Acetylcholine-activated ionic currents in parasympathetic neurons of bullfrog heart

1990 ◽  
Vol 63 (5) ◽  
pp. 1052-1059 ◽  
Author(s):  
N. Tateishi ◽  
D. K. Kim ◽  
N. Akaike
Keyword(s):  
Steady State ◽  
Rapid Change ◽  
Reversal Potential ◽  
External Solution ◽  
Ionic Currents ◽  
Hill Coefficient ◽  
Equilibrium Potential ◽  
Inward Rectification ◽  
Parasympathetic Neurons ◽  
State Component

1. The electrical and pharmacologic properties of acetylcholine (ACh)-induced current (IACh) were studied in the parasympathetic neurons isolated from bullfrog heart with the use of the concentration-clamp technique, which allows intracellular perfusion and rapid change of external solution within 2 ms under the single-electrode voltage-clamp condition. 2. The IACh consisted of an initial transient peak component and a successive steady-state plateau component. Both currents increased in a sigmoidal fashion with increasing ACh concentration. The dissociation constant (Kd value) and the Hill coefficient for each component were 2.2 X 10(-5) M and 1.6, respectively. 3. In the K(+)-free solution, the reversal potential (EACh) of IACh was close to the Na+ equilibrium potential (ENa). The current-voltage (I-V) relation showed inward rectification at positive potentials. 4. Nicotine mimicked only the peak component of IACh. However both peak and steady-state components were blocked nonselectively by the nicotinic blockers d-tubocurarine and hexamethonium. 5. Carbamylcholine (CCh) mimicked the steady-state component of IACh. The steady-state component was selectively inhibited by atropine at concentrations 1,000 times lower than that required for inhibition of the peak component. The steady state was blocked equally by either pirenzepine (M1 blocker) or AF-DX-116 (M2 blocker). 6. It was concluded that the IACh consisted of a peak component having double exponential activation and inactivation, mediated through the nicotinic actions, and a steady-state component having no inactivation, mediated through the muscarinic action.

Glycine-gated chloride current in acutely isolated rat hypothalamic neurons

1989 ◽  
Vol 62 (6) ◽  
pp. 1400-1409 ◽  
Author(s):  
N. Akaike ◽  
M. Kaneda
Keyword(s):  
Reversal Potential ◽  
External Solution ◽  
Hill Coefficient ◽  
Equilibrium Potential ◽  
Dependent Manner ◽  
Hypothalamic Neurons ◽  
Adult Rats ◽  
Concentration Dependent Manner ◽  
Current Decay ◽  
Continuous Application

1. Electrical and pharmacologic properties of glycine-induced currents were investigated in single hypothalamic neurons acutely isolated from young and adult rats by the use of a "concentration-clamp" technique, which allows both internal perfusion and rapid application of an external solution under single-electrode voltage-clamp. 2. The glycine-induced current reversed at the Cl- equilibrium potential (ECl), and a 10-fold decrease of extracellular Cl- with a large impermeable anion resulted in a 53 mV shift of the glycine reversal potential (EGly). 3. Glycine-induced Cl- currents (ICl) increased sigmoidally in a concentration-dependent manner with a Kd of 9 X 10(-5) M at a Hill coefficient of 1.8. Current inactivation occurred completely at all concentrations within 10 s. EGly remained unchanged during continuous application of glycine, suggesting that the inactivation process is because of desensitization. 4. The glycine-induced conductance exhibited a striking voltage dependency at membrane potentials more negative than -50 mV and reached a steady state value when hyperpolarized beyond -110 mV. 5. Both the activation and inactivation phases of glycine-induced ICl are described by double exponential (fast and slow components) functions with the concentrations used. All four time constants decreased with increasing glycine concentration. 6. The slow time constant of the current decay induced by glycine increased with depolarization and decreased with hyperpolarization, indicating that the rate of desensitization is considerably voltage dependent. The fast decay showed little voltage dependency. 7. Recovery of the glycine response after complete desensitization consisted of two components. 8. The blockade of the glycine response by strychnine and picrotoxin was noncompetitive.

