bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of intracellular pH on ATP‐sensitive K+ channels in mouse pancreatic beta‐cells.
Authors: Proks P , Takano M , Ashcroft F M
Published: December 2014

1. The effects of intracellular pH (pHi) on the ATP‐sensitive K+ channel (K+ATP channel) from mouse pancreatic beta‐cells were examined in inside‐out patches exposed to symmetrical 140 mM K+ solutions. 2. The relationship between channel activity and pHi was described by the Hill equation with half‐maximal inhibition (Ki) at pHi 6.25 and a Hill coefficient of 3.7. 3. Following exposure to pHi < 6.8, channel activity did not recover to its original level. Subsequent application of trypsin to the intracellular membrane surface restored channel activity to its initial level or above. 4. At ‐60 mV the relationship between pHi and the single‐channel current amplitude was described by a modified Hill equation with a Hill coefficient of 2.1, half‐maximal inhibition at pHi 6.48 and a maximum inhibition of 18.5%. 5. A decrease in pHi reduced the extent of channel inhibition by ATP: Ki was 18 microM at pH 7.2 and 33 microM at pH 6.4. The Hill coefficient was also reduced, being 1.65 at pH 7.2 and 1.17 at pH 6.4. 6. When channel activity was plotted as a function of ATP4‐ (rather than total ATP) there was no effect of pHi on the relationship. This suggests that ATP4‐ is the inhibitory ion species and that the effects of reducing pHi are due to the lowered concentration of ATP4‐. 7. Changes in external pH had little effect on either single‐channel or whole‐cell K+ATP currents. 8. The effects of pHi do not support a role for H+ in linking glucose metabolism to K+ATP channel inhibition in pancreatic beta‐cells.
Facilitatory effect of docosahexaenoic acid on N‐methyl‐D‐aspartate response in pyramidal neurones of rat cerebral cortex.
Authors: Nishikawa M , Kimura S , Akaike N
Published: December 2014

1. The effect of docosahexaenoic acid (DHA) on N‐methyl‐D‐aspartic acid (NMDA) responses in the presence of glycine was investigated in pyramidal neurons acutely dissociated from rat cerebral cortex in whole‐cell and single channel configurations. 2. DHA potentiated the NMDA‐induced response but reduced the non‐NMDA (kainate‐induced) response in a concentration‐dependent manner at a holding potential of ‐60 mV under voltage‐clamp conditions. 3. Arachidonic acid (AA) also potentiated the NMDA‐induced response in a manner similar to DHA. Oleic acid caused a slight potentiation. However, other polyunsaturated and saturated fatty acids had no such effects. 4. The facilitatory action of DHA on the NMDA‐induced response was not affected by adding inhibitors of cyclo‐oxygenase, lipoxygenase or phospholipase A2, suggesting that DHA may exert its facilitatory effect directly on the NMDA receptor. 5. The facilitatory action of DHA was observed in the presence of a saturating dose of NMDA. Moreover, a detailed analysis of the NMDA receptor‐operated single channel currents revealed that, in the presence of DHA, the open probability of the channel increased without changing the conductance, indicating that DHA may act by binding directly to a novel site on the NMDA receptor or by altering the lipid environment of the NMDA receptor and thereby potentiating the response to NMDA. 6. The results are discussed in terms of the possibility that DHA may play an important role in the genesis of long‐term potentiation, at least that involving the activation of NMDA receptors.
Contractile properties of single skinned fibres from the extraocular muscles, the levator and superior rectus, of the rabbit.
Authors: Lynch G S , Frueh B R , Williams D A
Published: December 2014

1. The superior rectus and levator palpebrae superioris (levator), two of the extraocular muscles, were dissected from the rabbit and stored in a glycerol‐based solution at ‐20 degrees C in order to prepare single, skinned fibres. 2. The Ca(2+)‐ and Sr(2+)‐activated isometric contractile properties were determined for individual extraocular muscle fibres. Fibres were separated into discrete groups or fibre types on the basis of their physiological characteristics. The superior rectus and levator muscles were both found to consist of fibres which exhibited similar contractile characteristics to fast‐ and slow‐twitch fibres from other mammalian muscle, including type I, type IIA and type IIB fibres. 3. As well as the existence of the normal, classical fibre types in extraocular muscle there were also a large number of fibres from both muscles which exhibited mixed fast‐ and slow‐twitch contractile characteristics within the single contracting unit. Of the fibres sampled, the mixed fibres comprised the second largest population (7/19, 37%) in the levator and the largest fibre population in the superior rectus (11/31, 35.5%). These results are consistent with histochemical and immunohistochemical reports in the literature which suggest the co‐existence of fast and slow myosin along the length of the extraocular muscle fibres. 4. Extraocular muscle fibres exhibited lower absolute maximum forces compared with other mammalian limb muscle fibres. However, when corrected for fibre cross‐sectional area, the maximum tension development was within the normal range for mammalian limb muscle fibres, except for one group (type IIA) of fast‐twitch fibres which exhibited significantly lower maximal tension. 5. The existence of a large proportion of fibres with composite fast‐ and slow‐twitch characteristics highlights the functional and morphological complexity of these muscles. It is postulated that the functional significance of these mixed fibres may be to provide or enhance the resolution for subtle, precise movements of the eye and eyelid.
The effect of glibenclamide on frog skeletal muscle: evidence for K+ATP channel activation during fatigue.
Authors: Light P E , Comtois A S , Renaud J M
Published: December 2014

