bluuebunny/crossref_metadata_2025 huggingface dataset:
Activation kinetics of calcium currents in bull‐frog sympathetic neurones.
Authors: Sala F
Published: December 2014

1. Calcium currents were recorded in dissociated bull‐frog sympathetic neurones (BSNs) through patch pipettes using discontinuous voltage clamp. Activation kinetics were examined by analysing turn‐on and turn‐off currents. 2. After short depolarizing pulses turn‐off tail currents were fitted with the sum of two exponentials. The fast component (time constant, tau approximately 240 microseconds at ‐40 mV) was undoubtedly due to the closure of calcium channels. The significance of a small and slower component is discussed. 3. Neither activation nor deactivation time courses changed as channels inactivated during progressively longer pulses or when the holding potential was less negative. No specific component was selectively suppressed by these manipulations. 4. Steady‐state activation of the Ca2+ current was described by the Boltzmann distribution raised to the second power. Currents had an apparent threshold at ‐30 mV and were half‐activated at +5 mV. 5. Calcium current turned on following m2 kinetics throughout the range of activation. The slowest time constant was around 1.2 ms between 0 and +10 mV. Turn‐on was faster at negative or more positive potentials. 6. The time course of decay of tail currents became progressively faster at more negative potentials. 7. The instantaneous current‐voltages (I‐V) curve was obtained from tail current measurements and fitted by a modified constant‐field equation. 8. The measured peak I‐V curve could be reconstructed from the activation curve and the instantaneous I‐V curve. 9. The activation kinetics of the calcium current in BSNs were consistent with the existence of a single kinetic class of channels and can be described with a simple m2 Hodgkin‐Huxley model.
Neurokinin A increases short‐circuit current across rat colonic mucosa: a role for vasoactive intestinal polypeptide.
Authors: Tien X Y , Wallace L J , Kachur J F , Won‐Kim S , Gaginella T S
Published: December 2014

1. Neurokinin A (NKA) is a mammalian tachykinin distributed principally in the nervous system, including the myenteric innervation of the gut. 2. NKA may be involved in neurogenic inflammation and as a modulatory factor in the diarrhoea associated with mucosal inflammation of inflammatory bowel disease (ulcerative colitis). 3. We evaluated the effect of NKA on the short‐circuit current ISC, assumed to reflect electrogenic chloride secretion, across muscle‐stripped rat colonic mucosa mounted in Ussing chambers. 4. Serosal addition of NKA produced a concentration‐dependent (0.1‐100 nM) increase in ISC with an EC50 (half‐maximal effective concentration) value of 7.5 nM. The maximum (mean +/‐ S.E.M.) increase in ISC (microA/cm2) for NKA was 111 +/‐ 10. 5. Tetrodotoxin (0.5 microM) and bumetanide (10 microM), but not atropine (1.0 microM), hexamethonium (100 microM) or pyrilamine (10 microM), significantly inhibited NKA‐induced increases in ISC. 6. The response to NKA was attenuated by 45 min pre‐treatment with antisera raised against vasoactive intestinal polypeptide (VIP). Moreover, prior desensitization to VIP attenuated the effect of NKA. 7. These studies suggest that NKA increases ISC in rat colon, in part, through a non‐cholinergic neural mechanism involving VIP.
Sodium‐ and calcium‐dependent conductances of neurones in the zebra finch hyperstriatum ventrale pars caudale in vitro.
Authors: Kubota M , Saito N
Published: December 2014

1. Intracellular recordings were made from zebra finch hyperstriatum ventrale pars caudale (HVc) neurones in in vitro slice preparations. 2. Small depolarizing current pulses elicited tonic firing while relatively large current pulses elicited an initial high‐frequency burst followed by tonic firing. Time‐dependent and fast‐activated inward rectification were observed when a hyperpolarizing current pulse was applied. 3. Fast action potentials were abolished by tetrodotoxin (TTX, 0.5 micrograms/ml). When Co2+ (2.2‐2.4 mM) was added to a low‐Ca2+ (0‐0.2 mM) solution, a plateau potential was elicited by a small depolarizing current pulse. These plateau potentials outlasted the applied current pulse and were abolished by TTX (0.5 micrograms/ml). 4. A long‐lasting after‐hyperpolarization (AHP), with a duration of tens of seconds, followed long trains of repetitive firing in control solution. Even in a low‐Ca2+ (0‐0.2 mM) solution containing Co2+ (2.2‐2.4 mM) the long‐lasting AHPs followed plateau potentials and were associated with an increase in input conductance. The long‐lasting AHP, as well as the plateau potential, was blocked by TTX (0.5 micrograms/ml). 5. While TTX abolished fast action potentials, two types of active responses with different thresholds were elicited by depolarizing current in the presence of TTX (0.5 micrograms/ml), particularly when tetraethylammonium (5 mM) was also added to the solution. Both the low‐threshold spike (LTS) and the high‐threshold spike (HTS) were abolished by Co2+ (2.2‐2.3 mM). 6. When Ba2+ (0.5‐1 mM) was added to a solution containing TTX (0.5 micrograms/ml), an LTS elicited by a depolarizing current pulse became larger in amplitude. At membrane potentials more positive than ‐55 mV, an HTS, but not the LTS, was elicited. 7. These results suggest that HVc neurones have three Na(+)‐dependent conductances: a fast Na+ conductance responsible for the fast spike, a persistent Na+ conductance responsible for the plateau potential, and a Na(+)‐activated K+ conductance responsible for the long‐lasting AHP; and that they have two Ca(2+)‐dependent conductances: a low‐threshold Ca2+ conductance responsible for the LTS, and a high‐threshold Ca2+ conductance responsible for the HTS.
The role of adenosine in the respiratory and cardiovascular response to systemic hypoxia in the rat.
Authors: Neylon M , Marshall J M
Published: December 2014

