Pre- and Postsynaptic Inhibitory Actions of Methionine-Enkephalin on Identified Bulbospinal Neurons of the Rat RVL

1998 ◽  
Vol 80 (4) ◽  
pp. 2003-2014 ◽  
Author(s):  
Abdallah Hayar ◽  
Patrice G. Guyenet
Keyword(s):  
Decay Time ◽  
Potassium Conductance ◽  
Outward Current ◽  
Cardiovascular Effects ◽  
Ventrolateral Medulla ◽  
Equilibrium Potential ◽  
Cellular Mechanisms ◽  
Patch Clamp Technique ◽  
Methionine Enkephalin ◽  
Postsynaptic Currents

Hayar, Abdallah and Patrice G. Guyenet. Pre- and postsynaptic inhibitory actions of methionine-enkephalin on identified bulbospinal neurons of the rat RVL. J. Neurophysiol. 80: 2003–2014, 1998. The effects of methionine-enkephalin (ME) on visualized bulbospinal neurons of the rostral ventrolateral medulla (RVL) were characterized in thin slices at 32°C using the whole cell patch-clamp technique. Thirty-five percent of the recorded neurons were found to be tyrosine hydroxylase immunoreactive (C1 neurons). In voltage-clamp recordings, ME (3 μM) induced an outward current in 66% of RVL bulbospinal neurons. A similar percentage of C1 and non-C1 neurons were opioid sensitive. The current induced by ME was inwardly rectifying, reversed close to the potassium equilibrium potential, and was blocked by barium. Most spontaneous postsynaptic currents recorded in these neurons were tetrodotoxin (TTX)-resistant miniature postsynaptic currents (mPSCs). Approximately, 75% of mPSCs had rapid kinetics (decay time = 4.7 ms) and were glutamatergic [miniature excitatory postsynaptic currents (mEPSCs)] because they were blocked by 6-cyano-7-nitroquinoxaline-2,3-dione (10 μM). The remaining mPSCs had much slower kinetics (decay time = 19.6 ms) and were GABAergic [miniature inhibitory postsynaptic currents (mIPSCs)] as they were blocked by gabazine (3 μM) but not by strychnine (3–10 μM). ME decreased the frequency of mEPSCs and mIPSCs by 69 and 43%, respectively. The inhibitory effects of ME were mimicked by the selective μ-opioid receptor agonist endomorphin-1 (EM, 3 μM) and were blocked by naloxone (1 μM). In the absence of TTX, excitatory PSCs evoked by focal electrical stimulation were isolated by application of gabazine and strychnine. EM reduced the amplitude of the evoked EPSCs by 41% without changing their decay time. We conclude that opioids inhibit the majority of RVL C1 and non-C1 bulbospinal neurons by activating a potassium conductance postsynaptically and by decreasing the presynaptic release of glutamate. These cellular mechanisms could explain the depressive cardiovascular effects and the sympathoinhibition produced by opioid transmitters in the RVL, in particular during hypotensive hemorrhage.

Nitric oxide mediates outward potassium currents in opossum esophageal circular smooth muscle

1996 ◽  
Vol 270 (6) ◽  
pp. G932-G938 ◽  
Author(s):  
J. Jury ◽  
K. R. Boev ◽  
E. E. Daniel
Keyword(s):  
Nitric Oxide ◽  
Smooth Muscle ◽  
Circular Muscle ◽  
Outward Current ◽  
Equilibrium Potential ◽  
Patch Clamp Technique ◽  
Pipette Solution ◽  
Whole Cell ◽  
Circular Smooth Muscle ◽  
Outward Currents

Single smooth muscle cells from the opossum body circular muscle were isolated and whole cell currents were characterized by the whole cell patch-clamp technique. When the cells were held at -50 mV and depolarized to 70 mV in 20-mV increments, initial small inactivating inward currents were evoked (-30 to 30 mV) followed by larger sustained outward currents. Depolarization from a holding potential of -90 mV evoked an initial fast inactivating outward current sensitive to 4-aminopyridine but not to high levels of ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA). The outward currents reversed near K+ equilibrium potential and were abolished when KCl was replaced by CsCl in the pipette solution. The sustained outward current was inhibited by quinine and cesium. High EGTA in the pipette solution reduced but did not abolish the sustained outward currents, suggesting that both Ca(2+)-dependent and -independent currents were evoked. The nitric oxide (NO)-releasing agents Sin-1 and sodium nitroprusside increased outward K+ currents. High levels of EGTA in the pipette solution abolished the increase in outward current induced by Sin-1. The presence of cyclopiazonic acid, an inhibitor of the sarcoplasmic reticulum (SR) Ca2+ pump, blocked the effects of NO-releasing agents. We conclude that NO release activates K+ outward currents in opossum esophagus circular muscle, which may depend on Ca2+ release from the SR stores.

