bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of inorganic phosphate on cross‐bridge kinetics at different activation levels in skinned guinea‐pig smooth muscle.
Authors: Osterman A , Arner A
Published: December 2014

1. The effects of inorganic phosphate (P(i)) on force, Ca(2+)‐force relationship, ATPase activity, maximal shortening velocity (Vmax) and rate of tension development were investigated in chemically skinned preparations of smooth muscle from the guinea‐pig taenia coli. 2. In maximally thiophosphorylated fibres, P(i) in the range 1‐40 mM inhibited isometric force, with a reduction of 20% at 20 mM P(i). 3. The relative force was similar at all [Ca2+], i.e. the Ca(2+)‐force relationship was not affected, when 20 mM P(i) was present. 4. After photolytic release of ATP from caged ATP in maximally thiophosphorylated fibres in the presence of 20 mM P(i), tension rose to a lower level but with a higher rate constant than in the absence of P(i). 5. Inorganic phosphate (20 mM) did not affect the ATP hydrolysis in fibres activated at intermediate [Ca2+] or by maximal thiophosphorylation. 6. Inorganic phosphate (20 mM) decreased force but did not influence Vmax in maximally activated fibres. At lower levels of activation by Ca2+, P(i) increased the Vmax and decreased force slightly without affecting the degree of myosin light chain phosphorylation. 7. We conclude that P(i) influences cross‐bridge reactions associated with force generation in smooth muscle. These reactions are not rate limiting for cross‐bridge turnover under isotonic or isometric conditions in maximally activated smooth muscle fibres, since P(i) did not influence Vmax or the rate of ATP turnover. 8. Since P(i) increased Vmax in submaximally activated muscles, we propose that, under these conditions, shortening velocity is rate limited by cross‐bridge states, reached early after attachment, which impose a mechanical resistance to shortening.
Decreased calcium currents in motor nerve terminals of mice with Lambert‐Eaton myasthenic syndrome.
Authors: Smith D O , Conklin M W , Jensen P J , Atchison W D
Published: December 2014

1. The effects of immunoglobulin G (IgG) from patients with Lambert‐Eaton myasthenic syndrome on Ca2+ currents in mammalian motor nerve terminals are unknown. Therefore, we recorded these currents in phrenic nerves of mice injected with serum from either LEMS patients, myasthenia gravis patients, or healthy control individuals. 2. In control preparations, the endplate currents induced by repetitive stimulation at > or = 20 Hz were depressed as expected. However, in the LEMS animals quantal content decreased and either depression did not occur or synaptic facilitation occurred. 3. Ca2+ currents were smaller in LEMS animals. At 0.5 Hz stimulation frequency, normalized Ca2+ currents in LEMS animals were 57 +/‐ 14% of those in control. At higher frequencies, Ca2+ currents become smaller in control but not in LEMS animals. 4. Ca2+ currents in controls were unaffected by addition of nifedipine but were reduced by 37% upon addition of omega‐conotoxin GVIA. In LEMS animals, however, the currents were depressed by 43% by nifedipine but were unaffected by omega‐conotoxin GVIA. Thus, LEMS is associated with reduced Ca2+ currents and a shift to dihydropyridine sensitivity.
Residual Ca2+ channel current modulation by megestrol acetate via a G‐protein alpha s‐subunit in rat hypothalamic neurones.
Authors: Costa A M , Spence K T , Plata-Salamán C R , ffrench‐Mullen J M
Published: December 2014

1. The inhibition of voltage‐activated Ca2+ channel currents by the orally active progesterone derivative, megestrol acetate (MA), was examined in freshly dissociated rat ventromedial hypothalamic nucleus (VMN) neurones using the whole‐cell voltage‐clamp technique with 10 mM Ba2+ as the charge carrier. 2. The steady‐state inhibition of the peak high‐threshold Ca2+ channel current evoked by depolarization from ‐80 to ‐10 mV by MA increased in a concentration‐dependent fashion. MA inhibited a fraction of the whole‐cell Ca2+ channel current while progesterone had no effect on the peak Ca2+ channel current (7% at 10 microM). The low‐threshold Ca2+ (T‐type) current, evoked from ‐100 to ‐30 mV, was unaffected by MA. 3. Intracellular dialysis with MA had no effect on the Ca2+ channel current. Concomitant extracellular perfusion of MA showed normal inhibitory activity, suggesting that the MA binding site can only be accessed extracellularly. 4. The high‐threshold Ca2+ channel current in VMN neurones was found to consist of four pharmacologically distinguishable components: an N‐type current, an L‐type current, a P‐type current, and a residual current. MA had no effect on the N‐, L‐ and P‐type Ca2+ channel currents, but inhibited the residual current. 5. In neurones isolated from cholera toxin‐treated animals, the MA‐induced inhibition of the Ca2+ channel current was significantly diminished, suggesting a G‐protein alpha S‐subunit involvement. 6. Treatment with antisense phosphothio‐oligodeoxynucleotides to the G alpha S‐subunit (antisense‐G alpha S) significantly reduced the MA‐induced inhibition of the Ca2+ channel current. Treatment with either sense‐G alpha S or antisense‐G alpha 11 had no effect, confirming a G alpha S‐subunit involvement. 7. These results suggest that appetite enhancement induced by MA in cachectic patients may in part be due to a novel central nervous system action, that is, inhibition of a fraction of the whole‐cell Ca2+ channel current to attenuate the firing of VMN neurones that may be involved in satiety mechanisms.
The effects of extracellular pH and calcium change on force and intracellular calcium in rat vascular smooth muscle.
Authors: Austin C , Wray S
Published: December 2014

