bluuebunny/crossref_metadata_2025 huggingface dataset:
Modulation of slow gating process of calcium channels by isoprenaline in guinea‐pig ventricular cells.
Authors: Ochi R , Kawashima Y
Published: December 2014

1. The mechanism of enhancement of Ca2+ current by isoprenaline was studied by recording single‐channel activity from cell‐attached patches on isolated guinea‐pig ventricular cells using patch pipettes containing 50 or 100 mM‐Ba2+. 2. Isoprenaline (100 nM) increased the amplitude of ensemble average currents by increasing the rate of non‐blank sweeps (availability). The current decay during 400 ms steps was significantly slowed by isoprenaline. However, the open probability for the non‐blank sweeps elicited by 100 ms steps was only slightly increased by the application of isoprenaline. 3. The durations of the available state (TS) and the unavailable state (TF) were estimated by the number of non‐blank and blank sweeps per run, respectively, applying repetitively 100 ms steps at 2 Hz. 4. At large negative holding potentials the distribution of TS was well fitted by an exponential curve, whose time constant was increased from 1.6 to 3.1 sweeps by 100 nM‐isoprenaline, while TF distributed approximately single exponentially with a time constant of 2.0 sweeps in control and 1.3 sweeps in the presence of the drug. 5. At depolarized holding potentials a slow voltage‐dependent component appeared in the histogram of TF and its time constant was markedly decreased by 100 nM‐isoprenaline. 6. The availability‐voltage relationship was simulated by the Boltzmann equation with a maximal value of 0.4 in the control. The maximal value was increased to 0.7 and the curve was shifted to a depolarizing direction by 7 mV by 100 nM‐isoprenaline. 7. Isoprenaline increased the availability of cardiac Ca2+ channels by increasing the forward rate constant and decreasing the backward rate constant in both voltage‐dependent and independent slow state transitions.
Comparison of intracellular pH transients in single ventricular myocytes and isolated ventricular muscle of guinea‐pig.
Authors: Bountra C , Powell T , Vaughan‐Jones R D
Published: December 2014

1. Intracellular pH was recorded (double‐barrelled pH‐selective microelectrodes) in single ventricular myocytes and whole papillary muscles isolated from guinea‐pig heart. Both preparations were acid‐loaded by various manoeuvres (addition and removal of external NH4Cl or CO2) in order that a comparison could be made of the size and speed of intracellular pH changes and hence of the apparent intracellular buffering power (beta). 2. For the same acid‐loading procedure, the size of intracellular pH (pHi) changes was about threefold larger in the isolated myocyte than in whole papillary muscle. The rate of initial acid loading as well as the subsequent rate of pHi recovery (caused by acid extrusion from the cell) were also threefold faster in the myocyte. Estimates of apparent intrinsic (non‐CO2) buffering power, based upon the size of pHi changes during acid loading, were 15‐20 mmol l‐1 for the myocyte and about 70 mmol l‐1 for whole muscle. This latter value is similar to previous estimates of beta in heart. 3. When acid extrusion was reduced by applying a high dose of amiloride (1 mmol l‐1), then the size of the pHi change during acid loading increased greatly in papillary muscle but changed much less in the myocyte; beta now appeared to be about 30 mmol l‐1 in whole muscle but remained essentially unchanged in the myocyte. 4. We conclude that previous values for beta in cardiac muscle have been greatly overestimated because of the presence of sarcolemmal acid extrusion. Paradoxically, this error in estimating beta is far less evident in the isolated myocyte. We suggest that this is because a much more rapid acid loading is achievable in the myocyte so that acid loading will be blunted less by acid extrusion than in whole muscle. We present a simple mathematical model that demonstrates this phenomenon. We conclude that beta in ventricular muscle is likely to resemble that measured in the isolated myocyte, i.e. 15‐20 mmol l‐1. 5. Slow acid loading in whole ventricular muscle will also affect the kinetics of pHi changes. The model indicates that the rate of pHi recovery from an acid load in papillary muscle does not reflect the pHi dependence of acid extrusion. Instead, it is heavily influenced by the slow rate of acid loading. This emphasises that great care should be taken when interpreting the kinetics of pHi changes in multicellular ventricular preparations.
Effects of metabolic inhibition on the membrane properties of isolated mouse primary sensory neurones.
Authors: Duchen M R
Published: December 2014

1. The patch‐clamp technique has been used to investigate the mechanisms that couple membrane excitability to metabolism in neurones isolated from mouse dorsal root ganglia. 2. Blockade of electron transport by cyanide (CN‐), reduction of the mitochondrial membrane potential with carbonyl cyanide p‐trifluoromethoxyphenyl hydrazone (FCCP), removal of glucose or inhibition of glycolysis with idoacetic acid (IAA), all increased a K+ conductance (gK), which could be sufficient to shunt action potentials. 3. The K+ conductance was reduced by incubation of cells in Ca2(+)‐free solutions or by increasing the Ca2+ buffering power of pipette‐filling solutions. The Ca2+ ionophore, ionomycin, also increased a K+ conductance, and current fluctuation analysis showed that the channels carrying the current induced by both ionomycin and by CN‐ had a similar mean conductance of circa 9 pS. Thus, increased gK was a Ca2(+)‐dependent K+ conductance, gK(Ca), reflecting a rise in resting [Ca2+]i. 4. The conductance was not affected by inclusion of ATP or an ATP‐regenerating system in the pipette, suggesting that the underlying rise in [Ca2+] is not due directly to loss of ATP, and confirming that the increased gK is not carried through ATP‐dependent K+ channels. 5. Voltage‐gated K+ currents evoked by membrane depolarization were increased by CN‐ or glucose removal. The current‐voltage relation of the increased gK mirrored the voltage dependence of Ca2+ entry, and thus reflects impaired cellular handling of the Ca2+ load imposed by depolarization. 6. The rise in [Ca2+]i and altered Ca2+ buffering capacity induced by metabolic blockade affected several other conductances: (i) a Ca2(+)‐dependent chloride current was increased. (ii) Both the low‐threshold transient and high‐threshold sustained voltage‐gated Ca2+ currents were attenuated and their thresholds were shifted in the hyperpolarizing direction. (iii) The inward current activated by hyperpolarization. IH, seen in large cells, was attenuated by either metabolic blockade or ionomycin. 7. The responses of these neurones to impaired metabolism thus depend largely on the effects of raised [Ca2+]i on the populations of channels expressed by the cells. These changes in membrane properties could account for some of the changes in neuronal behaviour seen during the clinical states of hypoxia or hypoglycaemia, underlying changes in central nervous system function.
Effects of metabolic blockade on the regulation of intracellular calcium in dissociated mouse sensory neurones.
Authors: Duchen M R , Valdeolmillos M , O'Neill S C , Eisner D A
Published: December 2014