Cl− Accumulation Does Not Account for the Depolarizing Phase of the Synaptic GABA Response in Hippocampal Pyramidal Cells

1999 ◽  
Vol 82 (2) ◽  
pp. 768-777 ◽  
Author(s):  
Katherine L. Perkins
Keyword(s):  
Guinea Pig ◽  
Short Interval ◽  
Outward Current ◽  
Pyramidal Cells ◽  
Equilibrium Potential ◽  
Inward Current ◽  
Postsynaptic Currents ◽  
Current Curve ◽  
Separate Channel ◽  
Channel Type

It has been proposed that the depolarizing phase of the biphasic synaptic GABA response could be mediated by HCO3 − passing through GABAA channels after dissipation of the transmembrane Cl− gradient due to intracellular Cl− accumulation. To test this hypothesis, giant GABA-mediated postsynaptic currents (GPSCs) were recorded from pyramidal cells in slices of adult guinea pig hippocampus in the presence of 4-aminopyridine. GPSCs consisted of an early outward current (GABAA component) followed by a late inward current (GABAD component). Spontaneous outward inhibitory postsynaptic currents (IPSCs) occurred during the GABADcomponent of the GPSC. GPSCs that were evoked 1–12 s after the preceding GPSC (short interval, siGPSCs) showed no GABADcomponent even though in many cells the amplitude of the siGPSC was greater than the amplitude of the GABAA component of the preceding spontaneous GPSC. In addition, the siGPSC evoked during the GABAD component of a spontaneous GPSC was an outward current. To test whether the siGPSC lacked a GABADcomponent because it was generated predominantly at the soma, where less of an increase in [Cl−]i would occur, picrotoxin was applied to the soma of the pyramidal cell. To the contrary, this focal application of picrotoxin caused less of a reduction in the amplitude of the siGPSC than in the amplitude of the GABAA component of the GPSC. Furthermore when a GPSC and siGPSC were evoked 10 s apart using identical stimuli, the area under the outward current curve was sometimes greater for the siGPSC than for the GPSC, and yet the siGPSC had no inward component. This result indicates that even when the location of Cl− entry was the same, more Cl− could enter the cell during the siGPSC than during the outward component of the GPSC and yet not lead to an inward current. In addition, when the second of two identical stimuli was applied during the inward GABAD component of the first evoked GPSC, the GABAA response it generated was always outward, demonstrating that the equilibrium potential for GABAA responses did not become more positive than the holding potential during a GPSC. Finally, evoking GPSCs at a hyperpolarized potential revealed that the siGPSC actually lacked a GABAD conductance. These results disprove the Cl− accumulation hypothesis of the synaptic depolarizing GABA response and suggest the possibility that a separate channel type may mediate the GABAD component of the GPSC.

Effect of substance P on C1 and other bulbospinal cells of the RVLM in neonatal rats

1997 ◽  
Vol 273 (2) ◽  
pp. R805-R813 ◽  
Author(s):  
Y. W. Li ◽  
P. G. Guyenet
Keyword(s):  
Substance P ◽  
Discharge Rate ◽  
Potassium Conductance ◽  
Input Resistance ◽  
Ventrolateral Medulla ◽  
Equilibrium Potential ◽  
Neonatal Rats ◽  
Spontaneous Firing Rate ◽  
Sympathetic Vasomotor Tone