1. In order to investigate the mechanism whereby changes in external pH (pHo) alter tone in rat mesenteric resistance vessels, we have made simultaneous measurements of tension and intracellular Ca2+ [Ca2+]i. Strips of mesenteric artery were loaded with the Ca(2+)‐sensitive indicator indo‐1 and superfused with physiological salt solution at pH 7.4 and 37 degrees C. 2. An increase of pHo from 7.4 to 7.9 produced an increase in tension. This was accompanied by an increase in [Ca2+]i in resting and high‐K(+)‐depolarized vessels. Acidification to 6.9 reduced tension and was associated with a fall in [Ca2+]i. Over the pHi range examined, 6.6‐7.9, parallel changes in [Ca2+]i and tension were found in K(+)‐activated vessels. 3. In contrast to the relatively slow change in [Ca2+]i, pHi and tension with change of pHo, depolarization produced rapid changes in [Ca2+]i and tension, consistent with a more direct action on Ca2+ mobilization. 4. Reducing the external [Ca2+] below 1 mM produced a pronounced fall in [Ca2+]i and force. Changes in [Ca2+]i, produced by alteration of external [Ca2+] (Cao2+) were used to examine the relation between [Ca2+]i and tension. A linear relation was found. Alteration of pHo to 6.9 or 7.9 did not significantly change this relation. When the tension data were normalized to their own maxima, no shift in the tension‐Ca2+ relation occurred, suggesting little or no effect of pH on the Ca2+ sensitivity of force production by the contractile proteins. 5. To determine further whether the changes in [Ca2+]i produced by alteration of pHo could account for all the changes observed in tension, [Ca2+]i was restored to control levels while maintaining an altered pHo. When this was done, restoration of [Ca2+]i led to restoration of force. Thus, in this preparation, the changes in [Ca2+]i produced by altering pHo in depolarized vessels can account for the changes in vascular tone.
Maturation of rat renal phosphate transport: effect of triiodothyronine.
Authors: Euzet S , Lelièvre-Pégorier M , Merlet-Bénichou C
Published: December 2014

1. Prevention of weaning prevents the increase in the carrier affinity for Na(+)‐cotransported phosphate (Pi) and the normal elevation of plasma 3,5,3'‐triiodothyronine (T3) which occur between postnatal days 14 and 21. 2. This study examines the possible role of T3 in the control of the maturation process of Pi transport. Clearance experiments and brush‐border membrane studies were performed on 14‐day‐old rats given T3. 3. The fractional excretion of Pi of T3‐treated rats was 33% lower compared with controls (P < 0.01). After Pi perfusion, it remained at a lower level, and the amount of Pi reabsorbed per minute, corrected for the glomerular filtration rate (RPi/GFR), was higher. 4. The membrane vesicles from 14‐day‐old rats given T3 showed a 30% increase in carrier affinity for Na(+)‐cotransported Pi. In addition to this maturational effect of T3, a 46% increase in Vmax was also observed. 5. The amount of immunoreactive Pi transporter, detected using anti‐(NaPi‐2) antiserum, was increased in T3‐treated rats. 6. Glucose transport, another Na(+)‐dependent transport system, was not altered by T3. 7. It is concluded that exogenous T3 given before the third postnatal week specifically induced precocious maturation of renal Pi transport in 14‐day‐old rats, suggesting that thyroid hormone is normally involved in this maturation.
Mesenteric blood pressure profile of conscious, freely moving rats.
Authors: Fenger-Gron J , Mulvany M J , Christensen K L
Published: December 2014

1. Blood pressure has been measured in the aorta and at four points in the mesenteric circulation of conscious, freely moving rats under physiological, resting conditions. 2. Using small polythene catheters, blood pressure was measured simultaneously in the aorta and either distally in the superior mesenteric artery (group A), at the base of a mesenteric arterial arcade (vessel diameter ca 100 microns) (group B), at the base of a mesenteric venous arcade (group C) or distally in the superior mesenteric vein (group D). Local blood flow distribution proximal and distal to the measurement point was restored after the cannulations through appropriate ligations. 3. In conscious animals 5‐17 h after surgery, systemic mean blood pressure was 121 +/‐ 2 mmHg. Local pressures at the four locations (as a percentage of systemic pressure) were: 95 +/‐ 1% in group A, 64 +/‐ 2% in group B, 13 +/‐ 1% in group C and 7 +/‐ 1% in group D. Thus, large arteries dissipated 5% of the total pressure drop, arcade small arteries 31%, the intramural circulation 51%, arcade veins 6% and the remaining veins plus the hepatic circulation 7%. 4. Immediately after surgery, the corresponding pressure drops were 4, 16, 66, 5 and 9%, respectively, thus emphasizing that the pressure profile can be profoundly affected by surgery and anaesthesia. 5. The data indicate that under resting conditions in conscious, freely moving rats, half the mesenteric vascular resistance resides outside the intramural circulation, primarily in the arcade small arteries.
Motor effects of stimulating the human cerebellar thalamus.
Authors: Ashby P , Lang A E , Lozano A M , Dostrovsky J O
Published: December 2014