1. Impaired intracellular Ca2+ concentration ([Ca2+]i) regulation may underlie alterations in neuronal function during hypoxia or hypoglycaemia and may initiate cell damage. We have used the Ca2(+)‐sensitive fluorophore, Fura‐2, to study the regulation of [Ca2+]i in neurones isolated from mouse dorsal root ganglia. Mean resting [Ca2+]i was 163 +/‐ 11 nM (mean +/‐ S.E.M., n = 38). 2. Depolarization by exposure to 20 or 30 mM‐K+ caused a rapid Co2(+)‐ and Cd2(+)‐sensitive rise in [Ca2+]i, which subsequently declined with a time course usually fitted by the sum of two exponential functions. 3. Interference with mitochondrial function (by CN‐ or FCPP) or with glycolysis (by glucose removal) all raised [Ca2+]i by up to 220%. Addition of FCCP in the presence of CN‐ further increased [Ca2+]i. The response to CN‐ was still seen in the absence of extracellular Ca2+, although it attenuated rapidly, indicating release from an intracellular store. 4. Either CN‐ or glucose removal increased the rise in [Ca2+]i induced by K+ 2‐ to 3‐fold and slowed recovery, suggesting interference with sequestration or extrusion of [Ca2+]i. 5. Resting [Ca2+]i rose when external Na+ was replaced by Li+ or N‐methyl‐D‐glucamine, demonstrating the presence of a Na(+)‐Ca2+ exchange process. However, Na+ replacement had only a slight effect on the handling of a Ca2+ load. 6. We conclude that (i) Ca2+ is released into the cytoplasm from intracellular organelles when energy supplies are reduced: (ii) that the extrusion or sequestration of Ca2+ entering the cell during electrical activity is rapidly impaired by interference with mitochondrial metabolism: and (iii) Na(+)‐Ca2+ exchange makes only a small contribution to intracellular Ca2+ homeostasis. 7. [Ca2+]i would thus be expected to rise in vivo during hypoxia or hypoglycaemia and may initiate alterations in neuronal function. However, if a rise in Ca2+ is an important cause of cell damage in cerebral hypoxaemia, the combination of excitation and hypoxia will lead to the largest increases in [Ca2+]i, while hypoxia alone appears to cause only a small increase in [Ca2+]i in quiescent cells.
Modes of action of local hypothalamic and skin thermal stimulation on salivary secretion in rats.
Authors: Kanosue K , Nakayama T , Tanaka H , Yanase M , Yasuda H
Published: December 2014

1. In urethane or ketamine‐anaesthetized rats, salivary secretion was observed when local brain sites or trunk skin were stimulated thermally or electrically. 2. Salivary secretion was facilitated by bilateral local brain warming. Sensitive sites were restricted to the preoptic area and anterior hypothalamus, but in a region distinct from a previously reported sensitive site for producing saliva‐spreading behaviour. 3. Unilateral warming of the preoptic area produced greater salivary secretion from the ipsilateral submandibular/sublingual salivary glands than from the contralateral glands. Electrical stimulation of the same sites elicited salivation only from the ipsilateral glands. 4. Trunk skin, not including the scrotum, was unilaterally cooled when spontaneous salivary secretion was observed in a hot environment. Salivary secretion from both sides was equally suppressed in response to the unilateral skin cooling. 5. We conclude that efferent signals from the anterior part of the hypothalamus project dominantly to the ipsilateral salivary gland for thermally induced salivary secretion. Thermal signals from the skin of either side of the trunk, on the other hand, appear to be integrated and to affect salivary secretion bilaterally.
The role of thyroid hormones in maturation of the adrenaline‐sensitive lung liquid reabsorptive mechanism in fetal sheep.
Authors: Barker P M , Strang L B , Walters D V
Published: December 2014

1. Following thyroidectomy at 106‐118 days fetal sheep were infused continuously with triiodothyronine (T3) from 110, 118, 125 or 131 days (n = 12) or with thyroxine (T4) from 118 days (n = 4) until the fetuses were delivered. Lung liquid secretion or absorption rates, heart rate, blood pressure and arterial blood gases were measured before and during 45 min periods of fetal infusions of adrenaline (n = 60) at 3‐8 day intervals. The effects of T3 or T4 replacement on the response to adrenaline were compared with data previously obtained in groups of euthyroid (control) and thyroidectomized (Tx) fetuses. 2. Fetuses infused with T4 (50 micrograms/day) following thyroidectomy had plasma T4 and T3 concentrations in the normal fetal range. Fetal plasma T3 levels in fetuses infused with T3 (60 micrograms/day) were at or above the high end of the normal range for full‐term fetuses. Those receiving 120 micrograms of T3 per day had levels equivalent to those normally seen in the postnatal T3 surge. 3. Normal maturation in the lung of the reabsorptive response of fetal lung liquid to adrenaline was seen in the fetuses infused with T3 or T4 from 118 days. A marginal advance in maturation was seen in fetuses infused with T3 from 110 days and a delay in maturation in those infused with T3 from 125 and 131 days.
Locomotion‐related variations in excitability of spino‐olivocerebellar paths to cat cerebellar cortical c2 zone.
Authors: Apps R , Lidierth M , Armstrong D M
Published: December 2014