Sixty-two bulbospinal neurons were recorded in the rostral ventrolateral medulla (RVLM) of neonatal rats using patch electrodes. Sixty-one percent of the recorded neurons identified by histology contained tyrosine-hydroxylase (C1 cells). Substance P increased the spontaneous firing rate of all recorded cells but had no effect on spike configuration. The peptide depolarized neurons that were silenced by injection of hyperpolarizing current and increased their input resistance. All cells (n = 12) were activated by a neurokinin (NK)1 receptor agonist but most were unaffected by an NK2- or an NK2-selective compound. In voltage clamp, substance P produced a current that was linearly related to the membrane voltage. This current reversed polarity close to the potassium equilibrium potential in 11 of 23 cells. It reversed at more hyperpolarized potentials or not at all in the rest of the cells. In conclusion, substance P upregulates the intrinsic discharge rate of C1 and other putative sympathoexcitatory cells in neonatal rats. This effect is mediated via NK1 receptors. The depolarization is mediated by a reduction in resting potassium conductance and possibly by an additional cationic conductance. These results support the possibility that substance P could play a role "in vivo" in setting the basal level of discharge of the vasomotor cells of RVLM and therefore in regulating sympathetic vasomotor tone.

Adenosine inhibition of synaptic transmission in the substantia gelatinosa

1994 ◽  
Vol 72 (4) ◽  
pp. 1611-1621 ◽  
Author(s):  
J. Li ◽  
E. R. Perl
Keyword(s):  
Synaptic Transmission ◽  
Dorsal Root ◽  
Potassium Conductance ◽  
Outward Current ◽  
Postsynaptic Membrane ◽  
Substantia Gelatinosa ◽  
Equilibrium Potential ◽  
Primary Afferent ◽  
Primary Afferent Fibers ◽  
Sustained Outward Current

1. We studied adenosine's action on synaptic transmission from primary afferent fibers to neurons of the substantia gelatinosa (SG) using tight-seal whole cell recordings in transverse slices of hamster spinal cord. Adenosine had two actions, hyperpolarization of the postsynaptic membrane and depression of the excitatory postsynaptic currents (EPSCs) evoked by dorsal root stimulation. 2. Under voltage clamp adenosine elicited a sustained outward current at a holding potential of -70 mV. The outward current was blocked by a combination of intracellular cesium and tetraethylammonium, an effect characteristic of potassium channels. The adenosine-induced current reversed at -97 +/- 6 (SD) mV, close to the potassium equilibrium potential. These observations suggest that adenosine activates a potassium conductance in SG neurons so as to inhibit primary afferent synaptic transmission postsynaptically. 3. Adenosine reduced the miniature EPSC frequency without significantly changing the amplitude. In contrast, the glutamate receptor competitive antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) substantially reduced the amplitudes of miniature EPSCs while producing a much smaller effect on the miniature frequency than adenosine. In evoked EPSCs adenosine reduced unitary content without reducing unitary amplitude. The effects on both miniature and evoked EPSCs suggest that adenosine inhibits synaptic currents by suppressing presynaptic transmitter release. 4. EPSCs evoked by dorsal root stimuli were subdivided into monosynaptic and polysynaptic categories. Adenosine at superfusion concentrations of 20-300 microM suppressed all polysynaptic EPSCs. Less than half of monosynaptic EPSCs were inhibited, usually those evoked by the slowest-conducting primary afferents. These observations were interpreted to indicate that a principal action of adenosine in SG is on interneuronal communication.

An outwardly rectifying K+ current active near resting potential in human retinal pigment epithelial cells

1995 ◽  
Vol 269 (1) ◽  
pp. C179-C187 ◽  
Author(s):  
B. A. Hughes ◽  
M. Takahira ◽  
Y. Segawa
Keyword(s):  
Retinal Pigment Epithelial ◽  
Retinal Pigment ◽  
Outward Current ◽  
Resting Potential ◽  
Equilibrium Potential ◽  
Delayed Rectifier ◽  
Patch Clamp Technique ◽  
Rpe Cells ◽  
Pigment Epithelial ◽  
K Current