1. The purpose of this study was to determine whether ATP‐sensitive K+ (K+ATP) channels are activated and contribute to the decrease in force during fatigue development in the sartorius muscle of the frog, Rana pipiens. Tetanic force (elicited by field stimulation), action potential and membrane conductance (using conventional microelectrodes), were measured in the presence and absence of glibenclamide, a K+ATP channel antagonist. Experiments were performed in bicarbonate‐buffered solutions at pH 7.2. 2. In unfatigued muscle 100 mumol l‐1 glibenclamide had no effect on the resting potential, the overshoot, the half‐depolarization time or the maximum rate of depolarization of action potentials, while the mean half‐repolarization time increased by 19 +/‐ 4% (+/‐ S.E.M.) and the maximum rate of repolarization decreased by 17 +/‐ 5%. 3. Fatigue was elicited using 100 ms tetanic contractions every 1 s for 3 min. In the absence of glibenclamide the mean half‐repolarization time increased from 0.57 +/‐ 0.05 to 0.89 +/‐ 0.05 ms during fatigue. The mean half‐repolarization times after fatigue, when muscle fibres were exposed to 100 mumol l‐1 glibenclamide either 60 min prior to fatigue or 60 s before the end of fatigue, were 1.16 +/‐ 0.08 and 1.17 +/‐ 0.07 ms respectively. Application of 100 mumol l‐1 glibenclamide after 5 min of recovery did not increase the half‐repolarization time, but decreased the rate of recovery compared to control values. 4. In unfatigued muscles, 100 mumol l‐1 glibenclamide did not affect the tetanic contraction. In the absence of glibenclamide, the mean tetanic force after fatigue was 11.0 +/‐ 0.9% of prefatigue values. Application of 100 mumol l‐1 glibenclamide 60 min before fatigue increased the rate of fatigue development as the mean tetanic force was 4.8 +/‐ 0.8% after 3 min of stimulation. The addition of 100 mumol l‐1 glibenclamide 60 s before the end of fatigue had no effect on tetanic force during this time compared to control. 5. In the absence of glibenclamide, muscles recovered 90.1 +/‐ 1.6% of their tetanic force after 100 min. Addition of 100 mumol l‐1 glibenclamide 60 min prior to fatigue significantly reduced the capacity of muscles to recover their tetanic force: after 100 min of recovery tetanic force was only 47.3 +/‐ 9.4% of the pre‐fatigue value.(ABSTRACT TRUNCATED AT 400 WORDS)
Enterotoxin Escherichia coli STa activates a nitric oxide‐dependent myenteric plexus secretory reflex in the rat ileum.
Authors: Rolfe V , Levin R J
Published: December 2014

1. Mucosally added enterotoxin Escherichia coli STa increased the electrogenic Cl‐ secretion measured as the short‐circuit current (Isc) across isolated muscle‐stripped and muscle‐unstripped rat mid‐ilea incubated in vitro. 2. Pretreatment with serosal L‐NAME (N omega‐nitro‐L‐arginine methyl ester) or tetrodotoxin (TTX) significantly reduced the maximum Isc and the duration of action of STa in the unstripped but not stripped ilea. D‐NAME (serosal), indomethacin or 5‐hydroxy‐tryptamine‐desensitization was ineffective on STa‐induced Isc in either stripped or unstripped ilea. 3. Serosal capsaicin reduced the maximum Isc of STa and its duration of action in unstripped ilea. 4. L‐Arginine induced a significantly larger increase in the Isc across unstripped ilea than across stripped ilea; this could be significantly reduced by serosal L‐NAME or TTX, although these were ineffective in stripped ilea. 5. Pretreatment of anaesthetized rats with I.P. L‐NAME suppressed the fluid secretion induced by luminal STa in ilea in vivo but had no effect on that induced by luminal carbachol. 6. Mucosal STa increased electrogenic Cl‐ secretion across intact rat ileum in vitro by activating a capsaicin‐sensitive, nitric oxide‐dependent myenteric plexus‐mediated secretory reflex. The suppression by L‐NAME of STa induced ileal fluid secretion in vivo probably involves the inhibition of this reflex.
Differential secretion of catecholamines in response to peptidergic and cholinergic transmitters in rat adrenals.
Authors: Guo X , Wakade A R
Published: December 2014

1. Rat adrenal medulla is stimulated by cholinergic and peptidergic transmitters released from splanchnic nerves. The peptidergic transmitter has been identified as vasoactive intestinal polypeptide (VIP) and its contribution in comparison to that of acetylcholine (ACh) is more prominent at low neuronal activity. The purpose of this study is to determine if ACh and VIP cause differential secretion of adrenaline and noradrenaline and whether the differential secretion also occurs when splanchnic nerves are stimulated at different frequencies. 2. Perfusion of the left adrenal gland with Krebs solution for several hours did not change adrenaline and noradrenaline contents (15.2 micrograms and 3.5 micrograms, respectively) and their ratio (4.4) from those of the unperfused right adrenal medulla (15.2 micrograms, 3.3 micrograms and 4.8, respectively). 3. Perfusion with ACh (10 microM for 4 min) resulted in the secretion of 109 ng of catecholamines and the ratio of adrenaline to noradrenaline was 3.8. Although the secretion increased with increased concentrations of ACh (30 and 100 microM), the ratios remained between 3 and 4. 4. Perfusion with VIP (10 microM for 4 min) resulted in the secretion of 27 ng of catecholamines and the ratio of adrenaline to noradrenaline was 9.7. A higher concentration of VIP (20 microM for 4 min) resulted in the secretion of greater amounts of catecholamines (102 ng) without significantly altering the ratio of adrenaline to noradrenaline (10.9). 5. Perfusion with as low as 0.01 microM pituitary adenylate cyclase‐activating polypeptide (PACAP) increased the secretion of catecholamines to 31 ng and the secretion increased in a dose‐dependent manner up to 0.3 microM.(ABSTRACT TRUNCATED AT 250 WORDS)
The reproducibility of the response of the human newborn to CO2 measured by rebreathing and steady‐state methods.
Authors: Cohen G , Henderson‐Smart D J
Published: December 2014