1. Cutaneous nerve stimulation was used to study the excitability of the spino‐olivocerebellar pathways (SOCPs) to the c2 zone of the paravermal cerebellar cortex in the cat. Non‐noxious single‐shock stimulation of the right and left superficial radial (SR) nerves via implanted cuff electrodes was used to evoke field potentials in the cerebellar cortex via the SOCPs. 2. The evoked potentials were recorded extracellularly either in lobule V of the anterior lobe (three cats) or within the paramedian lobule of the posterior lobe (one cat) with glass‐coated tungsten microelectrodes. Measurement of the amplitudes of the responses was used to monitor transmission in the SOCPs in cats at rest and during walking. 3. A total of eleven c2 recording sites were investigated in detail. At seven of these sites, responses were recorded both during locomotion and at rest. For all seven sites responses during locomotion were smaller, more variable in amplitude and less securely evoked (average reduction 59%). 4. At five out of the eleven recording sites (45%) the mean amplitude of responses elicited during different tenths of the step cycle fluctuated sufficiently that the largest response was more than twice the smallest. In the majority of these cases (4/5) the responses were largest in either mid‐stance or late swing. These fluctuations in response size occurred without parallel fluctuation in the amplitude of the peripheral nerve volley. At the remaining sites fluctuation of the cerebellar field size was less and in some cases practically absent. 5. At six recording sites it was possible to record the climbing fibre potentials evoked by stimulation of both the ipsilateral and contralateral superficial radial nerves. In all six cases the fluctuations in size of the response during locomotion occurred in phase, despite the fact that the two limbs move out of phase. 6. The probability that an individual stimulus would evoke any cerebellar response also varied between the different tenths of the step cycle and such variations occurred in parallel with the fluctuations in response size. This shows that the SOCP regulatory mechanism(s) must, at least in part, operate at a precerebellar level.
Two types of voltage‐dependent calcium current in rat somatotrophs are reduced by somatostatin.
Authors: Chen C , Zhang J , Vincent J D , Israel J M
Published: December 2014

1. Somatotrophs were obtained from rat pituitary glands after dissociation, separation and enrichment on a continuous gradient of bovine serum albumin at unit gravity. Somatotrophs were enriched up to 85% in the heavy fractions (F8 and F9). 2. After identification by reverse hemolytic plaque assay, patch‐clamp recording in the whole‐cell mode was performed on somatotrophs. 3. Under voltage‐clamp conditions, two types of Ca2+ currents were recorded. From a holding potential of ‐70 mV, depolarizing voltage steps to potentials more positive than ‐50 mV activated a current which rapidly inactivated and which was very sensitive to Ni2+ but not to Cd2+. This current corresponds to T‐type current. Depolarizing steps to potentials more positive than ‐30 mV from a holding potential of ‐40 mV triggered a current which slowly inactivated and which was very sensitive to Cd2+ but not to Ni2+. This current corresponds to L‐type current. 4. Application of somatostatin to the bath solution (10 nM) markedly reduced the amplitudes of both T‐ and L‐type currents. Somatostatin decreased the conductance of L‐type current without modifying its time‐ and voltage‐dependent inactivation but its activation was not affected. However, somatostatin decreased the conductance of T‐type currents, and also accelerated its time‐dependent inactivation. Half‐inactivation voltage of T‐type current was shifted from ‐52 to ‐63 mV by somatostatin but no change was obtained in the current activation curve. 5. All these modifications in Ca2+ currents were abolished by a pre‐treatment of the cultures with pertussis toxin (100 ng/ml, for 10 h). This pre‐treatment also blocked the inhibitory effect of somatostatin on high‐K(+)‐stimulated growth hormone release. 6. Our results show that somatostatin acts on somatotrophs by attenuating the voltage‐dependent Ca2+ currents. These effects may contribute to a somatostatin‐induced reduction in [Ca2+]i and the subsequent decline in growth hormone release.
Regulation of the intracellular free calcium concentration in single rat dorsal root ganglion neurones in vitro.
Authors: Thayer S A , Miller R J
Published: December 2014