1. Observations were made on seven human subjects with electrodes implanted in the cerebellar thalamus for the control of tremor. 2. Weak stimulation at 1‐2 Hz resulted in sudden lapses of posture of actively contracting contralateral limb muscles. Stronger stimuli caused muscle twitches even in relaxed muscles. 3. The stronger stimuli produced short latency facilitation of EMG activity in contralateral muscles; the pattern of muscle facilitation, the conduction velocity of the descending pathway and the estimated rise time of the underlying composite EPSP were consistent with direct activation of the corticospinal tract. 4. The lapses of posture produced by the weaker stimuli were associated with inhibition of ongoing EMG for up to 150 ms. This was due to the interruption of tonic drive to motoneurons rather than to their inhibition and was associated with depression of the motor evoked potential in relaxed muscles produced by weak anodal transcranial stimulation. The inhibition could be produced by 0.1 ms pulses, implying that a large‐diameter fibre system was being activated. 5. A number of mechanisms could contribute to the inhibition, including inhibition from the reticular nucleus of the thalamus.
Hypothalamic interaction between macrophage inflammatory protein‐1 alpha (MIP‐1 alpha) and MIP‐1 beta in rats: a new level for fever control?
Authors: Miñano F J , Fernández-Alonso A , Myers R D , Sancibrián M
Published: December 2014

1. The microinjection of macrophage inflammatory protein‐1 (MIP‐1 alpha; 5.0 and 25 pg) into the anterior hypothalamic, preoptic area (AHPOA) induced a slow onset; monophasic fever in rats that persisted for a long period. Microinjection of 25 pg MIP‐1 beta into the AHPOA induced a fever of rapid onset, whereas 5.0 pg MIP‐1 beta did not alter body temperature (Tb) significantly. When either MIP‐1 alpha or MIP‐1 beta was heated to 70 degrees C for 30 min prior to their injection, no pyrexic response was produced. 2. The concurrent microinjection of 25 pg MIP‐1 alpha and 25 pg MIP‐1 beta into the AHPOA attenuated the effects on Tb of either cytokine alone. However, pretreatment with either 5.0 pg MIP‐1 beta or 5.0 pg MIP‐1 alpha suppressed the febrile response induced by 25 pg MIP‐1 alpha or 25 pg MIP‐1 beta, given at the same site, respectively. 3. The present experiments show that MIP‐1 alpha and MIP‐1 beta are active individually and possess distinct differences in their evocation of a febrile response. Further, our results suggest a functional antagonism between MIP‐1 alpha and MIP‐1 beta that could represent a new level in the development of fever.
Voltage‐dependent interaction of open‐channel blocking molecules with gating of NMDA receptors in rat cortical neurons.
Authors: Antonov S M , Johnson J W
Published: December 2014

1. The mechanisms by which four adamantane derivatives (IEM‐1857, ‐1592, ‐1460 and ‐1754) block the open NMDA‐activated channel were studied at membrane voltages (Vm) from ‐170 to +30 mV. The rate constants of channel block (k+) and of channel unblock (k‐) were measured from the fully resolvable flicker of single‐channel currents induced by each compound. 2. The k+ of each compound exhibited a similar exponential dependence on voltage over the Vm range studied. 3. The k‐ of IEM‐1857 and IEM‐1592 over the Vm range studied, and of IEM‐1754 and IEM‐1460 from ‐30 to ‐90 mV, exhibited similar exponential dependencies on voltage. However, the k‐ of IEM‐1754 and IEM‐1460 at Vm values more hyperpolarized than ‐90 mV were much more steeply voltage dependent, suggesting that at these Vm values the two drugs can occupy a deeper binding site. 4. Each of the drugs induced a concentration‐dependent prolongation of the mean burst length at ‐90 mV, suggesting that while blocking they can interfere with channel closure. 5. The prolongation of mean burst length induced by the largest drug (IEM‐1857) increased with hyperpolarization. The increase was consistent at each Vm with the predictions of the sequential scheme of block, suggesting that channel closure is prevented when IEM‐1857 is bound. The prolongation of burst length induced by the smallest drug (IEM‐1754) was less than predicted by the sequential scheme and the deviation increased with hyperpolarization. 6. The IEM‐1857 concentration‐dependence of number of blockages per unit open time had a slope equal to k+ at ‐150 mV. The IEM‐1754 concentration‐dependence of number of blockages per unit open time revealed a slope about two times less than k+ for this compound at ‐150 mV. 7. The mean patch current was not significantly altered by 3 microM IEM‐1857 at Vm values from ‐90 to ‐150 mV, as expected of a drug that prevents channel closure when blocking. Mean patch current significantly decreased with hyperpolarization beyond ‐90 mV in the presence of 1 microM IEM‐1754. 8. The data suggest that there are two blocking sites at different depths within the NMDA‐activated channel. Channel closure is prevented when any of the IEM drugs occupy the shallow blocking site. Channel closure is permitted during occupation of a deeper blocking site that can be reached only by the smaller IEM drugs at hyperpolarized voltages.
Dopamine D1‐like receptor‐mediated presynaptic inhibition of excitatory transmission onto rat magnocellular basal forebrain neurones.
Authors: Momiyama T , Sim J A , Brown D A
Published: December 2014

