ channels was studied in chick dorsal root ganglion neurones.

In low extracellular [Ca2+] (< 10−8 M) and with Nao+ and Csi+ as the main charge carriers (120 mm), HVA Na+ currents started to activate at −40 mV, reached inward peak values near 0 mV and reversed at about +40 mV.

Addition of 30–500 μm Mg2+ to the bath caused a strong depression of inward Na+ currents that was voltage and dose dependent (KD=39 μm in 120 mm Na+ at −10 mV). The block was maximal at negative potentials (< −70 mV) and decreased with increasing positive potentials, suggesting that Mg2+ cannot escape to the cell interior.

Block of Ca2+ currents by Mg2+ was also voltage dependent, but by three orders of magnitude less potent than with Na+ currents (KD= 24 mm in 2 mm Ca2+ at −30 mV). The high concentration of Mg2+ caused a prominent voltage shift of channel gating kinetics induced by surface charge screening effects. To compensate for this, Mg2+ block of inward Ca2+ currents was estimated from the instantaneous I–V relationships on return from very positive potentials (+100 mV).

Inward Na+ and Ca2+ tail currents following depolarization to +90 mV were markedly depressed, suggesting that channels cleared of Mg2+ ions during strong depolarization are quickly re‐blocked on return to negative potentials. The kinetics of re‐block by Mg2+ was too fast (< 100 μs) to be resolved by our recording apparatus. This implies a rate of entry for Mg2+ > 1.45 × 108 M− s−1 when Na+ is the permeating ion and a rate approximately 3 orders of magnitude smaller for Ca2+.

Mg2+ unblock of HVA Na+ currents at +100 mV was independent of the size of outward currents, whether Na+, Cs+ or NMG+ were the main internal cations.

Consistent with the idea of a high‐affinity binding site for Ca2+ inside the channel, micromolar amounts of Ca2+ caused a strong depression of Na+ currents between −40 and 0 mV, which was effectively relieved with more positive as well as with negative potentials (KD= 0.7μm in 120 mm Na+ at −20 mV). In this case, the kinetics of re‐block could be resolved and gave rates of entry and exit for Ca2+ of 1.4 × 108 M−1 s−1 and 2.95 × 102 s−1, respectively.

The strong voltage dependence and weak current dependence of HVA channel block by divalent cations and the markedly different KD values of Na+ and Ca2+ current block by Mg2+ can be well described by a previously proposed model for Ca2+ channel permeation based on interactions between the permeating ion and the negative charges forming the high‐affinity binding site for Ca2+ inside the pore (Lux, Carbone & Zucker, 1990).
Measurement of sarcoplasmic reticulum Ca2+ content and sarcolemmal Ca2+ fluxes in isolated rat ventricular myocytes during spontaneous Ca2+ release
Authors: Díaz M.E. , Trafford A.W. , O'Neill S.C. , Eisner D.A.
Published: September 2004

Intracellular calcium concentration ([Ca2+]i) and Na+–Ca2+ exchange currents were measured in calcium‐overloaded voltage‐clamped rat ventricular myocytes loaded with the Ca2+sensitive fluorescent indicator indo‐1. Sarcoplasmic reticulum (SR) Ca2+ content was measured from the integral of the caffeineevoked current. In cells that had spontaneous SR Ca2+ release in 1 mm external Ca2+ concentration ([Ca2+]o), raising [Ca2+]o increased the frequency of release with no effect on SR Ca2+ content. In quiescent cells, increased [Ca2+]o produced spontaneous Ca2+ release associated with increased SR Ca2+ content. Further increase of [Ca2+]o had no effect on SR Ca2+ content. The amount of Ca2+ leaving the cell during each release was constant over a wide range of frequencies and [Ca2+]o values. It appears there is a maximum level of SR Ca2+ content, perhaps because spontaneous Ca2+ release results when the content reaches a threshold.