1. Simultaneous whole‐cell patch‐clamp and Fura‐2 microfluorimetric recordings of calcium currents (ICa) and the intracellular free Ca2+ concentration ([Ca2+]i) were made from neurones grown in primary culture from the dorsal root ganglion of the rat. 2. Cells held at ‐80 mV and depolarized to 0 mV elicited a ICa that resulted in an [Ca2+]i transient which was not significantly buffered during the voltage step and lasted long after the cell had repolarized and the current ceased. The process by which the cell buffered [Ca2+]i back to basal levels could best be described with a single‐exponential equation. 3. The membrane potential versus ICa and [Ca2+]i relationship revealed that the peak of the [Ca2+]i transient evoked at a given test potential closely paralleled the magnitude of the ICa suggesting that neither voltage‐dependent nor Ca2(+)‐induced Ca2+ release from intracellular stores made a significant contribution to the [Ca2+]i transient. 4. When the cell was challenged with Ca2+ loads of different magnitude by varying the duration or potential of the test pulse, [Ca2+]i buffering was more effective for larger Ca2+ loads. The relationship between the integrated ICa and the peak of the [Ca2+]i transient reached an asymptote at large Ca2+ loads indicating that Ca2(+)‐dependent processes became more efficient or that low‐affinity processes had been recruited. 5. Inhibition of Ca2+ influx with neuropeptide Y demonstrated that inhibition of a large ICa produced minor alterations in the peak of the [Ca2+]i transient, while inhibition of smaller currents produced corresponding decreases in the [Ca2+]i transient. Thus, inhibition of the ICa was reflected by a change in the peak [Ca2+]i only when submaximal Ca2+ loads were applied to the cell, implying that modulation of [Ca2+]i is dependent on the activation state of the cells. 6. Intracellular dialysis with the mitochondrial Ca2+ uptake blocker Ruthenium Red in whole‐cell patch‐clamp experiments removed the buffering component which was responsible for the more efficient removal of [Ca2+]i observed when large Ca2+ loads were applied to the cell. 7. When cells were superfused with 50 mM‐K+, [Ca2+]i transients recorded from the cell soma returned to control levels very slowly. Pharmacological studies indicated that mitochondria were cycling Ca2+ during this sustained elevation in [Ca2+]i. In contrast, [Ca2+]i transients recorded from cell processes returned to basal levels relatively rapidly. 8. Extracellular Na(+)‐dependent Ca2+ efflux did not significantly contribute to buffering [Ca2+]i transients in dorsal root ganglion neurone cell bodies.(ABSTRACT TRUNCATED AT 400 WORDS)
Electrophysiology of single heart cells from the rabbit tricuspid valve.
Authors: Anumonwo J M , Delmar M , Jalife J
Published: December 2014

1. The electrophysiology of single myocytes isolated from the rabbit tricuspid valve was studied using the patch‐clamp method (whole‐cell configuration). Cell dispersion was achieved by collagenase treatment, using the Langendorff retrograde perfusion procedure. 2. After isolation, and while incubating in the recovery (Kraftbrühe) solution, cells had clear striations and were mostly spindle‐shaped, or rod‐like (less than 10%), with length varying from 35 microns to over 150 microns, and diameter from 3 to 10 microns. 3. Upon exposure to Tyrode solution, the calcium‐tolerant cells were mostly rounded with smooth surfaces and well‐defined borders. The mean diameter of these cells was 15 +/‐ 5 microns (S.D., n = 9). A smaller percentage (about 30%) retained the original elongated shape. 4. Patch pipette recordings showed the presence of spontaneous activity in about 30% of round cells, and less frequently in elongated cells. Maximum diastolic potentials (MDPs) in the round cells averaged ‐82 +/‐ 6 mV, with a take‐off potential of ‐56 +/‐ 3 mV (n = 9), and an average maximum upstroke velocity (Vmax) value of 6.3 +/‐ 0.6 V/s (n = 4). In quiescent cells, the mean resting potential was 69 +/‐ 12 mV (n = 43). 5. Voltage clamp ramps revealed a steady‐state I‐V relation with a negative slope region. The mean input resistance value was 25 +/‐ 9 M omega (n = 16) for the elongated, and 883 +/‐ 481 M omega (n = 8) for the round cells. 6. Hyperpolarizing 5 s pulses (holding potential = ‐50 mV) occasionally revealed a slow, time‐dependent inward current whose peak increased progressively as a function of clamp potential. The slowly activating current was sensitive to caesium 2 mM), indicating its similarity to the so‐called ‘pacemaker current’ (iF). In alternate voltage‐ and current‐clamp experiments, blocking of iF did not stop pacemaker activity, but there was up to a fourfold increase in pacemaker cycle length. 7. In some cells, 5 s hyperpolarizing steps from a holding potential of ‐40 or ‐50 mV produced large, inwardly directed and voltage‐dependent current surges that decayed rapidly with time, similar to the inactivation described for the inward rectifier current, iK1. The current was very prominent at voltages more negative than ‐100 mV, and its decay process was best fitted by two time constants, one fast and one slow. For example, at ‐150 mV the time constants were 61 and 634 ms. The inward current was blocked by barium (1 mM).(ABSTRACT TRUNCATED AT 400 WORDS)
Suppression of cold‐induced thermogenesis in full‐term pregnant rats.
Authors: Imai‐Matsumara K , Matsumura K , Morimoto A , Nakayama T
Published: December 2014

1. Thermoregulation against cold exposure was studied in rats during pregnancy and early lactation, and compared with that of virgin rats. 2. When exposed to 0 degrees C for 60 min, rats which were within 24‐48 h of parturition (pre‐1‐day rats) and those within 24 h of parturition (pre‐0‐day rats) showed significantly larger falls of colonic temperature (Tco) than virgin rats. The temperature decrease was greatest in the pre‐0‐day rats, being 4.1 +/‐ 0.4 degrees C (mean +/‐ S.E.M.) at the end of cold exposure, compared with a decrease of 1.7 +/‐ 0.3 degrees C in the virgin rats. The tail skin temperature fell to 0 degrees C during cooling in all virgin rats and in pregnant rats at each gestational stage. 3. During cold exposure at 10 degrees C for 30 min, pre‐0‐day rats also showed significantly larger falls in Tco (1.8 +/‐ 0.6 degrees C) than virgin (0.4 +/‐ 0.2 degrees C), pre‐1‐week (0.8 +/‐ 0.3 degrees C), post‐0‐day (0.3 +/‐ 0.3 degrees C) or post‐1‐week rats (0.4 +/‐ 0.3 degrees C). Although body weights in pre‐0‐day rats were far larger than those in virgin rats, the increase in oxygen consumption per animal during cold exposure was 50% lower in pre‐0‐day rats (2.2 +/‐ 0.5 ml/min) than in virgin rats (5.3 +/‐ 0.3 ml/min). There was no difference in basal oxygen consumption per animal between the late pregnant and virgin rats. 4. Within 24 h after parturition, both the decrease of Tco and the increase of oxygen consumption during cold exposure returned to the values observed in virgin rats. 5. The present results demonstrate clearly that cold‐induced thermogenesis is significantly suppressed in rats at a late stage of pregnancy.
Corticospinal volleys evoked by anodal and cathodal stimulation of the human motor cortex.
Authors: Burke D , Hicks R G , Stephen J P
Published: December 2014

