ATP activates a cation conductance and Ca(2+)-dependent Cl- conductance in Hensen cells of guinea pig cochlea

1996 ◽  
Vol 271 (6) ◽  
pp. C1817-C1827 ◽  
Author(s):  
M. Sugasawa ◽  
C. Erostegui ◽  
C. Blanchet ◽  
D. Dulon
Keyword(s):  
Guinea Pig ◽  
Organ Of Corti ◽  
Reversal Potential ◽  
Initial Current ◽  
Fluorescence Measurements ◽  
Whole Cell Patch Clamp ◽  
Cation Conductance ◽  
Cl Channels ◽  
Hensen Cells ◽  
Cl Conductance

Simultaneous whole cell patch-clamp and indo 1 fluorescence measurements were used to characterize ATP-evoked membrane currents and intracellular Ca2+ concentration ([Ca2+]i) changes in isolated Hensen cells of the guinea pig organ of Corti. At negative holding potential, ATP activated a biphasic inward current and a concomitant increase in [Ca2+]i. The initial current activated within < 50 ms, showed a reversal potential near 0 mV and was reversibly inhibited by 30 microM suramin, suggesting this conductance was mediated by ATP-gated nonselective cation channels. The delayed ATP-activated current was mainly carried by Cl- as indicated by its shift in reversal potential when intracellular Cl- was replaced by gluconate. This Cl- conductance appeared to be Ca(2+)-activated secondarily to Ca2+ influx, since it required the presence of extracellular Ca2+ and was suppressed when an intracellular solution containing 10 mM 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid was used. In the absence of extracellular Ca2+, ATP still increased [Ca2+]i concomitant with a monophasic inward cation current, indicating Ca2+ release from intracellular stores. We conclude that Hensen cells have ionotropic and metabotropic P2 purinoceptors. They also have Ca(2+)-activated Cl- channels that can be activated by extracellular ATP, suggesting that purinoceptors in Hensen cells could play a regulatory role in ion and water balance of cochlear fluids.

The contribution of a Ca(2+)-activated Cl(−) conductance to amino-acid-induced inward current responses of ciliated olfactory neurons of the rainbow trout

2000 ◽  
Vol 203 (2) ◽  
pp. 253-262 ◽  
Author(s):  
K. Sato ◽  
N. Suzuki
Keyword(s):  
Rainbow Trout ◽  
Amino Acid ◽  
Reversal Potential ◽  
Amino Acid Mixture ◽  
Niflumic Acid ◽  
Flufenamic Acid ◽  
Acid Mixture ◽  
Inward Currents ◽  
Cl Channels ◽  
Cl Conductance

To determine whether amino-acid-induced inward currents of ciliated olfactory receptor neurons (ORNs) in rainbow trout (Oncorhynchus mykiss) include a Ca(2+)-activated Cl(−) conductance, we first studied changes in reversal potential and the current/voltage relationships of the responses of ORNs to an amino acid mixture (l-alanine, l-arginine, l-glutamate and l-norvaline; all 10 mmol l(−)(1)) with different concentrations of Na(+) and Cl(−) in the perfusion and recording pipette solutions. We also examined the effects of six different Cl(−) channel blockers on the responses of ORNs using a conventional whole-cell voltage-clamp technique. The amino acid mixture and one blocker were applied focally to the cilia of ORNs using a double-barrelled micropipette and a pressure ejection system. The expected shifts in reversal potential, indicating the contribution of the Ca(2+)-activated Cl(−) conductance, occurred in both positive and negative directions depending on the external and internal Na(+) and Cl(−) concentrations. Niflumic acid, flufenamic acid, NPPB [5-nitro-2-(3-phenylpropylamino)-benzonate] and DCDPC (3′, 5-dichlorodiphenylamine-2-carboxylate), at 0.5 mmol l(−)(1), reversibly blocked both the amino-acid-induced inward currents and the background activity in most ORNs. The effectiveness of these blocking agents varied from 77 to 91 % for ORNs perfused externally with standard Ringer's solution. SITS (4-acetamido-4′-isothiocyanatostilbene-2,2′-disulphonate), at 5.0 mmol l(−)(1), irreversibly inhibited the physiological response (100 % inhibition), whereas DIDS (4,4′-diisothiocyanatostilbene-2, 2′-disulphonate), at 5.0 mmol l(−)(1), had the smallest effect (45 %) of the inhibitors tested. The dose of niflumic acid inducing 50 % inhibition (IC(50)), determined specifically for the current component of the Ca(2+)-activated Cl(−) channels, was 70 μmol l(−)(1). Our results suggest that these blockers are not specific for Ca(2+)-activated Cl(−) channels and that the density of these channels varies between individual ORNs. Our results also show that the Ca(2+)-activated Cl(−) conductance plays an important role in olfactory transduction and allows fishes to adapt to various ionic environments.

