Persistent Sodium and Calcium Currents in Rat Hypoglossal Motoneurons

2003 ◽  
Vol 89 (1) ◽  
pp. 615-624 ◽  
Author(s):  
Randall K. Powers ◽  
Marc D. Binder
Keyword(s):  
Calcium Channels ◽  
Voltage Clamp ◽  
Developmental Stages ◽  
Calcium Currents ◽  
Neonatal Rat ◽  
Negative Slope ◽  
Channel Blockers ◽  
Whole Cell ◽  
Persistent Inward Currents ◽  
Inward Currents

Voltage-dependent persistent inward currents are thought to make an important contribution to the input–output properties of α−motoneurons, influencing both the transfer of synaptic current to the soma and the effects of that current on repetitive discharge. Recent studies have paid particular attention to the contribution of L-type calcium channels, which are thought to be widely distributed on both the somatic and the dendritic membrane. However, the relative contribution of different channel subtypes as well as their somatodendritic distribution may vary among motoneurons of different species, developmental stages, and motoneuron pools. In this study, we have characterized persistent inward currents in juvenile (10- to 24-day-old) rat hypoglossal (HG) motoneurons. Whole-cell, voltage-clamp recordings were made from the somata of visualized rat HG motoneurons in 300-μm brain stem slices. Slow (10 s), triangular voltage-clamp commands from a holding potential of −70 to 0 mV and back elicited whole-cell currents that were dominated by outward, potassium currents, but often showed a region of negative slope resistance on the rising phase of the command. In the presence of potassium channel blockers (internal cesium and external 4-aminopyridine and tetraethylammonium), net inward currents were present on both the rising and falling phases of the voltage-clamp command. A portion of the inward current present on the ascending phase of the command was mediated by TTX-sensitive sodium channels, whereas calcium channels mediated the remainder of the current. We found roughly the same relative contributions of P-, N-, and L-type channels to the calcium currents recorded at the soma that had previously been found in neonatal rat HG motoneurons. In most cells, the somatic voltage thresholds for calcium current onset and offset were similar and the peak current was largest on the ascending phase of the clamp command. However, about one-third of the cells exhibited a substantial clockwise current hysteresis, i.e., inward currents were present at lower voltages on the descending phase of the clamp command. In the same cells, 1-s depolarizing voltage-clamp commands were followed by prolonged tail currents, consistent with a prominent contribution from dendritic channels. In contrast to previous reports on turtle and mouse motoneurons, blocking L-type calcium channels did not eliminate these presumed dendritic currents.

scholarly journals Facilitation of Somatic Calcium Channels Can Evoke Prolonged Tail Currents in Rat Hypoglossal Motoneurons

2007 ◽  
Vol 98 (2) ◽  
pp. 1042-1047 ◽  
Author(s):  
Anna T. Moritz ◽  
Gregory Newkirk ◽  
Randall K. Powers ◽  
Marc D. Binder
Keyword(s):  
Voltage Clamp ◽  
Calcium Currents ◽  
Whole Cell ◽  
Persistent Inward Currents ◽  
Hypoglossal Motoneurons ◽  
Pharmacological Tests ◽  
Voltage Ramp ◽  
Voltage Dependent ◽  
Inward Currents ◽  
Whole Cell Recordings

Voltage-dependent persistent inward currents (PICs) make an important contribution to the input-output properties of alpha motoneurons. PICs are thought to be mediated by membrane channels located primarily on the dendrites as evidenced by prolonged tail currents following the termination of a voltage step and by a clockwise hysteresis in the whole cell inward currents recorded in response to depolarizing then repolarizing voltage ramp commands. We report here, however, that voltage-clamp currents with these same features can be generated in isolated somatic membrane patches from rat hypoglossal motoneurons. Long-lasting (200–800 ms) tail currents after 1-s voltage-clamp pulses were observed in nucleated patches from 16 of 23 cells. Further, these somatic PICs display “facilitation” in response to conditioning depolarization as previously observed in whole cell recordings from intact neurons. Pharmacological tests suggest that the PICs were primarily mediated by Cav1 channels. Our results show that many of the features of persistent calcium currents recorded from intact motoneurons do not necessarily reflect a remote dendritic origin but can also be ascribed to the intrinsic properties of their Cav1 channels.

scholarly journals Nonmodal gating of cardiac calcium channels as revealed by dihydropyridines.

