bluuebunny/crossref_metadata_2025_split huggingface dataset:
Calcium fluxes in mouse mammary tissue in vitro: intracellular and extracellular calcium pools
Authors: Neville M. C. , Peaker M. 
Published: December 2014

1. The total Ca content of the mammary gland increased from about 2 to 12 μmole/g tissue during the transition from pregnancy to lactation in the mouse. In tissue from lactating mice at least two thirds of the total Ca exchanged with external Ca in 6 hr. There was little non‐exchangeable Ca in tissues from pregnant mice.2. At 37 °C the time courses of influx and efflux of 45Ca in lactating tissues could be analysed by assuming three exponential components with rate constants of about 0·3, 0·06 and 0·005 min−1 and containing, respectively, 1·7, 1·5 and 4·7 μmole 45Ca/g tissue at the steady state.3. The rapidly effluxing component showed the time‐ and temperature‐dependence characteristic of bulk‐phase‐limited diffusion through the extracellular space. The diffusion coefficient was about one quarter of the self‐diffusion coefficient of Ca in aqueous solution, consistent with a tortuosity factor of about 2. A portion of the Ca in this component was displaced by La3+. The amount remaining in the presence of 3 m
m‐La3+ was close to that expected for free extracellular Ca. The rapid component was therefore interpreted as originating from an extracellular compartment containing both free and bound Ca.4. The rate of efflux of the intermediate component was slowed by a factor of ten when the temperature was decreased from 37 to 0 °C giving a Q10 of 2·7, expected for membrane transport. The slow component present at 37 °C was not displaced by EGTA or La3+, suggesting that it is not localized extracellularly. It was not apparent in the 0 °C efflux curves.5. The biphasic time course of uptake of ionophore (A23187)‐releasable 45Ca in particulate fractions obtained by homogenization and centrifugation of tissues which had been incubated with the isotope was consistent with the hypothesis that the two slower components of 45Ca flux originate from intracellular compartments. Mitochondrial uptake probably did not contribute significantly to Ca exchange in these tissues.6. 45Calcium fluxes in mammary tissues from pregnant mice also showed three components with rate constants similar to those found in tissues from lactating mice. The amount of Ca in each component was much smaller than in lactating tissue when compared on the basis of tissue weight.7. We conclude from these studies that: (i) intra‐ and extracellular Ca pools in mammary tissue can be distinguished on the basis of the temperature dependence of their fluxes and (ii) the transition from pregnancy to lactation is accompanied by large increases in both intra‐ and extracellular Ca pools in mammary alveolar cells.
Nucleoside translocation in sheep reticulocytes and fetal erythrocytes: a proposed model for the nucleoside transporter
Authors: Jarvis S. M. , Young J. D. 
Published: December 2014

1. Nucleoside transport by fetal erythrocytes from nucleoside‐permeable and nucleoside‐impermeable type new‐born lambs and by reticulocytes from adult sheep was compared with that of mature erythrocytes from adult sheep of the two phenotypes.2. Fetal cells and reticulocytes transported [U‐14C]uridine rapidly, with little difference between cells from the two types of sheep. Transport occurred by a saturable uptake mechanism with similar properties to that present in mature cells from adult nucleoside‐permeable type animals, except for an approximately 100‐fold higher Vmax.3. This increased translocation capacity was associated with increased numbers of high‐affinity [3H]nitrobenzylthioinosine binding sites (∼ 2000‐3000 sites/cell compared with approximately 20 sites/cell for mature nucleoside‐permeable sheep erythrocytes).4. The calculated transport capacity for each nucleoside translocation site is therefore similar in all cell types (140‐180 molecules/site. s at 25 °C, assuming that each transport site binds a single molecule of inhibitor). These values compare favourably with turnover estimates for the nucleoside transporter from human and pig erythrocytes.5. Loss of nucleoside transport activity after birth closely paralleled loss of [3H]nitrobenzylthioinosine binding sites and the progressive loss of fetal cells from the circulation. Similarly, reticulocyte maturation in vitro was also associated with rapid loss of both nucleoside transport capacity and inhibitor binding activity.6. p‐Chloromercuriphenylsulphonate and trypsin had no effect on [3H]nitrobenzylthioinosine binding to intact fetal cells. In contrast, both agents markedly inhibited binding to isolated ‘ghosts’ where both sides of the cell membrane were accessible to reagent. p‐Chloromercuriphenylsulphonate inhibition was markedly reduced in the presence of uridine, and reversed by addition of dithiothreitol.7. We conclude that nucleoside transport changes during ontogeny and reticulocyte maturation in the sheep as well as species differences in nucleoside transport capacity are regulated by variations in the numbers of functional transport sites per cell rather than by changes in the activity of a constant number of sites. It is also likely that the nucleoside carrier exhibits chemical asymmetry.8. A simple molecular model of the erythrocyte nucleoside transporter consistent with these and other known properties of the carrier is proposed.
Climbing fibre induced depression of both mossy fibre responsiveness and glutamate sensitivity of cerebellar Purkinje cells
Authors: Ito Masao , Sakurai Masaki , Tongroach Pavich 
Published: December 2014