Functional identification of a sarcolemmal chloride channel from bovine tracheal smooth muscle

1996 ◽  
Vol 271 (5) ◽  
pp. C1716-C1724 ◽  
Author(s):  
D. Salvail ◽  
A. Alioua ◽  
E. Rousseau
Keyword(s):  
Smooth Muscle ◽  
Lipid Bilayers ◽  
Reversal Potential ◽  
Tracheal Smooth Muscle ◽  
Disulfonic Acid ◽  
Equilibrium Potential ◽  
Functional Identification ◽  
Cl Channels ◽  
Cl Conductance

The biophysical and pharmacological characteristics of unitary Cl- currents from bovine tracheal smooth muscle cells were studied after reconstitution of microsomal vesicles into planar lipid bilayers. Two types of currents were recorded simultaneously in KCl buffer: the well-defined Ca(2+)-dependent K+ conductance [GK(Ca)] and a much smaller Cl- current, indicating that the Cl- channels under scrutiny originate from the same membrane as the GK(Ca)-type channels, the plasma membrane of airway smooth muscle (ASM) cells. The GK(Ca) activities were eliminated by the use of CsCl buffer. The average unitary Cl- conductance measured in 50 mM trans-250 mM cis CsCl was 77 +/- 6 pS (n = 21), and the reversal potential measured in various CsCl gradients followed the Cl- equilibrium potential as determined from the Nernst equation. In contrast with the previous reports describing the Ca2+ sensitivity of macroscopic ASM Cl- currents, this channel was found to be insensitive to cytoplasmic and extracellular Ca2+ levels. Phosphorylation cocktails, including protein kinases A, G, or C, did not alter the activity of the channel nor did changes in pH. Among a series of Cl- channel inhibitors, 4,4'-diisothiocyanostilbene-2, 2'-disulfonic acid [50% effective concentration (EC50) = 30 microM] and 5-nitro-2-(3-phenylpropylamino) benzoic acid (EC50 = 130 microM) were the most potent blockers of the current examined. The exact role of this surface Cl- conductance remains unclear, and its involvement in cellular activity needs further investigation.

Intracellular pH modulates the activity of chloride channels in isolated lacrimal gland acinar cells

1995 ◽  
Vol 268 (3) ◽  
pp. C647-C650 ◽  
Author(s):  
K. Park ◽  
P. D. Brown
Keyword(s):  
Intracellular Ph ◽  
Lacrimal Gland ◽  
Chloride Channels ◽  
Reversal Potential ◽  
Acinar Cells ◽  
Secretory Activity ◽  
Time Dependent ◽  
Whole Cell ◽  
Whole Cell Patch Clamp ◽  
Cl Channels

