GABA-Mediated Inhibition of Glutamate Release During Ischemia in Substantia Gelatinosa of the Adult Rat

2003 ◽  
Vol 89 (1) ◽  
pp. 257-264 ◽  
Author(s):  
Noriaki Matsumoto ◽  
Eiichi Kumamoto ◽  
Hidemasa Furue ◽  
Megumu Yoshimura
Keyword(s):  
Glutamate Release ◽  
Control Level ◽  
Outward Current ◽  
Substantia Gelatinosa ◽  
Slow Inward Current ◽  
Patch Clamp Technique ◽  
Inward Current ◽  
Adult Rat ◽  
Whole Cell Patch Clamp ◽  
Inward Currents

An ischemia-induced change in glutamatergic transmission was investigated in substantia gelatinosa (SG) neurons of adult rat spinal cord slices by use of the whole cell patch-clamp technique; the ischemia was simulated by superfusing an oxygen- and glucose-free medium (ISM). Following ISM superfusion, 21 of 37 SG neurons tested produced an outward current (23 ± 4 pA at a holding potential of −70 mV), which was followed by a slow and subsequent rapid inward current; the remaining neurons had only inward currents. During such a change in holding currents, spontaneous excitatory postsynaptic currents (EPSCs) were remarkably decreased in a frequency with time (half-decay time of the frequency: about 65 s). The frequency of spontaneous EPSCs was reduced to 28 ± 13% ( n = 37) of the control level during the generation of the slow inward current (about 4 min after the beginning of ISM superfusion) without a change in the amplitude of spontaneous EPSCs. When ISM was superfused together with either bicuculline (10 μM) or CGP35348 (20 μM; GABAA and GABAB receptor antagonists, respectively), spontaneous EPSC frequency reduced by ISM recovered to the control level and then the frequency markedly increased [by 325 ± 120% ( n = 22) and 326 ± 91% ( n = 17), respectively, 4 min after ISM superfusion]; this alteration in the frequency was not accompanied by a change in spontaneous EPSC amplitude. Superfusing TTX (1 μM)-containing ISM resulted in a similar recovery of spontaneous EPSC frequency and following increase (by 328 ± 26%, n = 12) in the frequency; strychnine (1 μM) did not affect ISM-induced changes in spontaneous EPSC frequency ( n = 5). It is concluded that the ischemic simulation inhibits excitatory transmission to SG neurons, whose action is in part mediated by the activation of presynaptic GABAAand GABAB receptors, probably due to GABA released from interneurons as a result of an ischemia-induced increase in neuronal activities. This action might protect SG neurons from an excessive excitation mediated by l-glutamate during ischemia.

Three types of slow inward currents as distinguished by melittin in 3-day-old embryonic heart

1988 ◽  
Vol 66 (8) ◽  
pp. 1017-1022 ◽  
Author(s):  
G. Bkaily ◽  
D. Jacques ◽  
T. Yamamoto ◽  
A. Sculptoreanu ◽  
M. D. Payet ◽  
...  
Keyword(s):  
Slow Inward Current ◽  
Heart Cells ◽  
Slow Inactivation ◽  
High Concentration ◽  
Patch Clamp Technique ◽  
Inward Current ◽  
Whole Cell Patch Clamp ◽  
Low Concentrations ◽  
Voltage Dependent ◽  
Inward Currents

Membrane slow inward currents of 3-day-old embryonic chick single heart cells were investigated using the whole-cell patch clamp technique. In a solution containing only Na+ ions and in the presence of tetrodotoxin and Mn2+, the inward current–voltage relationship presented two maxima, confirming the existence of two different voltage-dependent slow inward currents. The first type, a fast transient slow inward current (Isi (ft)), was activated from a holding potential of −80 mV and showed fast activation and inactivation. This current was highly sensitive to melittin (10−8 M) and insensitive to low concentrations of desmethoxyverapamil ((−)D888, 10−9–10−6 M). Depolarizing voltage steps from a holding a potential of −50 mV activated two components of the slow inward current, i.e., a slow and a sustained current (Isi(sts)) that showed a slow inactivation followed by a slow inactivation and a sustained component. Melittin at a high concentration (10−4 M) completely blocked the slow transient component (Isi(st)) and left unblocked the sustained component (Isi(s)). Both components (Isi(st) and Isi(s)) were blocked by verapamil (10−5 M) and low concentrations of (−)D888 (10−8–10−6 M).