1989 ◽  
Vol 93 (6) ◽  
pp. 1243-1273 ◽  
Author(s):  
A E Lacerda ◽  
A M Brown
Keyword(s):  
Calcium Channels ◽  
Single Channel ◽  
Calcium Currents ◽  
Neonatal Rat ◽  
Whole Cell ◽  
Frequency Distributions ◽  
Time Frequency ◽  
Channel Current ◽  
Open Time ◽  
Open States

The hypothesis that dihydropyridine (DHP)-sensitive calcium channels have three distinct modes of gating has been examined. The major prediction is that the relative frequencies among modes depend on DHP concentration while the kinetics within a mode do not. We tested this by studying whole-cell and single-channel calcium currents in neonatal rat and adult guinea pig cardiac myocytes in different concentrations of several DHPs. In the absence of DHPs calcium currents declined with time but the kinetics, which are the focus of this study, were unchanged. Open-time frequency distributions had insignificant numbers of prolonged openings and were well fit by single tau's. Agonist DHP stereoisomers produced concentration-dependent changes in whole-cell tail current tau's. The frequency distribution of single calcium channel current open times became biexponential and the tau's were concentration dependent. The average number of openings per trace of channels with customary open times increased with increases in DHP concentration. Latencies to first opening for the customary openings and for prolonged openings were shorter in the presence of DHPs. A second larger conductance is another important feature of DHP-bound single calcium channels. Thus DHPs not only caused prolonged openings; they produced numerous changes in the kinetics of customary openings and increased channel conductance. It follows that these effects of DHPs do not support the hypothesis of modal gating of calcium channels. The mode model is not the only model excluded by the results; models in which DHPs are allowed to act only or mainly on open states are excluded, as are models in which the effects are restricted to inactivated states. We suggest a different type of model in which cooperative binding of DHPs at two sites produces the essential changes in kinetics and conductance.

Postnatal development of pre- and postsynaptic GABAB-mediated inhibitions in the CA3 hippocampal region of the rat

1995 ◽  
Vol 73 (1) ◽  
pp. 246-255 ◽  
Author(s):  
J. L. Gaiarsa ◽  
V. Tseeb ◽  
Y. Ben-Ari
Keyword(s):  
Electrical Stimulation ◽  
Postnatal Development ◽  
Pyramidal Neurons ◽  
Developmental Stages ◽  
Hippocampal Slices ◽  
Phosphinic Acid ◽  
Calcium Currents ◽  
Neonatal Rat ◽  
Channel Blockers ◽  
Ca3 Pyramidal Neurons

1. Intracellular recordings were made from adult and neonatal rat hippocampal slices to study the postnatal development of GABAB-mediated inhibition in CA3 pyramidal neurons. 2. In the presence of glutamatergic receptor antagonists, direct electrical stimulation of the interneurons induced a biphasic GABAA- and GABAB-mediated inhibitory postsynaptic potential in adult [postnatal day (P) 30-P40] and young (P6-P8) CA3 pyramidal neurons. In contrast, in pups (P0-P3), electrical stimulation only induced a bicuculline-sensitive depolarizing GABAA synaptic potential. 3. The outward postsynaptic currents generated by bath-applications of baclofen (30 microM, 30 s) at P3 (78 +/- 60 pA, mean +/- SE) were 4 to 5 times smaller than those evoked between P6 (329 +/- 32 pA) and P30 (412 +/- 44 pA). At P0, baclofen failed to induce a postsynaptic current. 4. The outward currents generated by serotonin (50 microM, 30 s) and the A1 receptor agonist N-cyclopentyladenosine (40 microM, 30 s) ranged between 0 and 50 pA at P3 and between 200 and 400 pA at P6 and P30 (holding potential = -60 +/- 2 mV). 5. In the presence of potassium channel blockers, the amplitude of calcium current elicited by a depolarizing voltage step command (1 s) from a holding potential of -60 mV to a test potential of 0 mV was 2 +/- 0.15 nA at P6 (n = 9) and 0.73 +/- 0.14 nA at P3 (n = 8). Baclofen reversibly reduced the amplitude of calcium currents in young rats but not in pups. 6. Baclofen reversibly reduced the amplitude of the evoked GABAA-mediated and glutamatergic synaptic events at all developmental stages. These effects were dose dependent and antagonized by P-alpha 3-aminopropyl-P-diethoxymethyl-phosphinic acid (CGP) 35348 (500 microM). 7. We conclude that postsynaptic GABAB-mediated inhibition is absent or minimal during the first postnatal days in the CA3 region. In contrast, presynaptic GABAB inhibition is present at birth. We discuss the mechanisms and physiological consequences of these observations.