The ventilatory response (VR) of the full‐term newborn to CO2 was studied during quiet sleep using rebreathing and steady‐state methods. Rebreathing responses were obtained under normoxic (fractional inspired oxygen concentration, Fi,O2 = 0.21) and hyperoxic (Fi,O2 = 0.4) conditions. Ten infants were tested three to five times using each of the two rebreathing protocols and the results averaged. Overall, there was no significant difference between either the mean variability (coefficient of variation) or slope of the VR measured under normoxic and hyperoxic conditions. Four infants were studied using a steady‐state technique. There was marked test‐to‐test variability in VRs measured by this method. The results appear to indicate that the variability of the VR of the newborn to CO2 is not a result of chemoreflex changes in ventilation elicited in response to fluctuations in arterial PO2.
The effect of hypertonicity on force generation in tetanized single fibres from frog skeletal muscle.
Authors: Piazzesi G , Linari M , Lombardi V
Published: December 2014

1. We compared the tension transient that follows a step change in sarcomere length in normal Ringer solution with that in Ringer solution made hypertonic by the addition of 98 mM sucrose. Steps were applied on tetanized single muscle fibres during either the isometric plateau or the steady force response to lengthening at low speed. Sarcomere length was controlled on selected fibre segments by a striation follower. Analysis is limited to phase 1 (the tension change simultaneous with the length step, mainly due to cross‐bridge elasticity) and phase 2 (the quick phase of tension recovery, a manifestation of the cross‐bridge elementary force‐generating process). 2. At the isometric tetanus plateau the steady force is reduced by 19% in hypertonic solution, and the stiffness is slightly increased. During slow lengthening both steady force and stiffness are similar in normal solution and in hypertonic solution. In hypertonic solution the tension‐to‐stiffness ratio, a measure of the mean cross‐bridge extension before the step, is markedly reduced in isometric conditions (‐23%), but not during lengthening (‐2%). 3. The plots of instantaneous tension versus the length change during the step show that in hypertonic medium the elasticity of the fibre is almost undamped. Thus the increase in stiffness cannot be attributed to an increase in viscosity. 4. In isometric conditions (T2‐T1)/(Ti‐T1), the proportion of the initial tension drop recovered at the end of phase 2, is not affected by hypertonicity for releases of moderate and large size (> 2 nm) and is reduced for small releases (< 2 nm) and for stretches. The abscissa intercept of the relation (T2‐T1)/(Ti‐T1) versus step amplitude is the same in both media. During lengthening, for releases of small and moderate size, (T2‐T1)/(Ti‐T1) is 20% lower in hypertonic solution. For large releases the slope of the relation is lower so that the abscissa intercept is not changed. 5. The speed of quick tension recovery following a step length change imposed in isometric conditions is slightly depressed in hypertonic solution. The relation between speed of recovery and step amplitude maintains its shape and is shifted downwards. During lengthening, the speed of quick tension recovery in hypertonic solution is less dependent on step amplitude than in normal solution, as if a more linear viscoelasticity is responsible for a large fraction of residual recovery.(ABSTRACT TRUNCATED AT 400 WORDS)
Synergistic effect of adrenergic and muscarinic receptor activation on [Ca2+]i in rabbit ciliary body epithelium.
Authors: Farahbakhsh N A , Cilluffo M C
Published: December 2014

1. Changes in cytosolic free calcium concentration ([Ca2+]i) in response to cholinergic and adrenergic agents alone and in combination were investigated using fura‐2 fluorescence imaging in intact non‐pigmented epithelial cells of rabbit ciliary body. 2. Resting (‘baseline’) [Ca2+]i was 147 +/‐ 6 nM (mean +/‐ S.E.M.). Acetylcholine (ACh, 10 microM) doubled [Ca2+]i, and adrenaline (1 microM) increased it by about 36%. When ACh (10 microM) and adrenaline (1 microM) were applied together [Ca2+]i was transiently increased to 1160 +/‐ 160 nM, about 7 times the response induced by ACh alone. 3. Noradrenaline and 5‐bromo‐6‐(2‐imidazolin‐2‐yl‐amino)‐quinoxaline (UK 14304) had effects similar to adrenaline in enhancing the response to ACh. Phenylephrine (Phe) had a relatively smaller effect and none was observed for methoxamine and isoprenaline (Iso). 4. The response to ACh and adrenaline could be blocked by atropine (1 microM, 87 +/‐ 5%), yohimbine (1 microM, 73 +/‐ 8%), and to a lesser degree by prazosin (1 microM). Propranolol had no effect. 5. Lowering the extracellular calcium concentration to 3 nM dropped the baseline [Ca2+]i by half and reduced the response to ACh and adrenaline to a small and transient rise in [Ca2+]i. Addition of La3+ to Ca(2+)‐containing solution also lowered [Ca2+]i and largely reduced the response. 6. We conclude that simultaneous activation of muscarinic and alpha 2‐adrenergic receptors induces a large increase in [Ca2+]i, which is the result of both Ca2+ release and influx.
Mechanisms of vasoconstriction induced by endothelin‐1 in smooth muscle of rabbit mesenteric artery.
Authors: Yoshida M , Suzuki A , Itoh T
Published: December 2014