ATP-gated current in dissociated rat nucleus solitarii neurons

1992 ◽  
Vol 68 (3) ◽  
pp. 778-785 ◽  
Author(s):  
S. Ueno ◽  
N. Harata ◽  
K. Inoue ◽  
N. Akaike
Keyword(s):  
Purinergic Receptor ◽  
Reversal Potential ◽  
External Solution ◽  
Hill Coefficient ◽  
Inward Rectification ◽  
Inactivation Kinetics ◽  
Receptor Subtype ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Continuous Presence

1. The excitatory response of extracellularly applied ATP was investigated in freshly dissociated rat nucleus tractus solitarii neurons under whole-cell configuration using the ”concentration-clamp” technique. 2. At a holding potential of -70 mV, 100 microM ATP evoked inward current that was slowly desensitized in the continuous presence of ATP. The ATP-gated current increased in a concentration-dependent manner over the concentration range between 10 microM and 1 mM. The half-maximum concentration was 31 microM and the Hill coefficient was 1.2. 3. The potency of ATP analogues for the purinergic receptor was in the order of ATP = 2-methylthio-ATP much greater than ADP greater than alpha,beta-methylene ATP. Neither adenosine nor AMP evoked any responses. The order was consistent with a P2y receptor subtype. 4. The current-voltage relationship for the 100 microM ATP response showed a clear inward rectification at positive potentials beyond -50 mV. The reversal potential of the ATP-gated current was +13 mV. 5. The time constants of activation and inactivation of the ATP-gated current solution were dependent on the extracellular ATP concentration, and both kinetics became faster at higher ATP concentrations. 6. The ATP-gated current was also elicited in an external solution containing Ca2+ as a permeable cation. The inactivation kinetics in an external solution containing 75 mM Ca2+ were faster than those in an external solution with 150 mM Na+. 7. Calculated relative permeability ratios were PNa/PCs = 1.64 ([Na+]o = 30-150 mM), PCa/PCs = 2.17 ([Ca2+]o = 2 mM). Anions were not measurably permeable in this preparation.

ATP-induced inward current in neurons freshly dissociated from the tuberomammillary nucleus

1994 ◽  
Vol 71 (3) ◽  
pp. 868-873 ◽  
Author(s):  
K. Furukawa ◽  
H. Ishibashi ◽  
N. Akaike
Keyword(s):  
Purinergic Receptor ◽  
Reversal Potential ◽  
External Solution ◽  
Hill Coefficient ◽  
Inward Rectification ◽  
Dependent Manner ◽  
Induced Current ◽  
Excitatory Effect ◽  
Inward Current ◽  
Tuberomammillary Nucleus

1. Neurons in the tuberomammillary nucleus (TMN), which are considered to be histaminergic, were dissociated and their response to extracellularly applied ATP was investigated in the nystatin-perforated patch recording mode under voltage-clamp condition. 2. ATP induced a sustained inward current that was slowly desensitized at a holding potential of -60 mV. 3. The ATP response increased in a concentration-dependent manner. The half-maximum concentration (EC50) was 44 microM and the Hill coefficient was 1.8. 4. The potency of ATP analogues was in the order of ATP > or = 2-methylthio-ATP >> alpha, beta-methylene ATP > or = ADP. Neither adenosine nor AMP induced any response. The results suggest that the purinergic receptor in TMN neurons is P2y. 5. The current-voltage relationship for the 100 microM ATP showed a significant inward rectification at a potential more positive than -20 mV in an external solution with 150 mM Na+, but a significant rectification current was not observed in an external solution with 150 mM Cs+. The change in the reversal potential of the ATP response (EATP) to a 10-fold change of extracellular Na+ concentration was 56 mV, indicating that the ATP-induced current is highly selective for Na+ over Cl-. 6. The permeability ratio for cations was Na+:Li+:K+:Rb+: Cs+:Ca2+ = 2.16:1.36:1.68:1.54:1:2.55, indicating that the ATP-induced current is passing through the ligand-gated nonselective cation channel. 7. These results suggest that ATP has an excitatory effect on the TMN neurons by opening nonselective cation channels.

