bluuebunny/crossref_metadata_2025 huggingface dataset:
Acetylcholine receptor channels and their block by clonidine in cultured bovine chromaffin cells.
Authors: Cull‐Candy S G , Mathie A , Powis D A
Published: December 2014

1. Acetylcholine (10‐50 microM) applied to bovine chromaffin cells under whole‐cell voltage clamp produced an inward current and an increase in the noise level of the current trace. At concentrations greater than or equal to 20 microM the ACh current desensitized with time. Spectral analysis of the ACh‐induced increase in current noise showed that it could be fitted by a single Lorentzian component with a time constant, tau noise = 11 +/‐ 0.9 ms (Vm = ‐80 mV). 2. Clonidine (2‐30 microM) markedly reduced the size of the ACh‐induced whole‐cell current, and altered the shape of the noise spectrum. In the presence of clonidine, ACh noise spectra were fitted by two Lorentzian components with time constants which varied with clonidine concentration. The single‐channel conductance from noise (gamma = 24 pS) was unaltered by clonidine. 3. The reduction in the size of the whole‐cell ACh current, produced by clonidine, was not mimicked by adrenaline (at concentrations up to 60 microM). GABA‐induced whole‐cell currents and spectra of GABA noise were also unchanged suggesting that the effect of clonidine was specific for the ACh receptor channel. 4. When applied intracellularly (from the patch pipette) clonidine had no apparent influence on the whole‐cell ACh current. Furthermore, when clonidine was perfused over the cell surface at a concentration sufficient to block the whole‐cell ACh response, single ACh channels could still be recorded under the patch‐pipette tip (i.e. in cell‐attached patches, not exposed to clonidine). 5. Clonidine block showed little voltage dependence; the peak ACh current remained linearly dependent on clamp potential. In addition, the fast and slow time constants derived from ACh noise spectra in the presence of clonidine did not show the sort of dependence on antagonist concentration expected for simple channel block; this suggests that clonidine has a complex blocking action. 6. Single ACh channels recorded in outside‐out patches had a conductance of gamma = 39 +/‐ 0.7 pS, although a subpopulation of smaller and larger events also occurred in some patches. The mean single‐channel conductance in outside‐out patches was unaltered by clonidine. 7. Under normal conditions the kinetics of single ACh channels were more complex than suggested from noise analysis. Burst length distributions could be described by two exponential components with fast and slow time constants of tau f = 0.69 +/‐ 0.17 and tau s = 9.51 +/‐ 0.84 ms.(ABSTRACT TRUNCATED AT 400 WORDS)
Electrophysiological alternans and restitution during acute regional ischaemia in myocardium of anaesthetized pig.
Authors: Dilly S G , Lab M J
Published: December 2014

1. Alternate long and short action potential durations, or electrical alternans, has only been sporadically observed in ischaemic myocardium in situ. We systematically studied alternans in the latter to characterize the phenomenon, relate it to ventricular arrhythmia and suggest possible mechanisms. 2. Sixteen Landrace pigs were anaesthetized (Azaperone, N2O and O2), ventilated and the hearts exposed. A branch of the left coronary artery was ligated. Left intraventricular and systemic pressures were monitored. Monophasic action potentials were recorded simultaneously with up to five suction electrodes in and around the proposed ischaemia area. 3. A computer measured the duration of every action potential, at several phases of repolarization, throughout the first hour of ischaemia. This allowed the systematic study of the alternans. Measurements during defined stimulus protocols were also made for the construction of electrical restitution curves. 4. Alternans was found in all recordings within the ischaemic area and in two‐thirds of those in the ‘border’ area. There was no alternans in non‐ischaemic areas. 5. The alternans, when action potential duration was plotted for every beat, appeared as an oscillation which was pleomorphic. It could be: (a) stable for hundreds of beats; (b) switched or triggered (by one extraneous beat having a different cycle length) between one stable state with high and one with low or absent alternans; (c) damped; (d) undamped to take a crescendo form, sometimes preceding ventricular fibrillation. 6. The alternans in general showed an ill‐defined peak incidence between about 200 to 1500 beats after the onset of ischaemia, and a clearer late peak at about 3000 beats. These periods occurred at about 2‐7 min and 15‐40 min, corresponding to so‐called phase 1A and 1B arrhythmia respectively. Only the late peak was seen with triggered alternans. 7. The electrical restitution curve for the action potential duration during ischaemia when compared with curves, constructed with data from non‐ischaemic myocardium, showed a progressive depression in plateau, a reduction in magnitude and was flattened at 1 h. However, there was a reversal or reduction in decline at about 15‐45 min. 8. We propose that electrical alternans is a distinctive electrophysiological characteristic of ischaemic myocardium which may be causally related to ventricular arrhythmia and fibrillation, and that at least two mechanisms contribute to the alternans: (i) electrical restitution of the action potential and (ii) changes in intracellular calcium cycling.
Reflex tracheal contraction during pulmonary venous congestion in the dog.
Authors: Kappagoda C T , Man G C , Ravi K , Teo K K
Published: December 2014