1. The mechanism underlying the vasoconstriction induced by endothelin‐1 (ET‐1) was investigated by measuring the intracellular concentration of Ca2+ ([Ca2+]i), isometric force and phosphorylation of the myosin light chain (MLC) in endothelium‐denuded unskinned and beta‐escin‐treated skinned smooth muscle from resistance vessels of the rabbit mesentery. The role of protein kinase C (PKC) in the action of ET‐1 was studied in skinned smooth muscle using a synthetic peptide, PKC19‐36, which corresponds to the autoinhibitory domain of PKC. 2. ET‐1 (> 0.1 nM) induced slowly developing, maintained increases in [Ca2+]i and force. Nicardipine completely blocked the ET‐1‐induced increase in [Ca2+]i. BQ‐123 (an inhibitor of the ETA receptor) blocked the ET‐1‐induced contraction but IRL 1620 (Suc‐[Glu9,Ala11,15]‐ET‐1(8‐21), an agonist of the ETB receptor) failed to induce contraction. 3. In ionomycin‐ and 70 mM K(+)‐treated strips, ET‐1 shifted the [Ca2+]i‐force relationship to the left and enhanced the maximum amplitude of contraction induced by 2.6 mM Ca2+. In skinned smooth muscle treated with ionomycin, Ca2+ (0.1‐3 microM) increased both force and MLC phosphorylation, in a concentration‐dependent manner. ET‐1 with GTP shifted both the Ca(2+)‐force and Ca(2+)‐MLC phosphorylation relationships to the left without significant changes in the maximum responses. ET‐1 with GTP did not change the relationship between force and MLC phosphorylation. Similar effects were observed with phorbol 12,13‐dibutyrate (PDBu, an activator of PKC). These results indicate that the sensitivity of MLC phosphorylation to Ca2+ is enhanced both by ET‐1 with GTP and by PDBu. 4. PKC19‐36 (an inhibitor of PKC) modified neither the contraction nor MLC phosphorylation induced by 0.3 microM Ca2+ but blocked the PDBu‐induced enhancement of both these Ca(2+)‐induced responses. However, PKC19‐36 only partly inhibited the enhancement produced by ET‐1 with GTP on the Ca(2+)‐induced responses. PKC19‐36 did not modify the relationship between force and MLC phosphorylation in the presence either of ET‐1 with GTP or of PDBu. By contrast, BQ‐123, neomycin and guanosine 5'‐O‐(2‐thiodiphosphate) (GDP beta S) each abolished the ET‐1‐induced enhancement of the contraction induced by 0.3 microM Ca2+. 5. These results suggest that ET‐1 acts on the ETA receptor and increases Ca2+ influxes through an activation of the dihydropyridine‐sensitive Ca2+ channel, causing a long‐lasting and maintained contraction in resistance vessels of the rabbit mesentery.(ABSTRACT TRUNCATED AT 400 WORDS)
Characterization of the potassium channel from frog skeletal muscle sarcoplasmic reticulum membrane.
Authors: Wang J , Best P M
Published: December 2014

1. The sarcoplasmic reticulum (SR) membrane of skeletal muscle contains potassium channels which are thought to support charge neutralization during calcium release by providing a permeability pathway for counter‐ion movement. To describe the behaviour of the SR K+ channel under physiological conditions, single channel activity was recorded from excised patches of SR membrane. Patches were made from membrane blebs extruded from contracted muscle fibres whose surface membranes had been removed previously by mechanical dissection. 2. The channel was active over a large voltage range from ‐80 to +100 mV. The current‐voltage relationship of the channel was linear over most of this voltage range (slope conductance equal to 60 pS in 130 mM potassium), but showed rectification at voltages below ‐50 mV. 3. The activity of the channel (number of state transitions per unit time) was greater at positive voltages than at negative voltages. Analysis of dwell‐time distributions showed that the time spent in the open state is best fitted by a double Gaussian, suggesting that the channel possesses both a long (l)‐ and a short (s)‐lived open state with identical conductances. The dwell times for the two states were Ts = 0.3 ms and Tl = 2.6 ms at +90 mV and Ts = 0.1 ms and Tl = 15.1 ms at ‐40 mV. Thus, positive voltage decreased the long open time significantly which was consistent with the observed increase in channel activity at positive potentials. 4. The permeability sequence of the channel to various monovalent cations was deduced from the channel reversal potential under bi‐ionic conditions and was found to be: K+ > Rb+ > Na+ > Cs+ > Li+. 5. Channel activity was reduced when the patch was perfused with 1,10‐bis‐guanidino‐n‐decane (BisG10), a drug reported to block the SR K+ channel with high affinity. The drug concentration necessary to reduce the open probability (P(o)) by 50% was 19.8 microM at ‐40 mV and 338.2 microM at +50 mV. The zero voltage dissociation constant (Kd) was calculated to be 48 microM. 6. Pharmacological agents known to affect surface membrane K+ channels, such as 0.5 mM Ba2+ or 3.0 mM 4‐aminopyridine, were much less effective in blocking the channel than BisG10. Physiological calcium concentrations (pCa = 8.0 and 3.0) did not affect channel behaviour.4
The role of enteric inhibitory motoneurons in peristalsis in the isolated guinea‐pig small intestine.
Authors: Waterman S A , Costa M
Published: December 2014

1. Peristalsis is a co‐ordinated motor behaviour in which an anally propagated contraction of the circular muscle propels intraluminal contents. The role of excitatory motoneurons in peristalsis is well established; however the role of enteric inhibitory motoneurons is unknown. 2. A combination of a nitric oxide synthase inhibitor and apamin, which blocks relaxation of the circular muscle of guinea‐pig small intestine mediated by enteric inhibitory motoneurons, was used to investigate the role of inhibitory motoneurons in peristalsis in isolated segments of guinea‐pig small intestine. 3. N omega‐nitro‐L‐arginine methyl ester (L‐NAME, 400 microM) and N omega‐nitro‐L‐arginine (L‐NOArg, 100 microM) significantly reduced the threshold volume required to trigger emptying of the intestine. This effect was reversed by L‐arginine (4 mM) and L‐arginine alone increased the threshold volume for initiation of peristalsis. Sodium nitroprusside (0.1‐10 microM), which generates nitric oxide, also increased the threshold volume. L‐NAME, L‐NOArg, L‐arginine and sodium nitroprusside did not alter the maximal intraluminal pressure generated during emptying. Contraction of the longitudinal muscle during the initial phase of fluid infusion was significantly increased by L‐NAME and L‐NOArg and reduced by sodium nitroprusside (1 nM to 10 microM). 4. Apamin (0.5 microM) did not significantly alter the threshold volume necessary to initiate peristalsis or contraction of the longitudinal muscle. However, the maximal pressure generated when the intestine was emptying was significantly increased. Furthermore, short segments of circular muscle contracted apparently randomly, before peristaltic emptying was triggered. 5. A combination of L‐NAME and apamin completely disrupted peristalsis. Contractions of the circular muscle did not always start at the oral end. Stationary contractions as well as contractions propagating orally and anally were observed. 6. It is concluded that enteric inhibitory motoneurons are crucial for peristalsis to occur. They are important in setting the threshold at which peristaltic emptying is triggered, via nitric oxide. They are essential for the propagation of the circular muscle contraction, via an apamin‐sensitive mechanism of transmission. Contraction of the longitudinal muscle during peristalsis is partly inhibited by a nitric oxide‐mediated mechanism.
Effects on fluid and Na+ flux of varying luminal hydraulic resistance in rat colon in vivo.
Authors: Zammit P S , Mendizabal M , Naftalin R J
Published: December 2014

