bluuebunny/crossref_metadata_2025_split huggingface dataset:
Electrophysiological properties of sodium current subtypes in small cells from adult rat dorsal root ganglia
Authors: Rush A. M. , Bräu M. E. , Elliott A. A. , Elliott J. R. 
Published: September 2004

Whole‐cell and single‐channel Na+ currents were recorded from small (ca. 20 μm diameter) cells isolated from adult rat dorsal root ganglia (DRG). Currents were classified by their sensitivity to 0.3 μm tetrodotoxin (TTX), electrophysiological properties and single‐channel amplitude. Cells were classified according to the types of current recorded from them.
Type A cells expressed essentially pure TTX‐sensitive (TTX‐S) currents. Availability experiments with prepulse durations between 50 ms and 1 s gave a half‐available voltage (Vh) of around ‐65 mV but the availability curves often had a complex shape, consistent with multiple inactivation processes. Measured inactivation time constants ranged from less than 1 ms to over 100 s, depending on the protocol used.
Cell types B and C each had, in addition to TTX‐S currents, substantial and different TTX‐resistant (TTX‐R) currents that we have designated TTX‐R1 and TTX‐R2, respectively. TTX‐R1 currents had a 1 s Vh of ‐29 mV, showed little 1 Hz use dependence at ‐67 mV and recovered from the inactivation induced by a 60 ms depolarizing pulse with time constants of 1.6 ms (91 %) and 908 ms. They also exhibited slow inactivation processes with component time constants around 10 and 100 s. TTX‐R2 currents activated and inactivated at more negative potentials (1 s Vh= ‐46 mV), showed substantial 1 Hz use dependence and had inactivation (60 ms pulse) recovery time constants at ‐67 mV of 3.3 ms (58 %) and 902 ms.
Type D cells had little or no current in 0.3 μm TTX at a holding potential of ‐67 mV. Current amplitude increased on changing the holding potential to ‐107 mV. Type D cell currents had more hyperpolarized availability and I‐V curves than even TTX‐R2 currents and suggest the existence of TTX‐R3 channels.
In outside‐out patches with 250 mM external NaCl, the single‐channel conductance (γ) of TTX‐S channels was 19.5 pS and the potential for half‐maximal activation (Va) was ‐45 mV. One population of TTX‐R channels had a γ of 9.2 pS and a Va of ‐27 mV. A second population had a γ of 16.5 pS and a more negative Va of ‐42 mV. The latter population may underlie the type D cell current.
Small DRG cells express multiple Na+ currents with varied time constants and voltage dependences of activation and inactivation. Nociceptive cells still fire when chronically depolarized by an increased external K+ concentration. TTX‐R1 and TTX‐R2 Na+ channels may support that firing, while the range of inactivation time constants described here would increase the repertoire of DRG cell burst firing behaviour generally.
Cat cerebral arterial smooth muscle cells express cytochrome P450 4A2 enzyme and produce the vasoconstrictor 20‐HETE which enhances L‐type Ca2+ current
Authors: Gebremedhin Debebe , Lange Andrew R. , Narayanan Jayashree , Aebly Mikael R. , Jacobs Elizabeth R. , Harder David R. 
Published: September 2004

