bluuebunny/crossref_metadata_2025_split huggingface dataset:
Epoxyeicosatrienoic acids activate a high‐conductance, Ca2+‐dependent K+ channel on pig coronary artery endothelial cells
Authors: Baron A. , Frieden M. , Bény J.‐L. 
Published: September 2004

Epoxyeicosatrienoic acids (EETs) have been described as endothelium‐derived hyper‐polarizing factors (EDHFs), based on their stimulatory effects on smooth muscle K+channels. In order to reveal a putative autocrine effect of EETs on endothelial channels, we have studied the effects of the four EET regioisomers (5,6‐EET, 8,9‐EET, 11,12‐EET and 14,15‐EET) on the high‐conductance, Ca2+‐dependent K+ (BKCa) channel recorded in inside‐out patches of primary cultured pig coronary artery endothelial cells. Currents were recorded in the presence of either 500 nm or 1 μm free Ca2+ on the cytosolic side of the membrane.
In 81% of experiments, EETs at < 156 nm, applied on the cytosolic side of the membrane, transiently increased BKCa channel open state probability (P0) without affecting its unitary conductance, thus providing evidence for direct action of EETs, without involvement of a cytosolic transduction pathway.
The four EET regioisomers appeared to be equally active, multiplying the BKCa channel Po by a mean factor of 4.3 ± 0.6 (n= 15), and involving an increase in the number and duration of openings.
The EET‐induced increase in BKCa channel activity was more pronounced with low initial Po. When the BKCa channel was activated by 500 nm Ca2+, application of EETs increased the initial Po value of below 0.1 by a factor of 5. When the channel was activated by 1 μm Ca2+, application of EETs increased the initial Po value by a factor of 3.
Our results show that EETs potentiate endothelial BKCa channel activation by Ca2+. The autocrine action of EETs on endothelial cells, which occurs in the same concentration range as their action on muscle cells, should therefore fully participate in the vasoactive effects of EETs, and thus be taken into account when considering their putative EDHF function.
Amino terminal‐dependent gating of the potassium channel rat eag is compensated by a mutation in the S4 segment
Authors: Terlau Heinrich , Heinemann Stefan H. , Stühmer Walter , Pongs Olaf , Ludwig Jost 
Published: September 2004

Rat eag potassium channels (r‐eag) were expressed in Xenopus oocytes. They gave rise to delayed rectifying K+ currents with a strong Cole‐Moore effect.
Deletions in the N‐terminal structure of r‐eag either shifted the activation threshold to more negative potentials and slowed the activation kinetics (Δ2–190, Δ2–12 and Δ7–12) or resulted in a shift to more positive potentials and faster activation kinetics (Δ150–162).
The impact of the deletion Δ7–12 was investigated in more detail: it almost abolished the Cole‐Moore effect and markedly slowed down channel deactivation.
Unlike wild‐type channels, the deletion mutants Δ7–12 exhibited a rapid inactivation which, in combination with the slow deactivation, resulted in current characteristics which were similar to those of the related potassium channel HERG.
Both the slowing of deactivation and the inactivation induced by the deletion Δ7–12 were compensated by a single histidine‐to‐arginine change in the S4 segment, while this mutation (H343R) only had minor effects on the gating kinetics of the full‐length r‐eag channel.
These results demonstrate a functional role of the N‐terminus in the voltage‐dependent gating of potassium channels which is presumably mediated by an interaction of the N‐terminal protein structure with the S4 motif during the gating process.
Characterization of apical potassium channels induced in rat distal colon during potassium adaptation
Authors: Butterfield I. , Warhurst G. , Jones M. N. , Sandle G. I. 
Published: September 2004

Chronic dietary K+ loading stimulates an active K+ secretory process in rat distal colon, which involves an increase in the macroscopic apical K+ conductance of surface epithelial cells. In the present study, the abundance and characteristics of K+ channels constituting this enhanced apical K+ conductance were evaluated using patch clamp recording techniques.2. In isolated non‐polarized surface cells, K+ channels were seen in 9 of 90 (10%) cell‐attached patches in cells from control animals, and in 247 of 437 (57%) cell‐attached patches in cells from K+‐loaded animals, with a significant (P < 0.001) shift in distribution density. Similarly, recordings from cell‐attached patches of the apical membrane of surface cells surrounding the openings of distal colonic crypts revealed identical K+ channels in 1 of 11 (9%) patches in control animals, and in 9 of 13 (69%) patches in K+‐loaded animals.
In isolated surface cells and surface cells in situ, K+ channels had mean slope conductances of 209 ± 6 and 233 ± 14 pS, respectively, when inside‐out patches were bathed symmetrically in K2SO4 solution. The channels were sensitive to ‘cytosolic’ Ca2+ concentration, were voltage sensitive at ‘cytosolic’ Ca2+ concentrations encountered in colonic epithelial cells, and were inhibited by 1 mm quinidine, 20 mm TEA or 5 mm Ba2+ ions.
The data show that dietary K+ loading increases the abundance of Ca2+‐ and voltage‐sensitive large‐conductance K+ channels in the apical membrane of surface cells in rat distal colon. These channels constitute the enhanced macroscopic apical K+ conductance previously identified in these cells, and are likely to play a critical role in the active K+ secretory process that typifies this model of colonic K+ adaptation.
Contraction of the human diaphragm during rapid postural adjustments
Authors: Hodges P. W. , Butler J. E. , McKenzie D. K. , Gandevia S. C. 
Published: September 2004