1. In high decerebrate rabbits, cells were sampled extracellularly from the rostral flocculus. Purkinje cells were identified by their characteristic responses to stimulation of the contralateral inferior olive. Identification of basket cells was based on the absence of olivary responses and also on their location in the molecular layer adjacent to identified Purkinje cells. Mass field potentials in the flocculus were also studied.2. Single pulse stimulation of a vestibular nerve, either ipsilateral or contralateral, at a rate of 2/sec excited Purkinje cells with a latency of 3‐6 msec. This early excitation represents activation through vestibular mossy fibres, granule cells and their axons (parallel fibres). Similar early excitation also occurred in putative basket cells.3. Conjunctive stimulation of a vestibular nerve at 20/sec and the inferior olive at 4/sec, for 25 sec per trial, effectively depressed the early excitation of Purkinje cells by that nerve, without an associated change in spontaneous discharge. The depression recovered in about ten minutes. This recovery was followed by the onset of a slow depression lasting for an hour.4. Conjunctive vestibular—olivary stimulation produced no such depression in the following responses: early excitation in Purkinje cells induced from the vestibular nerve not involved in the conjunctive stimulation; early excitation in putative basket cells from either vestibular nerve; inhibition or rebound facilitation in Purkinje cells following the early excitation; vestibular‐evoked field potentials in the granular layer and white matter of the flocculus. These observations lead to the conclusion that the depression occurs specifically at parallel fibre—Purkinje cell synapses involved in conjunctive stimulation.5. Ionophoretic application of glutamate to Purkinje cells in conjunction with 4/sec olivary stimulation depressed the glutamate sensitivity of Purkinje cells; aspartate sensitivity was depressed to a much lesser degree. The depression diminished in about 10 min, but this recovery was succeeded by a slow depression lasting for an hour. The depression was seen only when glutamate sensitivity was relatively high, suggesting that the micro‐electrode was impinging onto Purkinje cell dendrites. These observations suggest that subsynaptic chemosensitivity of Purkinje cells to the putative neurotransmitter of parallel fibres is involved in the depression observed after conjunctive stimulation of a vestibular nerve and the inferior olive.6. The present results are consistent with the Marr‐Albus assumption concerning plasticity of cerebellar neuronal networks.
Long‐term synaptic enhancement and short‐term potentiation in rat fascia dentata act through different mechanisms
Authors: McNaughton Bruce L. 
Published: December 2014

1. The component processes contributing to post‐activation change in synaptic efficacy in the perforant pathway to the fascia dentata were studied in rats under sodium pentobarbitone anaesthesia.2. With low stimulus strength, which activated only a relatively small number of perforant path fibres, repetitive stimulation led to effects which had very similar characteristics to those observed at neuromuscular synapses under similar conditions. Paired shocks resulted in a short (∼ 100 ms) facilitation superimposed on a depression, possibly due to depletion of available transmitter, which recovered more slowly (∼ 4 s). Short trains of stimuli at 125‐250 Hz led to a longer lasting increase in synaptic strength which decayed to control levels with a double exponential time course. The two exponential components behaved like augmentation and potentiation at neuromuscular synapses, with time constants at 33 °C of about 5 s and about 90 s respectively.3. High‐intensity stimulus trains of identical frequency and duration led to an enhancement of synaptic strength which lasted for longer than 30 min.4. The paired shock depletion effect was increased in direct proportion to the amount of augmentation and potentiation present following low‐intensity stimulus trains. Following high‐intensity trains the paired shock depletion effect was increased by the same amount, and recovered with the same time course as following low‐intensity stimulus trains, even though there remained a significant enhancement of the synaptic response.5. The results are interpreted as indicating that augmentation and potentiation are due to an increase in the probability of transmitter release whereas long‐term enhancement acts through some other, as yet undetermined, mechanism. Following high‐intensity stimulation all three processes are activated.
Effects of extracellular potassium accumulation and sodium pump activation on automatic canine Purkinje fibres
Authors: Kline Richard P. , Kupersmith Joel 
Published: December 2014