scholarly journals Effect of Auxin (IAA) on the Fast Vacuolar (FV) Channels in Red Beet (Beta vulgaris L.) Taproot Vacuoles

2020 ◽  
Vol 21 (14) ◽  
pp. 4876
Author(s):  
Zbigniew Burdach ◽  
Agnieszka Siemieniuk ◽  
Waldemar Karcz
Keyword(s):  
Single Channel ◽  
Outward Current ◽  
K Channel ◽  
Single Channels ◽  
Patch Clamp Technique ◽  
Open Probability ◽  
Inward Current ◽  
Bath Solution ◽  
Outward Currents ◽  
Inward Currents

In contrast to the well-studied effect of auxin on the plasma membrane K+ channel activity, little is known about the role of this hormone in regulating the vacuolar K+ channels. Here, the patch-clamp technique was used to investigate the effect of auxin (IAA) on the fast-activating vacuolar (FV) channels. It was found that the macroscopic currents displayed instantaneous currents, which at the positive potentials were about three-fold greater compared to the one at the negative potentials. When auxin was added to the bath solution at a final concentration of 1 µM, it increased the outward currents by about 60%, but did not change the inward currents. The imposition of a ten-fold vacuole-to-cytosol KCl gradient stimulated the efflux of K+ from the vacuole into the cytosol and reduced the K+ current in the opposite direction. The addition of IAA to the bath solution with the 10/100 KCl gradient decreased the outward current and increased the inward current. Luminal auxin reduced both the outward and inward current by approximately 25% compared to the control. The single channel recordings demonstrated that cytosolic auxin changed the open probability of the FV channels at the positive voltages to a moderate extent, while it significantly increased the amplitudes of the single channel outward currents and the number of open channels. At the positive voltages, auxin did not change the unitary conductance of the single channels. We suggest that auxin regulates the activity of the fast-activating vacuolar (FV) channels, thereby causing changes of the K+ fluxes across the vacuolar membrane. This mechanism might serve to tightly adjust the volume of the vacuole during plant cell expansion.

Comparison of a Ca(2+)-gated conductance and a second-messenger-gated conductance in rat olfactory neurons

2000 ◽  
Vol 203 (3) ◽  
pp. 567-573
Author(s):  
Y. Okada ◽  
R. Fujiyama ◽  
T. Miyamoto ◽  
T. Sato
Keyword(s):  
Outward Current ◽  
Patch Clamp Technique ◽  
External Application ◽  
Olfactory Neurons ◽  
Internal Solution ◽  
Inward Current ◽  
Whole Cell Patch Clamp ◽  
Cation Conductance ◽  
Intracellular Ca ◽  
Sustained Outward Current

The effect of a rise in intracellular Ca(2+) concentration was analyzed in isolated rat olfactory neurons using a whole-cell patch-clamp technique. Intracellular dialysis of 1 mmol l(−)(1) Ca(2+) in a standard-K(+), low-Cl(−) internal solution (E(Cl)=−69 mV) from the patch pipette into the olfactory neurons induced a sustained outward current of 49+/−5 pA (N=13) at −50 mV in all the cells examined. The outward currents were inhibited by external application of 100 micromol l(−)(1) 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB). External application of a Ca(2+) ionophore, 3 micromol l(−)(1) ionomycin, induced an inward current in three of eight cells whose voltages were clamped using the gramicidin-perforated technique, but ionomycin elicited an outward current in the other five cells, suggesting that natural intracellular Cl(−) concentration in the olfactory neurons was heterogeneous. While intracellular dialysis of 50 micromol l(−)(1) inositol 1,4,5-trisphosphate (1,4,5-InsP(3)) in the standard-K(+), low-Cl(−) internal solution induced the NPPB-sensitive outward current in 31 % of cells, and 500 micromol l(−)(1) cAMP induced it in 21 % of cells, a large proportion of the cells displayed an inward current in response to 1,4,5-InsP(3) and cAMP. The results suggest that 1,4,5-InsP(3) and cAMP can elicit Ca(2+)-dependent Cl(−) conductance and Ca(2+)-independent cation conductance in rat olfactory neurons.