The response of the diaphragm to the postural perturbation produced by rapid flexion of the shoulder to a visual stimulus was evaluated in standing subjects. Gastric, oesophageal and transdiaphragmatic pressures were measured together with intramuscular and oesophageal recordings of electromyographic activity (EMG) in the diaphragm. To assess the mechanics of contraction of the diaphragm, dynamic changes in the length of the diaphragm were measured with ultrasonography.With rapid flexion of the shoulder in response to a visual stimulus, EMG activity in the costal and crural diaphragm occurred about 20 ms prior to the onset of deltoid EMG. This anticipatory contraction occurred irrespective of the phase of respiration in which arm movement began. The onset of diaphragm EMG coincided with that of transverses abdominis.Gastric and transdiaphragmatic pressures increased in association with the rapid arm flexion by 13.8 ± 1.9 (mean ±s.e.m.) and 13.5 ± 1.8 cmH2O, respectively. The increases occurred 49 ± 4 ms after the onset of diaphragm EMG, but preceded the onset of movement of the limb by 63 ± 7 ms.Ultrasonographic measurements revealed that the costal diaphragm shortened and then lengthened progressively during the increase in transdiaphragmatic pressure.This study provides definitive evidence that the human diaphragm is involved in the control of postural stability during sudden voluntary movement of the limbs.
Molecular constituents of maxi KCa channels in human coronary smooth muscle: predominant α+β subunit complexes
Authors: Tanaka Yoshio , Meera Pratap , Song Min , Knaus Hans‐Günther , Toro Ligia 
Published: September 2004

Human large‐conductance voltage‐ and calcium‐sensitive K+ (maxi KCa) channels are composed of at least two subunits: the pore‐forming subunit, α, and a modulatory subunit, β. Expression of the β subunit induces dramatic changes in α subunit function. It increases the apparent Ca2+ sensitivity and it allows dehydrosoyasaponin I (DHS‐I) to upregulate the channel.
The functional coupling of maxi KCa channel α and β subunits in freshly dissociated human coronary smooth muscle cells was assessed. To distinguish maxi KCa currents modulated by the β subunit, we examined (a) their apparent Ca2+ sensitivity, as judged from the voltage necessary to half‐activate the channel (V1/2), and (b) their activation by DHS‐I.
In patches with unitary currents, the majority of channels were half‐activated near –85 mV at 18 μm Ca2+, a value similar to that obtained when the human KCa channel α (HSLO) and β (HKVCaβ) subunits are co‐expressed. A small number of channels half‐activated around 0 mV, suggesting the activity of the α subunit alone.
The properties of macroscopic currents were consistent with the view that most pore‐forming α subunits were coupled to β subunits, since the majority of currents had values for V1/2 near to –90 mV, and currents were potentiated by DHS‐I.
We conclude that in human coronary artery smooth muscle cells, most maxi KCa channels are composed of α and β subunits. The higher Ca2+ sensitivity of maxi KCa channels, resulting from their coupling to β subunits, suggests an important role of this channel in regulating coronary tone. Their massive activation by micromolar Ca2+ concentrations may lead to a large hyperpolarization causing profound changes in coronary blood flow and cardiac function.
Activation of f‐channels by cAMP analogues in macropatches from rabbit sino‐atrial node myocytes
Authors: Bois P. , Renaudon B. , Baruscotti M. , Lenfant J. , DiFrancesco D. 
Published: September 2004