1. In rats anaesthetized with Saffan we have studied the effects of the adenosine receptor antagonists, theophylline and 8‐phenyltheophylline, upon the respiratory and cardiovascular responses evoked by 5 min periods of systemic hypoxia. 2. In the group of animals that were to receive theophylline (15 mg kg‐1 i.v.), arterial O2 pressure (Pa,O2) fell from 83 +/‐ 2 mmHg during air breathing to 38 +/‐ 3 or 34 +/‐ 3 mmHg during the 5th minute of two different control periods of hypoxia, while in the group that were to receive 8‐phenyltheophylline (10 mg kg‐1 i.v.), Pa,O2 fell from 83 +/‐ 1 to 53 +/‐ 2 mmHg. Neither drug significantly altered the levels of Pa,O2 reached during hypoxia. 3. During the control periods of hypoxia respiration increased, but the increase evoked at the 5th minute was significantly less than that evoked at the 2nd minute of hypoxia. This secondary waning of the hyperventilation was abolished by both drugs. 4. Similarly, both drugs attenuated the tendency for the hypoxia‐induced tachycardia to wane between the 2nd and 5th minute. 5. Further, both drugs substantially reduced both the hypoxia‐induced fall in arterial pressure and the increases in vascular conductance in hindlimb muscle, carotid vasculature and kidney. 6. Thus, we propose that in the rat the release of adenosine by hypoxic tissues makes a major contribution to the secondary decrease in respiration and heart rate that occurs during systemic hypoxia and to the accompanying vasodilatation in muscle and fall in arterial pressure. The effects of the adenosine antagonists on the carotid and renal vasculature are more equivocal and may be partly explained as a smaller autoregulatory dilatation to a smaller fall in systemic arterial pressure. 7. These results and proposals are discussed in relation to the conditions that are known to cause release of adenosine and in relation to its known effects upon the respiratory and cardiovascular systems.
Relation between muscarinic receptor cationic current and internal calcium in guinea‐pig jejunal smooth muscle cells.
Authors: Pacaud P , Bolton T B
Published: December 2014

1. The action of carbachol, which activates muscarinic receptors, was studied in single patch‐clamped cells where free internal calcium concentration in the cell (Cai2+) was estimated using the emission from the dye Indo‐1. Cells were dialysed with potassium‐free caesium solution to block any Ca(2+)‐activated K(+)‐current. 2. Carbachol applied to the cell evoked an initial peak in Cai2+ followed by a smaller sustained rise (plateau) upon which several oscillations in Cai2+ were often superimposed; the changes in inward, cationic current (icarb) followed changes in Cai2+ closely. Calcium entry blocker did not affect these responses. 3. The initial peak in Cai2+ produced by carbachol was due to calcium store release: it was essentially unchanged at +50 mV, and abolished by prior application of caffeine (10 mM) to the cell or by inclusion of heparin (which blocks D‐myoinositol 1,4,5‐trisphosphate receptors) in the pipette. In contrast, the rise in Cai2+ produced by ATP in rabbit ear artery smooth muscle cells was unaffected by caffeine or heparin as it was due to calcium entry into the cell. 4. The later sustained rise (plateau) in Cai2+ produced by carbachol was due to the entry of calcium into the cell down its electrochemical gradient as it was affected by changing the cell membrane potential or the calcium concentration of the bathing solution. As the sustained rise in Cai2+ produced by caffeine had similar properties, it was suggested that depletion of calcium stores can evoke an increased calcium entry into the cell through some pathway. 5. The cationic current evoked by carbachol was strongly dependent on Cai2+. It was small if any rise in Cai2+ due to calcium store release was prevented by the inclusion of heparin in the pipette solution and increased greatly if calcium entry was provoked through voltage‐dependent channels by applying a depolarizing pulse or if calcium was released from stores by caffeine. 6. In the longitudinal muscle of guinea‐pig small intestine, activation of muscarinic receptors by carbachol results in the opening of cationic channels; the resulting depolarization increases the frequency of action potential discharge and this determines the degree of contraction. Muscarinic receptor activation opens cationic channels by two mechanisms: release of stored calcium increases Cai2+ and this strongly potentiates a primary mechanism which may involve a G‐protein.
Resting and ADP‐evoked changes in cytosolic free sodium concentration in human platelets loaded with the indicator SBFI.
Authors: Sage S O , Rink T J , Mahaut‐Smith M P
Published: December 2014