1. The effect of pulmonary venous congestion on tracheal tone was studied in dogs anaesthetized with alpha‐chloralose. Pulmonary venous congestion was produced by partial obstruction of the mitral valve to increase left atrial pressure by 10 mmHg. Tracheal tone was measured in vivo by an isometric force displacement method. 2. Tracheal tone increased by 6.3 +/‐ 0.3 g from a control level of 91.6 +/‐ 2.8 g when left atrial pressure was increased by 10.5 +/‐ 0.3 mmHg. This response was abolished by cooling the cervical vagi to 8 degrees C at a point caudal to the origin of the superior laryngeal nerves. Also, sectioning the superior laryngeal nerves abolished this increase in tracheal tone. 3. Afferent activity recorded from rapidly adapting receptors of the airways increased significantly during pulmonary venous congestion. This increase in activity was abolished by cooling the vagi caudal to the recording site to 8‐9 degrees C. 4. Administration of propranolol (0.5 mg/kg) failed to abolish this increase in tracheal tone while atropine (3 mg/kg) did so. 5. Stimulation of left atrial receptors without an increase in left atrial pressure and stimulation of right atrial receptors with and without increases in right atrial pressure did not cause any change in tracheal tone. 6. It is suggested that pulmonary venous congestion is associated with a reflex increase in tracheal tone, the afferent limb of which is formed by pulmonary receptors discharging into myelinated fibres in the cervical vagi and the efferent limb by parasympathetic cholinergic fibres in the superior laryngeal nerves. The afferent receptors are likely to be the rapidly adapting receptors. This reflex may be of importance in the development of the respiratory symptoms associated with left ventricular failure.
Phosphorous compounds studied by 31P nuclear magnetic resonance spectroscopy in the taenia of guinea‐pig caecum.
Authors: Nakayama S , Seo Y , Takai A , Tomita T , Watari H
Published: December 2014

1. In the isolated taenia (0.4‐0.6 g) of guinea‐pig caecum, the intracellular phosphorous compounds and pH were investigated using 31P nuclear magnetic resonance (NMR) under various metabolic conditions. 2. The ratios of the intracellular concentration of phosphocreatine ([ PCr]) and inorganic phosphate [( Pi]) to nucleotide triphosphate ([ NTP]) were 1.71 +/‐ 0.14 and 0.58 +/‐ 0.11 (n = 25), respectively, in normal solution (32 degrees C). The intracellular pH estimated from the chemical shift of Pi was 7.05 +/‐ 0.06 (n = 25), agreeing well with those previously obtained. 3. In the absence of glucose, the [PCr] and [NTP] were decreased to almost a half after 150 min exposure to 40 mM‐K+ solution, while [Pi] was increased 3‐fold. These changes were much faster than the rate of decline in tension. When glucose was readmitted, the contractile response to K+ fully recovered in 50 min. However, this was accompanied with only a partial recovery of [PCr] and [Pi], but no recovery of [NTP]. The intracellular pH was lowered by about 0.2 of a unit, suggesting an increase in glycolysis. 4. In Ca2+‐free solution, respiratory inhibition with hypoxia or CN (1 mM) only decreased [PCr], leaving [NTP] nearly unchanged. On the other hand, respiratory inhibition in excess‐K+ solution containing Ca2+ (2.4 mM) severely depleted PCr and decreased [NTP] to 40%. Increasing glucose to 50 mM did not prevent these changes, although it increased tension development. 5. The simultaneous decrease of [NTP] and [PCr] during K+ contracture suggests that the activity of creatine phosphokinase is low. The recovery from respiratory inhibition was much better for [PCr] than for [NTP]. Slow, but perfect, recovery of all NTP peaks was produced by adding 1 mM‐adenosine to normal solution. 6. It was suggested that tension development is closely related to the turnover rate of ATP, and not to its concentration, and that deamination of adenosine is a limiting factor in the recovery of ATP after excessive consumption.
The kinetics of quantal releases during end‐plate currents at the frog neuromuscular junction.
Authors: Van der Kloot W
Published: December 2014

1. The preceding paper (Van der Kloot, 1988) described a method for estimating the timing of quantal releases during an end‐plate current. This period of elevated quantal release is called the early release period or ERP (Barrett & Stevens, 1972b). In the present paper, this deconvolution method is used to study the effects of varying quantal output by extracellular ions, stimulus patterns and drugs. 2. The data were obtained by voltage clamping end‐plates in low‐Ca2+ high‐Mg2+ solutions, or in solutions containing tubocurarine (measuring the decay of the miniature end‐plate currents (MEPCs) before curarization and assuming a value for MEPC amplitude after curarization). Data were also obtained by extracellular recording in Ca2+‐free solution, using a recording pipette filled with CaCl2 and regulating Ca2+ release with a bias current. The three approaches led to similar conclusions. 3. Quantal release rose during the ERP along a sigmoid curve and reached a maximum after about 1.4 ms at 10 degrees C. This is called the time to peak. Quantal release then fell, following an exponential time course with a time constant of about 1.2 ms (10 degrees C). This is called the time constant for decline. 4. The ERP was followed by further, elevated quantal release, at a much lower rate, which declined over a longer time course. This is called late release. The magnitude of late release appears to be almost independent of the magnitude of release during the ERP, although the deconvolution method is a poor one for determining late release. The remainder of the results therefore focus on the ERP. 5. Increasing [Ca2+]o increased quantal output, and the rate of quantal output. It did not change the time to peak or the time constant of decline. Similarly, replacing Ca2+ with Sr2+ did not alter the time course of the ERP. 6. Two‐pulse facilitation increased quantal output without changing the time to peak or the time constant of decline. 7. Quantal output was enhanced still more following a brief series of repetitive nerve stimulations. There was a lengthening of the time to peak; there was no change in the decline. The depression produced by longer series of repetitive stimulations did not change the time course of the ERP. 8. 4‐Aminopyridine (4‐AP) and dimethylsulphoxide (DMSO) increased quantal output and lengthened the time to peak, without altering the time constant for decline. 9. Adenosine decreased quantal output without altering the time course of the ERP.(ABSTRACT TRUNCATED AT 400 WORDS)
Slow‐to‐fast transformation of denervated soleus muscles by chronic high‐frequency stimulation in the rat.
Authors: Gorza L , Gundersen K , Lømo T , Schiaffino S , Westgaard R H
Published: December 2014