1. A new method of measuring fluid and ionic movements and the dehydrating power of the colon in vivo is described. A range of agarose gel cylinders, with calibrated hydraulic conductivities (Lp), were inserted into the lumen of the descending colon of anaesthetized rats. Fluxes of fluid, Na+ and K+ out of the gels were measured over a period of 60‐110 min. 2. Fluid absorption by the colon from 2.5% agarose gels was not slower than from solution without gel. Fluid absorption was inhibited by 66% when the agarose concentration was raised to 10%. In contrast 2.5% agarose gels caused a 73% (P < 0.001) reduction in water flow from rat ileum. 3. Increasing gel concentration to 10% or above caused the absorbate from the gels to become hypertonic (P < 0.001). 4. The measured suction pressure applied by the colonic hypertonic absorbate to the gels increased from 44 +/‐ 2.3 cmH2O (n = 23) with 2.5% agarose gels to 6713 +/‐ 960 cmH2O (n = 13) with 15% (P < 0.001). 5. Deoxycholate (2 mM) produced a decrease in fluid and Na+ absorption and reduced the suction pressure and power exerted by the colon.
The metabolic responses of human type I and II muscle fibres during maximal treadmill sprinting.
Authors: Greenhaff P L , Nevill M E , Soderlund K , Bodin K , Boobis L H , Williams C , Hultman E
Published: December 2014

1. Muscle biopsy samples were obtained from the vastus lateralis of six healthy volunteers before and after 30 s of treadmill sprinting. A portion of each biopsy sample was used for mixed‐fibre metabolite analysis. Single fibres were dissected from the remaining portion of each biopsy and were used for ATP, phosphocreatine (PCr) and glycogen determination. 2. Before exercise, PCr and glycogen contents were higher in type II fibres (79.3 +/‐ 2.7 and 472 +/‐ 35 mmol (kg dry matter (DM)‐1, respectively) compared with type I fibres (71.3 +/‐ 3.0 mmol (kg DM)‐1, P < 0.01 and 375 +/‐ 25 mmol (kg DM)‐1, P < 0.001, respectively). 3. Peak power output was 885 +/‐ 66 W and declined by 65 +/‐ 3% during exercise. Phosphocreatine and glycogen degradation in type II fibres during exercise (74.3 +/‐ 2.5 and 126.3 +/‐ 15.8 mmol (kg DM)‐1, respectively) was greater than the corresponding degradation in type I fibres (59.1 +/‐ 2.9 mmol (kg DM)‐1, P < 0.001 and 77.0 +/‐ 14.3 mmol (kg DM)‐1, P < 0.01, respectively). The decline in ATP during exercise was similar when comparing fibre types (P > 0.05). 4. Compared with previous studies involving similar durations of maximal cycling exercise, isokinetic knee extension and intermittent isometric contraction, the rates of substrate utilization recorded in type I fibres were extremely high, being close to the rapid rates observed in this fibre type during intense contraction with limb blood flow occluded.
Cotransport of K+, Cl‐ and H2O by membrane proteins from choroid plexus epithelium of Necturus maculosus.
Authors: Zeuthen T
Published: December 2014

1. The interaction between K+, Cl‐ and H2O fluxes was studied in the ventricular membrane of the choroid plexus epithelium from Necturus maculosus by means of ion‐selective microelectrodes. 2. Three experimental strategies were adopted: the osmolarity of the ventricular solution was increased abruptly by addition of (i) mannitol or (ii) KCl; (iii) Na+ in the ventricular solution was replaced isosmotically by K+. 3. The mannitol experiments showed that H2O had two pathways across the ventricular membrane. One was purely passive, with a water permeability, L'p, of 0.64 x 10(‐4) cm s‐1 (osmol l‐1)‐1. This operated in parallel with an ion‐dependent pathway of similar magnitude which was abolished in Cl(‐)‐free solutions. 4. When KCl was added there was a flow of H2O into the cell. Surprisingly, this took place despite the osmotic gradient which favoured an efflux of H2O. The effect was blocked by frusemide (furosemide), in which case KCl had the same effects as applications of NaCl or mannitol. 5. Replacement of Na+ with K+ caused an influx of H2O. This flux could proceed against osmotic gradients implemented by mannitol. 6. The present data and those of earlier publications show that the interdependence of the fluxes of K+, Cl‐ and H2O in the exit membrane can be described as cotransport. The fluxes have a fixed stoichiometry of 1:1:500, the flux of one species is able to energize the flux of the two others, and the transport exhibits saturation and is specific for K+ and Cl‐. 7. A molecular model based upon a mobile barrier in a membrane spanning protein gives an accurate quantitative description of the data.
A rise in the intracellular Ca2+ concentration of isolated rat supraoptic cells in response to oxytocin.
Authors: Lambert R C , Dayanithi G , Moos F C , Richard P
Published: December 2014