1. Excitatory postsynaptic currents (EPSCs) following focal afferent stimulation were recorded from patch‐clamped magnocellular neurones in a thin‐slice preparation of the rat basal forebrain. Evoked EPSCs had a mean decay time constant of 3.81 +/‐ 0.09 ms and were reversibly blocked by 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX, 5 microM). 2. Bath‐applied dopamine (DA) reduced evoked EPSC amplitude by up to 54.2 +/‐ 2.3% with an IC50 of 19.9 microM in normal Krebs solution (2.5 mM Ca2+, 1.2 mM Mg2+) without effect on postsynaptic holding current. 3. DA (30 microM) reduced the mean frequency of spontaneous miniature EPSCs recorded in 0.5 microM tetrodotoxin without affecting their mean amplitude, rise time or decay time constant. This effect was diminished by 100 microM Cd2+. 4. The effect of DA on evoked EPSCs was mimicked by the D1‐like receptor agonist, SKF 81297 (IC50 25.6 microM), but not by the D2‐like receptor agonist R(‐)‐TNPA (30 microM) or (‐)‐quinpirole (30 microM), and was antagonized by the D1‐like receptor antagonist R(+)‐SCH 23390 (estimated dissociation constant KB = 1.7 microM) but not by the D2‐like receptor antagonist S(‐)‐eticlopride (10 microM). 5. Forskolin (10 microM) reduced evoked EPSCs to approximately 60% of the control amplitude, and occluded the effect of subsequent application of DA. 6. These results suggest that glutamatergic afferents to magnocellular basal forebrain neurones possess presynaptic D1‐like DA receptors, and that activation of these receptors reduces excitatory glutamatergic transmission, probably via an adenylyl cyclase‐dependent pathway.
Histamine H3 receptor‐mediated depression of synaptic transmission in the dentate gyrus of the rat in vitro.
Authors: Brown R E , Reymann K G
Published: December 2014

1. The effects of histamine on excitatory synaptic transmission in the dentate gyrus region of rat hippocampal slices were examined using extracellular and whole‐cell patch‐clamp recording techniques. The GABAA receptor antagonist picrotoxin (50 microM) was present in the bath in all experiments. 2. Histamine (0.7‐70 microM) reversibly depressed field excitatory postsynaptic potentials (fEPSPs) or excitatory postsynaptic currents (EPSCs) recorded intracellularly by up to 30%. The presynaptic fibre volley and EPSC reversal potential were unaffected by histamine, as were responses following pressure ejection of the glutamate receptor agonist S‐alpha‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (S‐AMPA) into the slice. 3. Histamine (7 microM) depressed equally the AMPA and N‐methyl‐D‐aspartate (NMDA) components of the dual‐component EPSC, recorded at ‐40 mV. 4. In addition to depressing synaptic transmission, histamine also reduced the magnitude of paired‐pulse depression (PPD; 40 ms interpulse interval) of the medial perforant path EPSC. 5. Histamine depressed medial perforant path EPSCs more strongly than lateral perforant path EPSCs. Paired‐pulse facilitation (PPF; 40 ms interpulse interval) in the lateral perforant path was enhanced by histamine. 6. The effects of histamine on synaptic transmission and PPD were mimicked by the selective H3 receptor agonist R‐alpha‐methylhistamine (0.1‐10 microM) but not by the selective H2 receptor agonist dimaprit (10 microM). Similarly, the H3 receptor antagonist thioperamide (10 microM) blocked the effect of histamine whereas the H1 antagonist mepyramine (1 microM) and the H2 receptor antagonist cimetidine (50 microM) were ineffective. 7. Histamine actions on synaptic transmission and PPD were not occluded by application of the metabotropic glutamate agonist L‐2‐amino‐4‐phosphonobutyrate (AP4). 8. The results indicate that histamine depresses synaptic transmission in the dentate gyrus by binding to histamine H3 receptors located on perforant path terminals. The mechanism by which histamine depresses transmission is independent of that used by class III metabotropic glutamate receptors.
Two human paramyotonia congenita mutations have opposite effects on lidocaine block of Na+ channels expressed in a mammalian cell line.
Authors: Fan Z , George A L , Kyle J W , Makielski J C
Published: December 2014

1. Two mutant human skeletal muscle voltage‐gated Na+ channel alpha‐subunits (hSkM1), with mutations found in patients with hereditary paramyotonia congenita (T1313M on the III‐IV linker and R1448C on the outside of S4 of repeat IV), and wild‐type hSkM1 channels were expressed in a human embryonic kidney cell lines (tsA201) using recombinant cDNA. 2. Compared with wild‐type, both mutants exhibited altered inactivation phenotypes. Current decay was slowed for both, but voltage‐dependent availability from inactivation was shifted in the negative direction for R1448C and in the positive direction for T1313M. 3. The hypothesis that a local anaesthetic, lidocaine (lignocaine), binds primarily to the inactivated state to block the channel was reassessed by testing lidocaine block of these two mutants and the wild‐type channel. 4. T1313M showed reduced phasic block, but R1448C showed increased phasic block for trains of depolarizations. 5. Rest block (from ‐120 mV) was increased for R1448C (IC50 approximately equal to 0.2 mM) and decreased for T1313M (IC50 approximately equal to 1.3 mM) compared with wild‐type (IC50 approximately 0.5 mM), but these differences were diminished at a holding potential of ‐150 mV, suggesting that the differences were caused by binding to the inactivated state rather than a different affinity of lidocaine for the resting state. 6. Inactivated state affinity measured from lidocaine‐induced shifts in voltage‐dependent availability was reduced for T1313M (Kd = 63 microM) but little changed for R1448C (Kd = 14 microM) compared with wild‐type (Kd = 11 microM). Two pulse recovery protocols showed faster recovery from lidocaine block for T1313M and slower recovery for R1448C. Together these accounted for the opposite effects on lidocaine phasic block observed for the mutant channels. 7. Neither mutation is located at a putative lidocaine binding site in domain 4 S6, yet both affected lidocaine block. The data suggest that R1448C altered phasic lidocaine block mainly through altered kinetics, but T1313M altered block through a change in affinity for the inactivated state. These findings have implications for drug therapy of paramyotonia congenita, and also provide an insight into structural requirements for drug affinity.
Voltage‐activated proton current in eosinophils from human blood.
Authors: Gordienko D V , Tare M , Parveen S , Fenech C J , Robinson C , Bolton T B
Published: December 2014