From the relationship between [Ca2+]i and Na+–Ca2+ exchange current during a caffeine response, it is possible to estimate the changes in Na+–Ca2+ exchange current expected from a change of [Ca2+]i. The data show that the calcium oscillations contribute a significant fraction of the total extra Ca2+ efflux induced by increasing [Ca2+]o. Raising [Ca2+]o decreased the rate of calcium removal from the cell as measured from the rate of decay of the caffeine response, suggesting that both inhibition of Ca2+ efflux and increased Ca2+ entry account for the Ca2+ overload at elevated [Ca2+]o.

Inhibiting spontaneous SR Ca2+ release increases resting [Ca2+]i. The Ca2+ efflux is identical to that in the presence of release. It is concluded that spontaneous release of calcium, although potentially arrhythmogenic, is an effective way to activate Ca2+ efflux in overloaded conditions and minimizes any increase of diastolic tension.
A C‐terminal peptide of the GIRK1 subunit directly blocks the G protein‐activated K+ channel (GIRK) expressed in Xenopus oocytes
Authors: Luchian Tudor , Dascal Nathan , Dessauer Carmen , Platzer Dieter , Davidson Norman , Lester Henry A. , Schreibmayer Wolfgang
Published: September 2004

In order to find out the functional roles of cytosolic regions of a G protein‐activated, inwardly rectifying potassium channel subunit we studied block of GIRK channels, expressed in Xenopus laevis oocytes, by synthetic peptides in isolated inside‐out membrane patches.

A peptide (DS6) derived from the very end of the C‐terminus of GIRK1 reversibly blocked GIRK activity with IC50 values of 7.9 ± 2.0 or 3.5 ± 0.5 μg ml−1 (corresponding to 3.7 ± 0.9 or 1.7 ± 0.2 μmol l−1) for GIRK1/GIRK5 or GIRK1/GIRK4 channels, respectively.

Dose dependency studies of GIRK activation by purified βγ subunits of the G protein (Gβγ) showed that DS6 block of GIRK channels is not the result of competition of the peptide with functional GIRK channels for the available Gβγ.

Burst duration of GIRK channels was reduced, whereas long closed times between bursts were markedly increased, accounting for the channel block observed.

Block by the DS6 peptide was slightly voltage dependent, being stronger at more negative potentials.

These data support the hypothesis that the distal part of the carboxy‐terminus of GIRK1 is a part of the intrinsic gate that keeps GIRK channels closed in the absence of Gβγ.
Tetrodotoxin‐Sensitive Sodium Current in Sheep Lymphatic Smooth Muscle
Authors: Hollywood M. A. , Cotton K. D. , Thornbury K. D. , McHale N. G.
Published: September 2004

Fast inward currents were elicited in freshly isolated sheep lymphatic smooth muscle cells by depolarization from a holding potential of −80 mV using the whole‐cell patch‐clamp technique. The currents activated at voltages positive to −40 mV and peaked at 0 mV.

When sodium chloride in the bathing solution was replaced isosmotically with choline chloride inward currents were abolished at all potentials.

These currents were very sensitive to tetrodotoxin (TTX). Peak current was almost abolished at 1 μM with half‐maximal inhibition at 17 nM.

Examination of the voltage dependence of steady state inactivation showed that more than 90 % of the current was available at the normal resting potential of these cells (−60 mV).

The time course of recovery from inactivation was studied using a double‐pulse protocol and showed that recovery was complete within 100 ms with a time constant of recovery of 20 ms.

Under current clamp, action potentials were elicited by depolarizing current pulses. These had a rapid upstroke and a short duration and could be blocked with 1 μM TTX.

Spontaneous contractions of isolated rings of sheep mesenteric lymphatic vessels were abolished or significantly depressed by 1 μM TTX.
The effect of Ca2+–calmodulin‐dependent protein kinase II on cardiac excitation–contraction coupling in ferret ventricular myocytes
Authors: Li Li , Satoh Hiroshi , Ginsburg Kenneth S. , Bers Donald M.
Published: September 2004

The effect of Ca2+–calmodulin‐dependent protein kinase II (CaMKII) on excitation–contraction coupling (E–C coupling) was studied in intact ferret cardiac myocytes using the selective inhibitor KN‐93. KN‐93 decreased steady‐state (SS) twitch [Ca2+]i (by 51%), resting Ca2+ spark frequency (by 88%) and SS sarcoplasmic reticulum (SR) Ca2+ content evaluated by caffeine application (by 37.5%).