1. Double barrel potassium‐sensitive micro‐electrodes were used to measure fluctuations in extracellular potassium ion concentration in large automatic canine Purkinje fibres.2. Slow accumulations of potassium were seen in the extracellular space during prolonged beating. Following cessation of prolonged beating, a depletion of extracellular potassium ions was noted.3. The time course of these slow changes in extracellular potassium concentration were shown to be a function of the diffusion properties of the preparation, and the rate and degree of activation of the sodium pump.4. Action potential duration and maximum diastolic potential were simultaneously monitored during and after these periods of rapid stimulation, using conventional intracellular micro‐electrodes.5. Initial depolarization and later hyperpolarization of the maximum diastolic potential appeared to occur as a direct result of changes in extracellular potassium concentration and level of pump activation induced by the sudden and prolonged alteration in stimulus rate.6. Following prolonged stimulation, dramatic changes in the automatic beating rate were correlated with changes in extracellular potassium and pump rate, and their effects on the maximum diastolic potential and other parameters of the diastolic potential depolarization.7. For some locations of the potassium‐sensitive electrode tip, large fluctuations in extracellular potassium were seen during single beats. Alterations in action potential duration during abrupt rate changes appear to derive in part from modulation by these extracellular potassium concentration fluctuations altering net membrane currents. Slower shifts in action potential duration appear correlated to the degree of sodium pump activation.
Mechanisms of transmembrane calcium movement in cultured chick embryo ventricular cells
Authors: Barry William H. , Smith Thomas W. 
Published: December 2014

1. Uptake of calcium was studied in spontaneously contracting monolayers of cultured chick embryo ventricular cells. Ca exchange could be separated into two components: a rapid phase with a rate constant of 3·91/min, accounting for 1·6 nmol/mg protein; and a slower phase with a rate constant of 0·069/min, accounting for 2·7 nmol/mg protein.2. Negatively inotropic concentrations of the slow Ca channel blocker verapamil inhibited the rapid phase of Ca uptake partially, with a maximum inhibition of 30‐40% observed at concentrations of verapamil which completely inhibited contraction.3. The component of Ca uptake not inhibited by verapamil could be stimulated up to 25‐fold by elevation of intracellular Na concentration and reduction of extracellular Na concentration, and thus appeared to represent at least in part Ca uptake via Na—Ca exchange.4. Ca uptake by cultured cells could be almost completely inhibited by exposure to LaCl3, 1 mmol/l, within 5 s. This same concentration of La completely inhibited contraction within 5 s.5. During efflux of 45Ca from cells, exposure to La (1 mmol/l) slightly inhibited efflux initially with more marked inhibition of Ca influx after 3 min of La exposure. There was no evidence for a component of superficial La‐displaceable Ca, and thus the rapid phase of Ca uptake probably is due to an intracellular rapidly exchanging Ca pool. Efflux of Ca from this rapidly exchanging intracellular Ca pool was not significantly altered by exposure to Na‐free choline chloride solutions.6. We conclude that rapid Ca uptake in these cultured myocardial cells is due primarily to Ca influx via the slow Ca channel and via Na—Ca exchange. In the presence of physiological [Ca2+]i, efflux of Ca from this intracellular Ca pool does not appear to be due to Na—Ca exchange.
Evidence for the role of non‐quantal acetylcholine in the maintenance of the membrane potential of rat skeletal muscle
Authors: Bray J. J. , Forrest J. W. , Hubbard J. I. 
Published: December 2014