Cerebral arteries express cytochrome P450 4A enzymes (P450 4A) and produce 20‐ hydroxyeicosatetraenoic acid (20‐HETE), a potent constrictor of pial arterioles. It is not known which cell type in the vessel wall is responsible for the formation of 20‐HETE. We examined whether freshly isolated cerebral arterial muscle cells (VSMCs) express P450 4A and produce 20‐HETE. We also studied the effect of 20‐HETE on pressurized cerebral arteries and on whole‐cell L‐type Ca2+current (ICa) recorded in cat cerebral VSMCs.
Cat cerebral VSMCs incubated with [14C]arachidonic acid ([14C]AA) produced 20‐HETE (3.9 ± 1.1 pmol min−1 (mg protein)−1).
Reverse transcription‐polymerase chain reaction studies revealed that cat cerebral VSMCs express mRNA for P450 4A which metabolizes AA to 20‐HETE. Cloning and sequencing of the cDNA amplified from mRNA isolated from VSMCs showed > 96 % amino acid homology to the rat and human P450 4A2 and 4A3.
20‐HETE (1–300 nM) induced a concentration‐dependent constriction of cat cerebral arteries, which was inhibited by nifedipine.
Addition of 10 and 100 nM 20‐HETE to the bath increased peak ICa by 50 ± 3 and 100 ± 10 %, respectively. This effect was not influenced by altering the frequency of depolarization. 20‐HETE (100 nM) failed to increase ICa in the presence of nifedipine.
These results demonstrate that cat cerebral VSMCs express P450 4A enzyme, and produce 20‐HETE which activates L‐type Ca2+ channel current to promote cerebral vasoconstriction.
Angiotensin AT1 receptor‐mediated excitation of rat carotid body chemoreceptor afferent activity
Authors: Allen A. M. 
Published: September 2004

A high density of angiotensin II receptors was observed in the rat carotid body by in vitro autoradiography employing 125I‐[Sar1,Ile8]‐angiotensin II as radioligand. Displacement studies demonstrated that the receptors were of the AT1 subtype.
The binding pattern indicated that the AT1 receptors occurred over clumps of glomus cells, the principal chemoreceptor cell of the carotid body. Selective lesions of the sympathetic or afferent innervation of the carotid body had little effect on the density of receptor binding, demonstrating that the majority of AT1 receptors were intrinsic to the glomus cells.
To determine the direct effect of angiotensin II on chemoreceptor function, without the confounding effects of the vasoconstrictor action of angiotensin II, carotid sinus nerve activity was recorded from the isolated carotid body in vitro. The carotid body was superfused with Tyrode solution saturated with carbogen (95 % O2, 5 % CO2), maintained at 36 °C, and multi‐unit nerve activity recorded with a suction electrode.
Angiotensin II elicited a dose‐dependent excitation of carotid sinus nerve activity (maximum increase of 36 ± 11 % with 10 nM angiotensin II) with a threshold concentration of 1 nM. The response was blocked by the addition of an AT1 receptor antagonist, losartan (1 μM), but not by the addition of an AT2 receptor antagonist, PD123319 (1 μM).
In approximately 50 % of experiments the excitation was preceded by an inhibition of activity (maximum decrease of 24 ± 8 % with 10 nM angiotensin II). This inhibitory response was markedly attenuated by losartan but not affected by PD123319.
These observations demonstrate that angiotensin II, acting through AT1 receptors located on glomus cells in the carotid body, can directly alter carotid chemoreceptor afferent activity. This provides a means whereby humoral information about fluid and electrolyte homeostasis might influence control of cardiorespiratory function.
Low K+‐induced hyperpolarizations trigger transient depolarizations and action potentials in rabbit ventricular myocytes
Authors: Akuzawa‐Tateyama M. , Tateyama M. , Ochi R. 
Published: September 2004

The effects of large reductions of [K+]o on membrane potential were studied in isolated rabbit ventricular myocytes using the whole‐cell patch clamp technique.
Decreasing [K+]o from the normal level of 5.4 mm to 0.1 mm increased resting membrane potential (Vrest) from −75.6 ± 0.3 to −140.3 ± 1.9 mV (means ± s.e.m; n= 127), induced irregular, transient depolarizations with mean maximal amplitudes of 19.5 ± 1.5 mV and elicited action potentials in 56.7 % of trials. The action potentials exhibited overshoots of 37.9 ± 1.5 mV (n= 72) and sustained plateaux.
Addition of 0.1 mm La3+ in the presence of 0.1 mm[K+]o significantly increased Vrest but decreased the amplitude of transient depolarizations and suppressed the firing of action potentials.
Replacement of external Na+ or Cl− with N‐methyl‐D‐glucamine or aspartate, respectively, or internal dialysis with 10 mm EGTA or BAPTA had little effect on low [K+]o‐induced membrane potential changes.
Hyperpolarizing voltage clamp pulses to potentials between −110 and −200 mV activated irregular inward currents that increased in amplitude and frequency with increasing hyperpolarization and were depressed by 0.1 mm La3+.
The generation of transient depolarizations by low [K+]o can be explained as being a consequence of decreasing the inward rectifier K+ current (IK1) and the appearance of inward currents reflecting electroporation resulting from strong electric fields across the membrane.
Cytochrome P450: a novel system modulating Ca2+ channels and contraction in mammalian heart cells
Authors: Xiao Yong‐Fu , Huang Lin , Morgan James P. 
Published: September 2004