The action of the two diastereometric phosphorothioate derivatives of cAMP, Rp‐cAMPs and Sp‐cAMPs, was investigated on hyperpolarization‐activated ‘pacemaker’ current (if) recorded in inside‐out macropatches from rabbit sino‐atrial (SA) node myocytes.
When superfused on the intracellular side of f‐channels at the concentration of 10 μm, both cAMP derivatives accelerated if activation; their action was moderately less pronounced than that due to the same concentration of cAMP.
The measurement of the if conductance‐voltage relation by voltage ramp protocols indicated that both cAMP analogues shift the activation curve of if to more positive voltages with no change in maximal (fully activated) conductance.
Dose–response relationships of the shift of the if activation curve showed that both Rp‐cAMPs and Sp‐cAMPs act as agonists in the cAMP‐dependent direct f‐channel activation. Fitting data to the Hill equation resulted in maximal shifts of 9.6 and 9.5 mV, apparent dissociation constants of 0.82 and 5.4 μm, and Hill coefficients of 0.82 and 1.12 for Sp‐cAMPs and Rp‐cAMPs, respectively.
The activating action of Rp‐cAMPs, a known antagonist of cAMP in the activation of cAMP‐dependent protein kinase, confirms previously established evidence that f‐channel activation does not involve phosphorylation. These results also suggest that the cAMP binding site of f‐channels may be structurally similar to the cyclic nucleotide binding site of olfactory receptor channels.
Electrical resonance and Ca2+ influx in the synaptic terminal of depolarizing bipolar cells from the Goldfish retina
Authors: Burrone Juan , Lagnado Leon 
Published: September 2004

Whole‐cell recordings and fura‐2 measurements of cytoplasmic [Ca2+] were made in depolarizing bipolar cells isolated from the retinae of goldfish. The aim was to study the voltage signal that regulates Ca2+ influx in the synaptic terminal.
The current‐voltage relation was linear up to about −44 mV. At this threshold, the injection of 1 pA of current triggered a maintained ‘all‐or‐none’ depolarization to a plateau of −34 mV, associated with a decrease in input resistance and a damped voltage oscillation with a frequency of 50–70 Hz and initial amplitude of 4–10 mV. A second frequency component of 5–10 Hz was often observed. In a minority of cells the response to current injection was transient, recovering with an undershoot.
Unstimulated bipolar cells generated similar voltage signals, driven by current entering the cell through a non‐specific cation conductance that continuously varied in amplitude.
The threshold for activation of the Ca2+ current was −43 mV and free [Ca2+]i in the synaptic terminal rose during a depolarizing response. Simultaneous measurements of the fluorescence associated with the membrane marker FM1–43 demonstrated that these Ca2+ signals stimulated exocytosis. Regenerative depolarizations and associated rises in [Ca2+]i were blocked by inhibiting l‐type Ca2+ channels with 30 μm nifedipine.
Depolarization beyond −40 mV also elicited an outwardly rectifying K+ current. Blocking this current by replacing external Ca2+ with Ba2+ caused the voltage reached during a depolarizing response to increase to +10 mV.
The majority of the K+ current was blocked by 100 nm charybdotoxin, indicating that it was carried by large‐conductance Ca2+‐activated K+ channels. A transient voltage‐gated K2+current remained, which began to activate at −40 mV. High‐frequency voltage oscillations were blocked by 100 nm charybdotoxin, but low‐frequency oscillations remained.
These results indicate that the voltage response of depolarizing bipolar cells is shaped by l‐type Ca2+ channels, Ca+‐activated K+ channels and voltage‐dependent K+ channels. This combination of conductances regulates Ca2+ influx into the synaptic terminal and confers an electrical resonance on the bipolar cell.
Dual actions of tetracaine on intramembrane charge in amphibian striated muscle
Authors: Huang Christopher L.H. 
Published: September 2004