GABA-induced chloride current in rat isolated Purkinje cells

1989 ◽  
Vol 256 (6) ◽  
pp. C1153-C1159 ◽  
Author(s):  
M. Kaneda ◽  
M. Wakamori ◽  
N. Akaike
Keyword(s):  
Purkinje Cells ◽  
Reversal Potential ◽  
External Solution ◽  
Hill Coefficient ◽  
Equilibrium Potential ◽  
Maximal Response ◽  
Dependent Manner ◽  
Gamma Aminobutyric Acid ◽  
Concentration Jump ◽  
The Hill

Electrical and pharmacological properties of the gamma-aminobutyric acid (GABA)-induced current in the rat isolated cerebellar Purkinje cell bodies were studied using a concentration-jump method, which is termed as a "concentration-clamp" technique. This technique enables the rapid exchange of external solution around the neurons to be perfused internally under a voltage-clamp condition. The peak amplitude of GABA response increased sigmoidally with the increase of the concentration of GABA. The value of the GABA concentration that evokes a half-maximal response (Ka) was 5 X 10(-5) M, and the Hill coefficient was 1.8. The current-voltage relationship for the GABA response showed nonlinearity at membrane potentials more negative than -40 mV. The reversal potential of GABA-evoked current was close to the equilibrium potential of Cl- (ECl), indicating that the current elicited by GABA is carried by Cl-. Both the activation and inactivation phases of GABA-induced Cl- current (ICl) consisted of fast and slow components. These time constants in both phases decreased as the concentration of GABA increased. Strychnine and bicuculline inhibited the GABA-induced ICl in a dose-dependent manner, and the inhibition of the GABA response by bicuculline was competitive. Pentobarbital sodium augmented the GABA response and modified the inactivation phase. The augmentation of the GABA response by pentobarbital was more profound at lower concentrations of GABA and was accompanied by a change in the Hill coefficient from 2 to 1. The properties of the GABA response in cerebellar Purkinje cells were thought to be basically similar to those previously reported in other preparations.

Muscarinic receptor activation of potassium channels in rat dentate gyrus neurons

1993 ◽  
Vol 70 (4) ◽  
pp. 1544-1552 ◽  
Author(s):  
J. Nabekura ◽  
S. Ebihara ◽  
N. Akaike
Keyword(s):  
Dentate Gyrus ◽  
Reversal Potential ◽  
Outward Current ◽  
External Solution ◽  
Receptor Activation ◽  
Hill Coefficient ◽  
Equilibrium Potential ◽  
Maximal Response ◽  
Dependent Manner ◽  
Concentration Dependent Manner

1. The effects of acetylcholine (ACh) on granule cells freshly dissociated from rat dentate gyrus (DG) were studied using the nystatin perforated patch technique. This method allowed us to study ACh-induced currents (IACh) under voltage clamp without "run-down" of the ACh response. In some experiments, we used the conventional whole-cell method for intracellular application of drugs not permeable to cell membrane. 2. At a holding potential of -40 mV, ACh induced an outward current. The amplitude of IACh increased in a sigmoidal fashion with increasing ACh concentration. The half-maximal response and the Hill coefficient determined from the relation between ACh concentration and response were 4.98 x 10(-7) M and 1.70, respectively. 3. The reversal potential of IACh was close to the K+ equilibrium potential. The IACh was accompanied by an enhancement of the K+ current. 4. Muscarine and McN-A-343 mimicked the ACh response, whereas oxotremorine induced no response. 5. Muscarinic antagonists reversibly suppressed the IACh (10(-5) M) in a concentration-dependent manner, where the values of half-inhibition concentration (IC50) were 1.03 x 10(-6) M for pirenzepine and 2.21 x 10(-5) M for AF-DX-116. 6. Intracellular perfusion with GDP-beta S suppressed the IACh greatly. The IACh persisted in the neurons pretreated with an external solution containing pertussis toxin (IAP) for 18 h. 7. In the neurons perfused with Ca(2+)-free external solution containing 2 mM ethylene glycol-O,O'-bis (beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid and 10 mM Mg2+, the first application of ACh induced the IACh with an amplitude similar to that in the standard solution.(ABSTRACT TRUNCATED AT 250 WORDS)