1. Resting membrane potentials of rat diaphragm muscles cultured in Trowell T8 medium were measured in vitro. After 3 hr in culture the resting membrane potential of muscle fibres within 2·5 mm of nerve section (‘near’) was ‐68·3 ± 0·4 mV (nineteen preparations). This was significantly lower (P < 0·001) than the resting potential (‐74·0 ± 0·4 mV) measured in muscle fibres 8‐10 mm from the site of nerve section (‘far’) in the same preparations. A difference between the ‘near’ and the ‘far’ fibres was maintained in muscles cultured for 6 and 12 hr. Miniature end‐plate potentials were present in both ‘near’ and ‘far’ fibres cultured for 3 and 6 hr and ceased after 12‐15 hr.2. The presence of carbamylcholine (10−7 or 10−8 M) maintained the resting membrane potential of ‘near’ fibres close to that of ‘far’ fibres at 3, 6 and 12 hr. For example, at 3 hr in the presence of 10−8 M‐carbamylcholine the mean resting potential was 75·6 ± 0·5 mV in ‘near’ fibres and 76·1 ± 0·4 mV in ‘far’ fibres (four preparations). A similar effect was produced in preparations exposed to anticholinesterases: diisopropylphosphorofluoridate (DFP) (10−7 M), neostigmine (10−7 M) or physostigmine (10−5 M).3. Agents that blocked acetylcholine receptors had the reverse effect. In the presence of α‐bungarotoxin (1 μg/ml.) or d‐tubocurarine (10−5 M) the resting membrane potential of ‘far’ fibres was reduced to the level of ‘near’ fibres over the 24 hr period of observation. For example, at 3 hr in the presence of α‐bungarotoxin the mean resting potential was 67·2 ± 0·5 mV in ‘near’ fibres and 68·5 ± 0·6 mV in ‘far’ fibres (six preparations). The effect of d‐tubocurarine was reversible.4. When muscles were cultured in Ca2+‐free medium containing 1 mM‐EGTA and 10 mM‐Mg2+, there was no difference in membrane potential between ‘near’ and ‘far’ fibres and physostigmine (10−5 M) was ineffective in raising the membrane potential of ‘near’ fibres.5. It is suggested that non‐quantal acetylcholine released from nerve terminals maintains the membrane potential of muscle fibres through a Ca2+‐dependent mechanism.
Properties of miniature excitatory junctional currents at the locust nerve‐muscle junction
Authors: Cull‐Candy S. G. , Miledi R. 
Published: December 2014

1. Miniature excitatory junctional currents (m.e.j.c.s) were examined in conditions where inward current was carried mainly by Na+ (i.e. in normal medium, Ca2+‐free medium and Cl−‐free medium). M.e.j.c.s were also examined in isotonic Ca2+ where the inward post‐synaptic current was carried mainly by Ca2+.2. In normal medium, mean m.e.j.c. amplitude = 2.34±0.05 nA. The decay time constant of m.e.j.c.s (excluding a small percentage with abnormal shapes) was τm.e.j.c. = 2.62±0.11 msec (Vm = ‐80 mV, T = 22 °C). Decay‐time was not markedly changed in Ca2+‐free or Cl−‐free medium. τm.e.j.c. approaches the life‐time of glutamate activated junctional channels.3. Excitatory junctional currents, evoked by nerve impulses, decayed slightly faster than m.e.j.c.s obtained in the same fibres. Extracellularly recorded m.e.j.c.s and voltage‐clamped m.e.j.c.s were similar in time course.4. τm.e.j.c. decreased exponentially with membrane hyperpolarization. An e‐fold change was produced by 182.±24.8 mV change in Vm.5. The dependence of mean m.e.j.c. amplitude on clamp potential showed a slight non‐linearity at hyperpolarized levels. The equilibrium potential for transmitter action was close to 0 mV in normal solution as well as in Ca2+‐free and Cl−‐free solutions.6. The kinetics of junctional channels are altered in isotonic Ca2+. M.e.j.c. amplitude was reduced to about one‐third normal size; mean m.e.j.c. = 0.74±0.03 nA. The decay time becomes markedly briefer, τm.e.j.c. = 1.01±0.08 msec, indicating a reduction in mean channel life‐time (Vm = ‐80 mV, T = 22 °C).7. A population of slow time course and composite m.e.j.c.s appear when muscle fibres are hyperpolarized in isotonic Ca2+, thus producing a prolongation in mean τm.e.j.c.. This results from an influence of post‐synaptic membrane potential on presynaptic transmitter release. If such m.e.j.c.s are ignored the voltage dependence of τm.e.j.c. of the remaining events is abolished or even reversed indicating that voltage sensitivity of channel life‐time is altered in isotonic Ca2+. The equilibrium potential for transmitter action may be slightly more positive than normal.8. We estimate that a single packet of neurally released transmitter normally opens, on average, 250 ion channels at these junctions.
Induction and removal of inward‐going rectification in sheep cardiac Purkinje fibres
Authors: Carmeliet E. 
Published: December 2014