GABA-Mimetic Actions of Withania somnifera on Substantia Gelatinosa Neurons of the Trigeminal Subnucleus Caudalis in Mice

2013 ◽  
Vol 41 (05) ◽  
pp. 1043-1051 ◽  
Author(s):  
Hua Yin ◽  
Dong Hyu Cho ◽  
Soo Joung Park ◽  
Seong Kyu Han
Keyword(s):  
Patch Clamp ◽  
Receptor Antagonist ◽  
Withania Somnifera ◽  
Substantia Gelatinosa ◽  
Patch Clamp Technique ◽  
Pipette Solution ◽  
Whole Cell ◽  
Cell Patch Clamp ◽  
Whole Cell Patch Clamp ◽  
Inward Currents

The plant Withania somnifera (WS), also known as Ashwagandha, has been used widely in traditional medicine systems in India and Nepal (Ayurveda), and has been accepted to cure various ailments. In this study, the whole-cell patch clamp technique was performed to examine the mechanism of action of WS on the SG neurons of the Vc from mouse brainstem slices. In whole-cell patch clamp mode, methanol extract of Withania somnifera (mWS) induced short-lived and repeatable inward currents in all SG neurons tested (31.3±8.51 pA, n = 7) using a high chloride pipette solution. The mWS-induced inward currents were concentration dependent and maintained in the presence of tetrodotoxin (TTX), a voltage gated Na + channel blocker, CNQX, a non-NMDA glutamate receptor antagonist, AP5, an NMDA receptor antagonist and strychnine, a glycine receptor antagonist. The mWS induced currents were blocked by picrotoxin, a GABAA receptor antagonist. These results show that mWS has an inhibitory effects on SG neurons of the Vc through GABAA receptor-mediated activation of chloride ion channels, indicating that mWS contains compounds with sedative effects on the central nervous system. These results also suggest that mWS may be a potential target for modulating orofacial pain processing.

Biphasic modulation by galanin of excitatory synaptic transmission in substantia gelatinosa neurons of adult rat spinal cord slices

2011 ◽  
Vol 105 (5) ◽  
pp. 2337-2349 ◽  
Author(s):  
Hai-Yuan Yue ◽  
Tsugumi Fujita ◽  
Eiichi Kumamoto
Keyword(s):  
Spinal Cord ◽  
Synaptic Transmission ◽  
Outward Current ◽  
Substantia Gelatinosa ◽  
Rat Spinal Cord ◽  
Patch Clamp Technique ◽  
Similar Action ◽  
Membrane Hyperpolarization ◽  
Adult Rat ◽  
Adult Rat Spinal Cord

Although intrathecally administrated galanin modulates nociceptive transmission in a biphasic manner, this has not been fully examined previously. In the present study, the action of galanin on synaptic transmission in the substantia gelatinosa (SG) neurons of adult rat spinal cord slices was examined, using the whole cell patch-clamp technique. Galanin concentration-dependently increased the frequency of spontaneous excitatory postsynaptic current (EPSC; EC50 = 2.0 nM) without changing the amplitude, indicating a presynaptic effect. This effect was reduced in a Ca2+-free, or voltage-gated Ca2+ channel blocker La3+-containing Krebs solution and was produced by a galanin type-2/3 receptor (GalR2/R3) agonist, galanin 2–11, but not by a galanin type-1 receptor (GalR1) agonist, M617. Galanin also concentration-dependently produced an outward current at −70 mV (EC50 = 44 nM), although this appeared to be contaminated by a small inward current. This outward current was mimicked by M617, but not by galanin 2–11. Moreover, galanin reduced monosynaptic Aδ-fiber- and C-fiber-evoked EPSC amplitude; the former reduction was larger than the latter. A similar action was produced by galanin 2–11, but not by M617. Spontaneous and focally evoked inhibitory (GABAergic and glycinergic) transmission was unaffected by galanin. These findings indicate that galanin at lower concentrations enhances the spontaneous release of l-glutamate from nerve terminals by Ca2+ entry from the external solution following GalR2/R3 activation, whereas galanin at higher concentrations also produces a membrane hyperpolarization by activating GalR1. Moreover, galanin reduces l-glutamate release onto SG neurons from primary afferent fibers by activating GalR2/R3. These effects could partially contribute to the behavioral effect of galanin.

scholarly journals Divalent Cation Block of Inward Currents and Low-Affinity K+ Uptake in Saccharomyces cerevisiae

1999 ◽  
Vol 181 (1) ◽  
pp. 291-297 ◽  
Author(s):  
Stephen K. Roberts ◽  
Marc Fischer ◽  
Graham K. Dixon ◽  
Dale Sanders
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Patch Clamp ◽  
Divalent Cation ◽  
Divalent Cations ◽  
Outward Current ◽  
Alkali Cations ◽  
Patch Clamp Technique ◽  
Inward Current ◽  
Negative Membrane Potential ◽  
Inward Currents