Proton-induced sodium current in frog isolated dorsal root ganglion cells

1990 ◽  
Vol 63 (4) ◽  
pp. 805-813 ◽  
Author(s):  
N. Akaike ◽  
O. A. Krishtal ◽  
T. Maruyama
Keyword(s):  
Dorsal Root Ganglion ◽  
Dorsal Root ◽  
Ganglion Cells ◽  
Sodium Current ◽  
Rapid Change ◽  
External Solution ◽  
Hill Coefficient ◽  
Equilibrium Potential ◽  
Induced Current ◽  
Inactivation Curve

1. The proton-induced current was examined in isolated frog dorsal root ganglion (DRG) cells by the use of the "concentration-clamp" technique, which allows intracellular perfusion and rapid change of external solution with various pH (pHo) within 2 ms under single-electrode voltage-clamp condition. 2. Over one-half of the examined neurons showed no response for a "step" reduction of pHo even in a Ca2(+)-free external solution. In smaller neurons having a diameter less than 20 microns, the persistent and reliable proton-induced responses were obtained, though the current amplitude and the activation and inactivation varied considerably for each cell. 3. The decrease of external Na+ concentration ([Na+]o) reduced the proton response. The proton response reversed the direction and the Na+ equilibrium potential (ENa). 4. With decreasing pHo from 7.4, proton response increased in a sigmoidal fashion. The threshold was around pH 7.0 and the maximum response appeared at pH 5.2, whereas pKa and Hill coefficient were 6.0 and 1.97, respectively. 5. The activation and inactivation phases of the proton-induced current behaved as a single exponential function. The time constants of activation (tau a) and inactivation (tau i) were not affected by changing either the holding membrane potential (VH) or the low external Ca2+ concentration [( Ca2+]o) between 10(-6) and 5 X 10(-3) M. But the decrease of pHo up to 5.2 decreased both tau a and tau i in a saturable manner. 6. In the inactivation curve of proton-induced current obtained by decreasing pHo from various conditioning pHo to 5.5, half inactivation occurred at pHo 7.45.(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of a transient outward K+ current with inward rectification in canine ventricular myocytes

1998 ◽  
Vol 274 (3) ◽  
pp. C577-C585 ◽  
Author(s):  
Gui-Rong Li ◽  
Haiying Sun ◽  
Stanley Nattel
Keyword(s):  
Action Potential ◽  
Ventricular Myocytes ◽  
Reversal Potential ◽  
Outward Current ◽  
Equilibrium Potential ◽  
Inward Rectification ◽  
Transient Outward Current ◽  
Membrane Excitability ◽  
Voltage Dependent ◽  
Patch Technique

The threshold potential for the classical depolarization-activated transient outward K+ current and Cl− current is positive to −30 mV. With the whole cell patch technique, a transient outward current was elicited in the presence of 5 mM 4-aminopyridine (4-AP) and 5 μM ryanodine at voltages positive to the K+ equilibrium potential in canine ventricular myocytes. The current was abolished by 200 μM Ba2+ or omission of external K+([Formula: see text]) and showed biexponential inactivation. The current-voltage relation for the peak of the transient outward component showed moderate inward rectification. The transient outward current demonstrated voltage-dependent inactivation (half-inactivation voltage: −43.5 ± 3.2 mV) and rapid, monoexponential recovery from inactivation (time constant: 13.2 ± 2.5 ms). The reversal potential responded to the changes in[Formula: see text] concentration. Action potential clamp revealed two phases of Ba2+-sensitive current during the action potential, including a large early transient component after the upstroke and a later outward component during phase 3 repolarization. The present study demonstrates that depolarization may elicit a Ba2+- and[Formula: see text]-sensitive, 4-AP-insensitive, transient outward current with inward rectification in canine ventricular myocytes. The properties of this K+ current suggest that it may carry a significant early outward current upon depolarization that may play a role in determining membrane excitability and action potential morphology.