1. The resting membrane potential of freshly purified normodense human eosinophils bathed in and dialysed with quasi‐physiological solutions was ‐63 +/‐ 2 mV (n = 100). 2. In voltage‐clamp mode with quasi‐physiological internal and external solutions, voltage steps from the holding potential of ‐60 mV to levels positive to +20 mV resulted in development of a quasi‐instantaneous outward current and a slowly developing outward current. The instantaneous current was absent when the cells were bathed in and dialysed with K(+)‐free solution. 3. The slow outward current persisted following simultaneous replacement of K+, Na+ and most of the Cl‐ with largely impermeant ions (tetraethylammonium, N‐methyl‐D‐glucamine and methanesulphonate) and was augmented when the cell was dialysed with a solution of increased buffering capacity for protons. The observed reversal potential of the current closely followed the hydrogen equilibrium potential over a wide range of internal‐external pH combinations, indicating that the conductance underlying the slow outward current was highly selective for H+ ions. 4. Acidification of the pipette solution (increasing [H+]i) augmented the outward H+ current and shifted its activation range negatively, whilst acidification of the external solution had the opposite effect. The voltage dependence of the current is modulated by the transmembrane pH gradient so the only outward current could be activated. However, when the outward current was activated by a voltage step, rapid acidification of external solution produced an inward H+ current which rapidly deactivated. 5. The proton current was reversibly inhibited in a voltage‐dependent manner by extracellular application of Zn2+. The apparent dissociation constants were 8 nM (at +40 mV), 36 nM (at +70 mV) and 200 nM (at +100 mV). 6. The proton current was augmented by exposure to 10 microM arachidonic acid. This augmentation consisted of a shift of the voltage dependence of activation to more negative potentials and enhancement of maximum conductance (gH,max). The proton current recorded in eosinophils was significantly augmented under conditions of elevated cytosolic free calcium concentration ([Ca2+]i). The threshold level of [Ca2+]i associated with this effect lay between 0.1 and 1 microM and was not measurably affected by cytosolic acidification. 7. Eosinophils from human blood possess a voltage‐dependent H+ conductance (gH) which normally allows protons to move outwards only; raising [Ca2+]i was associated with augmentation of gH and intracellular acidification or arachidonate shifted its activation range negatively towards physiological potentials.
Spatially non‐uniform Ca2+ signals induced by the reduction of transverse tubules in citrate‐loaded guinea‐pig ventricular myocytes in culture.
Authors: Lipp P , Hüser J , Pott L , Niggli E
Published: December 2014

1. Ratiometric confocal microscopy and the whole‐cell patch clamp technique were used to simultaneously record intracellular Ca2+ transients and membrane currents from guinea‐pig ventricular myocytes. Intracellular dialysis with the low‐affinity Ca2+ buffer citrate enabled us to record and analyse Ca2+ transients caused by Ca2+ influx alone and by additional Ca2+ release from the sarcoplasmic reticulum (SR) in the same cell. 2. In freshly isolated adult myocytes (used within 1‐4 h of isolation) both types of Ca2+ transients (‘Ca2+ entry’ and ‘Ca2+ release’ transients) were spatially uniform regardless of the Ca2+ current (ICa) duration. In contrast, Ca2+ transients in short‐term cultured (1‐2 days) myocytes exhibited marked spatial inhomogeneities. ICa frequently evoked Ca2+ waves that propagated from either or both ends of the cardiac myocyte. Reduction of the ICa duration caused Ca2+ release that was restricted to one of the two halves of the cell. 3. Analysis of the Ca2+ entry signals in freshly isolated and short‐term cultured myocytes indicated that the spatial properties of the Ca2+ influx signal were responsible for the spatial properties of the triggered Ca2+ release from the SR. In freshly isolated ventricular myocytes Ca2+ influx was homogeneous while in short‐term cultured cells pronounced Ca2+ gradients could be found during Ca2+ influx. Spatial non‐uniformities in the amplitude of local Ca2+ entry transients were likely to cause subcellularly restricted Ca2+ release. 4. The changes in the spatial properties of depolarization‐induced Cai2+ signals during short‐term culture were paralleled by a decrease (to 65%) in the total cell capacitance. In addition, staining the sarcolemma with the membrane‐selective dye Di‐8‐ANEPPS revealed that, in cultured myocytes, t‐tubular membrane connected functionally to the surface membrane was reduced or absent. 5. These results demonstrate that the short‐term culture of adult ventricular myocytes results in the concomitant loss of functionally connected t‐tubular membrane. The lack of the t‐tubular system subsequently caused spatially non‐uniform SR Ca2+ release. Evidence is presented to show that in ventricular myocytes lacking t‐tubules non‐uniform SR Ca2+ release was, most probably, the result of inhomogeneous Ca2+ entry during ICa. These findings directly demonstrate the functional importance of the t‐tubular network for uniform ventricular Ca2+ signalling.
Effects of reduced muscle glycogen concentration on force, Ca2+ release and contractile protein function in intact mouse skeletal muscle.
Authors: Chin E R , Allen D G
Published: December 2014