Currents in freshly dissociated adult human retinal pigment epithelial (RPE) cells were studied using the perforated patch-clamp technique. The zero-current potential (V0) averaged -48.9 +/- 7.7 mV (n = 50). Depolarizing voltage pulses from -70 mV evoked an outward current that activated with first-order kinetics and that did not inactivate during prolonged depolarizations. Repolarizing the membrane potential produced tail currents that reversed near the K+ equilibrium potential, indicating that the sustained outward current was carried mainly by K+. The outwardly rectifying K+ conductance (gK) had an activation threshold voltage near -60 mV and was half-maximal at -37 mV. Approximately 25% of gK was active at the average V0. The K+ current was nearly completely blocked by 2 mM Ba2+ but was relatively insensitive to 20 mM tetraethylammonium. The kinetics, voltage dependence, and blocker sensitivity of this current clearly distinguish it from delayed rectifier K+ currents previously identified in RPE cells. We conclude that the sustained outward K+ current may help establish the resting potential of the apical and/or basolateral membranes and may also participate in K+ transport across the RPE.

Effects of glucose deprivation on NMDA-induced current and intracellular Ca2+ in rat substantia nigra neurons

1996 ◽  
Vol 75 (2) ◽  
pp. 740-749 ◽  
Author(s):  
Y. Nakashima ◽  
H. Ishibashi ◽  
N. Harata ◽  
N. Akaike
Keyword(s):  
Substantia Nigra ◽  
Outward Current ◽  
Glucose Deprivation ◽  
Equilibrium Potential ◽  
Dependent Manner ◽  
Induced Current ◽  
Patch Clamp Technique ◽  
Concentration Dependent Manner ◽  
Inward Current ◽  
Rat Substantia Nigra

1. The effects of glucose deprivation on N-methyl-D-asparate (NMDA)-induced current (INMDA) and the intracellular free Ca2+ concentration ([Ca2+]i) in the acutely dissociated rat substantia nigra neurons were investigated using the nystatin-perforated patch-clamp technique under voltage clamp and the microfluometry with a fluorescent probe, Indo-1. 2. Application of NMDA induced a peak and a successive steady-state inward current, and an outward current immediately after washout at a holding potential of -40 mV. The amplitudes of the three current components of INMDA were increased by increasing the concentrations of NMDA with half-maximum concentrations (EC50s) of 1.1 x 10(-4) M, 1.2 x 10(-4) M, and 1.6 x 10(-4) M, respectively. 3. The reversal potentials of the peak inward and outward currents were -4 +/- 3 (SE) mV and -76 +/- 2 mV, respectively. The latter was close to the theoretical K+ equilibrium potential (-82 mV). 4. The outward current was potentiated by increase in extracellular Ca2+ concentration and was blocked by Cs+ internal solution and suppressed by 5 x 10(-3) M tetraethylammonium chloride and 10(-7) M charybdotoxin, indicating that it was Ca(2+)-activated K+ current. 5. Application of NMDA increased [Ca2+]i in a concentration-dependent manner with an EC50 of 3.9 x 10(-5) M. 6. Depriving the external solution of glucose induced a slowly developing outward current and increased the basal level of [Ca2+]i. It also prolonged the NMDA-induced outward current without affecting the peak inward current, and prolonged the NMDA-induced increase in [Ca2+]i without changing the peak [Ca2+]i. 7. These findings suggest that the deprivation of glucose did not affect the NMDA-induced influx of Ca2+ into the cells, but it inhibited Ca2+ clearance by affecting the efflux of Ca2+ to the extracellular space, reuptake into the intracellular Ca2+ stores, and/or active extrusion from intracellular stores.

α2-Adrenoceptor-mediated presynaptic inhibition in bulbospinal neurons of rostral ventrolateral medulla

1999 ◽  
Vol 277 (3) ◽  
pp. H1069-H1080 ◽  
Author(s):  
Abdallah Hayar ◽  
Patrice G. Guyenet
Keyword(s):  
Presynaptic Inhibition ◽  
Rostral Ventrolateral Medulla ◽  
Ventrolateral Medulla ◽  
Patch Clamp Technique ◽  
Postsynaptic Currents ◽  
Outward Currents ◽  
Adrenoceptor Blocker ◽  
Brain Stem Slices ◽  
Synaptic Mechanisms