1. Adult soleus muscles were denervated and stimulated directly for 2‐130 days with ‘fast’ (short pulse trains at 100 Hz) or 'slow’ (continuously at 10 Hz, or long pulse trains at 15 Hz) stimulus patterns. 2. At the end of the period of stimulation isometric twitches and tetani and isotonic shortening velocities were measured. Frozen cross‐sections were later examined with antibodies against myosin heavy chains specific for adult fast, adult slow and fetal myosin. 3. Isometric twitch duration (twitch time‐to‐peak and half‐relaxation time) decreased during intermittent 100 Hz stimulation to values that were almost as fast as in the normal extensor digitorum longus (EDL) (95 and 94% transformation). The major part of the decrease occurred between 2 and 21 days after the onset of stimulation, and was accompanied by post‐tetanic potentiation of the twitch, 'sag’ in tension during an unfused tetanus, lower twitch/tetanus ratio and marked shifts to the right (higher frequencies) of the tension‐frequency curve of the muscle. In contrast, during 10 or 15 Hz stimulation the isometric twitch duration remained slow, the twitch continued to show post‐tetanic depression and absence of ‘sag’, while the twitch/tetanus ratio increased. 4. Denervation per se led to a slight increase and, then, after about a month, to a moderate and gradual decrease in twitch duration. The twitch/tetanus ratio increased markedly and post‐tetanic depression became less pronounced or disappeared. Muscle weight and particularly tetanic tension were markedly reduced and these reductions were to a large extent counteracted by electrical stimulation. 5. Implantation of sham electrodes had no effect on twitch duration of denervated or innervated control muscles, but reduced tetanic tension in the innervated control muscles. 6. Maximum isotonic shortening velocity of the whole muscle (mm/s) increased during intermittent 100 Hz stimulation to a value as fast as in the normal EDL (110% transformation). Since the muscle fibres also increased in length (35%) maximum intrinsic shortening velocity (fibre lengths/s) was only incompletely transformed (55%). The increase in Vmax occurred between 7 and 14 days after the onset of stimulation. 7. All the fibres stimulated intermittently at 100 Hz were strongly labelled with anti‐fast myosin and more than 90% were in addition weakly labelled by anti‐slow myosin. Weak and variable labelling with anti‐fast myosin was first detected 7 days after the onset of stimulation. In contrast, essentially all the fibres stimulated at 10 or 15 Hz showed no binding of anti‐fast but strong binding of anti‐slow myosin.(ABSTRACT TRUNCATED AT 400 WORDS)
Caffeine contracture in guinea‐pig ventricular muscle and the effect of extracellular sodium ions.
Authors: Kitazawa T
Published: December 2014

1. The mechanisms underlying the virtual absence of caffeine contracture in guinea‐pig heart in a Na+‐rich external solution were reinvestigated in small (50‐120 microns thick) bundles of intact and skinned papillary muscle fibres. 2. In Na+‐free solution, the peak tension of 30 mM‐caffeine contracture corresponded to the maximum tension of the skinned fibres, and was independent of changes in [Ca2+]o and [K+]o. In the presence of external Na+, the peak tension, which was at most several per cent of the maximum, was affected by [Ca2+]o, [Na+]o and [K+]o, and enhanced by Mn2+ and Ni2+. 3. In the absence of Ca2+, replacement of Na+ with K+ allowed caffeine to evoke a large contracture, showing that there was sufficient calcium stored in the cells under Na+‐rich conditions. After treatment with 30 mM‐caffeine in the Na+‐rich, Ca2+‐free solution, and upon replacement of all Na+ with Li+, caffeine was still able to produce a large contracture, which was dependent upon Ca2+ pre‐loading of the cells before the first caffeine treatment and upon the subsequent duration in the Na+‐free solution. 4. Replacement of Li+ with Na+ during the contracture led to rapid relaxation which was delayed by an increase in [Ca2+]o, depolarization by K+, and addition of La3+ and Mn2+. After Na+‐induced complete relaxation in the absence of Ca2+, upon removal of the drugs and Na+, subsequent application of caffeine to the cells evoked a large contracture without Ca2+ reloading. 5. In the skinned fibres, 30 mM‐caffeine increased the Ca2+ sensitivity of the contractile system and depressed the maximum tension. An increase in Na+ from 8.4 to 58.4 mM altered neither Ca2+ sensitivity nor the rate of tension development in the absence or presence of caffeine. 6. Increase in Na+ affected neither the rate nor the amount of Ca2+ uptake by the sarcoplasmic reticulum (SR) in the absence or presence of caffeine. Increasing Na+ slightly inhibited the caffeine‐induced Ca2+ release from the SR, but more than 10 mM‐caffeine produced SR Ca2+ depletion. 7. In the presence of a strong Ca2+ buffer, the steady level of Ca2+ uptake by the SR with 1 mM‐caffeine was equal to the amount of Ca2+ remaining in the SR just after the application of caffeine, indicating that Ca2+ release was not inactivated.(ABSTRACT TRUNCATED AT 400 WORDS)
Binocular summation in humans: evidence for a hierarchic model.
Authors: Frisén L , Lindblom B
Published: December 2014

1. Binocular summation was studied in human subjects using a batter of vision tests. Two tests assessed detection, another three acuity, one hyperacuity and one pattern recognition. 2. The magnitude of summation was consistent with, or exceeded, the level predicted from quadratic summation for both detection tests. 3. The summation factor was significantly smaller in the resolution tests than in the detection tests. Hyperacuity showed a large individual variation. 4. Spatial filtering of acuity targets did not influence summation. 5. No summation was found in the pattern recognition test. 6. It is argued that the degree of summation is related to the complexity of the visual task. A simple task yields a larger binocular summation than a more complex one. This may be related to the level of processing in the primary visual cortex.
Calcium‐activated non‐selective cation channel in ventricular cells isolated from adult guinea‐pig hearts.
Authors: Ehara T , Noma A , Ono K
Published: December 2014