1. In fifteen neurologically normal subjects, corticospinal volleys evoked by transcranial stimulation of the motor cortex were recorded from the spinal cord using epidural electrodes in the high‐thoracic and low‐thoracic regions during surgery to correct scoliosis. 2. Anodal stimulation at the vertex produced complex corticospinal volleys that could be recorded at both sites, with multiple waves analogous to the D and I waves documented in animal experiments. These volleys were of higher amplitude when the cathode was 7 cm lateral to the vertex rather than 7 cm anterior. There were no differences in conduction time between the two recording sites for D and I waves, when these waves could be identified at the low‐thoracic site. 3. Anodal stimuli of 150 V commonly produced a descending volley containing a single peak at both recording sites. Modest increases in stimulus intensity to 225‐375 V produced a peak 0.8 ms in advance of the wave of lowest threshold in thirteen subjects and, in seven subjects, further increases produced an additional peak 1.7 ms in advance of the first‐recruited wave. The early peaks increased in size with stimulus intensity, replacing the first‐recruited wave. These results suggest that the site of impulse initiation with electrical stimulation of the motor cortex shifts from superficial cortex to deep structures, approximately 5 and 10‐11 cm below the cortex. These sites are probably the internal capsule and the cerebral peduncle. 4. With cathode at the vertex and anode over the ‘hand area’ the response of lowest threshold occurred at the latency of the anodal D wave but could not be recorded at the low‐thoracic site, suggesting that it was generated by the anode over the ‘hand area’. Slightly higher intensities induced a ‘cathodal D wave’ and still higher intensities produced late peaks at latencies of anodal I waves. These cathodal D and I waves involved axons innervating lumbar segments. There was no evidence that cathodal stimulation preferentially produced I waves. Cathodal stimulation at the vertex with the anode 7 cm anteriorly produced similar results: D waves were produced at relatively low intensities, but I waves appeared at relatively high stimulus intensities if at all.(ABSTRACT TRUNCATED AT 400 WORDS)
Substance P increases catecholamine secretion from perfused rat adrenal glands evoked by prolonged field stimulation.
Authors: Zhou X F , Livett B G
Published: December 2014

1. We have studied the effect of substance P (SP) on catecholamine (CA) secretion evoked by prolonged field stimulation from the perfused rat adrenal gland in vitro. 2. Previous studies show that SP has an inhibitory effect on the nicotinic response in a number of different tissue preparations. In the present study, we found that SP at concentrations from 10(‐7) to 10(‐6) M markedly increased CA secretion evoked by prolonged high‐frequency field stimulation but not that evoked by low‐frequency field stimulation. 3. In the absence of field stimulation SP by itself had no direct effect on CA secretion. This indicates that SP acts as a neuromodulator rather than a neurotransmitter to increase CA secretion from the adrenal medulla. 4. The nicotinic receptors were still functional following a period of prolonged field stimulation (120 min) because CA secretion was markedly increased in response to the nicotinic agonist dimethylphenylpiperazinium (DMPP). 5. Capsaicin pre‐treatment reduced CA secretion evoked by prolonged field stimulation and the facilitatory action of SP on CA secretion lasted longer in these capsaicin‐pre‐treated rats than in controls, indicating that SP‐containing capsaicin‐sensitive fibres innervating the adrenal medulla may be involved in the regulation of CA secretion. 6. In parallel with the increase in CA secretion, 3H overflow from the splanchnic nerve pre‐labelled with [3H]choline was also increased by SP. The increase in CA secretion by SP lasted longer than the increase in 3H overflow. These results suggest that SP facilitates CA secretion from the adrenal gland at two levels: (1) pre‐synaptically by facilitating ACh release from splanchnic nerve terminals, and (2) post‐synaptically by modulating the nicotinic secretory response by protection against nicotinic desensitization of secretion. 7. The present studies provide further evidence that endogenous SP in the splanchnic nerve may modulate CA secretion during stress.
Phosphoinositide hydrolysis is not negatively regulated by protein kinase C in the peripheral tissues of rat and chick.
Authors: Bhave S V , Malhotra R K , Wakade T D , Wakade A R
Published: December 2014

1. Phorbol esters are known to inhibit phospholipase C‐mediated hydrolysis of membrane phosphoinositide. This inhibition is attributed to participation of protein kinase C (PKC) in a negative‐feedback control of phosphoinositide metabolism. We have tested this hypothesis by using different types of activators and inhibitors of PKC. 2. Phorbol‐12,13‐dibutyrate (PDB) inhibited the stimulatory effect of acetylcholine (ACh) on [3H]inositol monophosphate ([3H]IP) formation in cultured sympathetic neurons of the chick embryo and adrenal medulla of the rat. 3. Acetylcholine (ACh) and 5‐hydroxytryptamine (5‐HT) activated neuronal PKC by 3‐ to 8‐fold. The extent of PKC activation by 100 microM‐ACh was comparable to that of 100 nM‐PDB. Activation of PKC by pre‐incubation of sympathetic neurons with ACh (or 5‐HT) did not inhibit the stimulatory effects of ACh (or 5‐HT) on [3H]IP formation. 4. Pre‐treatment of sympathetic neurons or adrenal medulla with a PKC inhibitor H7 (1‐(5‐isoquinolinyl‐sulphonyl)‐2‐methyl‐piperazine) almost completely blocked activation of the enzyme induced by PDB, ACh or 5‐HT. However, blockade of PKC did not prevent the inhibitory effects of PDB on ACh‐induced [3H]IP formation. 5. Vasoactive intestinal polypeptide (VIP) and muscarine induced catecholamine secretion from the perfused adrenal medulla via formation of inositol‐1,4,5‐tirisphosphate (IP3). Phorbol‐12,13‐dibutyrate decreased muscarine‐induced catecholamine secretion. However, activation of PKC by VIP had no effect on muscarine‐induced catecholamine secretion and vice versa. 6. These results suggest that PKC is not negatively coupled to phosphoinositide hydrolysis in sympathetic neurons and chromaffin cells. Phorbol esters must have targets other than PKC to interfere with the phosphoinositide hydrolysis.
A group II‐activated ascending tract of lumbosacral origin in the cat spinal cord.
Authors: Harrison P J , Riddell J S
Published: December 2014