Proton-gated sodium current in parasympathetic ganglion cells of frog heart

1990 ◽  
Vol 63 (5) ◽  
pp. 1060-1067 ◽  
Author(s):  
D. K. Kim ◽  
N. Tateishi ◽  
N. Akaike
Keyword(s):  
Ganglion Cells ◽  
Sodium Current ◽  
External Solution ◽  
Extracellular Ph ◽  
Hill Coefficient ◽  
Equilibrium Potential ◽  
Frog Heart ◽  
Induced Current ◽  
Inactivation Curve ◽  
Parasympathetic Ganglia

1. The proton-gated current was investigated in whole-cell configuration of the neurons isolated from bullfrog heart parasympathetic ganglia with the use of the "concentration-clamp" technique, which combines intracellular perfusion and extremely rapid exchange of external solution within 1-2 ms, under a single-electrode voltage-clamp condition. 2. In all isolated neurons, a "step" decrease in extracellular pH (pHo) induced a transient inward current that was followed by a complete inactivation within 1 s. 3. The proton-gated current increased in a sigmoidal fashion as pHo decreased. In the external solution containing 2 mM Ca2+, the threshold of current activation was at pHo 7.4, and the maximum response appeared at pHo 6.5-5.5. The dissociation constant (Kd) and Hill coefficient were 7.1 and 3.2, respectively. 4. The proton-gated current was reduced by decreasing the extracellular Na+ concentration. In the absence of K+, the current produced by reduced extracellular pH reversed at the Na+ equilibrium potential (ENa), indicating that the current was carried by Na+. 5. Kinetics of both the activation and inactivation phases of proton-induced current were single exponential. The time constants of activation (tau a) and inactivation (tau i) had no potential dependence but decreased slightly by decreasing pHo. 6. In the inactivation curve of the proton-induced current obtained by decreasing pHo from various conditioning pHos to 6.5, the half-maximum inactivation occurred at pHo 7.75. 7. The proton-gated current was suppressed as the extracellular Ca2+ concentration [( Ca2+]o) increased from 0.1 to 10 mM, and the half inhibition occurred at greater than 10 mM [Ca2+]o.(ABSTRACT TRUNCATED AT 250 WORDS)

Alpha-adrenergic modulation of ionic currents in cultured parasympathetic neurons from rat intracardiac ganglia

1993 ◽  
Vol 69 (4) ◽  
pp. 1060-1070 ◽  
Author(s):  
Z. J. Xu ◽  
D. J. Adams
Keyword(s):  
Reversal Potential ◽  
Outward Current ◽  
Cell Voltage ◽  
Ionic Currents ◽  
Ca Channels ◽  
Adrenergic Agonists ◽  
Adrenergic Antagonist ◽  
Steady State Inactivation ◽  
Parasympathetic Neurons ◽  
Intracardiac Ganglia

1. Modulation of ionic conductances by alpha-adrenergic agonists was investigated in cultured parasympathetic neurons from rat intracardiac ganglia. Application of norepinephrine (NE, 25-100 microM) to the soma of isolated neurons reversibly reduced both the amplitude and duration of the Ca(2+)-dependent action potential evoked by injection of depolarizing current when Na+ and K+ currents were blocked pharmacologically. 2. In the whole-cell voltage-clamp mode, application of NE reversibly reduced the amplitude and rate of activation of Ca2+ current (ICa). The amplitude inhibition was greater at the peak of the current (55%) than at the end of a 700-ms pulse (20%). Maximal doses of NE produced only approximately 60% inhibition of peak ICa amplitude. 3. Inactivation of ICa was best fit by the sum of two exponential functions in the absence of NE, but was described by a single exponential function in the presence of NE. These results suggest that NE preferentially inhibited a fast inactivating component of the Ca2+ current in these parasympathetic neurons. 4. NE reversibly reduced the amplitude of Ba2+ tail currents through open Ca channels at all voltages from -40 to +150 mV with a slight shift in the activation curve determined from the current-voltage (I-V) relationship for the tail currents. NE did not change the voltage dependence of the steady-state inactivation of the calcium channels. 5. NE inhibited Ca2+ current either in the absence or presence of nifedipine but to a lesser extent in the presence of omega-conotoxin (omega-CGTX), suggesting that the Ca channels inhibited by NE are predominantly omega-CGTX sensitive. 6. The inhibition of ICa by NE was mimicked by the alpha 1-adrenergic agonists methoxamine and phenylephrine and potentiated in the presence of the alpha 2-adrenoceptor antagonist yohimbine (10 microM). NE inhibition of ICa was antagonized by bath application of the alpha-adrenergic antagonist phentolamine (1 microM), but not by prazosin (1-10 microM), yohimbine, or the beta-adrenergic antagonist propranolol (1 microM). Taken together, these results suggest that NE inhibition of Ca2+ current in rat parasympathetic cardiac neurons is mediated by an alpha-adrenergic receptor with properties that may differ from alpha 1- and alpha 2-adrenoceptors. 7. In approximately 35% of neurons studied, NE not only reduced depolarization-activated inward Ca2+ current but also increased an outward current, with a shift of the I-V curve and reversal potential to more negative voltages.(ABSTRACT TRUNCATED AT 400 WORDS)