Cytochrome P450 (P450) is a ubiquitous enzyme system that catalyses oxidative reactions of numerous endogenous and exogenous compounds. The modulatory effects of P450 on the L‐type Ca2+ current (ICa), intracellular free Ca2+ signals and cell shortening were assessed in adult rat single ventricular myocytes.
Bath administration of the imidazole antimycotics, clotrimazole, econazole and miconazole, which are potent P450 inhibitors, significantly suppressed cardiac ICa. While the Ca2+ channel antagonist nifedipine blocked ICa within 30 s, clotrimazole‐induced suppression of ICa required 5.1 ± 0.4 min (n= 14) to reach a steady low level. The suppression of ICa was dose dependent and recovered after washout of clotrimazole. Intracellular dialysis with the P450 antibody anti‐rat CYP1A2 also significantly reduced cardiac ICa.
Additional administration of the β‐adrenergic agonist isoprenaline (1 μM) or the membrane‐permeable 8‐bromo‐cAMP (2 mM) completely reversed the suppressant effects of clotrimazole and NaCN on ICa. In addition, intracellular dialysis with 2 mM cAMP abolished the P450 inhibitor‐induced suppression of ICa. Phosphorylation of the channel with hydrolysis‐resistant ATPγS prevented the suppressant effect of clotrimazole on ICa. Furthermore, dephosphorylation of the Ca2+ channel with intracellular dialysis with phosphatase types I and II reduced ICa by 85 ± 3 % and abolished clotrimazole‐induced suppression of ICa.
Extracellular administration of the phospholipase A2 inhibitors mepacrine and 4‐bromophenacyl bromide significantly suppressed ICa.
Clotrimazole, econazole, miconazole and CN− also significantly inhibited intracellular free Ca2+ signals and cell shortening in rat single ventricular myocytes.
Intracellular cAMP content was significantly reduced in isolated ventricular myocytes incubated with clotrimazole or CN−. Extracellular administration of 11, 12‐epoxyeicosatrienoic acid, one of the P450‐mediated metabolites of arachidonic acid, enhanced ICa and intracellular cAMP content. The epoxyeicosatrienoic acid also restored the amplitude of the reduced ICa in P450 antibody‐dialysed myocytes.
The present data suggest that cytochrome P450 modulates cardiac ICa and cell contraction, and the modulation may result from changes in intracellular levels of cAMP by P450‐ mediated metabolites of arachidonic acid.
Mutual occlusion of P2X ATP receptors and nicotinic receptors on sympathetic neurons of the guinea‐pig
Authors: Searl T. J. , Redman R. S. , Silinsky E. M. 
Published: September 2004

The interaction of ion channels activated by nicotinic receptor agonists with ion channels gated by extracellular ATP (i.e. P2X receptors) was studied on sympathetic neurons acutely dissociated from coeliac ganglia of the guinea‐pig. Patch clamp methods were used to measure the inward current generated through these non‐selective cationic channels under voltage clamp.
At the whole cell level, the specific nicotinic receptor agonists nicotine (5‐100 μM) or cytisine (50‐75 μM) and the P2X receptor agonists ATP (0.1‐7 μM) or α,β‐methylene ATP (6 μM) were examined separately and in the presence of the other receptor activator. When a nicotinic and P2X receptor agonist were applied together, mutually occlusive effects were generally observed. This occurred even with concentrations of agonists that in themselves generated little to no inward current.
The occlusive effects of nicotinic agonists on ATP‐gated currents were blocked by the nicotinic receptor/ion channel blocker hexamethonium (150 μM). The occlusive effects of ATP analogues on inward currents generated by nicotinic agonists were blocked by the P2X receptor antagonist suramin (100 μM).
Mutual occlusion of the effects of nicotinic agonists and ATP analogues were also observed when currents through single channels were studied in excised (outside‐out) patches.
The results suggest that nicotinic receptors and P2X ATP receptors do not act independently in these sympathetic neurons.
Laminar organization of epileptiform discharges in the rat entorhinal cortex in vitro
Authors: Lopantsev Valeri , Avoli Massimo 
Published: September 2004