1. Cytosolic free Na+ concentration, [Na+]i, was investigated in human platelets loaded with the fluorescent indicator SBFI (sodium‐binding benzofuran isophthalate). 2. SBFI fluorescence from platelet suspensions was measured at excitation wavelengths of 340 and 385 nm and the 340/385 nm fluorescence ratio was calibrated in terms of [Na+]i in situ. [Na+]i was set to known values by resuspending cells in media with various [Na+], in the presence of the Na(+)‐K+ ionophore, gramicidin. 3. Basal free [Na+]i was 5.5 +/‐ 0.3 mM (n = 50). This is considerably lower than estimates of total platelet Na+, suggesting that much intracellular Na+ is sequestered or bound. 4. ADP (40 microM) evoked a rise in [Na+]i from 6.4 +/‐ 0.7 to 18.3 +/‐ 1.1 mM (n = 8). The ADP‐evoked rise in [Na+]i was abolished when external Na+ was replaced with N‐methyl‐D‐glucamine. This indicates that the rise in [Na+]i was due to Na+ entry. 5. In platelets loaded with the fluorescent pH indicator, BCECF, 40 microM‐ADP was shown to evoke a fall in cytosolic pH (pHi) from 7.21 +/‐ 0.03 to 7.12 +/‐ 0.03 (n = 10). Three minutes after ADP addition pHi had only recovered to 7.15 +/‐ 0.03. The recovery was dependent on external Na+, suggesting it was mediated by Na(+)‐H+ exchange. However, this would only account for an increase in [Na+]i of approximately 0.5 mM, indicating most of the ADP‐evoked Na+ entry occurred by other mechanisms. 6. Stopped‐flow fluorimetry showed that the ADP‐evoked rise in [Na+]i commenced without measurable delay and peaked within 1 s. The initial kinetics were thus similar to those reported for ADP‐evoked rises in [Ca2+]i. 7. Cell‐attached patch‐clamp recordings showed that ADP evoked single‐channel inward currents when included in the pipette‐filling solution. The currents were similar whether Ca2+ was present or absent from the pipette. The slope conductance was 11 pS in the presence of external Ca2+ and 10 pS in its absence. Current‐voltage relationships were similar and the reversal potentials were close to 0 mV under both conditions. 8. SK & F 96,365 (20 microM), a blocker of receptor‐mediated Ca2+ entry in several non‐excitable cells, blocked the ADP‐evoked rise in [Na+]i. This compound has been shown to only partly block the biphasic ADP‐evoked rise in [Ca2+]i, being selective for the fast, receptor‐operated phase of entry. 9. These data suggest that ADP rapidly activates a channel in that platelet plasma membrane which is permeable to Na+ and divalent cations.
Response of the medullary respiratory network of the cat to hypoxia.
Authors: Richter D W , Bischoff A , Anders K , Bellingham M , Windhorst U
Published: December 2014

1. The effect of systemic hypoxia was tested in anaesthetized, immobilized, thoracotomized and artificially ventilated cats with peripheral chemoreceptor afferents either intact or cut. Extracellular recordings from different types of medullary respiratory neurones and intracellular recordings from stage 2 expiratory neurones were made to determine the hypoxia‐induced changes in neuronal discharge patterns and postsynaptic activity as an index for the disturbances of synaptic interaction within the network. 2. The general effect of systemic hypoxia was an initial augmentation of respiratory activity followed by a secondary depression. In chemoreceptor‐denervated animals, secondary depression led to central apnoea. 3. The effects of systemic hypoxia were comparable with those of cerebral ischaemia following occlusion of carotid and vertebral arteries. 4. In chemoreceptor‐denervated animals, all types of medullary respiratory neurones ceased spontaneous action potential discharge during hypoxia. 5. Reversal of inhibitory postsynaptic potentials (IPSPs) and/or blockade of IPSPs was seen after 2‐3 min of hypoxia. 6. During hypoxia, the membrane potential of stage 2 expiratory neurones showed a slight depolarization to ‐45 to ‐55 mV and then remained stable. 7. The neurone input resistance increased initially and then decreased significantly during central apnoea. 8. Rhythmogenesis of respiration was greatly disturbed. This was due to blockade of IPSPs and, in some animals, to more complex disturbances of phase switching from inspiration to expiration. 9. Central apnoea occurred while respiratory neurones were still excitable as shown by stimulus‐evoked orthodromic and antidromic action potentials. 10. The results indicate that the medullary respiratory network is directly affected by energy depletion. There is indication for a neurohumoral mechanism which blocks synaptic interaction between respiratory neurones in chemoreceptor‐intact animals.
Gating of L‐type Ca2+ channels in embryonic chick ventricle cells: dependence on voltage, current and channel density.
Authors: Mazzanti M , DeFelice L J , Liu Y M
Published: December 2014

1. L‐type calcium channels in embryonic chick heart ventricle have voltage‐dependent, time‐variant kinetics when they conduct inward currents carried by 20 mM‐Ba2+. Depolarizing the membrane from ‐20 to 20 mV increases mean open time from 1.4 to 4.2 ms. Mean open time increases monotonically with voltage. The single‐channel conductance, 18 +/‐ 2 pS, is approximately linear over this voltage range, and the extrapolated reversal potential is 38 +/‐ 5 mV. 2. In cell‐attached patches with five or more L‐type Ca2+ channels in the patch, the currents elicited by 500 ms depolarizing steps, from a ‐80 mV holding potential, inactivate rapidly and have large tail currents. In the same patch, currents from a ‐40 mV holding potential are smaller, inactivate more slowly, and have practically no tail currents. 3. In cell‐attached patches containing one of two L‐type Ca2+ channels, currents from ‐80 or ‐40 mV are virtually identical, and they are similar to the currents from multichannel patches held at ‐40 mV. 4. The voltage‐dependent, time‐variant kinetics of individual L‐type Ca2+ channels are unaltered if the patch is removed from the cell and forms an inside‐out configuration. In these experiments the internal membrane was bathed with an artificial, intracellular‐like solution containing no phosphorylating enzymes or substrates. 5. Cells bathed in 20 mM‐Ba2+ solutions and held at ‐80 mV have currents with an early phase that inactivates in tens of milliseconds, a late phase that inactivates in hundreds of milliseconds, and a large, slow tail current. Currents from ‐40 mV have only the late phase and practically no tails. However, if the maximum current is less than 0.1 pA pF‐1, records from either ‐80 or ‐40 mV are virtually identical, and they are similar to currents from cells with higher channel density held at ‐40 mV. Furthermore, if cells are stimulated before full recovery from inactivation, the reduced current is accompanied by slower inactivation. 6. Whole‐cell currents in 1.5 mM‐Ca2+ solutions are entirely abolished by addition of 20 microM‐nifedipine, and they are enhanced 2‐3 times by addition of 30 microM‐cyclic AMP and 3 mM‐ATP to the whole‐cell recording electrode. The whole‐cell currents in 20 mM‐Ba2+ solutions are also completely blocked by 20 microM‐nifedipine, regardless of kinetics or holding potential. Thus, by definition, the cells we are studying contain only L‐type channels.(ABSTRACT TRUNCATED AT 400 WORDS)
Spontaneous release of nitric oxide inhibits electrical, Ca2+ and mechanical transients in canine gastric smooth muscle.
Authors: Ozaki H , Blondfield D P , Hori M , Publicover N G , Kato I , Sanders K M
Published: December 2014