1. The purpose of this study was to examine the effects of reduced glycogen concentration on force, Ca2+ release and myofibrillar protein function during fatigue in skeletal muscle. Force and intracellular free Ca2+ concentration ([Ca2+]i) were measured in single mammalian skeletal muscle fibres during fatigue and recovery. Glycogen was measured in bundles of 20‐40 fibres from the same muscle under the same conditions. 2. Fatigue was induced by repeated maximum tetani until force was reduced to 30% of initial. This was associated with a reduction in muscle glycogen to 27 +/‐ 6% of control values. In fibres allowed to recover for 60 min in the presence of 5.5 mM glucose (n = 6), tetanic (100 Hz) force recovered fully but tetanic [Ca2+]i remained at 82 +/‐ 8% of initial values. This prolonged depression in Ca2+ release was not associated with decreased muscle glycogen since glycogen had recovered to pre‐fatigue levels (157 +/‐ 42%). 3. To examine the responses under conditions of reduced muscle glycogen concentration, fibres recovered from fatigue for 60 min in the absence of glucose (n = 6). After glucose‐free recovery, the decreases in tetanic force and [Ca2+]i were only partially reversed (to 64 +/‐ 8% and 57 +/‐ 7% of initial values, respectively). These alterations were associated with a sustained reduction in muscle glycogen concentration (27 +/‐ 4% of initial values). 4. In another set of fibres, fatigue was followed by 50 Hz intermittent stimulation for 22.6 +/‐ 4 min. With this protocol, tetanic force and [Ca2+]i partially recovered to 76 +/‐ 9% and 55 +/‐ 6% of initial levels, respectively. These changes were associated with a recovery of muscle glycogen (to 85 +/‐ 10%). 5. During fatigue, Ca2+ sensitivity and maximum Ca(2+)‐activated force (Fmax) were depressed but these alterations were fully reversed when muscle glycogen recovered. When glycogen did not recover, Ca2+ sensitivity remained depressed but Fmax partially recovered. The altered myofibrillar protein function is probably due to alterations in inorganic phosphate levels or other metabolites associated with reduced levels of muscle glycogen. 6. These data indicate that the reductions in force, Ca2+ release and contractile protein inhibition observed during fatigue are closely associated with reduced muscle glycogen concentration. These findings also suggest that the changes in Ca2+ release associated with fatigue and recovery have two components‐one which is glycogen dependent and another which is independent of glycogen but depends on previous activity.
Potassium currents in acutely isolated human hippocampal dentate granule cells.
Authors: Beck H , Clusmann H , Kral T , Schramm J , Heinemann U , Elger C E
Published: December 2014

1. Properties of voltage‐ and Ca(2+)‐dependent K+ currents were investigated in thirty‐four dentate granule cells acutely isolated from the resected hippocampus of eleven patients with therapy‐refractory temporal lobe epilepsy (TLE). 2. When intracellular Ca2+ was strongly buffered with 11.5 mM EGTA‐1 mM Ca2+ in the recording pipette, K+ currents (IK) with a slow activation and biexponential time‐dependent decay could be elicited, which showed a threshold for activation around ‐30 mV. 3. A contribution of Ca(2+)‐dependent K+ currents became apparent with intracellular solution containing 1 mM BAPTA‐0.1 mM Ca2+. Superfusion of low‐Ca2+ extracellular solution blocked 43% of outward currents in this recording configuration. Outward current components could also be blocked by substituting 5 mM Ba2+ for extracellular Ca2+ (78%), or by application of 100 microM Cd2+ (25%). 4. The Ca(2+)‐dependent K+ currents could be pharmacologically subdivided into two components. One component was sensitive to 500 microM tetraethylammmonium (TEA; 41%) and 10 nM charybdotoxin (CTX; 47.2%). The blocking effects of 10 nM CTX and 500 microM TEA were not additive, suggesting that both agents block the same conductance. A second, smaller outward current component was blocked by 50 nM apamin (13%). 5. A transient A‐type K+ current could be observed in six neurones and showed a fast monoexponential time‐dependent inactivation with a steady‐state voltage dependence that was distinct from that of IK. The A‐type current was blocked by 4‐aminopyridine (4‐AP) but not by TEA or low‐Ca2+ solution. 6. We conclude that outward currents in human hippocampal dentate granule cells can be separated into at least four types by their kinetic and pharmacological properties. These include at least one voltage‐dependent current similar to those observed in mammalian hippocampal neurones, and two Ca(2+)‐dependent K+ currents that most probably correspond to SK‐ and BK‐type currents. A classical A‐type current could be detected in some patients with Ammon's horn sclerosis (AHS) but not in patients with lesion‐associated TLE.
Brain‐derived neurotrophic factor and nerve growth factor potentiate excitatory synaptic transmission in the rat visual cortex.
Authors: Carmignoto G , Pizzorusso T , Tia S , Vicini S
Published: December 2014

1. The effect of brain‐derived neurotrophic factor (BDNF) and nerve growth factor (NGF) on excitatory synaptic transmission in the developing visual cortex was studied by whole‐cell patch‐clamp recordings from rat brain slices. 2. Both neurotrophins induced a rapid increase in the amplitude of impulse‐evoked excitatory postsynaptic currents (EPSCs). BDNF also increased the frequency of spontaneous EPSCs. 3. Analysis of the currents revealed that alpha‐amino‐3‐hydroxy‐5‐methyl‐isoxazole propionic acid (AMPA) and N‐methyl‐D‐aspartate (NMDA) receptor‐mediated components contributing to the EPSC peak amplitude were equally potentiated by the neurotrophins. 4. When synaptic transmission was studied by minimal stimulation of intracortical afferents, neurotrophins induced a decrease in the occurrence of release failures. 5. A number of neurones were insensitive to the effects of the neurotrophins, possibly related to the considerable heterogeneity of neuronal types and to the uneven distribution of neurotrophin receptors in the visual cortex. 6. The probability of neurotransmitter release represents a rapidly modifiable synaptic feature by which neurotrophins can potentiate the efficacy of excitatory synaptic transmission in the visual cortex.
Inhibition of nitroxidergic nerve function by neurogenic acetylcholine in monkey cerebral arteries.
Authors: Toda N , Ayajiki K , Okamura T
Published: December 2014