1. Intracellular Ca2+ concentration ([Ca2+]i) was monitored in single cells isolated from adult rat supraoptic (SO) nuclei. The great majority of cells (85%) were neurones and most were immunoreactive to oxytocin or to vasopressin (AVP). 2. The resting [Ca2+]i of the majority (80%) of the neurones remained stable while 20% of the neurones displayed spontaneous [Ca2+]i oscillations which disappeared in low‐Ca2+ (100 nM) EGTA buffer. 3. Addition of 100 nM oxytocin increased the [Ca2+]i in both stable and oscillating cells. Two types of responses were observed: (i) a sustained response with [Ca2+]i being maintained at an elevated level and (ii) a brief response with [Ca2+]i quickly returning to a near‐resting level. Responses were reproducible, dose dependent and blocked with a specific oxytocin antagonist. 4. Removal of extracellular Ca2+ did not block the oxytocin response. In EGTA buffer, application of thapsigargin (200 nM) onto oxytocin‐sensitive cells induced an increase in [Ca2+]i and inhibited the oxytocin response. These effects were not induced by other intracellular Ca2+ mobilizers such as tBuBHQ (see Methods) or caffeine. 5. In conclusion, half of the SO cells respond to oxytocin with a rise in [Ca2+]i. The effect is mediated by oxytocin receptors and results from release of Ca2+ from thapsigargin‐sensitive stores.
Simultaneous expression of cardiac and skeletal muscle isoforms of the L‐type Ca2+ channel in a rat heart muscle cell line.
Authors: Mejía-Alvarez R , Tomaselli G F , Marban E
Published: December 2014

1. We have investigated the identity of the L‐type Ca2+ channels present in the H9c2 myoblast line derived from embryonic rat ventricle. To this end, we characterized macroscopic and unitary Ba2+ currents through Ca2+ channels, and looked for specific genetic messages encoding different L‐type Ca2+ channel isoforms. 2. The macroscopic Ba2+ current (recorded in 10 mM BaCl2) revealed two components with different time courses of activation. The fast component (IBa,fast) activates with a time constant of 23 +/‐ 12 ms (at +10 mV), while the slow component activates with a time constant of 125 +/‐ 12 ms (at +10 mV). 3. Single‐channel recordings revealed the presence of two independent channels with conductance values of 11 and 25 pS (in 70 mM Ba2+). These values are identical to those reported previously for skeletal muscle and cardiac Ca2+ channels, respectively. 4. The mean ensemble current from the 11 pS channel reproduced the time course of the slow component observed at the macroscopic level, while the 25 pS ensemble time course paralleled that of the fast component. 5. Reverse transcriptase polymerase chain reaction (PCR) with alpha 1‐isoform‐specific primers revealed the presence of two distinct transcripts in H9c2 cells. The sequences of the PCR products showed a high degree of homology with the corresponding segments of the rabbit cardiac and skeletal muscle L‐type Ca2+ channel isoforms. Adult rat skeletal and cardiac muscle expressed only one type of transcript. 6. H9c2 cells appear to be unique in that they simultaneously express both skeletal muscle and cardiac isoforms of the L‐type Ca2+ channel alpha 1‐subunit. Thus, the H9c2 cell line may prove to be useful when studying the regulation of subtype‐specific Ca2+ channel gene expression.
Neurally maintained hypersecretion in undernourished rat intestine activated by E. coli STa enterotoxin and cyclic nucleotides in vitro.
Authors: Nzegwu H C , Levin R J
Published: December 2014

1. The electrogenic secretory responses of stripped jejuna and ilea from chronically undernourished rats (50% control diet for 21 days) to the bacterial enterotoxin Escherichia coli STa, measured as the short‐circuit current in vitro, show an enhanced maximum secretion (ISC, max) with a prolonged duration compared with fed intestine. 2. The ISC, max is unaffected by pretreatment of the intestine in vitro with hexamethonium, atropine, procaine or indomethacin, or by desensitization to 5‐hydroxytryptamine (5‐HT), while the prolonged duration is unaffected by atropine, indomethacin or 5‐HT desensitization but is reduced by hexamethonium and procaine. 3. Both 8‐bromo‐cyclic GMP and dibutyryl cyclic AMP added serosally activate the enhanced ISC, max and its maintenance. Pretreatment with tetrodotoxin had no effect on the initial ISC, max but prevented its maintenance. 4. Bethanechol, dimethyl phenyl piperazinium, vasoactive intestinal polypeptide, 5‐HT and luminal propionate all induced the characteristic hypersecretory activity in the undernourished intestine compared with the fed state, but none could activate the maintenance circuit to prolong their transient responses. 5. Maintenance of the induced hypersecretory activity is the first example of induction of the neural control of intestinal secretion by the dietary intake level and illustrates the plasticity of the enteric nervous system.
Spatial integration of local transmitter responses in motoneurones of the turtle spinal cord in vitro.
Authors: Skydsgaard M , Hounsgaard J
Published: December 2014

1. Integration of responses to local activation of transmitter receptors in the dendrites of motoneurones was investigated in a slice preparation of the turtle spinal cord. Membrane‐active substances were applied from up to three independent iontophoresis electrodes during intracellular recording from the cell body. 2. Responses to glutamate could be evoked from dendrites closer than 20 microns from the tip of the glutamate electrode. The effects of other substances were more widespread. 3. In normal medium the configuration of a glutamate response was affected by time‐dependent anomalous rectification. In the presence of muscarine the sum of glutamate responses from two different dendrites recruited a voltage‐sensitive plateau potential. 4. The response to glutamate from one dendrite could be attenuated by local application of gamma‐aminobutyric acid (GABA) without effects on soma conductance or glutamate responses from other dendrites. 5. The response to glutamate from one dendrite could be selectively enhanced by local application of tetraethylammonium (TEA) or N‐methyl‐D‐aspartate (NMDA) without effects on soma conductance or glutamate responses from other dendrites. 6. NMDA could convert a tonic glutamate response from one dendrite into a phasic response without affecting the configuration of glutamate responses from other dendrites. 7. The effects of TEA and NMDA were facilitated by depolarization and reduced by hyperpolarization. 8. We conclude that the cable structure of motoneurones and the distribution of synapses and voltage‐sensitive ion channels provide relative autonomy to non‐linear synaptic processing and modulation in confined dendritic regions.
Limited independent flexion of the thumb and fingers in human subjects.
Authors: Kilbreath S L , Gandevia S C
Published: December 2014