ABSTRACT We have used the patch clamp technique to characterize whole-cell currents in spheroplasts isolated from a trk1Δ trk2Δstrain of Saccharomyces cerevisiae which lacks high- and moderate-affinity K+ uptake capacity. In solutions in which extracellular divalent cation concentrations were 0.1 mM, cells exhibited a large inward current. This current was not the result of increasing leak between the glass pipette and membrane, as there was no effect on the outward current. The inward current comprised both instantaneous and time-dependent components. The magnitude of the inward current increased with increasing extracellular K+and negative membrane potential but was insensitive to extracellular anions. Replacing extracellular K+ with Rb+, Cs+, or Na+ only slightly modulated the magnitude of the inward current, whereas replacement with Li+ reduced the inward current by approximately 50%, and tetraethylammonium (TEA+) and choline were relatively impermeant. The inward current was blocked by extracellular Ca2+ and Mg2+ with apparentKi s (at −140 mV) of 363 ± 78 and 96 ± 14 μM, respectively. Furthermore, decreasing cytosolic K+ increased the magnitude of the inward current independently of the electrochemical driving force for K+influx, consistent with regulation of the inward current by cytosolic K+. Uptake of 86Rb+ by intacttrk1Δ trk2Δ cells was inhibited by extracellular Ca2+ with a Ki within the range observed for the inward current. Furthermore, increasing extracellular Ca2+ from 0.1 to 20 mM significantly inhibited the growth of these cells. These results are consistent with those of the patch clamp experiments in suggesting that low-affinity uptake of alkali cations in yeast is mediated by a transport system sensitive to divalent cations.

Calcium Inward Currents in Insect Muscle Fibers

1972 ◽  
Vol 50 (11) ◽  
pp. 1114-1116 ◽  
Author(s):  
H. Washio
Keyword(s):  
Muscle Fiber ◽  
Muscle Fibers ◽  
Outward Current ◽  
Complete Removal ◽  
Membrane Current ◽  
Slow Inward Current ◽  
Inward Current ◽  
Insect Muscle ◽  
Inward Currents ◽  
Ca Ions

The membrane current in a locust muscle fiber was characterized by an early but slow inward current followed by a small late outward current in a solution containing Ca and tetraethylammonium (TEA). The maximum inward current decreased with decreasing Ca concentration, almost disappearing upon complete removal of Ca. The inward current has so far not been found in the standard saline. The results indicate that the inward currents are carried by Ca ions.

scholarly journals Adenosine Inhibits GABAergic and Glycinergic Transmission in Adult Rat Substantia Gelatinosa Neurons

2004 ◽  
Vol 92 (5) ◽  
pp. 2867-2877 ◽  
Author(s):  
Kun Yang ◽  
Tsugumi Fujita ◽  
Eiichi Kumamoto
Keyword(s):  
Cyclic Amp ◽  
Substantia Gelatinosa ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Adult Rat ◽  
Whole Cell Patch Clamp ◽  
Pain Transmission ◽  
Adenosine Receptor Agonist ◽  
Adult Rat Spinal Cord ◽  
Dose Dependent

The effect of adenosine on inhibitory postsynaptic currents (IPSCs) was examined in substantia gelatinosa (SG) neurons of adult rat spinal cord slices by using the whole cell patch-clamp technique. Adenosine reversibly reduced the amplitude of GABAergic and glycinergic electrically evoked IPSCs (eIPSCs) in a dose-dependent manner (EC50 = 14.5 and 19.1 μM, respectively). The A1 adenosine-receptor agonist N6-cyclopentyladenosine also reduced the eIPSCs, whereas the A1 antagonist 8-cyclopentyl-1,3-dimethylxanthine reversed the inhibition produced by adenosine. In paired-pulse experiments, the ratio of the second to first GABAergic or glycinergic eIPSC amplitude was increased by adenosine, whereas the response of SG neurons to exogenous GABA or glycine was unaffected. Adenosine reduced the frequency of GABAergic and glycinergic spontaneous IPSCs without changing their amplitude. This reduction in frequency disappeared in the presence of a K+-channel blocker (4-aminopyridine) but not in the absence of Ca2+. The inhibition by adenosine disappeared in the presence of cyclic-AMP analog (8-Br-cyclic AMP) and adenylate-cyclase activator (forskolin) but not protein-kinase C (PKC) activator (phorbol-12,13-dibutyrate). We conclude that adenosine suppresses inhibitory transmission in SG neurons by activating presynaptic A1 receptors and that this action is mediated by K+ channels and cyclic AMP but not by Ca2+ channels and PKC. This inhibitory action of adenosine probably contributes to the modulation of pain transmission in the SG.