1. A class of Ca2+‐activated non‐selective cation channel was identified in ventricular cells, which were dissociated from adult guinea‐pig hearts using collagenase. 2. Under cell‐attached conditions the patch electrode filled with a Na+‐rich solution recorded no obvious single‐channel current at the resting membrane potential. Subsequent superfusion of the ventricular cell with a Na+‐free Tyrode solution induced an inward‐going single‐channel current as well as contracture of the cell. Kinetics of this channel were not affected by varying the membrane potential. 3. Single‐channel currents showing a conductance similar to those observed in the cell‐attached patches were recorded in isolated inside‐out membrane patches when the inner side of the membrane was exposed to a free Ca2+ concentration ([Ca2+]i) higher than 0.3 microM. The slope conductance of the channel was 14.8 +/‐ 2.9 pS (mean +/‐ S.D., n = 17) at 20‐25 degrees C. 4. The reversal potential examined in the inside‐out patch was about 0 mV irrespective of the Na+‐rich, K+‐rich, Li+‐rich or Cs+‐rich solutions on either side of the membrane, thereby indicating that the channel was almost equally permeable to these cations. 5. The open probability of the channel was increased by raising [Ca2+]i with the maximum value of 0.93 +/‐ 0.17 (n = 4) at about 10 microM [Ca2+]i. The dose‐response relation was fitted to the saturation kinetics with a Hill coefficient of 3.0 and a half‐maximum concentration of 1.2 microM [Ca2+]i. 6. The gating kinetics were complex; both the open and closed time histograms showed at least two exponential components with time constants of 3.8 +/‐ 1.3 ms and 140 +/‐ 110 ms for open time and 1.8 +/‐ 1.1 ms and 14.9 +/‐ 5.3 ms for closed time (n = 4) at 10 microM [Ca2+]i. Reduction of [Ca2+]i resulted in both a decrease of the time constant of the slow component in the open time histogram and an increase of the two time constants of the closed time histogram. 7. Contribution of the channel to the whole‐cell current was discussed based on an estimation of the channel density, presumably about 0.04 approximately 0.4/microns 2. Maximum activation of the channel would produce 7.2 approximately 72 nS of membrane conductance, which would explain the reported magnitude of the Ca2+‐mediated background conductance of the single myocyte. The channel may also contribute, at least in part, to the transient inward current which develops in Ca2+‐overloaded cardiac cells.
Calcium‐independent increase of transmitter release at frog end‐plate by trinitrobenzene sulphonic acid.
Authors: Kijima H , Tanabe N
Published: December 2014

1. Application of an amino‐residue‐modifying reagent, 2,4,6‐trinitrobenzene‐1‐sulphonic acid (TNBS), to the frog neuromuscular junction in high‐magnesium Ringer solution rapidly increased both the amplitude of nerve‐evoked end‐plate potentials (EPPs) and the frequency of miniature end‐plate potentials (MEPPs). These showed a similar initial time course and reached a maximum 3‐7 min and about 10 min, respectively, after the start of application of 2 mM‐TNBS. Then, the EPP amplitude decreased, while the MEPP frequency maintained its plateau value. The increase in transmitter release and the decrease in EPP amplitude by TNBS may have been due to different modes of action. 2. The distribution of MEPP amplitude was unchanged by TNBS treatment. 3. The carbachol‐induced postsynaptic potential and the extracellularly recorded presynaptic action current were not affected by TNBS treatment for up to 30 min, indicating that the change in EPP amplitude produced by TNBS was not due to either a postsynaptic effect or a change in action potential at the presynaptic terminal. 4. The frequency of MEPPs was increased by TNBS application even when Ca2+ was omitted from the external Ringer solution or when a specific calcium channel blocker, synthetic omega‐conotoxin, was added. This indicates that Ca2+ inflow to the nerve terminal is not necessary for TNBS action. 5. When a calcium chelator, BAPTA, was loaded into the presynaptic nerve terminal, the facilitation of EPPs by trains of nerve stimuli was scarcely observed. This suggested that the cytosolic free Ca2+ in the presynaptic terminal was buffered by BAPTA. Under this condition, the amplitudes of EPPs were increased by TNBS application to the same extent as in the control without BAPTA, but were accompanied by little facilitation. The MEPP frequency was also increased by TNBS to the same extent as in the control. These results suggest strongly that augmentation of transmitter release by TNBS was not due to an increase in cytosolic Ca2+ concentration. 6. These observations suggest that TNBS might react with specific protein(s) on the outer surface of the presynaptic membrane and accelerate the exocytosis of synaptic vesicles.
Reversal potential of the calcium current in bull‐frog atrial myocytes.
Authors: Campbell D L , Giles W R , Hume J R , Noble D , Shibata E F
Published: December 2014