1. We investigated whether human subjects can activate selectively flexor pollicis longus (FPL) and digital portions of flexor digitorum profundus (FDP). These muscles were selected because they are the only flexors of the distal phalanges. 2. Electromyographic activity (EMG) was recorded with intramuscular electrodes from one digital component of the deep flexors (‘test’) while subjects lifted weights by flexing the distal interphalangeal joint of the other digits in turn (‘lifting’ digits). Only recording sites at which single motor units were recruited selectively at low forces were used. The weights lifted represented 2.5‐50% of the maximal voluntary contraction (MVC). We measured the lowest weight lifted which produced phasic and tonic coactivation in the ‘test’ muscle. 3. The extent of coactivation varied with the ‘distance’ between the test and lifting digits although no significant difference occurred in the pattern of coactivation thresholds among the digital flexors. The extent of coactivation increased when angular displacement or velocity at the distal interphalangeal joint of the lifting digit increased but was not critically dependent on restraint of the hand. 4. Because mechanical ‘connections’ could interfere with the ability to move a distal phalanx independently, the arms of nine cadavers were studied. The separation of tendons between the thumb (FPL) and the index portion of FDP, and between the index and middle portions of FDP, usually extended more proximally in the forearm than separation between the tendons to the middle and ring fingers and between the ring and little fingers. Direct intertendinous links were also noted. 5. It is not possible to direct a sufficiently focal motor command to flex selectively the distal joint of the fingers and thumb when forces exceeding 2.5% MVC are generated. For the middle, ring and little fingers in particular, movement of adjacent digits may also involve ‘in‐series’ mechanical links between adjacent components of FDP.
The action of Na+ as a cofactor in the inhibition by cytoplasmic protons of the cardiac Na(+)‐Ca2+ exchanger in the guinea‐pig.
Authors: Doering A E , Lederer W J
Published: December 2014

1. Na(+)‐Ca2+ exchange current was activated in giant excised patches of guinea‐pig cardiac sarcolemma by raising the intracellular sodium concentration ([Na+]i). When the pHi was simultaneously acidified to 6.4, the current was transient, dropping by 80% in 30 s. 2. Pre‐exposure to a pHi of 6.4 for 15 s reduced the peak Na(+)‐Ca2+ exchange current without altering the decay rate or steady‐state current. Recovery from proton inhibition was seen when [Na+]i was removed for 9 s. 3. A mathematical model of Na(+)‐Ca2+ exchange function reproduced the experimental results. In addition, two model‐dependent predictions were seen experimentally. (i) [Na+]i‐dependent ‘inactivation’ of Na(+)‐Ca2+ exchange may arise from pHi effects. We observed experimentally that pre‐exposure to acidic pHi can remove the transient current component attributed to [Na+]i‐dependent ‘inactivation’. (ii) self‐exchange should be inhibited by acidification. This has been observed by other investigators. 4. We have hypothesized that there are two components to inhibition of the Na(+)‐Ca2+ exchanger by intracellular protons, and that one is enhanced by increased [Na+]i (Doering & Lederer, 1993b). This hypothesis is supported by the data presented here and by a model of Na(+)‐Ca2+ exchange behaviour in which binding of intracellular sodium to the exchanger enhances the affinity of the exchanger for inhibitory intracellular protons.
Calcium dependence of quantal secretion from visualized sympathetic nerve varicosities on the mouse vas deferens.
Authors: Macleod G T , Lavidis N A , Bennett M R
Published: December 2014

1. The effects of calcium on the secretion of quanta recorded from single varicosities on the surface of the mouse vas deferens has been determined. 2. If recordings were made from two adjacent varicosities and the [Ca2+]o increased from a low value (1 mM), then the increase in the mean quantal content of the endplate potential (m(e)) was almost entirely due to an increase in the binomial probability for secretion (pe). At higher [Ca2+]o there was an increasing tendency for the binomial parameter (ne) to increase from 1 to 2. When ne increased there was very little change in pe, indicating that the new release site recruited from the other varicosity has a relatively low probability for secretion. 3. If recordings were restricted to single varicosities and the [Ca2+]o increased in the range from 1 to 4 mM then the increases in m(e) were almost always due to an increase in pe. The gradient relating the log of m(e) to the log of [Ca2+]o for [Ca2+]o of 1.0‐1.5 mM was in the range 3.2‐5.4 (mean 4.2). 4. Test impulses gave a similar proportional increase in m(e) following a conditioning impulse at all varicosities from which recordings were made. 5. Facilitation of m(e) declined exponentially with an increase in the test‐conditioning interval from 0.5 to 4 s. The time constant for this decline was about 6 s. 6. The results indicate that single release sites show a similar fourth power dependency on [Ca2+]o and facilitate to about the same degree following a conditioning impulse.
Development of spinal reflex pathways from muscle afferents to motoneurones in chick embryos devoid of descending inputs.
Authors: Ozaki S , Kudo N
Published: December 2014