1. Modification by endogenous or exogenous acetylcholine and vasoactive intestinal polypeptide (VIP) of vasodilatation mediated by nitric oxide (NO) released from nitroxidergic nerves was studied in isolated monkey cerebral arteries. In arterial strips denuded of endothelium, transmural electrical stimulation (2‐20 Hz) produced relaxations that were abolished by tetrodotoxin. 2. The relaxation response was attenuated by acetylcholine, and the attenuation was reversed by atropine. Attenuation was also observed with AF‐DX 116, an antagonist of the muscarinic acetylcholine receptor subtype, M2. NO‐induced relaxation was not affected by acetylcholine. Neurogenic relaxation was also inhibited by physostigmine and potentiated by atropine. 3. VIP in concentrations that elicited slight relaxation did not alter the response to nerve stimulation. In the strips showing tachyphylaxis to VIP, the neurogenic response was not inhibited. 4. Histochemical studies of whole‐mount preparations revealed nerve fibres with NO synthase and VIP immunoreactivity, and also acetylcholinesterase, suggesting the presence of perivascular nitroxidergic, VIPergic and cholinergic innervation. 5. It is concluded that the actions of nitroxidergic nerve fibres on the monkey cerebral artery are inhibited by nerve‐released acetylcholine acting on prejunctional muscarinic receptors, possibly of the M2 subtype. Despite the presence of VIP immunoreactive nerve fibres and the ability of exogenous VIP to relax the artery, there is no evidence supporting either a prejunctional modulation of nitroxidergic nerve function by VIP or a role for VIP as a vasodilatory neurotransmitter.
Role of submandibular and sublingual saliva in maintenance of taste sensitivity recorded in the chorda tympani of rats.
Authors: Matsuo R , Yamauchi Y , Morimoto T
Published: December 2014

1. To evaluate the role of saliva in the maintenance of taste sensitivity, the activities in the rat chorda tympani innervating taste buds in the anterior part of the tongue were analysed. The effects of chronic extirpation of the submandibular and sublingual salivary glands were tested and compared with results after chronic oral administration of artificial saliva. 2. Removal of the salivary glands sharply decreased chorda tympani responses to four different taste stimuli by 7 days post‐desalivation, while a stable response to cold water was observed by at least 28 days. 3. This selective decrease in taste responses was considerably recovered by 7‐day‐oral injection of artificial saliva (containing NaHCO3, KCl and/or mucin) or distilled water. However, the injection of the salt‐containing artificial saliva induced significantly larger sucrose and smaller NaCl, HCl and quinine responses than did the injection of distilled water. 4. In our salivary manipulations, an alteration in the number of the functional sweet receptors was suggested by the cross‐adaptation technique using NaHCO3, whereas sensitivity to the epithelial sodium transport blocker, amiloride, was stable in the NaCl response. 5. Salivary water and electrolytes which may participate in forming the external environment of the taste receptor cells modulated taste sensitivity in the chorda tympani.
Long‐term facilitation of ventilation following repeated hypoxic episodes in awake goats.
Authors: Turner D L , Mitchell G S
Published: December 2014

1. This study tested two hypotheses: (1) that episodic hypoxia elicits long‐term facilitation (LTF) in respiratory neurons that is manifest as an increase in ventilation in awake goats; and (2) that LTF causes complex changes in respiratory pattern which are responsible for the increase in ventilation. 2. Each goat participated in two protocols. In the first, inspired gas mixtures were alternated between isocapnic normoxia and hypoxia (arterial partial pressure of oxygen, Pa,O2 = 47 mmHg) for ten cycles. Each hypoxic episode lasted 3 min and normoxic intervals were 5 min. Ventilatory variables were measured during the last minute of each episode and periodically for up to 1 h following the last hypoxic episode. The second, sham protocol was undertaken at least 2 weeks later and was identical to the first, except that isocapnic hypoxia was replaced with normoxia. 3. Inspired ventilation (VI) increased during the first isocapnic hypoxic episode and reached progressively higher levels in subsequent hypoxic episodes. VI also increased progressively among normoxic intervals, such that by the tenth normoxic interval, it had increased 68% relative to the comparable sham value (P < 0.05). Respiratory frequency (FR), tidal volume and mean inspiratory flow all contributed to the augmented VI during both isocapnic normoxia and hypoxia. The increase in VI lasted up to 40 min after the final hypoxic episode, with an increased FR making the greatest contribution. The persistent increase in VI strongly suggests that episodic hypoxia elicits LTF in respiratory neurons in the awake goat. Complex changes in respiratory pattern underpin the ventilatory manifestation of LTF.
The immunophilin FK506‐binding protein modulates Ca2+ release channel closure in rat heart.
Authors: Xiao R P , Valdivia H H , Bogdanov K , Valdivia C , Lakatta E G , Cheng H
Published: December 2014