scholarly journals MicroRNA-204 Is Necessary for Aldosterone-Stimulated T-Type Calcium Channel Expression in Cardiomyocytes

2018 ◽  
Vol 19 (10) ◽  
pp. 2941 ◽  
Author(s):  
Riko Koyama ◽  
Tiphaine Mannic ◽  
Jumpei Ito ◽  
Laurence Amar ◽  
Maria-Christina Zennaro ◽  
...  
Keyword(s):  
Calcium Channels ◽  
Molecular Mechanisms ◽  
Calcium Currents ◽  
Neonatal Rat ◽  
Messenger Rnas ◽  
Screening Assay ◽  
Rat Cardiomyocytes ◽  
Ventricular Cardiomyocytes ◽  
Channel Expression ◽  
Beating Frequency

Activation of the mineralocorticoid receptor (MR) in the heart is considered to be a cardiovascular risk factor. MR activation leads to heart hypertrophy and arrhythmia. In ventricular cardiomyocytes, aldosterone induces a profound remodeling of ion channel expression, in particular, an increase in the expression and activity of T-type voltage-gated calcium channels (T-channels). The molecular mechanisms immediately downstream from MR activation, which lead to the increased expression of T-channels and, consecutively, to an acceleration of spontaneous cell contractions in vitro, remain poorly investigated. Here, we investigated the putative role of a specific microRNA in linking MR activation to the regulation of T-channel expression and cardiomyocyte beating frequency. A screening assay identified microRNA 204 (miR-204) as one of the major upregulated microRNAs after aldosterone stimulation of isolated neonatal rat cardiomyocytes. Aldosterone significantly increased the level of miR-204, an effect blocked by the MR antagonist spironolactone. When miR-204 was overexpressed in isolated cardiomyocytes, their spontaneous beating frequency was significantly increased after 24 h, like upon aldosterone stimulation, and messenger RNAs coding T-channels (CaV3.1 and CaV3.2) were increased. Concomitantly, T-type calcium currents were significantly increased upon miR-204 overexpression. Specifically repressing the expression of miR-204 abolished the aldosterone-induced increase of CaV3.1 and CaV3.2 mRNAs, as well as T-type calcium currents. Finally, aldosterone and miR-204 overexpression were found to reduce REST-NRSF, a known transcriptional repressor of CaV3.2 T-type calcium channels. Our study thus strongly suggests that miR-204 expression stimulated by aldosterone promotes the expression of T-channels in isolated rat ventricular cardiomyocytes, and therefore, increases the frequency of the cell spontaneous contractions, presumably through the inhibition of REST-NRSF protein.

scholarly journals Mobilization of intracellular calcium by intracellular flash photolysis of caged dihydrosphingosine in cultured neonatal rat sensory neurones.

1998 ◽  
Vol 45 (2) ◽  
pp. 311-326 ◽  
Author(s):  
A Ayar ◽  
N M Thatcher ◽  
U Zehavi ◽  
D R Trentham ◽  
R H Scott
Keyword(s):  
Flash Photolysis ◽  
Reversal Potential ◽  
Neonatal Rat ◽  
Xenon Lamp ◽  
Patch Clamp Technique ◽  
Pipette Solution ◽  
Sensory Neurones ◽  
Whole Cell ◽  
Inward Currents ◽  
Current Activation