1. Voltage clamp recordings of the calcium current (ICa) in single myocytes which were enzymatically isolated from bull‐frog atrium show that a genuine reversal of the current flowing through Ca2+ channels can be recorded (ef. Reuter & Scholz, 1977; Lee & Tsien, 1982, 1984; Campbell, Giles & Shibata, 1988c). In normal 2.5 mM [Ca2+]0 Ringer solution this apparent reversal potential (Erev) is near +50 mV, a value well below the predicted thermodynamic Ca2+ equilibrium potential (ECa). 2. None the less, Erev shifts with variations in extracellular divalent ion concentrations (Ca2+, Sr2+ and Ba2+) according to the predictions of a Nernstian divalent cation electrode, i.e. approximately 29 mV per 10‐fold change in the external concentration of divalent ion. 3. The existing theoretical analysis of this Erev has been extended in order to clarify its interpretation with regard to the selectivity characteristics of ICa. 4. The apparent reversal potential is analysed using a form of the constant field equation which has been modified to include (i) simultaneous monovalent and divalent cation movements and (ii) the presence of a surface potential (V'). This equation can be solved to yield an explicit expression for Erev. The effects of V' on apparent permeability ratios for the Ca2+ channel Erev are demonstrated. 5. In combination, our experimental results and calculations suggest that: (i) previous estimates of V' which were used to describe permeability (P) ratios of Ca2+ channels in various cardiac preparations may be in error, (ii) in normal [Ca2+]o the PNa/PCa ratio is very small, and (iii) PCa/PK must be greater than 1000. An analysis of the relative selectivity of the channel for divalent cations compared to K+ shows that PCa greater than PSr greater than PBa, assuming that PK remains the same after the divalent substitutions. 6. The Ca2+ channel in bull‐frog atrial cells is thus much more selective for Ca2+ ions than had previously been estimated; in particular, inward flow of monovalent cations (e.g. Na+) through these channels does not contribute significantly to the observed ICa. The physiological implications of this high selectivity for Ca2+ ions are discussed.
Increase by trifluoperazine in calcium sensitivity of myofibrils in a skinned fibre from frog skeletal muscle.
Authors: Kurebayashi N , Ogawa Y
Published: December 2014

1. Since it has been demonstrated that trifluoperazine (TFP) increases the affinity for Ca2+ of troponin C as well as calmodulin, the effect of TFP was examined on the Ca2+‐induced tension in mechanically skinned fibres isolated from frog skeletal muscle and on Ca2+‐dependent ATPase activity of myofibrils from similar frog skeletal muscle. 2. Lower concentrations of TFP increased the Ca2+ sensitivity of myofibrils without a change in the maximum tension, giving rise to a less steep tension‐pCa relationship. This effect was reversible although thorough washes were necessary. The drug also enhanced myofibrillar ATPase activity, not only at low Ca2+ concentrations but also at saturating high Ca2+ concentrations. The increased affinity of troponin C for Ca2+ is difficult to accept as the sole explanation for the stimulatory effect of TFP. 3. Half of the maximum stimulating effect was obtained between 10 and 30 microM‐TFP, which is similar to the reported apparent inhibition constant (Ki) for calmodulin‐dependent enzyme reactions. However, the stimulating effect of TFP cannot be attributed to its inhibition of calmodulin because of the finding that this effect was independent of Ca2+. Earlier published results (e.g. Klee & Vanaman, 1982) also support this conclusion. 4. Studies on myofibrillar ATPase activity suggest that the stimulating effect of TFP is not identical in its underlying action with those of caffeine and quercetin, which are also known as Ca2+‐sensitizing drugs, having a similar eventual effect on tension development. 5. Higher concentrations of TFP decreased the maximum tension induced by high concentrations of Ca2+, while enhancing the tension in the presence of low concentrations of Ca2+. Analogous findings for ATPase activity were also made. TFP concentration for half the maximum depression was about 10 times higher than that for half the maximum stimulation. This suggests that different site(s) are involved in the stimulatory and inhibitory effects of TFP, although there may be some sites in common. 6. Discussion favours the stimulating effects of TFP as being caused considerably by the affected molecular interactions among myosin, actin, tropomyosin and troponin.
Corticotrophin‐releasing factor, vasopressin and pro‐opiomelanocortin mRNA responses to stress and opiates in the rat.
Authors: Lightman S L , Young W S
Published: December 2014

1. Rats underwent either: (1) acute or chronic morphine or naloxone administration; (2) simple morphine withdrawal or naloxone‐precipitated withdrawal in morphine‐dependent animals; or (3) stress from I.P. administration of hypertonic saline. 2. Quantitative in situ hybridization histochemistry was performed using synthetic oligonucleotide probes for corticotrophin‐releasing factor (CRF), vasopressin, pro‐opiomelanocortin (POMC), dynorphin, enkephalin and oxytocin mRNAs. The paraventricular and supraoptic nuclei were examined in all studies and the arcuate nucleus and pituitary gland in the acute withdrawal study. 3. Neither acute nor chronic morphine administration altered either (a) hypothalamic parvocellular or magnocellular CRF mRNA, or (b) anterior pituitary or pars intermedia POMC mRNA. 4. Naloxone‐precipitated morphine withdrawal resulted in a marked increase in parvocellular (but not magnocellular) CRF mRNA within 4 h and levels remained elevated through 24 h. There was no change in arcuate nucleus or pars intermedia POMC mRNA, but in the anterior pituitary there was a delayed increase, significant at 24 h. 5. Simple morphine withdrawal without the use of naloxone did not result in any change in CRF mRNA but there were increases in magnocellular vasopressin and dynorphin mRNA, presumably related to decreased water intake. 6. Intraperitoneal hypertonic saline stress also resulted in a marked accumulation of both parvocellular CRF and vasopressin mRNA without any concomitant change in magnocellular vasopressin mRNA. Increased translation of CRF mRNA was also evidenced by increased immunoreactive CRF detected by immunocytochemistry.
Luminal propionate‐induced secretory response in the rat distal colon in vitro.
Authors: Yajima T
Published: December 2014