Increasing extracellular Ca2+ concentration ([Ca2+]o) to 5 mm in KN‐93 restored SR Ca2+ load and Ca2+ spark frequency towards that in control (2 mm Cao2+), but SS twitch [Ca2+]i was still significantly depressed by KN‐93.

KN‐93 decreased Ca2+ transient amplitude of SS twitches much more strongly than the amplitude of post‐rest (PR) twitches. In the control, the time constant (τ) of [Ca2+]i decline of SS twitches was faster than that for PR twitches. This stimulation‐dependent acceleration of [Ca2+]i decline was abolished by KN‐93.

Voltageclamp experiments demonstrated that KN‐93 significantly inhibited sarcolemmal L‐type Ca2+ current (ICa) during repetitive pulses by slowing recovery from inactivation. This may explain the preferential action of KN‐93 to suppress SS vs. PR twitches.

In KN‐93, even when both ICa and SR Ca2+ load were matched to the control levels by manipulation of conditioning voltage‐clamp pulses, contraction and twitch Ca2+ transients were still both significantly depressed (to 39 and 49% of control, respectively).

Since KN‐93 reduced SR Ca2+ release channel (RyR) activity during E–C coupling, even for matched SR Ca2+ load and trigger ICa, we infer that endogenous CaMKII is an important modulator of E–C coupling in intact cardiac myocytes. Effects of KN‐93 on ICa and SS twitch [Ca2+]i decline also indicate that endogenous CaMKII may have stimulatory effects on ICa and SR Ca2+ uptake.
Synergistic activation of guinea‐pig cardiac cystic fibrosis transmembrane conductance regulator by the tyrosine kinase inhibitor genistein and cAMP
Authors: Shuba Lesya M. , McDonald Terence F.
Published: September 2004

The regulation of cardiac Cl− current (IC1) by tyrosine and serine/threonine phosphorylation was examined in guinea‐pig and rat ventricular myocytes. The protein tyrosine kinase (PTK) inhibitor genistein (GST) and phosphotyrosine phosphatase (PTP) inhibitor sodium orthovanadate (VO4) were used to modify tyrosine phosphorylation, whereas forskolin (FSK), cAMP, and other agents were used to modify cytoplasmic cAMP concentration and protein kinase A (PKA) phosphorylation.

Low concentrations (0.1 μm) of FSK did not activate the PKA‐regulated cystic fibrosis transmembrane regulator (CFTR) ICl in guinea‐pig ventricular myocytes, but strongly potentiated activation of an ICl by 20–100 μm GST. The potentiation did not occur when GST was replaced by PTK‐inactive daidzein, and it was strongly inhibited by 1 mm VO4.

Potentiation by 0.1 μm FSK was linked to a small stimulation of the adenylate cyclase–cAMP–PKA pathway. The potentiation was not mimicked by inactive 1,9‐dideoxyforskolin, and was inhibited by muscarinic stimulation (ACh) and by a PKA inhibitor. Internal application of a cAMP solution that alone was too weak to activate CFTR ICl strongly potentiated the activation of ICl by 50 μm GST and occluded potentiation by 0.1 μm FSK.

The foregoing suggests that potentiated ICl flows through cAMP‐dependent CFTR channels. In agreement with this interpretation, GST did not increase ICl when CFTR was maximally activated by a high concentration (5 μm) of FSK and okadaic acid, and neither GST nor GST plus FSK activated an ICl in CFTR‐deficient rat myocytes. The lack of effect in rat myocytes was not due to the absence of functional, channel‐relevant PKA and PTK–PTP systems, because (as in guinea‐pig myocytes) L‐type Ca2+ current (ICa,L) was stimulated by FSK and inhibited in a VO4‐reversible manner by GST.