The effects of intracellular pH (pHi) on Ca(2+)-activated Cl- currents in rat lacrimal gland acinar cells were examined. Cl- currents were recorded by conventional whole cell patch-clamp methods using K(+)-free and Na(+)-free solutions. pHi was varied by using electrode solutions with pH at 6.8, 7.3, or 7.8, and Ca2+ activity was buffered at 100 nM with ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid. Increasing pH from 6.8 to 7.8 was found to increase whole cell currents. The currents observed exhibited time-dependent activation at depolarizing potentials and time-dependent inactivation at hyperpolarizing potentials (pH 7.8). This behavior is characteristic of Ca(2+)-activated Cl- channels in lacrimal gland cells. The selectivity of the current was examined at pH 7.8 by removing Cl- from the bath solution. This maneuver caused a positive shift in the reversal potential, as expected for a Cl(-)-selective current. Thus increasing pHi appears to activate Ca(2+)-activated Cl- channels. The possibility that an increase in pHi may help sustain Cl- channel activity during secretory activity is discussed.

Extracellular H+ modulates acetylcholine-activated nonselective cation channels in guinea pig ileum

1995 ◽  
Vol 268 (1) ◽  
pp. C162-C170 ◽  
Author(s):  
R. Inoue ◽  
Y. Waniishi ◽  
Y. Ito
Keyword(s):  
Guinea Pig ◽  
Reversal Potential ◽  
Type Equation ◽  
Hill Coefficient ◽  
Patch Clamp Technique ◽  
Physiological Importance ◽  
Cationic Current ◽  
Whole Cell Patch Clamp ◽  
Pka Value ◽  
External Ph

The effects of external H+ on the acetylcholine-induced inward current (nonselective cationic current; InsACh) in guinea pig ileal smooth muscle were investigated using the conventional whole cell patch-clamp technique. When the external pH (pHo) was lowered, the amplitude of InsACh was increased, with no significant change in the reversal potential or no detectable induction of other ionic permeabilities. The dose-response curve for this effect was best described by a Hill-type equation with an apparent pKa value of 7.4 and a Hill coefficient of approximately 1. The effect of pHo was associated with a shift of the steady-state activation curve for InsACh; the half-maximum activation potential became more negative on lowering pHo. Similar results were obtained when InsACh was activated by intracellularly applied guanosine 5'-O-(3-thiotriphosphate). These results indicate that the external H+ activity is an efficient regulator of InsACh channel, and this may have a physiological importance for controlling the muscarinic receptor-mediated contractions in this muscle.

Permeability of time-dependent K+ channel in guinea pig ventricular myocytes to Cs+, Na+, NH4+, and Rb+

1990 ◽  
Vol 259 (5) ◽  
pp. H1448-H1454 ◽  
Author(s):  
R. W. Hadley ◽  
J. R. Hume
Keyword(s):  
Guinea Pig ◽  
Ventricular Myocytes ◽  
Reversal Potential ◽  
Outward Current ◽  
Time Dependent ◽  
K Channel ◽  
Delayed Rectifier ◽  
Patch Clamp Technique ◽  
Whole Cell Patch Clamp ◽  
Outward Currents

Currents through time-dependent K+ channels (also referred to as IK or the delayed rectifier) were studied with the whole cell patch-clamp technique in isolated guinea pig ventricular myocytes. IK measurements were restricted to the examination of deactivation tail currents. Substitution of various monovalent cations for external K+ produced shifts of the reversal potential of IK. These shifts were used to calculate permeability ratios relative to K+. The permeability sequence for the IK channels was K+ = Rb+ greater than NH4+ = Cs+ greater than Na+. Time-dependent outward currents were also examined when the myocytes were dialyzed with Cs+ instead of K+. A sizeable time-dependent outward current, quite similar to that seen with K+ dialysis, was demonstrated. This current was primarily carried by intracellular Cs+, as the reversal potential of the current shifted 46 mV per 10-fold change of external Cs+ concentration. The significance of Cs+ permeation through IK channels is discussed with respect to the common use of Cs+ in isolating other currents.