1. The stimulatory action of propionate on colonic electrolyte transport and involvement of the enteric reflex in this was studied in vitro using an Ussing chamber in the rat. The short‐circuit current (Isc) and bidirectional fluxes of Na+ and Cl‐ were measured. Mucosa‐submucosa preparations, containing the submucosal nerve, from the distal colon were used in most cases. 2. Mucosal application of propionate caused transient increases in the transmural potential difference, with the mucosal side negative, Isc and conductance. Serosal application of the acid had no effect. 3. Adaptation of the Isc response occurred when the acid was applied to the bathing solution cumulatively without washing out the first dose. If tissues were washed and held more than 20 min before the next application, the response was almost completely restored. 4. The increase in Isc in response to propionate was concentration dependent, with a 50% effective concentration of approximately 7 x 10(‐5) M. 5. Two other short‐chain fatty acids (SCFAs), n‐butyrate and n‐valerate, but not acetate, increased Isc when added to the mucosal bathing solution. 6. Bumetanide (3 x 10(‐5) M) and the serosal chloride‐free condition, but not amiloride (10(‐4) M), inhibited the responses of Isc to propionate. Propionate‐stimulated Cl‐ secretion resulted mainly from an increase in unidirectional serosal‐to‐mucosal Cl‐ movement. Propionate did not affect the Na+ flux. 7. Tetrodotoxin (10(‐7) M), somatostatin (10(‐7) M) and hexamethonium (10(‐4) M) inhibited the propionate‐evoked increase in Isc by 40, 70 and 30%, respectively. 8. Atropine (10(‐5) M) also inhibited the Isc‐increase response to propionate more than 90%. 9. Pre‐treatment (2 min) of the mucosal surface with procaine (5 x 10(‐4) M) inhibited the propionate‐evoked increase in Isc by 90%. 10. The results suggest that luminal propionate transiently stimulated the colonic chloride secretory response that is not due to direct action on colonocytes, but due in large part to release of acetylcholine at neuro‐colonocyte junctions, probably via an enteric reflex involving a mucosal sensory mechanism, cholinergic motor nerves and submucosal ganglia.
Inositol trisphosphate‐induced membrane potential oscillations in Xenopus oocytes.
Authors: Berridge M J
Published: December 2014

1. The role of inositol 1,4,5‐trisphosphate (Ins1,4,5P3) in controlling the membrane potential oscillations induced by acetylcholine in Xenopus oocytes was investigated by studying the effect of injecting Ins1,4,5P3. 2. Perfusing Xenopus oocytes with low concentrations of acetylcholine (less than or equal to 1 x 10(‐7) M) induced regular oscillations in membrane potential. The frequency of these oscillations accelerated as the concentration of acetylcholine was increased. 3. Ionophoretic application of low doses of Ins1,4,5P3 stimulated membrane depolarization in the form of an initial brief spike which was followed by a burst of oscillations when the amount of Ins1,4,5P3 injected was increased. 4. When low doses of Ins1,4,5P3 were injected at 30 s intervals, there was rapid desensitization of the early response which recovered if the interval between injections was extended to 2 min or longer. 5. In comparison to the vegetal pole, the animal pole was much more sensitive to Ins1,4,5P3. This localization of Ins1,4,5P3 sensitivity in the animal pole may contribute to the electrical field which surrounds Xenopus oocytes. 6. A model is presented to explain these oscillations based on the phenomenon of calcium‐induced calcium release. It is proposed that Ins1,4,5P3 releases calcium from an Ins1,4,5P3‐sensitive pool which is then periodically taken up and released by an Ins1,4,5P3‐insensitive pool. It is the overloading of this Ins1,4,5P3‐insensitive pool which may provide the trigger to spontaneously release calcium back into the cytoplasm.
Guanine nucleotide‐ and inositol 1,4,5‐trisphosphate‐induced calcium release in rabbit main pulmonary artery.
Authors: Kobayashi S , Somlyo A P , Somlyo A V
Published: December 2014

1. The effects of activation of guanine nucleotide‐binding protein (G protein) by guanine nucleotides or sodium fluoride on the release of intracellular Ca2+ and on tension development were determined in chemically skinned strips of rabbit main pulmonary arteries (MPA). Ca2+ movements were monitored with Fura‐2, as the change in free Ca2+ concentration in the bath medium surrounding the skinned MPA. 2. Sodium fluoride or non‐hydrolysable analogues of GTP, guanosine 5'‐[gamma‐thio]triphosphate (GTP gamma S) and guanosine 5'‐[beta,gamma‐imido]triphosphate (GMP‐PNP), induced sustained and dose‐dependent contraction of skinned MPA. GTP (100 microM) induced transient contraction of skinned MPA. GTP gamma S did not contract intact MPA. We also confirmed that inositol 1,4,5‐trisphosphate (InsP3) released sufficient Ca2+ to induce contraction of skinned, but not intact, MPA. 3. Guanosine 5'‐[beta‐thio]diphosphate (GDP beta S), a non‐hydrolysable analogue of GDP that competitively inhibits the binding of guanine nucleotides to G proteins, inhibited the contractions induced by GTP gamma S. Neomycin (1 mM) inhibited the GTP gamma S‐induced contractions, but also, to a lesser extent, contractions induced by caffeine. 4. Depletion of Ca2+ from the sarcoplasmic reticulum (SR) or treatment with Triton X‐100 inhibited the GTP gamma S‐induced contractions. The effects of Ca2+ depletion was reversible, while that of Triton X‐100 was irreversible. GTP gamma S (up to 100 microM) had no apparent effect on the pCa‐tension curve of freeze‐glycerinated MPA. 5. GTP gamma S‐ or InsP3‐induced contractions occurred in the presence of 20 mM‐procaine, while this agent completely blocked the contraction induced by caffeine. 6. Both GTP gamma S and InsP3 induced an increase in the Fura‐2 fluorescence signal of the bath medium surrounding the skinned MPA, indicating that GTP gamma S releases intracellular Ca2+. The release of Ca2+ induced by GTP gamma S was inhibited by GDP beta S. 7. During the initial phasic contraction induced by GTP gamma S, added InsP3 had little or no additive effect, in contrast to its additive effect during the latter sustained contraction induced by GTP gamma S.(ABSTRACT TRUNCATED AT 400 WORDS)
Effect of acetylstrophanthidin on twitches, microscopic tension fluctuations and cooling contractures in rabbit ventricle.
Authors: Bers D M , Bridge J H
Published: December 2014