1. Electrophysiological investigations have revealed a population of ascending tract neurones originating in the lumbosacral enlargement, with input from group II muscle afferents of the cat hindlimb. 2. Single‐unit microelectrode recordings were made in the lateral funiculus at L6, from the axons of thirty‐four ascending tract neurones. All of the axons were antidromically activated by stimulation of the ipsilateral lateral funiculus at Th13 and, whenever tested (eight units), at C1. 3. Conduction velocities of the axons, between the L6 and Th13 segment, ranged from 33 to 92 m s‐1 (mean 61 m s‐1). 4. All of the ascending tract neurones were discharged following electrical stimulation of muscle nerves at group II strength, but not by weaker stimuli in the group I range. Most of the investigated neurones were excited by group II afferents of more than one muscle nerve. In addition, a proportion of the units tested could also be discharged by cutaneous and by joint afferents. 5. Responses to natural stimuli were investigated in eighteen ascending tract neurones discharged by electrical stimulation of group II afferents in the gastrocnemius‐soleus (GS) and plantaris (P1) nerves which were dissected free in continuity with their muscles. Seven units were spontaneously active. Eight units responded to isometric contraction of the GS/P1 muscles with a discharge occurring mainly on the falling phase of muscle tension. Nine units increased their discharge frequency in response to stretching of the muscles and five units responded to mechanically probing the muscles with a blunt instrument. 6. The final termination sites of this group of ascending tract neurones has yet to be determined. Initial attempts (three units) to antidromically activate the neurones from the cerebellum have been unsuccessful. Other likely areas of termination in the brain stem are considered.
The effects of bumetanide, amiloride and Ba2+ on fluid and electrolyte secretion in rabbit salivary gland.
Authors: Lau K R , Howorth A J , Case R M
Published: December 2014

1. In order to distinguish between models of anion secretion, the effects of transport inhibitors on saliva flow rate and electrolyte composition were studied during the plateau phase of secretion in rabbit mandibular salivary glands. 2. Bumetanide, an inhibitor of Na+,K+,2Cl‐ co‐transport, inhibited flow rate (by 60%) and reduced Cl‐ concentration. K+ and HCO3‐ concentrations were increased. Forskolin, an adenylate cyclase activator which inhibits ductal transport, did not significantly affect this pattern of changes. 3. Amiloride, used at concentrations that would inhibit Na(+)‐H+ exchange, inhibited flow rate (by 30%). Cl‐ concentration was initially increased before subsequently decreasing at the same time as HCO3‐ concentration increased. These concentration changes can probably be attributed to ductal transport. When amiloride was applied to glands perfused with nominally HCO3‐ ‐free solutions, inhibition of flow rate was rapid and almost complete. 4. When amiloride and bumetanide were both present in the perfusate, flow rate was inhibited by 92%. The pattern of electrolyte changes was not significantly different from that observed in the presence of bumetanide alone. 5. Inhibition of K+ channel activity using Ba2+ also inhibited flow rate. Cl‐ concentration was increased as was K+ concentration. HCO3‐ concentration was not increased. 6. The anion exchange inhibitor DIDS (4,4'‐diisothiocyanatostilbene‐2,2'‐disulphonic acid) had no effect on either flow rate or electrolyte concentration. It did, however, elicit secretion in the absence of acetylcholine. 7. The data suggest that Na(+)‐H+ and Cl‐ ‐HCO3‐ exchangers are unlikely to be involved in fluid and electrolyte secretion in these glands as suggested by some authors. Most of the data can be explained by postulating the existence of non‐specific anion channels in the apical membranes of the acinar cells.
The effects of lyotropic anions on charge movement, calcium currents and calcium signals in frog skeletal muscle fibres.
Authors: Delay M , Garcia D E , Sanchez J A
Published: December 2014

1. Intramembrane charge movement and Ca2+ currents were monitored in voltage‐clamped segments of frog skeletal muscle fibres using the triple‐Vaseline‐gap technique. Calcium signals were measured in current‐clamped fibres using either of the indicators Arsenazo III or Antipyrylazo III. 2. Non‐linear capacitative currents (charge 1) were obtained using a subtraction procedure which employed either a ‐20 mV control pulse from a holding potential of ‐100 mV or alternatively a control pulse to +80 mV in depolarized fibres. The amount of charge mobilized depended on voltage according to a two‐state Boltzmann function. The total charge (Qmax) was increased by ca 100% and the steepness parameter (k) by ca 70% when a +80 mV control pulse was used. 3. Thiocyanate (SCN‐) and other lyotropic anions reversibly shifted the voltage dependence of mobilized charge towards negative potentials. Qmax was not significantly affected. ‘Off’ charge tails were greatly prolonged by lyotropic anions. 4. Extracellularly applied lyotropic anions affected the dihydropyridine‐sensitive Ca2+ current by shifting the I‐V relation toward more negative voltages and delaying deactivation of the tail currents. 5. The effects of lyotropic anions did not depend on whether the anion was introduced intracellularly or extracellularly. 6. Extracellular SCN‐ reversibly increased the peak amplitude and rate of rise of Ca signals, and decreased the latent period between stimulation and onset of the Ca signal. 7. It is concluded that lyotropic anions have similar effects on Ca2+ currents and on charge movement.
Acute effects of neomycin on slowly adapting type I and type II cutaneous mechanoreceptors in the anaesthetized cat and rat.
Authors: Baumann K I , Hamann W , Leung M S
Published: December 2014