1. The synaptic connections of reflex pathways between hindlimb muscle afferents and motoneurones were investigated in chicken embryos. Descending inputs to the lumbar spinal cord were eliminated by removing two to four segments of the thoracic spinal cord at embryonic day 2 (E2; gap operation). Intracellular recordings from motoneurones innervating the lateral gastrocnemius (LG) muscle were performed in isolated spinal cord preparations of normal and gap‐operated embryos at E14‐E15. 2. In both normal and gap‐operated embryos, homonymous and synergistic muscle nerve stimulation evoked EPSPs in LG motoneurones at a short and fixed latency, suggesting that they were evoked monosynaptically. EPSPs from synergistic muscle afferents were much smaller than those from homonymous muscle afferents. The volleys from the antagonistic muscle nerve evoked IPSPs at a longer central latency than found for EPSPs in both embryos. 3. The maximal amplitudes of homonymous and synergistic EPSPs in gap‐operated embryos were 1.3 and 1.7 times, respectively, larger than in normal embryos. Homonymous EPSPs were observed in all LG motoneurones examined, but synergistic EPSPs were more common in gap‐operated than in normal embryos. 4. Antagonistic IPSPs were more common in motoneurones of gap‐operated embryos than in those of normal embryos. Homonymous and synergistic muscle nerve stimulation also elicited IPSPs in LG motoneurones in both normal and gap‐operated embryos. IPSPs evoked both by homonymous and by synergistic muscle nerve stimulation were more common in gap‐operated than in normal embryos. 5. The spatial pattern of reflex pathways from hindlimb muscle afferents to motoneurones in chick embryos devoid of both supraspinal and long propriospinal inputs to the lumbar spinal cord is similar to that in normal embryos. However, both mono‐ and polysynaptic connections in these pathways are enhanced by the blockade of descending projections.
Inhibition of inwardly rectifying K+ current by external Ca2+ ions in freshly isolated rabbit osteoclasts.
Authors: Yamashita N , Ishii T , Ogata E , Matsumoto T
Published: December 2014

1. Regulation of membrane potential by extracellular Ca2+ concentration ([Ca2+]o) was examined in freshly isolated rabbit osteoclasts. 2. The resting membrane potential of osteoclasts was close to the K+ equilibrium potential in 1 mM Ca2+ medium. An elevation of [Ca2+]o caused membrane depolarization, accompanied by a decrease in the membrane conductance. 3. The inwardly rectifying K+ current observed under voltage clamp was dose‐dependently inhibited by an elevation of [Ca2+]o, which explained the membrane depolarization caused by high [Ca2+]o. 4. Other divalent cations also inhibited the inwardly rectifying K+ current with the following order of potency: Ca2+ < Ni2+ < or = Co2+ < Cd2+. 5. In the presence of intracellular GTP gamma S the inwardly rectifying K+ current was irreversibly inhibited by [Ca2+]o, whereas the inhibition of the inwardly rectifying K+ current was greatly attenuated by intracellular application of GDP beta S. 6. Pertussis toxin (PTX) treatment did not abolish the inhibition of the inwardly rectifying K+ current caused by [Ca2+]o. 7. These results suggest that inwardly rectifying K+ channels in osteoclasts were regulated by a PTX‐insensitive G‐protein, which was coupled to the putative Ca2+ receptor or sensor on the cell membrane.
Gradation of Ca(2+)‐induced Ca2+ release by voltage‐clamp pulse duration in potentiated guinea‐pig ventricular myocytes.
Authors: Isenberg G , Han S
Published: December 2014

1. This study tests the hypothesis that whole‐cell cardiac SR Ca2+ release is graded by recruitment of independent ‘release units’. Structurally, an individual release unit may comprise ca four sarcolemmal L‐type Ca2+ channels, adjacent ryanodine‐sensitive sarcoplasmic reticulum (SR) Ca2+ release channels and the junctional gap between them. After depolarization, the first opening of a single L‐type Ca2+ channel of the unit provides sufficient Ca2+ influx to increase local [Ca2+] beyond the threshold activating Ca(2+)‐induced Ca2+ release (CICR), which amplifies local [Ca2+] until all release channels of the unit are active. This all‐or‐none activation does not spread to other release units. Gradation of whole‐cell Ca2+ release is predicted to correlate with the cumulative probability density distribution of first latency of L‐type Ca2+ channels or the activation time course of the calcium current, ICa. 2. Guinea‐pig ventricular myocytes were potentiated by paired voltage‐clamp pulses (1 Hz, 2 mM [Ca2+]o, 40 microM K5‐indo‐1, 36 degrees C). When the cellular Ca2+ load was at a steady high level, cytosolic calcium concentration ([Ca2+]c) transients were measured in response to test pulses of varied pulse duration (PD, 1‐180 ms) and amplitude (‐20, 0, 20 and 50 mV). The maximal rate of rise (RRmax) of the [Ca2+]c transient was used as an indicator for SR Ca2+ release. 3. Fast [Ca2+]c transients due to 4 ms pulses to 0 or 50 mV were blocked by 1 mM cadmium suggesting that these Ca2+ release signals are triggered by Ca2+ influx through L‐type Ca2+ channels and not by Ca2+ influx through Na(+)‐Ca2+ exchange. 4. RRmax increased with longer PD along the sigmoidal curve [1‐exp(‐PD/tau)]kappa(exponent k: 2 < k < 3). The time constant, tau, resembled the activation time constant of whole‐cell ICa (Cs(+)‐dialysed cells). A PD longer than a limiting duration did not modify RRmax. That is, inactivation of ICa was not reflected in the duration dependence. 5. Single L‐type Ca2+ channels (cell‐attached patches, 36 degrees C, ‐20 mV, 3.6 mM CaCl2 and 1 microM Bay K 8644 in patch pipette) opened with a waiting time the cumulative probability distribution of which resembled the duration dependence of RRmax, suggesting that the first opening of L‐type Ca2+ channels determines whether the corresponding release unit contributes to the [Ca2+]c transient activated during a short voltage‐clamp pulse. 6. The time constant, tau, of the duration dependence was shorter at positive than at negative potentials.(ABSTRACT TRUNCATED AT 400 WORDS)