Interictal and ictal epileptiform discharges induced by 4‐aminopyridine (4AP, 50 μM) were studied in the rat lateral entorhinal cortex with field potential and intracellular recordings in an in vitro slice preparation. Both types of discharge disappeared in layer II, but continued to occur in layers IV‐VI after a knife cut separation was made at approximately 600 μm from the pia (n= 4 slices).
Interictal depolarizations recorded in layer IV‐VI cells (amplitude, 29.4 ± 8.6 mV; duration, 386 ± 177.4 ms, means ± s.d.; n= 17) were capped by action potential bursts, while smaller interictal depolarizations in layer II cells (amplitude, 11.7 ± 5.8 mV; duration, 192.6 ± 47.9 ms; n= 10) were associated with single action potentials and were terminated by a hyperpolarization. Ictal discharges were initiated by an interictal discharge; they were characterized by a depolarization of 31.5 ± 6.2 mV (n= 12) in layer IV‐VI and 11.6 ± 3.5 mV (n= 7) in layer II neurones.
Slow, presumptive Ca2+‐mediated spikes occurred in layer II (n= 4) and IV‐VI (n= 6) cells loaded with the Na+ channel blocker QX‐314 (50 mM). These events were synchronized with population spikes during interictal and ictal discharges, and were abolished by Ni2+ (1 mM, n= 4 cells) along with the 4AP‐induced synchronous activity.
The N‐methyl‐D‐aspartate (NMDA) receptor antagonist 3,3‐(2‐carboxypiperazine‐4‐yl)‐propyl‐1‐phosphonate (CPP, 10 μM) abolished ictal discharges and reduced interictal depolarizations in layer IV‐VI neurones (n= 4). The non‐NMDA receptor antagonist 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX, 10 μM) abolished both interictal and ictal activity (n= 4 cells).
These findings provide evidence for a role played by NMDA‐mediated mechanisms in the generation of epileptiform discharges in the entorhinal cortex. Lack of an NMDA‐mediated component along with presence of inhibition in layer II neurones results in attenuation of epileptiform activity at this site. Moreover Ca2+‐mediated spikes may contribute to the appearance of epileptiform discharges in this model.
ADP and inositol trisphosphate evoke oscillations of a monovalent cation conductance in rat megakaryocytes
Authors: Hussain Jamila F. , Mahaut‐Smith Martyn P. 
Published: September 2004

A combination of conventional whole‐cell patch clamp recordings and fura‐2 fluorescence photometry was used to study the membrane currents during oscillations of intracellular Ca2+concentration ([Ca2+]i) in single rat megakaryocytes.At a holding potential of ‐60 mV, in NaCl external saline and KCl internal saline with low levels of Ca2+buffering, 10 μmADP evoked [Ca2+]ioscillations and simultaneous Ca2+‐gated K+currents at a frequency of 3–10 spikes min−1. A smaller inward current was also activated, with a time course that identified this component as the inositol 1,4,5‐trisphosphate (IP3)‐activated monovalent cation current previously demonstrated in rat megakaryocytes.Cs+replacement of internal K+combined with 100 nM external charybdotoxin (CTX) abolished the outward currents and revealed that an inward current was also transiently activated during each [Ca2+]ispike. This underlying conductance was permeable to Na+and Cs+, but possessed little or no permeability to Cl−or divalent cations.Intracellular dialysis with IP3(5‐50 μm) activated the monovalent cationic conductance prior to release of Ca2+from intracellular stores. The [Ca2+]iincrease was associated with a second phase of cationic current, implying that both IP3and Ca2+can activate this conductance. Buffering of [Ca2+]iwith BAPTA abolished the second phase of current, leaving monophasic spikes of inward current, often occurring at regular intervals.These data demonstrate that a monovalent cation current, which results in Na+influx under normal ionic conditions, oscillates in response to ADP receptor stimulation due to activation by both IP3and [Ca2+]i. This provides a route for long‐term Na+entry in the megakaryocyte following stimulation of receptors coupled to phospholipase C activation and may play a role in cell shape change.
NMDA and GABAAreceptors in the rat Kölliker‐Fuse area control cardiorespiratory responses evoked by trigeminal ethmoidal nerve stimulation
Authors: Dutschmann Mathias , Herbert Horst 
Published: September 2004