ADH-evoked [Cl-]i-dependent transient in whole cell current of distal nephron cell line A6

1995 ◽  
Vol 268 (1) ◽  
pp. F64-F72 ◽  
Author(s):  
T. Nakahari ◽  
Y. Marunaka
Keyword(s):  
Cell Line ◽  
Arginine Vasotocin ◽  
Distal Nephron ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
A6 Cells ◽  
Whole Cell Patch Clamp ◽  
Cation Conductance ◽  
Cell Current ◽  
Cl Conductance

The effect of antidiuretic hormone (ADH) on a distal nephron cell line (A6) was studied using the whole cell patch-clamp technique. A6 cells were cultured on a permeable support filter for 10-14 days in media containing 10% fetal bovine serum without supplemental aldosterone. In the unstimulated condition A6 cells had very small conductances of Na+,K+, and Cl-. Arginine vasotocin (AVT, 140 mU/ml, 280 nM) evoked a "transient" increase in whole cell currents as did dibutyryl-adenosine 3',5'-cyclic monophosphate (5 mM). These transients consisted of two components; one was the nonselective cation conductance, and the other was the Cl- conductance. Activation of these conductances was dependent on intracellular Cl- concentration ([Cl-]i). At low [Cl-]i (< or = 50 mM) both conductances were activated, whereas when [Cl-]i was 80 mM, only the Cl- conductance was activated. At high [Cl-]i (125 mM), both conductances were inhibited. It seems likely that the [Cl-]i maintained at a low level (< or = 50 mM) is an important requirement for A6 cells to respond to AVT.

Low frequency electromagnetic radiation and hearing

2009 ◽  
Vol 123 (11) ◽  
pp. 1204-1211 ◽  
Author(s):  
J Morales ◽  
M Garcia ◽  
C Perez ◽  
J V Valverde ◽  
C Lopez-Sanchez ◽  
...  
Keyword(s):  
Guinea Pig ◽  
Electromagnetic Fields ◽  
Control Animal ◽  
Guinea Pigs ◽  
Low Frequency ◽  
Organ Of Corti ◽  
Outer Hair Cells ◽  
Morphological Alteration ◽  
Electric Response ◽  
Hearing Level

AbstractObjective:To analyse the possible impact of low and extremely low frequency electromagnetic fields on the outer hairs cells of the organ of Corti, in a guinea pig model.Materials and methods:Electromagnetic fields of 50, 500, 1000, 2000, 4000 and 5000 Hz frequencies and 1.5 µT intensity were generated using a transverse electromagnetic wave guide. Guinea pigs of both sexes, weighing 100–150 g, were used, with no abnormalities on general and otic examination. Total exposure times were: 360 hours for 50, 500 and 1000 Hz; 3300 hours for 2000 Hz; 4820 hours for 4000 Hz; and 6420 hours for 5000 Hz. One control animal was used in each frequency group. The parameters measured by electric response audiometer included: hearing level; waves I–IV latencies; wave I–III interpeak latency; and percentage appearance of waves I–III at 90 and 50 dB sound pressure level intensity.Results:Values for the above parameters did not differ significantly, comparing the control animal and the rest of each group. In addition, no significant differences were found between our findings and those of previous studies of normal guinea pigs.Conclusion:Prolonged exposure to electromagnetic fields of 50 Hz to 5 KHz frequencies and 1.5 µT intensity, produced no functional or morphological alteration in the outer hair cells of the guinea pig organ of Corti.