1. In canine antrum, rhythmic electrical activity consists of a rapid upstroke phase followed by a plateau depolarization. In response to slow waves, cytosolic Ca2+ ([Ca2+]cyt) and tension increased. 2. Addition of sodium nitroprusside (SNP, 0.5 microM) decreased the amplitude of the plateau phase of slow waves without significant effects on the upstroke depolarization. SNP also inhibited changes in [Ca2+]cyt and tension associated with the plateau potential. SNP induced a negative chronotropic effect at concentrations above 0.1 microM. 3. Similar to the effects of SNP, illumination of muscles during slow waves with ultraviolet (UV) light caused premature repolarization. UV illumination is known to release NO in some tissues. 4. L‐NG‐monomethyl‐arginine (L‐NMMA, 300 microM), Methylene Blue (MB, 5 microM) and oxyhaemoglobin (oxy‐Hb, 5 microM) increased the force of contractions. In contrast, L‐arginine (L‐Arg, 300 microM) decreased contractile force and antagonized the effects of L‐NMMA. 5. During the upstroke phase, SNP caused a small reduction in [Ca2+]cyt and a large reduction in force, suggesting that SNP caused a decrease in Ca2+ sensitivity. 6. In muscles permeabilized by alpha‐toxin, cyclic GMP (100 microM) and UV illumination inhibited Ca(2+)‐induced contraction (at pCa 5.5). 7. These data suggest that NO or NO‐related compounds are spontaneously released in gastric muscles. These agents have two effects on excitation‐contraction coupling: (i) inhibition (directly and/or indirectly) of the voltage‐dependent Ca2+ channels that participate in the plateau phase of slow waves, and (ii) reduction in the Ca2+ sensitivity of the contractile element.
Effects of 2‐amino‐4‐phosphonobutyric acid on cells in the distal layers of the tiger salamander's retina.
Authors: Hare W A , Owen W G
Published: December 2014

1. We studied the effects of 2‐amino‐4‐phosphonobutyric acid (APB) on the response properties of rods, horizontal cells and bipolar cells in the isolated, perfused retina of the tiger salamander, Ambystoma tigrinum. A concentration of 100 microM was found to be sufficient to elicit maximal effects. 2. Rods hyperpolarized slightly upon exposure to 100 microM‐APB and their response amplitudes were slightly reduced. The amplitude of the cone‐generated component of the rod's response to 700 nm light was not significantly affected by APB. 3. Horizontal cells hyperpolarized by 2‐5 mV upon exposure to 100 microM‐APB. The rod‐driven component of the horizontal cell response increased in amplitude while the cone‐driven component decreased in amplitude. APB thus causes an increase in voltage gain between rods and horizontal cells and a decrease in cone/horizontal cell gain. These findings can be explained in terms of an APB‐induced reduction in transmitter release from the cones. 4. APB at a concentration of 100 microM caused an increase in the length constant of the horizontal cell syncytium. Our analysis shows this to be due primarily to a 50% reduction in the coupling impedance between the cells of the syncytium. 5. The effects of APB on off‐centre bipolar cells were qualitatively similar to those on horizontal cells. APB increased the amplitudes of rod‐driven responses and reduced those of cone‐driven responses. The length constants, both of the receptive field centre and of the surround, were increased and the strength of the surround relative to the centre was reduced by about 20%. 6. APB abolished the depolarizing light responses of the receptive field centres of on‐centre bipolar cells. A hyperpolarizing response remained whose spatial properties were similar to those of the receptive field surround. We believe this response to reflect a direct (feedforward) input to on‐centre bipolar cells from horizontal cells.
Central changes in processing of mechanoreceptive input in capsaicin‐induced secondary hyperalgesia in humans.
Authors: Torebjörk H E , Lundberg L E , LaMotte R H
Published: December 2014

1. Capsaicin, the algesic substance in chilli peppers, was injected intradermally in healthy human subjects. A dose of 100 micrograms given in a volume of 10 microliters caused intense pain lasting for a few minutes after injection and resulted in a narrow area of hyperalgesia to heat and a wide surrounding area of hyperalgesia to mechanical stimuli (stroking) lasting for 1‐2 h. 2. Nerve compression experiments with selective block of impulse conduction in myelinated (A) but not in unmyelinated (C) fibres indicated that afferent signals in C fibres contributed to pain from capsaicin injection and to heat hyperalgesia, whereas conduction in afferent A fibres was necessary for the perception of mechanical hyperalgesia. 3. Electrical intraneural microstimulation normally eliciting non‐painful tactile sensations was accompanied by pain when the sensation was projected to skin areas within the region of mechanical hyperalgesia induced by capsaicin injection. 4. The threshold for pain evoked by intraneural microstimulation was reversibly lowered and pain from suprathreshold stimulation was exaggerated during the period of mechanical hyperalgesia, regardless of lidocaine anaesthesia of the cutaneous innervation territory of the stimulated fibres. 5. The results indicate that hyperalgesia to stroking on a skin area surrounding a painful intradermal injection of capsaicin is due to reversible changes in the central processing of mechanoreceptive input from myelinated fibres which normally evoke non‐painful tactile sensations.
Dopamine D2 receptor stimulation differentially affects voltage‐activated calcium channels in rat pituitary melanotropic cells.
Authors: Keja J A , Stoof J C , Kits K S
Published: December 2014