1. We have measured the effect of the aglycone acetylstrophanthidin (ACS) on twitches, cooling contractures and microscopic tension fluctuations in rabbit ventricular muscle. 2. Both developed twitches and cooling contractures are strengthened by applications of ACS in the range 1‐4 microM. This positive inotropy averages 150‐160% of control (zero ACS) in both twitches and cooling contractures. Cooling contracture magnitude is assumed to reflect the availability of sarcoplasmic reticulum (SR) Ca2+ for contraction (Bridge, 1986). We infer that ACS increases the availability of SR Ca2+ by enlarging SR Ca2+ stores and this may contribute to the positive inotropy. 3. However, twitches appear to increase at lower concentrations of ACS than those required to increase cooling contractures. This observation suggests that the initial ACS inotropy may be achieved without an increase in SR Ca2+. Furthermore, low doses of ACS produce positive inotropy in the presence of 10.0 mM‐caffeine where cooling contractures are abolished. This also suggests that positive inotropy occurs in the absence of SR Ca2+ accumulation. 4. Rest decay of both cooling contractures and twitches is significantly slowed in 4 and 8 microM‐ACS. We infer that ACS slows the rate of decline of SR Ca2+ available for contraction by slowing the rate at which Ca2+ is lost from the cell during rest. This suggests that ACS produces a net slowing of Ca2+ efflux during activity which in the absence of altered Ca2+ influx will result in net Ca2+ gain and presumably enlarged SR Ca2+ stores. 5. Increasing the concentration of ACS (6‐10 microM) results in a decline in developed twitch tension, total tension and an increase in rest tension. Measurement of microscopic tension fluctuations indicates that as developed twitches decline, the root mean square (r.m.s.) of the tension fluctuations increases in a reciprocal manner. This supports the suggestion of others that the decline in developed twitch tension and the appearance of tension fluctuations are causally related. 6. Although ACS (6‐10 microM) causes a decline in twitch tension, rapid cooling contractures remain elevated. We suggest that in the presence of Ca2+ oscillations the magnitude of cooling contractures reflects the sum of cytosolic Ca2+ and Ca2+ that is available for release. If microscopic tension fluctuations do represent Ca2+ moving between the SR and cytosol the sum of SR and cytosolic Ca2+ and hence cooling contracture might not decline.(ABSTRACT TRUNCATED AT 400 WORDS)
The dependence of motoneurone membrane potential on extracellular ion concentrations studied in isolated rat spinal cord.
Authors: Forsythe I D , Redman S J
Published: December 2014

1. Intracellular recordings from ninety‐nine motoneurones have been made in an in vitro hemisected spinal cord preparation. Their mean resting membrane potential in normal artificial cerebrospinal fluid (CSF) was ‐71 +/‐ 0.5 mV (+/‐ S.E.M.). The mean amplitude of the action potential was 84.0 +/‐ 1.4 mV (n = 50), and the mean input conductance was 101 +/‐ 7 nS (n = 49). 2. Both membrane potential and input conductance were sensitive to changes in [K+]o, [Na+]o, [Cl‐]o and [Ca2+]o. 3. Replacement of extracellular Ca2+ by Mn2+ resulted in less than 1 mV hyperpolarization and a decrease in input conductance from 102 +/‐ 7 to 93 +/‐ 6 nS (n = 15). 4. At high [K+]o (greater than 10 mM) the membrane potential followed the potential predicted by the Nernst equation for K+ ions with a slope of 58 mV per 10‐fold change in [K+]o. At low [K+]o (less than 10 mM) there was significant deviation from K+ equilibrium potential (EK). 5. [K+]i was found to be 106 mM when estimated from the reversal potential of the after‐hyperpolarization of the antidromic action potential. 6. The reversal potential of the recurrent inhibitory postsynaptic potential (IPSP) in normal CSF was used to calculate [Cl‐]i. This was 6.6 mM, which is less than would be expected if Cl‐ was passively distributed, indicating the presence of an outwardly directed Cl‐ pump. 7. Decreasing [Cl‐]o from control (134 mM) to 4 mM resulted in a depolarization of 6.9 +/‐ 0.9 mV and a decrease in input conductance from 102 +/‐ 5 to 90 +/‐ 5 nS (n = 14) in 3 mM [K+]o. 8. Decreasing [Na+]o from 156 to 26 mM by substitution with choline resulted in a 6.2 +/‐ 0.5 mV hyperpolarization and a decrease in input conductance from from 102 +/‐ 4 to 76 +/‐ 4 nS (n = 5) in 3 mM [K+]o. 9. The input conductances for Na+, Cl‐ and K+ at the resting potential were calculated. After allowing for a microelectrode leak conductance, the relative input conductances were gNa/gK = 0.13 and gCl/gK = 0.25.
Long‐term potentiation at nicotinic synapses in the rat superior cervical ganglion.
Authors: Briggs C A , McAfee D A
Published: December 2014