The synergistic activation of CFTR by low concentrations of FSK and GST cannot be explained by either a GST‐induced elevation of cAMP concentration or inhibition of serine/threonine phosphatase. Rather, it appears to be due to tyrosine dephosphorylation that facilitates PKA‐mediated phosphorylation of the channels.
Swelling‐induced Cl− current in guinea‐pig atrial myocytes: inhibition by glibenclamide
Authors: Sakaguchi Masayuki , Matsuura Hiroshi , Ehara Tsuguhisa
Published: September 2004

Whole‐cell currents were recorded from guinea‐pig atrial myocytes using the patch‐clamp technique under conditions designed to block K+ channels, Ca2+ channels and electrogenic transporters.

Exposure of atrial myocytes to the hyposmotic external solution (Na+ reduction to about 70% of control) resulted in hyposmotic cell swelling which was associated with activation of an outwardly rectifying Cl− current (ICl,swell).

Whereas the activation of ICl,swell was not significantly affected by replacement of ATP in the pipette solution with the non‐hydrolysable ATP analogue 5′‐adenylyl‐imidodiphosphate (AMP‐PNP), its activation was greatly reduced in cells dialysed with an ATP‐free pipette solution, thus indicating that the activation process of ICl,swell requires the presence of intra‐cellular ATP, but not its hydrolysis.

Bath application of glibenclamide produced a concentration‐dependent block of ICl,swell with a half‐maximal inhibitory concentration (IC50) of 60.0 μm and a Hill coefficient of 2.1. The maximal effect (100% inhibition) was obtained with 500 μm glibenclamide. The steady‐state inhibition showed little voltage dependence, while glibenclamide at concentrations of more than 100 μm inhibited the outward ICl,swell more rapidly than the inward ICl,swell. The glibenclamide inhibition was fully reversible after removal of the drug, even when a maximal effect (full inhibition) was achieved at a high drug concentration (500 μm).

These results show that (i) glibenclamide is one of the most potent inhibitors of guinea‐pig atrial ICl,swell and (ii) atrial ICl,swell and the cystic fibrosis transmembrane conductance regulator (CFTR) Cl− currents are almost equally sensitive to inhibition by glibenclamide.
A quantitative analysis of the activation and inactivation kinetics of HERG expressed in Xenopus oocytes
Authors: Wang Shimin , Liu Shuguang , Morales Michael J. , Strauss Harold C. , Rasmusson Randall L.
Published: September 2004

The human etherà‐go‐go‐related gene (HERG) encodes a K+o channel that is believed to be the basis of the delayed rectified current, IKr, in cardiac muscle. We studied HERG expressed in Xenopus oocytes using a two‐electrode and cut‐open oocyte clamp technique with [K+]o of 2 and 98 mm.

The time course of activation of the channel was measured using an envelope of tails protocol and demonstrated that activation of the heterologously expressed HERG current (IHERG) was sigmoidal in onset. At least three closed states were required to reproduce the sigmoid time course.

The voltage dependence of the activation process and its saturation at positive voltages suggested the existence of at least one relatively voltage‐insensitive step. A three closed state activation model with a single voltage‐insensitive intermediate closed state was able to reproduce the time and voltage dependence of activation, deactivation and steady‐state activation. Activation was insensitive to changes in [K+]o.

Both inactivation and recovery time constants increased with a change of [K+]o from 2 to 98 mm. Steady‐state inactivation shifted by ∼30 mV in the depolarized direction with a change from 2 to 98 mm K*o

Simulations showed that modulation of inactivation is a minimal component of the increase of this current by [K+]o, and that a large increase in total conductance must also occur.
Differential Effects of Endothelin‐1 on Basal and Isoprenaline‐Enhanced Ca2+ Current in Guinea‐Pig Ventricular Myocytes
Authors: Thomas George P. , Sims Stephen M. , Karmazyn Morris
Published: September 2004

We examined the effect of endothelin‐1 (ET‐1) on basal and isoprenaline‐enhanced L‐type Ca2+ current (ICa,L) in guinea‐Pig ventricular myocytes under nystatin‐perforated patch configuration.