The rostral ventrolateral medulla (RVLM) controls sympathetic tone via excitatory bulbospinal neurons. It is also the main target of α2-adrenoceptor (α2-AR) agonists used for treatment of hypertension. In this study, we examined the synaptic mechanisms by which α2-AR agonists may inhibit the activity of RVLM bulbospinal neurons. We recorded selectively from RVLM bulbospinal neurons in brain stem slices of neonate rats (P5–P21) using the patch-clamp technique (holding potential −70 mV). α2-ARs were activated by norepinephrine (NE, 30 μM) in the presence of the α1-adrenoceptor blocker prazosin. NE induced modest outward currents (5–28 pA) in 70% of the cells that were blocked by barium and by the α2-AR antagonist 2-methoxyidazoxan. The magnitude of this current was not correlated with the tyrosine hydroxylase immunoreactivity of the neurons. Mono- and oligosynaptic excitatory postsynaptic currents (EPSCs) or monosynaptic inhibitory postsynaptic currents (IPSCs) were evoked by focal electrical stimulation. In all cells, NE decreased the amplitude of the evoked EPSCs in the absence or presence of barium (49 and 70%) and decreased the amplitude of the evoked IPSCs (64 and 59%). The effect of NE on EPSC amplitude was blocked by 2-methoxyidazoxan. Focal stimulation produced a 1- to 2-s EPSC afterdischarge (probably due to activation of interneurons) that was 53% inhibited by NE. In the presence of tetrodotoxin, NE decreased the frequency of miniature EPSCs by 74%. In short, α2-AR stimulation produces weak postsynaptic responses in RVLM bulbospinal neurons and powerful presynaptic inhibition of both glutamatergic and GABAergic inputs. Thus the inhibition of RVL bulbospinal neurons by α2-AR agonists in vivo results from a combination of postsynaptic inhibition, disfacilitation, and disinhibition.

Activation of GABAB receptors increases a potassium conductance in rat bulbospinal neurons of the C1 area

1996 ◽  
Vol 271 (5) ◽  
pp. R1304-R1310 ◽  
Author(s):  
Y. W. Li ◽  
P. G. Guyenet
Keyword(s):  
Gabab Receptor ◽  
Outward Current ◽  
Gabab Receptors ◽  
Neonatal Rat ◽  
Ventrolateral Medulla ◽  
Equilibrium Potential ◽  
Inward Rectification ◽  
Dependent Manner ◽  
Gamma Aminobutyric Acid

In anesthetized rats, iontophoresis of the gamma-aminobutyric acid (GABAB)-receptor agonist and antispastic drug baclofen inhibits the bulbospinal vasomotor neurons of the rostral ventrolateral medulla (RVLM). The present study was carried out to determine whether C1 adrenergic and other bulbospinal neurons of the RVLM have postsynaptic GABAB receptors. Retrogradely labeled RVLM bulbospinal neurons (n = 52) were recorded in 120-micron-thick slices from neonatal rat brain (3-10 days old). Most neurons (48/52) were tonically active (3 +/- 0.6 spikes/s). Twenty-six neurons were recovered histologically, and 18 of them were immunoreactive for tyrosine hydroxylase (TH). In current clamp, baclofen (0.3-10 microM) hyperpolarized RVLM bulbospinal cells in a dose-dependent manner (16 +/- 0.5 mV hyperpolarization by 3 microM baclofen; n = 19) and decreased input resistance by 40% (n = 10). In voltage clamp (1 microM tetrodotoxin present; holding potential: -40 to -60 mV), 3 microM baclofen induced an outward current of 21 +/- 2 pA (n = 29). This current exhibited inward rectification and reversed polarity close to the K+ equilibrium potential (external K+ from 2.5 to 10 mM). The current induced by baclofen was reduced 90% by 0.1-0.2 mM BaCl2 (n = 6) and was blocked reversibly by the selective GABAB-receptor antagonist CGP-55845A (0.5-1 microM; n = 6). All histologically verified TH-immunoreactive cells (n = 18) were sensitive to baclofen. In summary, RVLM bulbospinal neurons including C1 adrenergic cells possess GABAB receptors. Activation of these receptors increases an inwardly rectifying K+ conductance. This effect reduces the intrinsic firing frequency of RVLM vasomotor neurons "in vitro" and may contribute to the sympatholytic action of baclofen "in vivo."

scholarly journals Prototypical Imidazoline-1 Receptor Ligand Moxonidine Activates Alpha2-Adrenoceptors in Bulbospinal Neurons of the RVL