The effects of graded concentrations of tetracaine on the steady‐state and kinetic properties of intramembrane charge were examined in intact voltage‐clamped amphibian muscle fibres.
The micromolar tetracaine concentrations that were hitherto reported to abolish Ca2+ transients in skeletal muscle failed to affect significantly the steady‐state charge. Maximal reductions of such intramembrane charge required relatively high, 1–2 mm, concentrations of tetracaine.
The plots of maximum charge against tetracaine concentration suggested a saturable 1:1 drug binding that spared a fixed amount of tetracaine‐resistant (qβ) charge but inhibited a discrete fraction of susceptible (qγ) charge with a KD between 0.1 and 0.2 mm.
The qβ charge thus isolated by 2 mm tetracaine was conserved through a wide range of applied test voltages and pulse durations and regardless of whether the imposed transition from the holding potential (−90 mV) to the test potential took place in one or more steps.
Similarly, ‘on’ and ‘off’qβ currents that were elicited by voltage steps from fixed conditioning to varying test levels mapped onto non‐linear phase‐plane trajectories that nevertheless depended uniquely upon voltage. In contrast, the currents that followed voltage steps made from varying prepulse levels to fixed −90 or −20 mV test potentials mapped onto identical qβ phase‐plane trajectories that were independent of the prepulse history.
The charge movements that followed strong depolarizing voltage clamp steps to test potentials in the range −50 to 0 mV approximated simple monotonic decays that could empirically be described by a single time constant. Nevertheless, a complete inhibition of a tetracaine‐sensitive (qγ) charge movement by 2 mm tetracaine that left only qγ charge, sharply altered both the magnitude and the voltage dependence of these time constants. This establishes a distinct contribution of the qγ species to overall charge kinetics even at such test voltages.
Under such a criterion for the voltage dependence of charging kinetics, even the micromolar (0.05‐0.2mm) tetracaine concentrations that failed to markedly alter the steady‐state charge consistently increased the charging time constants yet did not influence their voltage sensitivity.
These findings demonstrate the existence of separate kinetic and steady‐state effects of tetracaine on intramembrane charge movements, at micromolar and millimolar anaesthetic concentrations, respectively. These parallel earlier effects of tetracaine that have been reported upon the transient and sustained components of sarcoplasmic reticular Ca2+ release. They also establish that maximally effective concentrations of tetracaine isolate a single distinct species of conserved (qβ) intramembrane charge.
A novel slow hyperpolarization‐activated potassium current (IK(SHA)) from a mouse hippocampal cell line
Authors: Wischmeyer Erhard , Karschin Andreas 
Published: September 2004

A slow hyperpolarization‐activated inwardly rectifying K+ current (IK(SHA)) with novel characteristics was identified from the mouse embryonic hippocampus x neuroblastoma cell line HN9.10e.
The non‐inactivating current activated negative to a membrane potential of −80 mV with slow and complex activation kinetics (τact≈ 1–7 s) and a characteristic delay of 1–10 s (−80 to −140 mV) that was linearly dependent on the membrane potential.
Tail currents and instantaneous open channel currents determined through fast voltage ramps reversed at the K+ equilibrium potential (EK) indicating that primarily K+, but not Na+, permeated the channels.
I
K(SHA) was unaffected by altering the intracellular Ca2+ concentration between ∼0 and 10 μm, but was susceptible to block by 5 mm extracellular Ca2+, Ba2+ (K1= 0.42 mm), and Cs+ (K1= 2.77 mm)
In cells stably transformed with M2 muscarinic receptors, IK(SHA) was rapidly, but reversibly, suppressed by application of micromolar concentrations of muscarine.
At the single channel level K(SHA) channel openings were observed with the characteristic delay upon membrane hyperpolarization. Analysis of unitary currents revealed an inwardly rectifying I–V profile and a channel slope conductance of 7 pS. Channel activity persisted in the inside‐out configuration for many minutes.
It is concluded that IK(SHA) m HN9.10e cells represents a novel K+ current, which is activated upon membrane hyperpolarization. It is functionally different from both classic inwardly rectifying Ikir currents and other cationic hyperpolarization‐activated 7H currents that have been previously described in neuronal or glial cells.
Synapse Formation and Hypoxic Signalling in Co‐Cultures of Rat Petrosal Neurones and Carotid Body Type 1 Cells
Authors: Zhong Huijun , Zhang Min , Nurse Colin A. 
Published: September 2004