1. The nature of the signal that terminates the release of Ca2+ from the cardiac sarcoplasmic reticulum has remained elusive. This study was intended to examine whether FK506‐binding protein (FKBP), which is tightly associated to the ryanodine receptor (RyR)/Ca2+ release channel, plays a role in the termination of Ca(2+)‐induced Ca2+ release (CICR) in heart. 2. Confocal microscopy and the Ca2+ indicator fluo‐3 were used to visualize the elementary release events, i.e. ‘Ca2+ sparks’ in rat ventricular myocytes under resting or voltage‐clamped conditions. Additionally, electrophysiological single‐channel recordings, at constant [Ca2+] or during [Ca2+] steps produced by photorelease of caged Ca2+, were obtained from rat cardiac RyRs incorporated in planar lipid bilayers. 3. Inhibition of FKBP by the immunosuppressants FK506 or rapamycin increased the duration of spontaneous or depolarization‐evoked Ca2+ sparks 6‐ to 7‐fold. In addition, Ca2+ sparks were seen with two‐level amplitudes, corresponding to full and half normal spark amplitude. 4. FK506 potentiated and prolonged electrically stimulated [Ca2+]i transients and contractions, but did not affect the amplitude and kinetics of the L‐type Ca2+ channel current. 5. In planar lipid bilayers, FK506 (15 microM) prolonged approximately 7‐fold the mean open lifetime of reconstituted single RyRs, induced the appearance of long‐lasting subconductance states, and markedly slowed the spontaneous decay of RyR activity elicited by fast and sustained Ca2+ stimuli. The time constant of the spontaneous decay of activity increased from 1.8 s in control to > or = 20 s in the presence of FK506. 6. We conclude that FKBP may afford an intrinsic mechanism to terminate RyR openings and it may thus exert a negative feedback on CICR in heart cells.
Assessment of Weld Residual Stress Measurement Precision: Mock-Up Design and Results for the Contour Method
Authors: Olson Mitchell D. , Hill Michael R. , Willis Eric , Peterson Artie G. , Patel Vipul I. , Muránsky Ondrej
Published: December 2014

Recent experimental work has shown residual stress measurements in welded material to be difficult. To better assess the precision of residual stress measurement techniques, a measurement article was designed to allow repeated measurements of a nominally identical stress field. The measurement article is a long 316L stainless steel plate containing a machine-controlled eight-pass slot weld. Measurements of weld direction residual stress made with the contour method found high tensile stress in the weld and heat-affected zone, with a maximum near 450 MPa and compressive stress away from the weld, a typical residual stress profile for constrained welds. The repeatability standard deviation of repeated contour method residual stress measurements was found to be less than 20 MPa at most spatial locations away from the boundaries of the plate. The repeatability data in the weld are consistent with those from a previous repeatability experiment using the contour method in quenched aluminum bars. A finite-element simulation and neutron diffraction measurements were performed for the same weld and provided results consistent with the contour method measurements. Much of the material used in the work remains available for use in assessing other residual stress measurement techniques, or for an interlaboratory reproducibility study of the contour method.
The Impact of International Football Matches on Primary Sponsors and Shareholder Wealth
Authors: Bouchet Adrien , Doellman Thomas W. , Troilo Michael , Walkup Brian R.
Published: December 2014

The effect of sponsorship on the stock market returns of the sponsoring companies has been previously studied, but the internationalizing aspect of sponsorship has been overlooked. We examine returns to shareholders for firms sponsoring international football matches using an event study analysis. We find that there are cumulative abnormal returns to stockholders of sponsoring firms of international matches 10 days after the match and 20 days after the match. This finding is robust across several different event-study methods. We also find this general pattern across different professional football leagues, as well as a positive effect on returns by sponsoring high-profile football clubs. We theorize that the elapsed time until the effect on the stock price is the result of building brand awareness before a shift in the price becomes evident. These findings add nuance to the literature on sponsorship and event studies, which is almost exclusively domestic in character.
The imprint of atmospheric evolution in the D/H of Hesperian clay minerals on Mars
Authors: Mahaffy P. R. , Webster C. R. , Stern J. C. , Brunner A. E. , Atreya S. K. , Conrad P. G. , Domagal-Goldman S. , Eigenbrode J. L. , Flesch G. J. , Christensen L. E. , Franz H. B. , Freissinet C. , Glavin D. P. , Grotzinger J. P. , Jones J. H. , Leshin L. A. , Malespin C. , McAdam A. C. , Ming D. W. , Navarro-Gonzalez R. , Niles P. B. , Owen T. , Pavlov A. A. , Steele A. , Trainer M. G. , Williford K. H. , Wray J. J. ,   the MSL Science Team
Published: December 2014

Of water and methane on Mars

The Curiosity rover has been collecting data for the past 2 years, since its delivery to Mars (see the Perspective by Zahnle). Many studies now suggest that many millions of years ago, Mars was warmer and wetter than it is today. But those conditions required an atmosphere that seems to have vanished. Using the Curiosity rover, Mahaffy
et al.
measured the ratio of deuterium to hydrogen in clays that were formed 3.0 to 3.7 billion years ago. Hydrogen escapes more readily than deuterium, so this ratio offers a snapshot measure of the ancient atmosphere that can help constrain when and how it disappeared. Most methane on Earth has a biological origin, so planetary scientists have keenly pursued its detection in the martian atmosphere as well. Now, Webster
et al.
have precisely confirmed the presence of methane in the martian atmosphere with the instruments aboard the Curiosity rover at Gale crater.

Science
, this issue p.
412
, p.
415
; see also p.
370
Sarcoma botryoides in an infant
Authors: van Sambeeck S J , Mavinkurve-Groothuis AMC , Flucke U , Dors N
Published: December 2014

A 17-month-old girl with no medical history presented at our emergency room with abnormal vaginal bleeding and vaginal tissue loss with a “grape bunch” appearance. Physical examination showed no abnormalities, but gynaecological examination showed abnormal vaginal tissue protruding through the vagina introitus. Given the typical clinical presentation, the age of the girl and the location and aspect of the lesion, there was a high suspicion of the botryoid variant of embryonal rhabdomyosarcoma of the vagina. Histology of a biopsy of the lesion was consistent with embryonal rhabdomyosarcoma. As no metastases were detected, the girl received chemotherapy. This case report describes the importance of early recognition of the typical clinical symptoms of sarcoma botryoides, since a rapid diagnosis followed by treatment is necessary to prevent death.