2000 ◽  
Vol 83 (2) ◽  
pp. 766-776 ◽  
Author(s):  
Abdallah Hayar ◽  
Patrice G. Guyenet
Keyword(s):  
Outward Current ◽  
Ventrolateral Medulla ◽  
Agonist Activity ◽  
Neonatal Rats ◽  
Vasomotor Tone ◽  
Postsynaptic Currents ◽  
Whole Cell Recordings ◽  
Brain Stem Slices ◽  
Sympathetic Vasomotor Tone ◽  
Stem Slices

Moxonidine is an antihypertensive drug that lowers sympathetic vasomotor tone by stimulating either alpha2-adrenergic (α2-AR) or imidazoline I1 receptors within the rostral ventrolateral medulla (RVL). In this study, we investigated the effects of moxonidine (10 μM) on RVL neurons in brain stem slices of neonatal rats. We recorded mainly from retrogradely labeled RVL bulbospinal neurons (putative presympathetic neurons) except for some extracellular recordings. Prazosin was used to block alpha1-adrenoceptors. Moxonidine inhibited the extracellularly recorded discharges of all spontaneously active RVL neurons tested (bulbospinal and unidentified). This effect was reversed or blocked by the selective α2-AR antagonist SKF 86466 (10 μM). In contrast, the I1 imidazoline ligand AGN 192403 (10 μM) had no effect on the spontaneous activity. In whole cell recordings (holding potential −70 mV), moxonidine produced a small and variable outward current (mean 7 pA). This current was observed in both tyrosine hydroxylase–immunoreactive and other bulbospinal neurons and was blocked by SKF 86466. Excitatory postsynaptic currents (EPSCs) evoked by focal electrical stimulation were isolated by incubation with gabazine and strychnine, and inhibitory postsynaptic currents (IPSCs) were isolated with 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). Moxonidine reduced the amplitude of the evoked EPSCs (EC50 = 1 μM; 53% inhibition at 10 μM) but not their decay time constant (5.6 ms). The effect of moxonidine on EPSCs persisted in barium (300 μM) and was reduced ∼80% by SKF 86466. Moxonidine also reduced the amplitude of evoked IPSCs by 63%. In conclusion, moxonidine inhibits putative RVL presympathetic neurons both presynaptically and postsynaptically. All observed effects in the present study are consistent with an α2-AR agonist activity of moxonidine.

Choline chloride activates time-dependent and time-independent K+ currents in dog atrial myocytes

1994 ◽  
Vol 266 (1) ◽  
pp. C42-C51 ◽  
Author(s):  
B. Fermini ◽  
S. Nattel
Keyword(s):  
Choline Chloride ◽  
Outward Current ◽  
Time Dependent ◽  
Equilibrium Potential ◽  
Delayed Rectifier ◽  
Atrial Myocytes ◽  
Patch Clamp Technique ◽  
Voltage Dependent ◽  
K Current ◽  
K Currents

Using the whole cell configuration of the patch-clamp technique, we studied the effect of isotonic replacement of bath sodium chloride (NaCl) by choline chloride (ChCl) in dog atrial myocytes. Our results show that ChCl triggered 1) activation of a time-independent background current, characterized by a shift of the holding current in the outward direction at potentials positive to the K+ equilibrium potential (EK), and 2) activation of a time- and voltage-dependent outward current, following depolarizing voltage steps positive to EK. Because the choline-induced current obtained by depolarizing steps exhibited properties similar to the delayed rectifier K+ current (IK), we named it IKCh. The amplitude of IKCh was determined by extracellular ChCl concentration, and this current was generally undetectable in the absence of ChCl. IKCh was not activated by acetylcholine (0.001-1.0 mM) or carbachol (10 microM) and could not be recorded in the absence of ChCl or when external NaCl was replaced by sucrose or tetramethylammonium chloride. IKCh was inhibited by atropine (0.01-1.0 microM) but not by the M1 antagonist pirenzepine (up to 10 microM). This current was carried mainly by K+ and was inhibited by CsCl (120 mM, in the pipette) or barium (1 mM, in the bath). We conclude that in dog atrial myocytes, ChCl activates a background conductance comparable to ACh-dependent K+ current, together with a time-dependent K+ current showing properties similar to IK.

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