1. In sheep cardiac Purkinje fibres superfused with K‐free, Na‐free medium, the membrane potential can be stable either at a low negative level (‐50 mV) or at a high negative level (‐100 mV). The mechanism underlying the existence of these two stable potential levels was investigated using the two‐micro‐electrode voltage‐clamp technique.2. By applying a voltage clamp of a certain duration at an appropriate level the membrane potential could be shifted from one stable level to the other. The shift was observed in Cl‐free medium, excluding a redistribution of Cl as a possible explanation.3. Currents during and following a voltage step and their change with amplitude and duration of the voltage step could not be explained on the basis of depletion or accumulation of K ions in the narrow extracellular clefts.4. Instantaneous currents determined from the high negative resting level showed a high conductance and a pronounced inward rectification, while measurements from the low negative resting level indicated a low conductance and absence of inward rectification. The steady‐state current—voltage relation was dependent on the holding potential and showed memory or hysteresis.5. Estimation of the conductance by superimposed short voltage‐clamp pulses showed an increase in conductance during a hyperpolarizing clamp from the low negative level and a decrease in conductance during a depolarizing clamp from the high negative level. The time‐dependent current during a hyperpolarizing clamp from the low negative level reversed direction at a potential level corresponding to EK, assuming a cleft K concentration of about 1 mM. In the presence of 0·1 mM‐Ba the time‐dependent current was abolished.6. The results suggest that the shift between the two stable levels is due to a time‐dependent conductance change in the K inward rectifier channel, iK1. The existence of memory excludes activation or de‐activation only depending on the voltage gradient. Interaction of extracellular K ions with a site in the membrane is proposed as the activating mechanism.
Calcium channel and calcium pump involved in oscillatory hyperpolarizing responses of L‐strain mouse fibroblasts
Authors: Okada Yasunobu , Tsuchiya Wakoh , Yada Toshihiko 
Published: December 2014

1. In fibroblastic L cells, spontaneously repeated hyperpolarizing responses (oscillation of membrane potential) and hyperpolarizing responses evoked by electrical stimuli were suppressed by the external application of a K+ channel blocker, nonyltriethylammonium (C9). This hydrophobic TEA‐analogue also inhibited the hyperpolarization induced by intracellular Ca2+ injection.2. Quinine or quinidine, known inhibitors of the Ca2+‐activated K+ channel of red cells, instantaneously inhibited these hyperpolarizations. Thus, these hyperpolarizations are likely to be caused by the operation of Ca2+‐sensitive K+ channels.3. Azide, which is known to inhibit the mitochondrial Ca2+ uptake in fibroblasts, and caffeine, dantrolene Na and oxalate, which affect the microsomal Ca2+ transport, did not exert any effects upon the electrical potential profiles.4. On the other hand, Ca2+ channel blockers (nifedipine, D 600 and Co2+) suppressed the hyperpolarizing responses, but not the hyperpolarizations produced by intracellular Ca2+ injection, suggesting that the calcium ions responsible for the hyperpolarizing responses are mainly derived from outside the cell through Ca2+ channels.5. Flavones of plant origin, which are known to inhibit Ca2+‐ATPase, prolonged the duration of the hyperpolarizing phase of the oscillation or produced a sustained hyperpolarization.6. It is concluded that the Ca2+ channel and the Ca2+ pump play essential roles in the generation of the hyperpolarizing response and of the membrane potential oscillation in L cells, and that these hyperpolarizations are brought about by a transient elevation of cytosolic Ca2+ level which, in turn, activates Ca2+‐dependent K+ channels.