Proteinase-Activated Receptor-1 Activation Presynaptically Enhances Spontaneous Glutamatergic Excitatory Transmission in Adult Rat Substantia Gelatinosa Neurons

2009 ◽  
Vol 102 (1) ◽  
pp. 312-319 ◽  
Author(s):  
T. Fujita ◽  
T. Liu ◽  
T. Nakatsuka ◽  
E. Kumamoto
Keyword(s):  
Sensory Processing ◽  
Substantia Gelatinosa ◽  
Activation Mechanism ◽  
Patch Clamp Technique ◽  
Nociceptive Transmission ◽  
Adult Rat ◽  
Excitatory Transmission ◽  
Whole Cell Patch Clamp ◽  
Proteinase Activated Receptors ◽  
Adult Rat Spinal Cord

Proteinase-activated receptors (PARs) have a unique activation mechanism in that a proteolytically exposed N-terminal region acts as a tethered ligand. A potential impact of PAR on sensory processing has not been fully examined yet. Here we report that synthetic peptides with sequences corresponding to PAR ligands enhance glutamatergic excitatory transmission in substantia gelatinosa (SG) neurons of adult rat spinal cord slices by using the whole cell patch-clamp technique. The frequency of spontaneous excitatory postsynaptic current (EPSC) was increased by PAR-1 agonist SFLLRN-NH2 (by 47% at 1 μM) with small increases by PAR-2 and -4 agonists (SLIGKV-NH2 and GYPGQV-OH, respectively; at >3 μM); there was no change in its amplitude or in holding current at −70 mV. The PAR-1 peptide action was inhibited by PAR-1 antagonist YFLLRNP-OH. TFLLR-NH2, an agonist which is more selective to PAR-1 than SFLLRN-NH2, dose-dependently increased spontaneous EPSC frequency (EC50 = 0.32 μM). A similar presynaptic effect was produced by PAR-1 activating proteinase thrombin in a manner sensitive to YFLLRNP-OH. The PAR-1 peptide action was resistant to tetrodotoxin and inhibited in Ca2+-free solution. Primary-afferent monosynaptically evoked EPSC amplitudes were unaffected by PAR-1 agonist. These results indicate that PAR-1 activation increases the spontaneous release of l-glutamate onto SG neurons from nerve terminals in a manner dependent on extracellular Ca2+. Considering that sensory processing within the SG plays a pivotal role in regulating nociceptive transmission to the spinal dorsal horn, the PAR-1-mediated glutamatergic transmission enhancement could be involved in a positive modulation of nociceptive transmission.

Participation of Ca currents in colonic electrical activity

1989 ◽  
Vol 257 (3) ◽  
pp. C451-C460 ◽  
Author(s):  
P. D. Langton ◽  
E. P. Burke ◽  
K. M. Sanders
Keyword(s):  
Slow Waves ◽  
Ca Current ◽  
Patch Clamp Technique ◽  
Inward Current ◽  
Whole Cell Patch Clamp ◽  
Mechanical Threshold ◽  
Inward Currents ◽  
Window Current ◽  
The Relationship ◽  
Ca Currents

Canine colonic myocytes were studied with the whole cell patch-clamp technique. In 1.8 mM Ca2+, inward currents were evoked by depolarization. Currents activated positive to -50 mV, peaked at approximately 0 mV, and reversed at approximately +50 mV. Inward current was potentiated by high external Ca2+ concentration and BAY K8644 and was decreased by low external Ca2+, nifedipine, and Mn2+, indicating that the current was carried by Ca2+. Overlap of the activation-inactivation properties indicated a "window current" range (-40 to -20 mV) in which inward current might be sustained for long durations at potentials achieved during electrical slow waves. Voltage-clamp protocols simulating physiological depolarizations elicited sustained inward currents. Maximum changes in intracellular Ca2+ resulting from sustained inward currents were calculated, which suggested that depolarizations at the level of slow waves may increase cell Ca2+ sufficiently to cause contraction. The data suggest that electrical slow waves in colonic myocytes are due in part to inward Ca current. This current appears to be sufficient to explain the relationship between slow waves and contractions and provides an explanation for the mechanical threshold in colonic muscles.

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