1. Slowly adapting type I (SAI) and type II (SAII) mechanoreceptors in the skin were studied in anaesthetized cats and rats employing mechanical stimuli every 30 s. Individual stimuli rose within 200 ms to a plateau force which was kept constant through a feedback control unit for 2000 ms. 2. In cats, close arterial infusion of neomycin (2.5 mg/min) as sulphate was given through a side branch into the femoral blood stream for 5, 10 or 20 min at a rate of 0.025 ml/min. At other times saline was infused at the same rate. 3. After 20 min of neomycin infusion (total 50 mg) nervous discharge of cat SAI receptors was suppressed to about 30% of the control responses before neomycin infusion. Nervous responses were reduced more profoundly during the plateau phase of stimulation than during the dynamic phase. The interspike interval histogram was severely distorted. 4. In contrast, cat SAII receptors maintained about 70% of their control response after 20 min of neomycin infusion. The interspike interval histogram showed an orderly shift towards longer intervals maintaining its normal shape. 5. In rats, intradermal microinfusion of neomycin (30 micrograms/min) through a glass micropipette into the immediate vicinity of the receptor under investigation resulted in severe transient suppression of SAI receptor responses to about 10% of the control level. Receptor responses recovered almost completely about 1 h after the end of neomycin application. 6. It is concluded that the observed differences between the two types of slowly adapting mechanoreceptors are consistent with the hypothesis that the SAI receptor functions as a secondary sensory receptor, with a synaptic link between the Merkel cell and the primary afferent neurone.
Chloride‐sensitive nature of the adrenaline‐induced current in guinea‐pig cardiac myocytes.
Authors: Matsuoka S , Ehara T , Noma A
Published: December 2014

1. Ionic selectivity of an adrenaline‐induced current was investigated in single guinea‐pig ventricular cells by recording whole‐cell currents using the patch clamp technique combined with internal perfusion. Other ionic currents and exchange currents known in ventricular cells were suppressed by appropriate inhibitors and the adrenaline‐induced current was defined as a difference between currents obtained in the presence and absence of adrenaline. 2. The adrenaline‐induced current was time independent and its I‐V relation showed saturation of the inward current in the negative voltage range. 3. The reversal potential was approximately ‐20 mV with 140 mM‐NaCl external solution and Cs(+)‐rich internal solution containing 51 mM‐Cl‐. Replacing Na+ with various monovalent and divalent cations (Li+, K+, Rb+, Cs+, Ca2+, Sr2+ and Ba2+) produced no appreciable change in the reversal potential. 4. Varying the external Cl‐ concentration ([Cl‐]o) in exchange for aspartate or benzenesulphonate greatly changed the reversal potential. The relationship between the reversal potential and log[Cl‐]o indicated a slope of 59.5 or 53.6 mV per tenfold change in [Cl‐]o in the presence of 51 or 102 mM‐Cl‐ in the internal solution, respectively. 5. Anion substitutions did not appreciably affect the I‐V relation before application of adrenaline, suggesting that the cell membrane had a low Cl‐ conductance in the control state. 6. 4.4'‐Dinitrostilbene‐2‐2'‐disulphonic acid (DNDS: 1‐10 mM), a specific inhibitor of membrane chloride permeability, depressed the adrenaline‐induced current without changing the reversal potential. 7. The results suggest strongly that the adrenaline‐induced current is carried mainly by Cl‐. However, the development of this current appears to depend also on external cations, since the magnitude of the adrenaline response varied depending on the external cations species, with no response in Tris‐HCl or TEA‐Cl solution. The external cations may facilitate the adrenaline response with a sequence of efficacy of Na+ greater than K+, Rb+ greater than Cs+, Li+, divalent cations.
Halothane increases Ca2+ efflux via Ca2+ channels of sarcoplasmic reticulum in chemically skinned rat myocardium.
Authors: Herland J S , Julian F J , Stephenson D G
Published: December 2014

1. A method has been developed to study Ca2+ fluxes across the sarcoplasmic reticulum (SR) of chemically (saponin) skinned myocardium without interference from the SR Ca2+ pump. 2. Exposure of rat cardiac trabeculae to a solution containing 50 micrograms/ml saponin for 10 min or longer caused an SR Ca2+ efflux which was not blocked by Ruthenium Red (RRed) and did not require the presence of nucleotides. 3. Exposure of the saponin‐treated cardiac preparation to 11 mM‐AMP, when the SR Ca2+ pump was not active, enhanced Ca2+ release from the SR by a mechanism which was blocked by 10 microM‐RRed. 4. The amount of Ca2+ loaded by the 10 min saponin‐treated trabeculae was maintained constant for at least 3 min when the preparations were transferred to low [Ca2+] solutions (0.1 mM‐EGTA; pCa greater than 7.5) containing ATP. This indicated that the Ca2+ pump can efficiently recycle Ca2+ lost from the SR under these conditions. 5. Halothane (0.47 and 1.89 mM) reversibly increased the rate of Ca2+ release from the SR regardless of whether or not the SR Ca2+ pump was active. This effect was more marked at 1.89 mM than at 0.47 mM. RRed (10 microM) completely blocked the Ca2+ release induced by both concentrations of halothane. 6. The presence of nucleotide (11 mM‐AMP) did not affect the halothane‐induced Ca2+ release when the Ca2+ pump was inactive. 7. Exposure of cardiac preparations to solutions containing more than 5 mM‐halothane irreversibly damaged the ability of the SR to load Ca2+. 8. The results suggest that at lower doses (0.47 and 1.89 mM) halothane specifically and reversibly stimulates Ca2+ efflux via the RRed‐sensitive SR Ca2(+)‐release channel by a mechanism which does not require the presence of nucleotides or relatively high [Ca2+]. The results also suggest that AMP and halothane act independently and non‐synergistically to increase Ca2+ efflux through the same SR Ca2(+)‐release channel. At higher doses (greater than 5 mM) halothane irreversibly damages the SR membrane, presumably by disrupting the lipid bilayer.
Strong Pasteur effect in rabbit corneal endothelium preserves fluid transport under anaerobic conditions.
Authors: Riley M V , Winkler B S
Published: December 2014