Electrical stimulation (10 s) of the ethmoidal nerve (EN5) evokes the nasotrigeminal reflex responses, including apnoea, bradycardia and rise in arterial blood pressure. In the present study, we examined the involvement ofN‐methyl‐D‐aspartate (NMDA), AMPA/kainate, (γ‐aminobutyric acidA(GABAA) and glycine receptors in the Kölliker‐Fuse (KF) nucleus in the mediation of the nasotrigeminal reflex responses.Unilateral injections (n= 6) of 50‐100 nl of the NMDA receptor antagonist AP5 into the KF area led to a significant blockade of the EN5‐evoked respiratory depression and bradycardia. Injections placed into the midlevel of the KF area were most effective (80‐90 % blockade). The rise in arterial blood pressure remained unaffected.Unilateral injections (n= 6) of the AMPA/kainate receptor antagonist CNQX into the KF area failed to block EN5‐evoked autonomic responses significantly.Unilateral injections (n= 5) of the GABAAreceptor antagonist bicuculline enhanced the EN5‐evoked respiratory depression and bradycardia. The effect persisted for up to 30 s after stimulation. Bicuculline injections into the midlevel of the KF area were most effective. The increase in arterial blood pressure remained unaffected.Unilateral injections (n= 5) of the glycine receptor antagonist strychnine into the KF area did not produce any significant effects on EN5‐evoked autonomic responses.Our results suggest that the KF area represents a mandatory relay for the nasotrigeminally induced apnoea and bradycardia which are predominantly mediated by NMDA receptors in the KF. Furthermore, it appears that KF neurons are under a potent GABAergic inhibitory control. The EN5‐evoked rise in arterial blood pressure was not altered by any of the drugs and, therefore, appears not to be mediated via the KF.
Streptomycin inhibition of myogenic tone, K+‐induced force and block of L‐type calcium current in rat cerebral arteries
Authors: Miller A. L. , Langton P. D. 
Published: September 2004

Streptomycin has been demonstrated to inhibit mechanosensitive conductances in a wide variety of cell types, including muscle. The action of streptomycin on rat cerebral arteries that exhibit pressure‐induced myogenic response was investigated.
Pressure‐induced tone, measured using isobaric myography, in isolated pressurized cerebral arteries was reversibly and concentration‐dependently inhibited by streptomycin with an IC50 of 2.6 mM.
Isometric K+‐induced force, measured using isometric myography, is supported by voltage‐gated Ca2+ entry. Streptomycin reversibly and concentration‐dependently inhibited isometric force with an IC50 of 1.71 mM.
Voltage‐gated macroscopic inward Ca2+ channel currents were recorded from freshly isolated rat basilar myocytes. These were reversibly and concentration‐dependently inhibited by streptomycin with an IC50 of 1.79 and 0.47 mM when 10 and 1.8 mM CaCl2, respectively, was used as the charge carrier.
These data suggest that streptomycin inhibits myogenic tone and K+‐induced isometric force largely by blockade of L‐type, dihydropyridine‐sensitive Ca2+ channels. In conclusion, streptomycin is not useful in the investigation of stretch‐activated channels which may underlie the myogenic response of rat small cerebral arteries.
ATP‐activated cation currents in single guinea‐pig hepatocytes
Authors: Capiod Thierry 
Published: September 2004