To investigate synaptic mechanisms mediating chemosensory signalling in the carotid body, we developed co‐cultures of chemoreceptor type 1 cell clusters and dissociated petrosal neurones (PNs) from 7‐ to 14‐day‐old rat pups and tested for functional connectivity in CO2–HCO3− ‐or Hepes‐buffered medium at ∼35 °C.
When cultured without type 1 cells, PNs were almost always quiescent (n= 104) and unresponsive to hypoxia (PO2= 5–25 mmHg) during perforated patch, whole‐cell recordings of membrane potential or voltage‐activated currents; in contrast, many PNs (77 out of 170) that were juxtaposed to type 1 cell clusters in co‐culture displayed spontaneous activity, comprising spikes and subthreshold potentials (SSPs) that resembled synaptic potentials.
Additional tests suggested that de novo chemical synapses developed between PNs and type 1 cell clusters in vitro. For example: (i) the spontaneous activity was reversibly suppressed by substituting low calcium‐high magnesium in the bath; (ii) SSPs had variable amplitudes and persisted following action potential blockade with TTX (1 μm); (iii) the interval distribution between successive spontaneous events appeared random; and (iv) the frequency of spontaneous potentials was diminished (reversibly) by the nicotinic antagonist hexamethonium (100 /tM), suggesting contributions from the spontaneous release of ACh.
Many complexes of ‘juxtaposed’ PNs and type 1 clusters were physiologically functional, since exposure to hypoxia caused a reversible depolarization and/or increased spike discharge in ∼30% of such neurones (n= 140). The hypoxia‐induced spike discharge persisted in the presence of the dopamine D2 receptor blocker spiperone (10–50 μm; n= 5); however, this discharge was reversibly inhibited by 100–200 μm hexamethonium, suggesting that it was mediated, at least in part, by ACh acting through nicotinic receptors.
The hypoxia‐induced spike discharge and frequency of spontaneous potentials in co‐cultured PNs were reversibly suppressed when the buffer was switched from C02‐HC03∼ to Hepes (10 mm) at pH 7.4;′further, ‘functional’ PNs that displayed spontaneous activity and/or hypoxia‐induced responses in co‐culture were encountered more frequently in CO2–HCO3− (≥ 40 %) than in Hepes (≤26 %) buffer.
We conclude that functional chemical synapses can develop de novo in cultures of carotid body type 1 cells and PNs and that ACh is probably an important excitatory neuro‐transmitter secreted from type 1 cells during hypoxic chemotransduction in the rat carotid body.
Functional nicotinic ACh receptors on interneurones in the rat hippocampus
Authors: Jones Susan , Yakel Jerrel L. 
Published: September 2004

Neuronal nicotinic ACh receptors (nAChRs) were studied in the rat hippocampal slice preparation using whole‐cell patch‐clamp recording techniques.
Responses to ACh (100 μm) were detected on inhibitory interneurones in the CA1 field of the hippocampus proper and in the dentate gyrus, but not on principal excitatory neurones in either region. The different neuronal types were identified based on their morphology and location.
ACh excited interneurones in the hippocampus and dentate gyrus in current‐clamp recordings. In voltage‐clamp recordings, ACh‐activated inward currents were recorded from interneurones in the presence of blockers of synaptic transmission and the muscarinic ACh receptor antagonist atropine. The zero current potential for this response to ACh was near 0 mV.
The effect of ACh was mimicked by the nAChR‐selective agonists nicotine (100 μm) and 1,1‐dimethyl‐4‐phenyl‐piperazinium iodide (DMPP, 100 μm). The response to ACh was reversibly antagonized by the neuronal nAChR antagonist mecamylamine (10 μm). The nAChR α7 subunit‐selective antagonists α‐bungarotoxin (100 nm) and methyllycaconitine (10 nm) also inhibited the response to ACh.
These observations demonstrate the presence of functional nAChRs on inhibitory interneurones in the rat hippocampus. Thus, a novel mechanism by which ACh can regulate neuronal activity in the hippocampus is revealed.
Calcium action potentials restricted to distal apical dendrites of rat neocortical pyramidal neurons
Authors: Schiller Jackie , Schiller Yitzhak , Stuart Greg , Sakmann Bert 
Published: September 2004

Simultaneous whole‐cell voltage and Ca2+ fluorescence measurements were made from the distal apical dendrites and the soma of thick tufted pyramidal neurons in layer 5 of 4‐week‐old (P28–32) rat neocortex slices to investigate whether activation of distal synaptic inputs can initiate regenerative responses in dendrites.
Dual whole‐cell voltage recordings from the distal apical trunk and primary tuft branches (540–940 μm distal to the soma) showed that distal synaptic stimulation (upper layer 2) evoking a subthreshold depolarization at the soma could initiate regenerative potentials in distal branches of the apical tuft which were either graded or all‐or‐none. These regenerative potentials did not propagate actively to the soma and axon.
Calcium fluorescence measurements along the apical dendrites indicated that the regenerative potentials were associated with a transient increase in the concentration of intracellular free calcium ([Ca2+]i) restricted to distal dendrites.
Cadmium added to the bath solution blocked both the all‐or‐none dendritic regenerative potentials and local dendritic [Ca2+]i transients evoked by distal dendritic current injection. Thus, the regenerative potentials in distal dendrites represent local Ca2+ action potentials.
Initiation of distal Ca2+ action potentials by a synaptic stimulus required coactivation of AMPA‐ and NMDA‐type glutamate receptor channels.
It is concluded that in neocortical layer 5 pyramidal neurons of P28–32 animals glutamatergic synaptic inputs to the distal apical dendrites can be amplified via local Ca2+ action potentials which do not reach threshold for axonal AP initiation. As amplification of distal excitatory synaptic input is associated with a localized increase in [Ca2+]i these Ca2+ action potentials could control the synaptic efficacy of the distal cortico‐cortical inputs to layer 5 pyramidal neurons.