1. The hydration and transparency of the mammalian cornea are maintained by a metabolically dependent fluid transport system located in the endothelial cell layer. The purpose of the study was to determine whether this pump activity is dependent upon aerobic or anaerobic metabolism. 2. The ability of the cornea, superfused in vitro with a bicarbonate‐Ringer solution containing glucose and adenosine, to maintain normal hydration (thickness) when respiration is inhibited has been examined in intact and de‐epithelialized preparations and correlated with glycolytic activity and cellular concentrations of ATP. 3. In respiring intact and de‐epithelialized corneas thickness was maintained for periods up to 5 h during superfusion with the control Ringer solution. 4. KCN (10(‐3) M) or antimycin A (10(‐6) M) caused the intact cornea to swell at 15 +/‐ 3 microns h‐1, whereas the de‐epithelialized tissue maintained normal thickness under these conditions. This swelling of the intact tissue appears to be due to the osmotic effect of increased epithelial lactate production under anaerobic conditions. 5. Pre‐swollen de‐epithelialized corneas deturgesced fully to original thickness at a rate of 43 +/‐ 7 microns h‐1 under aerobic conditions, but with KCN or antimycin they deturgesced at only 65% of that rate and generally failed to return to their original thickness. 6. Ouabain (10(‐4) M) caused de‐epithelialized corneas to swell in the presence of KCN or antimycin, as it did under aerobic conditions, showing that maintenance of hydration or deturgescence are pump‐dependent processes under both conditions. 7. Lactate production was markedly stimulated by KCN or antimycin in intact and de‐epithelialized preparations, but not in the stroma alone. The magnitude of the Pasteur effect was calculated to be 5‐fold in the endothelium and 2.5‐fold in the epithelium. Ouabain inhibited anaerobic lactate production in the endothelium by 50%. 8. ATP content of the epithelium following 5 h superfusion was 22.0 nmol cm‐2 in control (aerobic) corneas, but fell to 1.9 nmol cm‐2 in the presence of 10(‐3) M‐KCN, whereas the endothelial value fell only from 1.1 to 0.7 nmol cm‐2 under these conditions. 9. Omission of glucose from the medium containing KCN or antimycin caused immediate swelling of tissues and a rapid decline of ATP content to less than 1% of that in control conditions.(ABSTRACT TRUNCATED AT 400 WORDS)
Oxygen consumption for K+ uptake during post‐stimulatory activation of Na+, K(+)‐ATPase in perfused rat mandibular gland.
Authors: Murakami M , Miyamoto S , Imai Y
Published: December 2014

1. In order to determine the stoichiometry of K+ uptake and ATP consumption by Na+, K(+)‐ATPase in the isolated, perfused mandibular gland of the rat, oxygen consumption and net K+ uptake from the vascular side were measured during the recovery period following K+ depletion by stimulation with acetylcholine in combination with ouabain. 2. Acetylcholine (10(‐6) M) induced fluid secretion and an initial, transient release of K+ from the gland. Addition of ouabain suppressed salivary fluid secretion and caused an additional, dose‐dependent, transient release of K+. 3. With acetylcholine (10(‐6) M), the oxygen consumption of the gland increased to 62.4 +/‐ 4.2 microliters/(g min) from a resting value of 12.9 +/‐ 1.2 microliters/(g min). The increased oxygen consumption was suppressed by ouabain, in a dose‐dependent fashion. 4. Withdrawal of acetylcholine and ouabain induced a net uptake of K+ and, simultaneously, an increase in oxygen consumption. The cumulative K+ uptake and the increment of oxygen consumption during the recovery period were dependent on the concentration of used ouabain. 5. The rates of K+ uptake and ATP hydrolysis were compared during recovery, assuming that six moles of ATP were hydrolysed for each mole of oxygen consumed. The data obtained during the later phase of the recovery lay on a single straight line with a slope of 1.8 for each of the various concentrations of ouabain, suggesting that the relationship between K+ uptake and energy consumption was linear. 6. Assuming the K+:ATP stoichiometry of the Na+, K(+)‐ATPase to be 2:1, K+ would appear to be transported with ca 90% efficiency by Na+, K(+)‐ATPase in the rat mandibular gland. Using 1.8‐2.0 as a K+/ATP stoichiometry, the rate of primary active K+ uptake and the corresponding passive K+ efflux during secretion were estimated to be 20‐22 mumol/(g min) from the oxygen consumption and the net K+ flux. 7. The passive K+ efflux was estimated, from the initial rate of K+ release caused by addition of 10(‐3) M‐ouabain, to be 30 mumol/g min). The discrepancy between the estimated active K+ uptake and passive K+ release (8‐10 mumol/(g min] could be attributed to a secondary active K+ uptake process such as Na(+)‐K(+)‐2Cl‐symport.