1. Whole‐cell voltage clamp recordings were made from 141 rat pituitary melanotropic cells in short‐term, serum‐free, primary culture. The effects of the dopamine D2 receptor agonist, LY 171555, on sodium, potassium and barium currents were investigated. 2. Application of 1 microM‐LY 171555 did not affect the inward sodium and outward potassium currents. 3. Application of LY 171555 reversibly inhibited barium currents, with the strongest inhibition on the early inward current. The effect was dose dependent (IC50 = 4 x 10(‐8) M), maximal inhibition of the total current was 30% and the LY 171555‐induced block (1 microM) was reversibly antagonized by (+/‐)sulpiride (4 microM). 4. Using barium‐selective saline solutions, different types of barium current (T, N, and two L components) were identified on the basis of their voltage‐dependent kinetics. Their relative amplitudes differed between cells. 5. The T‐type current activated at potentials positive to ‐60 mV, reaching peak amplitude between ‐20 and ‐10 mV. At ‐30 mV, this current was inhibited up to 30% by 1 microM‐LY 171555. The time constants of activation (10‐3 ms) and inactivation (50‐20 ms) as well as the voltage dependence of inactivation (potential of half‐maximal inactivation (H), ‐61 mV; slope factor (S), 4.9 mV) were not affected by LY 171555 application. 6. A rapidly inactivating (time constants 100‐50 ms), high threshold current component was identified as an N‐type current. This current activated at command potentials positive to ‐30 mV and reached a maximal amplitude at +10 mV. The steady‐state inactivation was described by a single Boltzmann equation with H = ‐65 mV and S = 11.7 mV. Application of 1 microM‐LY 171555 completely suppressed this current. 7. The slowly inactivating (time constants > 1500 ms), high‐threshold, L‐type current displayed the same voltage dependence of activation as the N current. The voltage dependence of inactivation was modelled by the sum of two Boltzmann equations (L1: H1 = ‐45 mV, S1 = 13.0 mV; L2:H2 = ‐11 mV, S2 = 6.0 mV), indicating the existence of two L channel populations. Neither time course, nor voltage dependence of inactivation were influenced by LY 171555. However, LY 171555 induced a slow‐down in the time course of activation, which necessitated the use of two time constants to model the activation kinetics. One of these (approximately 2 ms) was also observed under control conditions.(ABSTRACT TRUNCATED AT 400 WORDS)
A study of glycogen depletion and the fibre‐type composition of cat skeleto‐fusimotor units.
Authors: Barker D , Scott J J , Stacey M J
Published: December 2014

1. We have used the glycogen‐depletion technique, combined with myofibrillar ATPase (mATPase) staining for muscle fibre type, to study the fibre‐type composition of four skeleto‐fusimotor (beta) units in cat peroneus tertius, namely, one beta dynamic (beta d) unit and three beta static (beta s) units. 2. Depletion of glycogen was observed in serial cross‐sections of thirty‐four beta‐unit extrafusal muscle fibres of various types traced from origin to insertion. No fibre was depleted of glycogen throughout its length; depletion was restricted to a number of zones, usually about five. Oxidative (type I) and oxidative‐glycolytic (type IIA) fibres were depleted for a significantly greater proportion of their total length than glycolytic IIB fibres. 3. The fibre‐type composition of the beta d unit was determined by tracing its fibres from end to end. The muscle unit consisted of one intrafusal bag1 fibre and ninety‐three extrafusal muscle fibres comprising seventy‐six type I fibres, eleven IIC fibres, and six fibres that changed from IIC to I during the course of their length (IIC/I fibres). The extrafusal fibre‐type composition was thus 81.7% I plus 18.3% IIC and IIC/I. 4. The three beta s units (beta s1, beta s2, beta s3) were all fast‐contracting and fatigued rapidly. Identification of their extrafusal fibre types, made in 1 mm2 areas sampled from different parts of each unit, gave mixed compositions as follows: beta s1, IIB + 6.7% IIA; beta s2, IIB + 5.8% IIA; beta s3, IIB + 29.9% IIA. The intrafusal component of each unit included either one or two long chain fibres. 5. In a discussion of the results, the fact that the continuous stimulation of extrafusal muscle fibres does not deplete them of glycogen throughout their length is examined in relation to the work of others who have assumed that it did. With regard to the finding of mixed extrafusal fibre types in the beta units, a distinction is drawn between minimal (around 5%) and moderate mixing. It is suggested that minimal mixing may occur in any motor unit as the outcome of endplate degeneration with foreign replacement, but that moderate mixing indicates an on‐going process of conversion from one fibre type to another which in the adult may prove to occur only among beta units.
Interhemispheric inhibition of the human motor cortex.
Authors: Ferbert A , Priori A , Rothwell J C , Day B L , Colebatch J G , Marsden C D
Published: December 2014

1. Using two magnetic stimulators, we investigated the effect of a conditioning magnetic stimulus over the motor cortex of one hemisphere on the size of EMG responses evoked in the first dorsal interosseous (FDI) muscle by a magnetic test stimulus given over the opposite hemisphere. 2. A single conditioning shock to one hemisphere produced inhibition of the test response evoked from the opposite hemisphere when the conditioning‐test interval was 5‐6 ms or longer. We shall refer to this as interhemispheric inhibition. However, the minimum latency of inhibition observed using surface EMG responses may have underestimated the true interhemispheric conduction time. Single motor unit studies suggested values 4‐7 ms longer than the minimum interval observed with surface EMG. 3. Interhemispheric inhibition was seen when the test muscle was active or relaxed. Increasing the intensity of the conditioning stimulus increased the duration of inhibition: increasing the intensity of the test stimulus reduced the depth of inhibition. 4. The conditioning coil had to be placed on the appropriate area of scalp for inhibition to occur. The effect of the conditioning stimulus was maximal when it was applied over the hand area of motor cortex, and decreased when the stimulus was moved medial or lateral to that point. 5. The inhibitory effect on the test stimulus probably occurred at the level of the cerebral cortex. In contrast to the inhibition of test responses evoked by magnetic test stimuli, test responses evoked in active FDI by a small anodal electric shock were not significantly inhibited by a contralateral magnetic conditioning stimulus. Similarly, H reflexes in relaxed forearm flexor muscles were unaffected by conditioning stimuli to the ipsilateral hemisphere. However, inhibition was observed if the experiment was repeated with the muscles active.
Stretch‐activated anion currents of rabbit cardiac myocytes.
Authors: Hagiwara N , Masuda H , Shoda M , Irisawa H
Published: December 2014