The ability of dihydrosphingosine to release Ca2+ from intracellular stores in neurones was investigated by combining the whole cell patch clamp technique with intracellular flash photolysis of caged, N-(2-nitrobenzyl)dihydrosphingosine. The caged dihydrosphingosine (100 microM) was applied to the intracellular environment via the CsCl-based patch pipette solution which also contained 0.3% dimethylformamide and 2 mM dithiothreitol. Cultured dorsal root ganglion neurones from neonatal rats were voltage clamped at -90 mV and inward whole cell Ca2+-activated currents were recorded in response to intracellular photorelease of dihydrosphingosine. Intracellular photorelease of dihydrosphingosine (about 5 microM) was achieved using a Xenon flash lamp. Inward Ca2+-activated currents were evoked in 50 out of 57 neurones, the mean delay to current activation following photolysis was 82+/-13 s. The responses were variable with neurones showing transient, oscillating or sustained inward currents. High voltage-activated Ca2+ currents evoked by 100 ms voltage step commands to 0 mV were not attenuated by photorelease of dihydrosphingosine. Controls showed that alone a flash from the Xenon lamp did not activate currents, and that the unphotolysed caged dihydrosphingosine, and intracellular photolysis of 2-(2-nitrobenzylamino) propanediol also did not evoke responses. The dihydrosphingosine current had a reversal potential of -11+/-3 mV (n = 11), and was carried by two distinct Cl- and cation currents which were reduced by 85% and about 20% following replacement of monovalent cations with N-methyl-D-glucamine or application of the Cl- channel blocker niflumic acid (10 microM) respectively. The responses to photoreleased dihydrosphingosine were inhibited by intracellular application of 20 mM EGTA, 10 microM ryanodine or extracellular application of 10 microM dantrolene, but persisted when Ca2+ free saline was applied to the extracellular environment. Intracellular application of uncaged dihydrosphingosine evoked responses which were attenuated by photolysis of the caged Ca2+ chelator Diazo-2. Experiments also suggested that extracellular application of dihydrosphingosine can activate membrane conductances. We conclude that dihydrosphingosine directly or indirectly mobilises Ca2+ from ryanodine-sensitive intracellular stores in cultured sensory neurones.

Voltage-Sensitivity of Motoneuron NMDA Receptor Channels Is Modulated by Serotonin in the Neonatal Rat Spinal Cord

2001 ◽  
Vol 86 (3) ◽  
pp. 1131-1138 ◽  
Author(s):  
Jason N. MacLean ◽  
Brian J. Schmidt
Keyword(s):  
Spinal Cord ◽  
Nmda Receptor ◽  
Membrane Voltage ◽  
Neonatal Rat ◽  
Negative Slope ◽  
Voltage Sensitivity ◽  
Spinal Motoneurons ◽  
Rat Spinal Cord ◽  
Whole Cell ◽  
Nonlinear Membrane

Both N-methyl-d-aspartate (NMDA) and serotonin (5-HT) receptors contribute to the generation of rhythmic motor patterns in the rat spinal cord. Co-application of these chemicals is more effective at producing locomotor-like activity than either neurochemical alone. In addition, NMDA application to rat spinal motoneurons, synaptically isolated in tetrodotoxin, induces nonlinear membrane behavior that results in voltage oscillations which can be blocked by 5-HT antagonists. However, the mechanisms underlying NMDA and 5-HT receptor interactions pertinent to motor rhythm production remain to be determined. In the present study, an in vitro neonatal rat spinal cord preparation was used to examine whether NMDA receptor-mediated nonlinear membrane voltage is modulated by 5-HT. Whole-cell recordings of spinal motoneurons demonstrated that 5-HT shifts the region of NMDA receptor-dependent negative slope conductance (RNSC) of the current-voltage relationship to more hyperpolarized potentials and enhances whole-cell inward current. The influence of 5-HT on the RNSC was similar to the effect on the RNSC of decreasing the extracellular Mg2+concentration. The results suggest that 5-HT may modulate this form of membrane voltage nonlinearity by regulating Mg2+ blockade of the NMDA ionophore.