1. Nicotinic fast excitatory postsynaptic potentials (fast EPSPs) were recorded intracellularly from postganglionic neurones in the isolated rat superior cervical ganglion. 2. An hours‐long potentiation of the fast EPSP could be induced by brief tetanic stimulation of the preganglionic nerve (5 Hz for 5 s to 20 Hz for 20 s). While long‐term potentiation (LTP) can be detected in every ganglion by extracellular techniques, LTP was induced in only two‐thirds of the nicotinic synaptic responses. 3. Muscarinic blockade with atropine did not prevent LTP of the fast EPSP. 4. LTP of the fast EPSP did not correlate with changes in input resistance nor cell potential, as recorded in the soma. 5. The formation of nicotinic LTP appeared to depend upon stimulation of the nerve terminals. Non‐synaptic tetanic depolarization of the postganglionic neurone, effected by injecting depolarizing current pulses through the intracellular microelectrode, was not sufficient. LTP could be induced by synaptic tetani in two‐thirds of the same neurones. 6. The response to exogenous 1,1‐dimethyl‐4‐phenylpiperazinium (DMPP), a selective nicotinic agonist, was not increased during nicotinic synaptic LTP. This was true whether DMPP was applied by pressure‐ejection from an extracellular micropipette during intracellular recording, or by brief superfusion during sucrose‐gap recording of postganglionic responses. 7. Responses to exogenous acetylcholine and carbachol were increased during nicotinic LTP when these non‐selective cholinergic agonists were applied by pressure‐ejection during intracellular recording. However, the potentiation of the fast EPSP was always at least twofold greater than the potentiation of the response to these exogenous agonists. 8. Potentiation of the responses to acetylcholine and carbachol may have been due to long‐term enhancement of muscarinic responses. Thus, no postsynaptic basis for nicotinic LTP was uncovered in these studies.
Modification of L‐type calcium current by intracellularly applied trypsin in guinea‐pig ventricular myocytes.
Authors: Hescheler J , Trautwein W
Published: December 2014

1. The L‐type Ca2+ current was recorded in guinea‐pig ventricular myocytes by the patch clamp technique in the whole‐cell configuration. The modification of the current by intracellular application of proteases was studied. 2. During the first phase of action, trypsin, an endopeptidase, increased the amplitude of Ca2+ current about 3‐fold. 3. Thereafter, there was a drastic slowing of the inactivation time course of the enhanced Ca2+ current. The half‐time of inactivation increased from a control value of about 25 ms to values larger than 200 ms. 4. Cell dialysis with carboxypeptidase A, an exopeptidase, also enlarged the amplitude of Ca2+ current, but did not affect the kinetics of Ca2+ current. Leuaminopeptidase did not modify the Ca2+ current. 5. The hypothesis that Ca2+ channels are affected by the protease is supported by the fact that alterations of the extracellular Na+ or K+ concentration did not influence the modification of the membrane current. Another argument for the involvement of Ca2+ channels is that the modified membrane current could be blocked by inorganic and organic Ca2+ channel blockers (e.g. 10 microM‐Cd2+, 100 microM‐La3+ or 1 microM‐D600). 6. Although the actions of trypsin and maximal concentrations of isoprenaline on the amplitude of the Ca2+ current were not additive, the slowing of inactivation by trypsin occurred independently from beta‐adrenergic stimulation. 7. The effect of trypsin on the Ca2+ current could not be blocked by intracellular 5'‐adenylyl‐imidodiphosphate (AMP‐PNP) or Rp‐adenosine 3'5'‐monothionophosphate (Rp‐cAMPS), both of which are known to suppress the cyclic AMP‐dependent phosphorylation of the Ca2+ channel. 8. It was concluded that trypsin may directly modify the membrane protein which forms the Ca2+ channel. Since the increment in peak Ca2+ current resembled the action of cyclic AMP‐dependent phosphorylation, it may be related to the removal of a ‘chemical’ inactivation gate which is normally controlled by phosphorylation. The slowing of the time course of Ca2+ current inactivation by trypsin could be due to a modification of the voltage‐dependent inactivation gate. Alternatively, the endopeptidase might remove an internal Ca2+ binding site normally responsible for Ca2+‐dependent inactivation.
A submucosal mechanism for catecholamine‐induced increases in fluid absorption in rabbit ileum in vitro.
Authors: Ahsan M A , Naftalin R J , Smith P M
Published: December 2014

1. The effects of clonidine and dopamine on water movements across the mucosal and serosal surfaces of rabbit ileum have been investigated using a high‐resolution method for monitoring water flows in vitro. 2. Theophylline (10 mM) and carbamyl choline (10 microM) caused a reduction in fluid inflow across the mucosal surface and a smaller decrease in fluid outflow across the serosal surface. Addition of the alpha 2‐adrenergic agonist clonidine or dopamine fully reversed the theophylline, or carbamyl choline‐induced decrease in mucosal inflow in a dose‐related manner. 3. The effects of clonidine on mucosal inflow are blocked by the alpha 2‐adrenergic antagonist, yohimbine. Yohimbine was much less effective than pimozide or d‐butaclamol in blocking the effect of dopamine on mucosal inflow. These findings support the view that there are separate alpha 2‐adrenergic and dopaminergic receptors. 4. The hydraulic conductance (Lp) of the serosal surface was measured directly from the change in serosal exit flow following addition of 2 mosmol kg‐1 of polyethylene glycol (molecular mass 20,000 Da) to the serosal bathing solution. Theophylline reduced the Lp by 35%. Clonidine (1 microM) added to theophylline‐treated tissues increased the Lp by 66%. This effect was prevented by yohimbine (1 microM). 5. The effects of theophylline, clonidine and dopamine on the permeability of the mucosal and serosal surfaces of the tissue to [3H]mannitol were measured. These showed that theophylline increased the rate of labelled mannitol loss across the mucosal surface but reduced the mannitol permeability across the serosal surface. This latter effect was reversed by clonidine and dopamine. 6. Changes in transepithelial electrical potential difference (PD), short‐circuit current and resistance were monitored. Theophylline caused a rapid increase in PD and short‐circuit current and a slower increase in resistance. Clonidine (5 microM) reversed the effects on PD and resistance but was without significant effect on short‐circuit current. The results suggest that a major component of secretagogue‐induced reduction in fluid transport in vitro is due to mechanical changes in the submucosa, probably induced by modulation of neurotransmitter release within the tissue.