BICUCULLINE/BACLOFEN-INSENSITIVE GABA RESPONSE IN CRUSTACEAN NEURONES IN CULTURE

1994 ◽  
Vol 191 (1) ◽  
pp. 167-193
Author(s):  
C Jackel ◽  
W Krenz ◽  
F Nagy
Keyword(s):  
Reversal Potential ◽  
Membrane Conductance ◽  
Gamma Aminobutyric Acid ◽  
Action Potential Firing ◽  
Patch Clamp Technique ◽  
Conformationally Restricted ◽  
Whole Cell Patch Clamp ◽  
Gabac Receptor ◽  
Potential Firing ◽  
Sr 95531

Neurones were dissociated from thoracic ganglia of embryonic and adult lobsters and kept in primary culture. When gamma-aminobutyric acid (GABA) was applied by pressure ejection, depolarizing or hyperpolarizing responses were produced, depending on the membrane potential. They were accompanied by an increase in membrane conductance. When they were present, action potential firing was inhibited. The pharmacological profile and ionic mechanism of GABA-evoked current were investigated under voltage-clamp with the whole-cell patch-clamp technique. The reversal potential of GABA-evoked current depended on the intracellular and extracellular Cl- concentration but not on extracellular Na+ and K+. Blockade of Ca2+ channels by Mn2+ was also without effect. The GABA-evoked current was mimicked by application of the GABAA agonists muscimol and isoguvacine with an order of potency muscimol&gt;GABA&gt;isoguvacine. cis-4-aminocrotonic acid (CACA), a folded and conformationally restricted GABA analogue, supposed to be diagnostic for the vertebrate GABAC receptor, also induced a bicuculline-resistant chloride current, although with a potency about 10 times lower than that of GABA. The GABA-evoked current was largely blocked by picrotoxin, but was insensitive to the GABAA antagonists bicuculline, bicuculline methiodide and SR 95531 at concentrations of up to 100 &micro;mol l-1. Diazepam and phenobarbital did not exert modulatory effects. The GABAB antagonist phaclophen did not affect the GABA-induced current, while the GABAB agonists baclophen and 3-aminopropylphosphonic acid (3-APA) never evoked any response. Our results suggest that lobster thoracic neurones in culture express a chloride-conducting GABA-receptor channel which conforms to neither the GABAA nor the GABAB types of vertebrates but shows a pharmacology close to that of the novel GABAC receptor described in the vertebrate retina.

Dual effects of external magnesium on action potential duration in guinea pig ventricular myocytes

1995 ◽  
Vol 268 (6) ◽  
pp. H2321-H2328 ◽  
Author(s):  
S. Zhang ◽  
T. Sawanobori ◽  
H. Adaniya ◽  
Y. Hirano ◽  
M. Hiraoka
Keyword(s):  
Guinea Pig ◽  
Action Potential ◽  
Decay Time ◽  
Action Potential Duration ◽  
Time Course ◽  
Ventricular Myocytes ◽  
Membrane Surface ◽  
Surface Charges ◽  
Whole Cell Patch Clamp ◽  
K Current

Effects of extracellular magnesium (Mg2+) on action potential duration (APD) and underlying membrane currents in guinea pig ventricular myocytes were studied by using the whole cell patch-clamp method. Increasing external Mg2+ concentration [Mg2+]o) from 0.5 to 3 mM produced a prolongation of APD at 90% repolarization (APD90), whereas 5 and 10 mM Mg2+ shortened it. [Mg2+]o, at 3 mM or higher, suppressed the delayed outward K+ current and the inward rectifier K+ current. Increases in [Mg2+]o depressed the peak amplitude and delayed the decay time course of the Ca2+ current (ICa), the latter effect is probably due to the decrease in Ca(2+)-induced inactivation. Thus 3 mM Mg2+ suppressed the peak ICa but increased the late ICa amplitude at the end of a 200-ms depolarization pulse, whereas 10 mM Mg2+ suppressed both components. Application of 10 mM Mg2+ shifted the voltage-dependent activation and inactivation by approximately 10 mV to more positive voltage due to screening the membrane surface charges. Application of manganese (1-5 mM) also caused dual effects on APD90, similar to those of Mg2+, and suppressed the peak ICa with slowed decay. These results suggest that the dual effects of Mg2+ on APD in guinea pig ventricular myocytes can be, at least in part, explained by its action on ICa with slowed decay time course in addition to suppressive effects on K+ currents.

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