ET‐1 at concentrations of 1, 5 and 10 nM had little effect on basal ICa,L. However, ICa,L enhanced by isoprenaline (500 nM) was significantly attenuated by 5 nM ET‐1 by more than 50%. This effect was reversed upon washout. ICa,L enhanced by forskolin was also decreased by ET‐1.

The inhibitory effect of ET‐1 against isoprenaline was completely blocked by the ETA receptor antagonist BQ‐123 (1 μm). In myocytes incubated with pertussis toxin (PTX, 2 μg ml−1) for 5 h, ET‐1 did not inhibit isoprenaline‐enhanced ICa,L.

Although ET‐1 has been shown to activate specific protein kinase C (PKC) isoforms, a significant inhibitory effect of ET‐1 was maintained in the presence of the PKC inhibitor bisindolylmaleimide (20 nm). The nitric oxide (NO) donor SIN‐1 (10 μm) attenuated but failed to prevent the ET‐1 effect.

In summary, our results demonstrate that ET‐1 is devoid of any significant effects on basal ICa,L. However, it exerts a potent inhibitory effect against isoprenaline‐enhanced ICa,L. This effect is mediated through ETA receptors coupled to PTX‐sensitive G‐proteins and occurs in the presence of PKC inhibition and NO generation.
Activation of a novel non‐selective cation channel by alloxan and H2O2 in the rat insulin‐secreting cell line CRI‐G1
Authors: Herson P. S. , Ashford M. L. J.
Published: September 2004

Alloxan and its auto‐oxidation product hydrogen peroxide (H2O2) irreversibly depolarize insulinoma cells by opening a non‐selective cation channel. The channel opened is characterized by a linear current–voltage relation with a conductance of approximately 70 pS and very slow kinetics (of the order of seconds).

Cells are protected against the alloxan‐induced channel opening and consequent cell depolarization by the presence of H2O2 and hydroxyl radical scavengers.

The free radical‐activated non‐selective cation channel is not operative in isolated patches but can be activated by the application of β‐NAD to the cytoplasmic aspect of the membrane.
Outward potassium currents of supraoptic magnocellular neurosecretory cells isolated from the adult guinea‐pig
Authors: Hlubek Michael D. , Cobbett Peter
Published: September 2004

Several types of whole‐cell outward K+ current recorded from magnocellular neurosecretory cells (MNCs) dissociated from the supraoptic nucleus of the adult guinea‐pig were identified on the basis of their voltage dependence, kinetics, pharmacology and Ca2+ dependence.

The predominant K+ current evoked from a holding potential of −40 mV was slowly activating, long‐lasting, tetraethylammonium (TEA) sensitive and showed little steady‐state inactivation. Also, this current was reduced by extracellular Cd2+. These data suggest that in supraoptic MNCs classical Ca2+‐insensitive, delayed rectifier channels (Kv) and Ca2+‐sensitive, non‐inactivating channels (KCa) both contribute to the sustained current.

A transient, low‐threshold K+ current, which was 4‐aminopyridine (4‐AP) sensitive and showed significant steady‐state inactivation, was evoked along with the sustained current from a holding potential of −90 mV. Based on these characteristics, this current corresponds to the A‐current (IK(A)) described in other neurons.

I
K(A) was activated when Ca2+ influx was blocked or when Ca2+ was absent from the extracellular medium, suggesting that Ca2+ influx is not necessary for activation of the current.

In many recordings, a transient 4‐AP‐insensitive outward current was evoked from a holding potential of −40 mV. This high‐threshold transient K+ current was abolished by extracellular Cd2+ or TEA and was absent when extracellular Ca2+ was replaced by Sr2+, suggesting that it is a transient Ca2+‐dependent K+ current.