Whole-cell voltage-clamp study of the fading of GABA-activated currents in acutely dissociated hippocampal neurons

1986 ◽  
Vol 56 (1) ◽  
pp. 1-18 ◽  
Author(s):  
J. R. Huguenard ◽  
B. E. Alger
Keyword(s):  
Steady State ◽  
Voltage Clamp ◽  
Time Course ◽  
Reversal Potential ◽  
Cell Voltage ◽  
Membrane Properties ◽  
Equilibrium Potential ◽  
Spinal Cord Neurons ◽  
Whole Cell ◽  
Whole Cell Voltage Clamp

The lability of the responses of mammalian central neurons to gamma-aminobutyric acid (GABA) was studied using neurons acutely dissociated from the CA1 region of the adult guinea pig hippocampus as a model system. GABA was applied to the neuronal somata by pressure ejection and the resulting current (IGABA) recorded under whole-cell voltage clamp. In initial experiments we examined several basic properties of cells in this preparation. Our data confirm that passive and active membrane properties are similar to those which characterize cells in other preparations. In addition, GABA-dependent conductance (gGABA), reversal potential (EGABA), and the interaction of GABA with pentobarbital and bicuculline all appeared to be normal. Dendritic GABA application could cause depolarizing GABA responses, and somatic GABA application caused hyperpolarizations due to chloride (Cl-) movements. Repetitive brief applications (5-15 ms) of GABA (10(-5) to 10(-3) M) at a frequency of 0.5 Hz led to fading of successive peaks of IGABA until, at a given holding potential, a steady state was reached in which IGABA no longer changed. Imposing voltage steps lasting seconds during a train of steady-state GABA responses led initially to increased IGABA that then diminished with maintenance of the step voltage. The rate of decrease of IGABA at each new holding potential was independent of the polarity of the step in holding potential but was highly dependent on the rate of GABA application. Application rates as low as 0.05 Hz led to fading of IGABA, even with activation of relatively small conductances (5-15 nS). Since IGABA evoked by somatic GABA application in these cells is carried by Cl-, the Cl- equilibrium potential (ECl) is equal to the reversal potential for IGABA, i.e., to EGABA. The fading of IGABA with changes in holding potential can be almost entirely accounted for by a shift in ECl resulting from transmembrane flux of Cl- through the GABA-activated conductance. Maneuvers that prevent changes in the intracellular concentration of Cl-ions, [Cl-]i, including holding the membrane potential at EGABA during repetitive GABA application or buffering [Cl-]i with high pipette [Cl-], prevent changes in EGABA. Desensitization of the GABA response (an actual decrease in gGABA) occurs in these neurons during prolonged application of GABA (greater than 1 s) but with a slower time course than changes in EGABA. Whole-cell voltage-clamp techniques applied to tissue-cultured spinal cord neurons indicated that rapid shifts in EGABA result from repetitive GABA application in these cells as well.(ABSTRACT TRUNCATED AT 250 WORDS)

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