Whole cell calcium currents in acutely isolated olfactory bulb output neurons of the rat

1996 ◽  
Vol 75 (3) ◽  
pp. 1138-1151 ◽  
Author(s):  
X. Wang ◽  
J. S. McKenzie ◽  
R. E. Kemm
Keyword(s):  
Olfactory Bulb ◽  
Bay K 8644 ◽  
Calcium Currents ◽  
Neonatal Rat ◽  
Bathing Solution ◽  
Whole Cell ◽  
Peak Current ◽  
Maximum Peak ◽  
Slope Factor ◽  
Output Neurons

1. Voltage-gated whole cell Ca2+ currents have been investigated in olfactory bulb (OB) output neurons acutely isolated from neonatal rats. 2. Identification of OB output neurons, mitral or tufted cells, was based on morphology and size and validated by their retrograde labeling with rhodamine or Fast Blue. Of labeled neurons, 45% exhibited either phasic or nonphasic spontaneous firing that was blocked by 10(-7) M tetrodotoxin, 0.5 mM Cd2+, or 1 mM Co2+ in the bathing solution. 3. Whole cell Ca2+ currents displayed holding potential sensitivity indicative of low voltage-activated (LVA) and high voltage-activated (HVA) currents, which exhibited similar dependence on extracellular Ca2+ concentration and could be completely abolished by bathing in 500 microM Cd2+ or in Ca(2+)-free solution. 4. A T-type LVA Ca2+ current, detected in 65% of OB output neurons tested, was activated by depolarizing to -57 mV from holding potential -86 mV and fully inactivated at holding potentials more positive than -60 mV. It was permeated equally by 2.6 mM Ca2+, Sr2+ and Ba2+. The half-activation potential was -35 mV with a slope factor of 7 mV. Depolarizing to -26 mV from different holding potentials in a 2.6-mM Ca2+ solution gave a steady-state half-inactivation potential of -82 mV with a slope factor of 10.7 mV. This LVA current was not sensitive to 5 microM omega-conotoxin (omega-CgTx) or 5 microM Bay K 8644 and was resistant to block by 30 microM Cd2+, by 50 microM verapamil or by 5 microM nifedipine. 5. HVA Ca2+ currents, detected in 97% of OB output cells, activated at around -30 to -20 mV, with maximum peak current at approximately 4 mV in 2.6 mM Ca2+ external solution. They showed similar permeability to 2.6 mM Ca2+ and Sr2+, but the maximum peak current was increased 40% in 2.6 mM Ba2+. Depolarizing to 4 mV from different holding potentials yielded a half-inactivation potential of -67 mV with a slope factor of 13.2 mV. Two components, as suggested by their sensitivities to 5 microM Bay K 8644, nifedipine. omega-CgTx and to voltage, may resemble the L-type and N-type currents described in other neural preparations. However, 5 microM omega-CgTx seemed to block both components, being more effective at more positive potentials. There was a residual component of Cd(2+)-sensitive current not affected by cumulative addition of nifedipine and omega-CgTx. 6. omega-Agatoxin IVA (omega-Aga), a selective P-type Ca2+ channel blocker, had no detectable effect at 50 or 200 nM and 1 microM doses on whole cell Ca2+ currents elicited by 200-ms voltage steps to 4 mV from holding potential -86 mV. 7. We conclude that both LVA and HVA Ca2+ currents exist in neonatal rat OB output neurons, showing distinct kinetic and pharmacological characteristics. The HVA Ca2+ currents contain at least two components, probably resembling L- and N-type currents. Another fast-inactivating HVA component, insensitive to nifedipine, omega-CgTx and omega-Aga, could represent the newly established R-type Ca2+ current.

Functional Expression of L-, N-, P/Q-, and R-Type Calcium Channels in the Human NT2-N Cell Line

2000 ◽  
Vol 84 (6) ◽  
pp. 2933-2944 ◽  
Author(s):  
Torben R. Neelands ◽  
Anthony P. J. King ◽  
Robert L. Macdonald
Keyword(s):  
Calcium Channel ◽  
Cell Line ◽  
Functional Expression ◽  
Calcium Currents ◽  
Channel Blockers ◽  
Patch Clamp Technique ◽  
Teratocarcinoma Cell ◽  
Inactivation Curve ◽  
Whole Cell ◽  
Channel Currents