We conclude that the presence of multiple types of K+ current may, in part, underlie the complex firing patterns of oxytocinergic and vasopressinergic MNCs.
Characterization of a nicotinamide–adenine dinucleotide‐dependent cation channel in the CRI‐G1 rat insulinoma cell line
Authors: Herson P. S. , Dulock K. A. , Ashford M. L. J.
Published: September 2004

Cell‐free excised membrane patches were used to examine the properties of a novel nicotinamide–adenine dinucleotide (β‐NAD+)‐activated ion channel in the rat insulin‐secreting cell line, CRI‐G1.

In inside‐out recordings, β‐NAD+ (0.05–1.0 mm) induced the appearance of a channel characterized by extremely slow kinetics, with mean open times in the range of seconds. The estimated EC50 for activation was 114 μm. Channel activity declined with time (run‐down) following activation by β‐NAD+ in excised patches and this was not prevented by intra‐cellular application of trypsin.

The single channel current–voltage relationship was linear with a conductance of 74 pS in symmetrical NaCl. The channel appears equally permeable to Na+, K+ and Cs+, exhibits an appreciable permeability to Ca2+, Mg2+ and Ba2+, but excludes anions.

The channel displays an unusual voltage sensitivity, with an abrupt increase in open‐state probability at depolarized voltages.

Channel opening, in the presence of β‐NAD+, required both Ca2+ and Mg2+ to be present at the internal side of the membrane. Activation by Ca2+ required a concentration of at least 10 μm and was maximal at 0.1 mm. Ba2+ did not substitute for Ca2+ in inducing channel activity nor did it inhibit activation by Ca2+. Increasing the concentration of intracellular Mg2+ stabilized the open state of NAD+‐activated channels.

The non‐selective cation channel reported here differs in its gating and modulatory characteristics from non‐selective cation channels described in other tissues. This channel may play a role in the pathophysiological reponses of β‐cells to oxidative stress.
GABAA receptor‐mediated IPSCs in rat thalamic sensory nuclei: patterns of discharge and tonic modulation by GABAB autoreceptors
Authors: Le Feuvre Y. , Fricker D. , Leresche N.
Published: September 2004

The patterns of discharge of spontaneous GABAA‐mediated inhibitory postsynaptic currents (sIPSCs), originating from the nucleus reticularis thalami (NRT), and their modulation by GABAg autoreceptors, were studied in rat thalamocortical (TO) neurones using whole‐cell voltage‐clamp recordings in brain slices.

sIPSCs were recorded in all ventro‐basal (VB) and dorsal lateral geniculate (LGN) neurones. In VB neurones, in the presence of tetraethylammonium (TEA, 5 mm), these sIPSCs can occur in bursts at frequencies of either 0.1 or 1–2 Hz. In the presence of tetrodotoxin (TTX), these bursting activities are replaced by the continuous discharge of miniature IPSCs (mIPSCs), recorded in the absence of TEA, at a frequency of 4 Hz. The kinetic properties of mIPSCs were similar in VB and LGN TC neurones.

In VB TC neurones the GABAB receptor agonist (±)‐baclofen, at a concentration of 0.05 μm, decreased the mIPSC frequency by 22% without affecting their amplitude distribution. Increasing the (±)‐baclofen concentration to 1 and 10 μm caused similar reductions (41 and 47%, respectively) in the mIPSCs frequency: these values were significantly different from the one observed with 0.05 μm (±)‐baclofen. In LGN TC neurones, where mIPSCs originate from both NRT and local interneurone terminals, 1 μm (±)‐baclofen produced a 66% reduction in the mIPSC frequency.

The GABAB receptor antagonist CGP55845A (50 nm) not only blocked the baclofen‐mediated decrease in mIPSC frequency, but also produced a 52% increase in the mIPSC frequency compared with control in three out of seven neurones. Application of CGP55845A (50–500 nm) alone produced a 77 % increase in the mIPSC frequency in three out of nine VB neurones, and in the LGN, CGP55845A (100 nm) produced a 53% increase in four out of nine neurones. CGP55845A (100 nm) also reversibly increased the amplitude of evoked GABAA IPSCs by 74 and 57 % in three out of three VB and three out of five LGN neurones, respectively.