Responses of single guinea‐pig liver cells to the application of external ATP were studied using the whole‐cell voltage clamp technique.
When the cells were loaded with 5 mM EGTA in the absence of K+ and Cl− in both internal and external solutions, application of ATP (0.03‐100 μM) elicited a large cation‐selective inward current at negative holding potentials. The current densities at the peak of the response to 100 μM ATP were 4.5 ± 0.5 pA pF−1 (mean ± s.e.m., n= 18) in the presence of Na+ and Ca2+ in the external medium and 3.3 ± 0.7 pA pF−1 (n= 6) with Ca2+ as the major permeant ion.
Divalent cations, when added during the response to ATP in the presence of Na+ and Ca2+, exerted different effects: CdSO4 (2 mM) totally and NiSO4 (2 mM) partially blocked the inward current whereas MnSO4 (2 mM) did not block it. The ATP‐activated conductance was permeable to all the divalent cations tested in this study, i.e. Ca2+, Cd2+, Ni2+, Mn2+ and Mg2+. No response to ATP was observed in the absence of external cations.
The activation of the inward current was not maintained in the continuous presence of ATP. The effect of Ca2+ ions on the desensitization of the response was studied in different external solutions. The decline in the amplitude of the inward current after the peak was fitted with a single exponential with a time constant of about 2 s for pure Ca2+, Cd2+ or Ni2+ currents, 3 s for Mg2+ or Mn2+ and 4 s in the presence of both Na+ and Ca2+.
Under more physiological conditions, the entry of Ca2+ evoked after the stimulation of P2X purinoceptors was associated with an increase in fluo‐3 fluorescence and a marked reduction in the delay before the mobilization of internal Ca2+ stores following the activation of P2Y purinoceptors.
Fundamental calcium release events revealed by two‐photon excitation photolysis of caged calcium in guinea‐pig cardiac myocytes
Authors: Lipp Peter , Niggli Ernst 
Published: September 2004

In cardiac muscle, ‘Ca2+ sparks’ have been proposed to underlie Ca2+‐induced Ca2+ release (CICR), and to result from openings of clusters of Ca2+ channels (ryanodine receptors; RyRs) located in the sarcoplasmic reticulum membrane.
To investigate the elementary nature of these Ca2+ signals directly, a diffraction‐limited point source of Ca2+ was created in single cardiac myocytes by two‐photon excitation photolysis of caged Ca2+. Simultaneously, concentration profiles of released Ca2+ were imaged at high temporal and spatial resolution with a laser‐scanning confocal microscope.
This approach enabled us to generate and detect photolytic Ca2+ signals that closely resembled the Ca2+ sparks occurring naturally, not only in amplitude and size, but also in their ability to trigger additional Ca2+ sparks or Ca2+ waves.
Surprisingly, at low photolytic power minuscule events with estimated Ca2+ release fluxes 20‐40 times smaller than those calculated for a typical Ca2+ spark were directly resolved. These events appeared to arise from the opening of a more limited number of RyRs (possibly one) or from RyRs exhibiting a different gating mode and may correspond to the elusive ‘Ca2+ quark’.
The Ca2+ quark represents the fundamental Ca2+ release event of excitable cells implementing hierarchical Ca2+ signalling systems with Ca2+ release events of various but distinct amplitude levels (i.e. Ca2+ quarks, Ca2+ sparks and full cellular Ca2+ transients).
A graded recruitment of nanoscopic Ca2+ release domains (i.e. Ca2+ quarks) exhibiting variable degrees of spatial coherence and coupling may then build up intermediate Ca2+ signalling events (i.e. Ca2+ sparks). This mechanism suggests the existence of Ca2+ sparks caused by gating of a variable fraction of RyRs from within an individual cluster. Additional mobilization of a variable number of these Ca2+ sparks enables cardiac cells to show graded cellular Ca2+ transients. Similar recruitment processes may underlie regulation of Ca2+ signalling on the cellular level in general.