1. Stretch‐activated anion currents were studied in sino‐atrial and atrial cells using the whole‐cell patch clamp technique. With continuous application of positive pressure (5‐15 cmH2O) through the patch clamp electrode, the cell was inflated and the membrane conductance was increased. 2. Voltage clamp steps revealed that the stretch‐activated currents had time‐independent characteristics. The increase in membrane conductance was reversible on subsequent application of negative pressure to the electrode. 3. The reversal potential of the stretch‐activated currents was shifted by 60 mV for a 10‐fold change in intracellular Cl‐ concentration, while it was unaffected by replacement of Na+ in the extracellular solution by N‐methyl‐D‐glucamine. Cell superfusion with Cl(‐)‐deficient solution (10 mM Cl‐) reduced the amplitude of outward current. These findings indicate that the stretch‐activated conductance is Cl‐ selective. 4. The sequence of anion permeability through the stretch‐activated conductance was determined to be I‐(1.7) > NO3‐(1.5) > Br‐(1.2) > Cl‐(1.0) > and F‐(0.6). SCN‐ appeared to be more permeant than I‐. 5. The stretch‐activated conductance was reduced by the Cl‐ channel blockers, 4,4'‐dinitrostilbene‐2,2'‐disulphonic acid disodium salt, 4‐acetamido‐4'‐isothiocyanatostilbene‐2,2'‐disulphonic acid or anthracene‐9‐carboxylate (9‐AC). Administration of furosemide or bumetanide had no effect. 6. The stretch‐activated Cl‐ current was recorded even though intracellular Ca2+ ions were chelated by including 10 mM EGTA in the pipette solution. Neither the specific peptide inhibitor of cyclic AMP‐dependent protein kinase (50 microM), nor the non‐selective blocker of protein kinases, H‐7 (20 microM), was effective in reducing the stretch‐activated Cl‐ current, suggesting that the stretch‐activated Cl‐ current is a novel type of cardiac Cl‐ current, which shows a different modulatory mechanism from that of other cardiac Cl‐ currents.
The actions of baclofen on neurones and synaptic transmission in the nucleus tractus solitarii of the rat in vitro.
Authors: Brooks P A , Glaum S R , Miller R J , Spyer K M
Published: December 2014

1. Intracellular and whole‐cell patch recordings were made from sixty‐seven neurones located in the nucleus tractus solitarii (NTS) in transverse slices of rat brainstem. 2. Baclofen at concentrations of 2‐20 microM caused hyperpolarization from normal resting membrane potentials (Vm). This response was associated with a decrease in input resistance (Rm) tested by current pulses in discontinuous current clamp mode when membrane potential was restored to control level by current injection. In single electrode discontinuous voltage clamp mode, baclofen at these concentrations caused a small (< 50 pA) outward current associated with increased membrane conductance measured by voltage steps from holding potentials (Vh) of ‐50 or ‐60 mV. Current‐voltage relations at these Vhs and the results of varying Vh between ‐50 and ‐110 mV during responses to baclofen gave a reversal potential of ‐73 mV. The amplitudes of baclofen responses were related to K+ concentration tested by comparing responses in media containing 1‐24 mM extracellular K+, indicating that postsynaptically baclofen acts via a K+ conductance. 3. These effects were still apparent in the presence of tetrodotoxin (which did not abolish all spontaneous synaptic activity) and also in medium containing a combination of Co2+, the excitatory amino acid antagonist 6‐cyano‐7‐nitro‐quinoxaline‐2,3‐dione (CNQX) and the GABAA antagonist bicuculline which blocked synaptic activity. 4. The amplitude and frequency of spontaneous postsynaptic potentials (spPSPs) and spontaneous postsynaptic currents (spPSCs) were reduced by baclofen at concentrations (1 microM or less) which had no effect on membrane potential or holding current in current or voltage clamp recordings respectively. 5. The amplitude of evoked excitatory (evEPSPs/evEPSCs) and inhibitory (evIPSPs/evIPSCs) synaptic events elicited by electrical stimulation in the vicinity of the tractus solitarius (TS) was reduced by low concentrations of baclofen (250 nM‐1 microM) which did not produce discernible postsynaptic responses. 6. In order to examine the effects of baclofen on excitatory synaptic events without contamination with inhibitory events, stimulation of the TS was carried out in the presence of bicuculline. Conversely to investigate actions on purely inhibitory synaptic responses experiments were carried out with CNQX in the bathing solution. Inhibitory synaptic responses could still be evoked, presumably by stimulation of interneurones in the vicinity of the TS. IPSPs/IPSCs were more sensitive to baclofen than EPSPs/EPSCs. 7. The effects of baclofen on membrane potential or holding current and PSP/PSCs were antagonized by 2‐hydroxysaclofen (400 microM) confirming that baclofen was acting at gamma‐aminobutyric acid (GABA)B receptors.(ABSTRACT TRUNCATED AT 400 WORDS)
Inward rectification of acetylcholine‐elicited currents in rat phaeochromocytoma cells.
Authors: Ifune C K , Steinbach J H
Published: December 2014