The biophysical and pharmacological properties of voltage-gated calcium channel currents in the human teratocarcinoma cell line NT2-N were studied using the whole cell patch-clamp technique. When held at −80 mV, barium currents ( I Bas) were evoked by voltage commands to above −35 mV that peaked at +5 mV. When holding potentials were reduced to −20 mV or 5 mM barium was substituted for 5 mM calcium, there was a reduction in peak currents and a right shift in the current-voltage curve. A steady-state inactivation curve for I Ba was fit with a Boltzmann curve ( V 1/2 = −43.3 mV; slope = −17.7 mV). Maximal current amplitude increased from 1-wk (232 pA) to 9-wk (1025 pA) postdifferentiation. Whole cell I Bas were partially blocked by specific channel blockers to a similar extent in 1- to 3-wk and 7- to 9-wk postdifferentiation NT2-N cells: 10 μM nifedipine (19 vs. 25%), 10 μM conotoxin GVIA (27 vs. 25%), 10 μM conotoxin MVIIC (15 vs. 16%), and 1.75 μM SNX-482 (31 vs. 33%). Currents were completely blocked by 300 μM cadmium. In the presence of nifedipine, GVIA, and MVIIC, ∼35% of current remained, which was reduced further by SNX-482 (7–14% of current remained), consistent with functional expression of L-, N-, and P/Q-calcium channel types and one or more R-type channel. The presence of multiple calcium currents in this human neuronal-type cell line provides a potentially useful model for study of the regulation, expression and cellular function of human derived calcium channel currents; in particular the R-type current(s).

scholarly journals Serotonin Facilitates a Persistent Calcium Current in Motoneurons of Rats With and Without Chronic Spinal Cord Injury

2007 ◽  
Vol 97 (2) ◽  
pp. 1236-1246 ◽  
Author(s):  
X. Li ◽  
K. Murray ◽  
P. J. Harvey ◽  
E. W. Ballou ◽  
D. J. Bennett
Keyword(s):  
Spinal Cord ◽  
Calcium Currents ◽  
High Dose ◽  
Chronic Injury ◽  
Persistent Inward Currents ◽  
Cord Injury ◽  
Inward Currents ◽  
Acute Spinal Rats ◽  
Chronic Spinal Rats

In the months after spinal cord transection, motoneurons in the rat spinal cord develop large persistent inward currents (PICs) that are responsible for muscle spasticity. These PICs are mediated by low-threshold TTX-sensitive sodium currents (Na PIC) and L-type calcium currents (Ca PIC). Recently, the Na PIC was shown to become supersensitive to serotonin (5-HT) after chronic injury. In the present paper, a similar change in the sensitivity of the Ca PIC to 5-HT was investigated after injury. The whole sacrocaudal spinal cord from acute spinal rats and spastic chronic spinal rats (S2 level transection 2 mo previously) was studied in vitro. Intracellular recordings were made from motoneurons and slow voltages ramps were applied to measure PICs. TTX was used to block the Na PIC. For motoneurons of chronic spinal rats, a low dose of 5-HT (1 μM) significantly lowered the threshold of the Ca PIC from −56.7 ± 6.0 to −63.1 ± 7.1 mV and increased the amplitude of the Ca PIC from 2.4 ± 1.0 to 3.0 ± 0.73 nA. Higher doses of 5-HT acted similarly. For motoneurons of acute spinal rats, low doses of 5-HT had no significant effects, whereas a high dose (about 30 μM) significantly lowered the threshold of the L-Ca PIC from −58.5 ± 14.8 to −62.5 ± 3.6 mV and increased the amplitude of the Ca PIC from 0.69 ± 1.05 to 1.27 ± 1.1 nA. Thus Ca PICs in motoneurons are about 30-fold supersensitive to 5-HT in chronic spinal rats. The 5-HT–induced facilitation of the Ca PIC was blocked by nimodipine, not by the Ih current blocker Cs+ (3 mM) or the SK current blocker apamin (0.15 μM), and it lasted for hours after the removal of 5-HT from the nCSF, even increasing initially after removing 5-HT. The effects of 5-HT make motoneurons more excitable and ultimately lead to larger, more easily activated plateaus and self-sustained firing. The supersensitivity to 5-HT suggests the small amounts of endogenous 5-HT below the injury in a chronic spinal rat may act on supersensitive receptors to produce large Ca PICs and ultimately enable muscle spasms.

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