Application of GABA (1.5–5 μm) decreased the mIPSC frequency in VB TC neurones by a similar extent (48%) as 1–10 μm (±)‐baclofen.

In the presence of 100 μm Cd2+, (±)‐baclofen still decreased the mIPSC frequency by about 40%, indicating that the effect of presynaptic GABAB receptor activation on spontaneous GABA release did not occur through a reduction of voltage‐dependent Ca2+ currents.

Cd2+ (100 μm) decreased the amplitude of both mIPSCs and isoguvacine‐induced current by 30 and 19%, respectively, indicating an effect of this divalent cation on postsynaptic GABAA receptors.

We conclude that GABAB autoreceptors are present on the GABAergic terminals within the thalamic sensory nuclei and that these receptors can be tonically activated by the ambient GABA.
Metabotropic synaptic regulation of intrinsic response properties of turtle spinal motoneurones
Authors: Delgado‐Lezama Rodolfo , Perrier Jean‐Francois , Nedergaard Steen , Svirskis Gytis , Hounsgaard Jørn
Published: September 2004

The effect of a brief train of electric stimuli in the dorsolateral funiculus on the intrinsic response properties of turtle motoneurones was investigated in transverse sections of the spinal cord in vitro.

Even when glutamatergic, GABAergic and glycinergic ionotropic synaptic transmission was blocked by antagonists of AMPA, NMDA, glycine and GABA receptors, dorsolateral funiculus (DLF) stimulation induced a facilitation of plateau potentials during current clamp and the underlying inward current in voltage clamp. This facilitation lasted more than 10 s.

The plateau potential and the facilitation by DLF stimulation was absent in the presence of 10 μm nifedipine. The DLF‐induced facilitation was reduced by antagonists of 5‐HT1a, group 1 metabotropic glutamate receptors and muscarine receptors.

These findings suggest that the intrinsic properties of spinal motoneurones are dynamically regulated by afferent synaptic activity. These afferents can be of spinal and extraspinal origin. Continuous regulation of intrinsic response properties could be a mechanism for motor flexibility.
Protein kinase A‐dependent and ‐independent stimulation of exocytosis by cAMP in mouse pancreatic B‐cells
Authors: Renström E. , Eliasson Lena , Rorsman P.
Published: September 2004

The mechanisms by which cAMP stimulates Ca2+‐dependent insulin secretion were investigated by combining measurements of whole‐cell Ca2+ currents, the cytoplasmic free Ca2+ concentration ([Ca2+]i) and membrane capacitance in single mouse B‐cells maintained in tissue culture.

Cyclic AMP stimulated exocytosis > 4‐fold in whole‐cell experiments in which secretion was evoked by intracellular dialysis with a Ca2+‐EGTA buffer with a [Ca2+]i of 1.5μm. This effect was antagonized by inhibitors of protein kinase A (PKA).

Photorelease of cAMP from a caged precursor potentiated exocytosis at Ca2+ concentrations which were themselves stimulatory (≥60 nm), but was without effect in the complete absence of Ca2+.

Elevation of intracellular cAMP (by exposure to forskolin) evoked a 6‐fold PKA‐dependent enhancement of the maximal exocytotic response (determined as the maximum increase in cell capacitance that could be elicited by a train of depolarizations) in perforated‐patch whole‐cell recordings.

Exocytosis triggered by single depolarizations in standard whole‐cell recordings was strongly potentiated by cAMP, but in this case the effect was unaffected by PKA inhibition.

When exocytosis was triggered by Ca2+ released from Ca2+‐NP‐EGTA (‘caged Ca2+), cAMP exerted a dual stimulatory effect on secretion: a rapid (initiated within 80 ms) PKA‐independent phase and a late PKA‐dependent component.

We conclude that cAMP stimulates insulin secretion both by increasing the release probability of secretory granules already in the readily releasable pool and by accelerating the refilling of this pool.