1. Currents elicited by acetylcholine (ACh) were studied in the rat phaeochromocytoma cell line, PC12, using patch‐clamp techniques. 2. Whole‐cell ACh‐elicited currents are inwardly rectifying and intracellular Mg2+ can play a role in determining the extent of whole‐cell current rectification. Increasing the intracellular Mg2+ concentration, [Mg2+]i, augmented the rectification. The effects of increased [Mg2+]i on the whole‐cell current can be explained by the block of receptor channels by Mg2+. 3. In the nominal absence of internal divalent cations, however, a substantial degree of rectification remains. This rectification is probably not due to divalent cations, as buffering the external Mg2+ concentration to 50 microM and the internal concentration to nominally 0 Mg2+ did not reduce the rectification. The remanent rectification was not due to block by the main permeant cation, Na+. Using K+ or Cs+ as the main monovalent cation inside the cell did not diminish the rectification. Neither replacing the pH buffer, HEPES, with phosphate buffer nor increasing the intracellular pH removed the rectification. 4. For ACh receptor channels in excised patches, the voltage dependence of the probability of being open (Popen) stemmed mainly from the voltage dependence of the channel burst duration. The channel opening rate was relatively voltage independent. The weak voltage dependence displayed by the channel burst duration was insufficient to account for the reduced whole‐cell outward current at positive potentials. The mean burst duration of the channel did not have a simple logarithmic relationship with voltage. 5. In the absence of intracellular Mg2+, the instantaneous current‐voltage relationship for whole‐cell currents was linear suggesting that the I‐V relationship of single channels in perfused cells is linear and does not contribute to the rectification of the whole‐cell current. 6. In perfused cells, receptor channels had a low steady‐state probability of being open at positive potentials compared to channels in excised patches. Voltage jumps to positive potentials revealed a process in perfused cells which could account for the low Popen. Relaxations of agonist‐induced current at +40 mV had a large, exponentially decaying component that quickly closed channels (rate constant, tau, approximately 400 microseconds). The mechanism responsible for this decay could explain the rectification that remains in the absence of intracellular divalent cations.(ABSTRACT TRUNCATED AT 400 WORDS)
Na+ current densities and voltage dependence in human intercostal muscle fibres.
Authors: Ruff R L , Whittlesey D
Published: December 2014

1. Voltage‐clamp Na+ currents (INa) were studied in human intercostal muscle fibres using the loose‐patch‐clamp technique. 2. The fibres could be divided into two groups based upon the properties of INa. The two groups of fibres were called type 1 and type 2. 3. Both type 1 and type 2 fibres demonstrated fast and slow inactivation of INa. 4. Type 1 fibres had lower INa on the endplate border and extrajunctional membrane than type 2 fibres and required larger membrane depolarizations to inactivate Na+ channels by fast or slow inactivation of INa. 5. Type 2 fibres had a higher ratio of INa at the endplate border compared to extrajunctional membrane than Type 1 fibres. 6. Measurement of membrane capacitance suggested that the increase in INa at the endplate border was due to increased Na+ channel density. 7. Histochemical staining of some fibres suggested that type 1 fibres were slow twitch and type 2 fibres were fast twitch. 8. Differences in the properties of Na+ channels between fast‐ and slow‐twitch fibres may contribute to the ability of fast‐twitch fibres to operate at high firing frequencies and slow‐twitch fibres to be tonically active.
Shear stress‐induced calcium transients in endothelial cells from human umbilical cord veins.
Authors: Schwarz G , Callewaert G , Droogmans G , Nilius B
Published: December 2014

1. Changes of the free cytosolic Ca2+ concentration induced by shear stress were measured in Fura‐2 acetoxymethyl ester‐loaded endothelial cells from human umbilical cord veins. 2. We were able to induce Ca2+ transients in almost every cell by blowing a stream of physiological solution onto a single endothelial cell thereby inducing shear stress between 0 and 50 dyn cm‐2. The Ca2+ response could be graded by varying the shear stress, and reached a half‐maximal value at a shear stress of 30 dyn cm‐2. 3. The shear stress responses critically depended on the extracellular Ca2+ concentration and were absent in a Ca(2+)‐free solution. Repetitive application of short pulses of shear stress induced cumulative effects because of the slow decay of the shear stress Ca2+ responses (time constants 82.3 +/‐ 17.8 s from twenty‐five cells). Application of a depolarizing high potassium solution to reduce the driving force for Ca2+ entry decreased the Ca2+ transients in some of the cells. 4. Application of shear stress in the presence of other divalent cations, such as nickel, cobalt or barium, always produced substantial changes in the ratio of the 390/360 nm fluorescence signal, indicating influx of these cations and subsequent quenching of the Fura‐2 fluorescence. 5. Shear stress responses in the presence of 10 mM Ca2+ were completely blocked by application of 1 mM La3+. 6. Incubation of the cells with the phorbol ester 12‐O‐tetradecanoyl phorbol‐13‐acetate (TPA) did not alter the shear stress response, but completely blocked histamine‐induced Ca2+ transients. 7. Small submaximal shear stress potentiated the Ca2+ transients induced by histamine. 8. We conclude that shear stress‐dependent Ca2+ signals are induced by an influx of calcium that is not modulated via protein kinase C and not activated by membrane depolarization. The influx pathway is also permeable to divalent cations such as Ni2+, Co2+ and Ba2+, but is blocked by La3+.
Effects of specific carotid body and brain hypoxia on respiratory muscle control in the awake goat.
Authors: Smith C A , Engwall M J , Dempsey J A , Bisgard G E
Published: December 2014

1. We assessed the effects of specific brain hypoxia on the control of inspiratory and expiratory muscle electromyographic (EMG) activities in response to specific carotid body hypoxia in seven awake goats. We used an isolated carotid body perfusion technique that permitted specific, physiological, steady‐state stimulation of the carotid bodies or maintenance of normoxia and normocapnia at the carotid bodies while varying the level of systemic, and therefore, brain oxygenation. 2. Isolated brain normocapnic hypoxia of up to 1.5 h duration increased inspired minute ventilation (VI) by means of increases in both tidal volume (VT) and respiratory frequency (fR). Electromyographic activities of both inspiratory and expiratory muscles were augmented as well. These responses were similar to those produced by low levels of whole‐body normoxic hypercapnia. We conclude that moderate levels of brain hypoxia (Pa,O2 approximately 40 mmHg) in awake goats caused a net stimulation of ventilatory motor output. 3. Hypoxic stimulation of the carotid bodies alone caused comparable increases in VT and fR, and EMG augmentation of both inspiratory and expiratory muscles whether the brain was hypoxic or normoxic. These responses were quite similar to those obtained over a wide range of whole‐body normoxic hypercapnia. We conclude that the integration of carotid body afferent information is not affected by moderate brain hypoxia in awake goats. 4. We found no evidence for an asymmetrical recruitment pattern of inspiratory vs. expiratory muscles in response to carotid body hypoxia or in response to brain hypoxia alone. 5. Our data support the concept that moderate brain hypoxia results in a net stimulation of respiratory motor output. These findings question the significance of ‘central hypoxic depression’ to the regulation of breathing under physiological levels of hypoxaemia in the awake animal.
The effect of endotoxin‐induced fever on thermoregulation in the newborn rabbit.
Authors: Hull D , Vinter J , McIntyre J
Published: December 2014

1. Metabolic responses of endotoxin‐injected newborn rabbits were measured in a closed circuit calorimeter at a constant environmental temperature within their thermoneutral range. Oxygen consumption and colonic temperatures were then measured over a range of environmental temperatures from 21.0 to 40.0 degrees C and the responses of endotoxin‐injected rabbits compared with non‐injected litter mates. 2. To measure their preferred thermal environment, endotoxin‐injected and non‐injected litter mates were allowed to settle on a thermal gradient and their colonic temperatures measured. 3. In a constant environmental temperature of 37 degrees C, rises in oxygen consumption and colonic temperature following endotoxin injection were found to be biphasic. Oxygen consumption rose from 21 ml kg‐1 min‐1 to a maximum 35 ml kg‐1 min‐1 and colonic temperature rose from 39.0 to 39.8 degrees C. 4. The maximal rate of oxygen consumption was the same in both injected and non‐injected animals, 53 ml kg‐1 min‐1, being provoked at the same environmental temperature of 24 degrees C. Minimal rates of oxygen consumption were also similar for the two groups but in the injected animals they were achieved at an environmental temperature of 39 degrees C, 2 degrees C higher than for non‐injected animals. 5. We conclude that newborn rabbits challenged with a pyrogen have both behavioural and physiological responses. The thermogenic response is consistent with a change in sensitivity to feedback information rather than a simple shift in the central thermoregulatory set point. We find no evidence to support the view that in the febrile response a higher body temperature necessitates an increase in metabolic rate, the so‐called Q10 effect.
Modulation of K+ and Ca2+ channels by histamine H1‐receptor stimulation in rabbit coronary artery cells.
Authors: Ishikawa T , Hume J R , Keef K D
Published: December 2014

1. The modulation of whole‐cell K+ and Ca2+ currents by stimulation of histamine H1‐receptors in freshly isolated single smooth muscle cells from the rabbit coronary artery was characterized using the patch‐clamp technique at 35 degrees C. Single‐channel K+ currents were also analysed using the cell‐attached patch configuration. 2. The histamine H1‐receptor agonist, 2‐(2‐aminoethyl)pyridine (AEP) (0.1 mM), increased the amplitude of voltage‐activated inward Ba2+ currents, recorded using the perforated‐patch recording technique, which could be completely blocked by the dihydropyridine antagonist, nicardipine (1 microM). 3. Whole‐cell outward K+ currents in rabbit coronary artery cells could be classified into at least two components: (a) a slowly inactivating, 4‐aminopyridine (4‐AP)‐sensitive low‐noise current, and (b) a non‐inactivating, tetraethylammonium (TEA)‐sensitive high‐noise current. 4. AEP (0.1 mM) caused changes in whole‐cell outward K+ currents which depended upon membrane voltage. Specifically: (a) AEP enhanced the amplitude of outward currents at voltages between ‐30 and 0 mV, and (b) AEP decreased the outward currents at more positive potentials. 5. The removal of extracellular Ca2+ caused little inhibition of the effects of AEP on K+ currents, whereas the depletion of intracellular Ca2+ stores by pretreatment with ryanodine and caffeine prevented the effects of AEP on K+ channels. Moreover, acute exposure to ryanodine (10 microM) or thapsigargin (1 microM), a Ca(2+)‐ATPase inhibitor, caused voltage‐dependent changes in the outward currents similar to those observed with AEP. These results suggest that the voltage‐dependent effects of AEP on K+ currents are mainly mediated by release of Ca2+ from intracellular stores. 6. The dual stimulatory and inhibitory effect of AEP on whole‐cell K+ currents was shown to be due to a differential effect on two distinct types of K+ channels. The stimulatory effect observed over the voltage range ‐30 to 0 mV was prevented by pretreatment of cells with low concentrations of TEA (1 mM), whereas the inhibitory effect observed at positive potentials was prevented by pretreatment of cells with 4‐AP (3 mM). 7. Single‐channel recordings revealed two types of unitary K+ currents with conductances of 225 and 70 pS in the cell‐attached configuration with symmetrical K+ solutions (150 mM K+ in pipette‐150 mM K+ in bath). Bath application of AEP (0.1 mM) caused a marked increase in the open probability of the large conductance channels.(ABSTRACT TRUNCATED AT 400 WORDS)