The leguminous lectin of Lonchocarpus araripensis promotes antinociception via mechanisms that include neuronal inhibition of Na+ currents

The antiarrhythmic agent flecainide appears beneficial for painful congenital myotonia and LQT-3/ΔKPQ syndrome. Both diseases manifest small but persistent late Na+ currents in skeletal or cardiac myocytes. Flecainide may therefore block late Na+ currents for its efficacy. To investigate this possibility, we characterized state-dependent block of flecainide in wild-type and inactivation-deficient rNav1.4 muscle Na+ channels (L435W/L437C/A438W) expressed with β1 subunits in Hek293t cells. The flecainide-resting block at −140 mV was weak for wild-type Na+ channels, with an estimated 50% inhibitory concentration (IC50) of 365 μM when the cell was not stimulated for 1,000 s. At 100 μM flecainide, brief monitoring pulses of +30 mV applied at frequencies as low as 1 per 60 s, however, produced an ∼70% use-dependent block of peak Na+ currents. Recovery from this use-dependent block followed an exponential function, with a time constant over 225 s at −140 mV. Inactivated wild-type Na+ channels interacted with flecainide also slowly at −50 mV, with a time constant of 7.9 s. In contrast, flecainide blocked the open state of inactivation-deficient Na+ channels potently as revealed by its rapid time-dependent block of late Na+ currents. The IC50 for flecainide open-channel block at +30 mV was 0.61 μM, right within the therapeutic plasma concentration range; on-rate and off-rate constants were 14.9 μM−1s−1 and 12.2 s−1, respectively. Upon repolarization to −140 mV, flecainide block of inactivation-deficient Na+ channels recovered, with a time constant of 11.2 s, which was ∼20-fold faster than that of wild-type counterparts. We conclude that flecainide directly blocks persistent late Na+ currents with a high affinity. The fast-inactivation gate, probably via its S6 docking site, may further stabilize the flecainide-receptor complex in wild-type Na+ channels.

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The distribution of low-threshold TTX-resistant Na+ currents in rat trigeminal ganglion cells

Neuroscience ◽

10.1016/j.neuroscience.2012.07.012 ◽

2012 ◽

Vol 222 ◽

pp. 205-214 ◽

Cited By ~ 8

Author(s):

R.S. Scroggs

Keyword(s):

Trigeminal Ganglion ◽

Ganglion Cells ◽

Na Currents ◽

Low Threshold

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Abstract W MP41: Potentiation of Gaba-Mediated Synaptic Inhibition in the Recovery Phase: A Novel Therapeutic Target for Stroke

Stroke ◽

10.1161/str.45.suppl_1.wmp41 ◽

2014 ◽

Vol 45 (suppl_1) ◽

Author(s):

Takeshi Hiu ◽

Tonya Bliss ◽

Jeanne Paz ◽

Eric Wang ◽

Zoya Farzampour ◽

...

Keyword(s):

Pyramidal Cells ◽

Recovery Phase ◽

Synaptic Activity ◽

Stroke Recovery ◽

Functional Changes ◽

Post Stroke ◽

Array Tomography ◽

Neuronal Inhibition ◽

Gaba Signaling ◽

Novel Therapeutic

Background: Stroke is a major cause of disability yet pharmacotherapy targeting the recovery phase is lacking. Cortical circuit reorganization adjacent to the stroke site promotes recovery, thus elucidating mechanisms that promote this plasticity could lead to new therapeutics. Tonic neuronal inhibition, mediated by extrasynaptic GABA A receptors,inhibits post-stroke recovery. However, effects of phasic (synaptic) GABA signaling - which promotes plasticity during development - are unknown. Here we use a combined approach of i) array tomography to determine the composition of GABA synapses in the post-stroke mouse brain, ii) electrophysiology to determine whether stroke leads to functional changes in GABA-mediated phasic inhibition, and (iii) treatment with zolpidem, an FDA-approved GABA agonist, to modulate recovery. Results: We found, using array tomography, a 1.7-fold increase in the number of GABAergic synapses containing the α1 receptor subunit in layer 5 of the peri-infarct cortex (synapse number/μm 3 : 0.039±0.006 (control) vs 0.064±0.006 (stroke); P<0.01), but not in layer 2/3. There was an associated increase in spontaneous inhibitory post-synaptic currents (sIPSC) specific to layer 5 pyramidal neurons (sIPSC charge (fC): -403±27.8 (control) vs -724±166 (stroke); p=0.03). This effect was transient, occurring during the onset of functional recovery. To test whether the increased phasic inhibitory GABAergic signaling promotes stroke recovery, we treated animals with zolpidem, an agonist with high affinity for α1 subunit-containing GABA A receptors. Low dose zolpidem increased GABA A phasic signaling in layer 5 pyramidal cells and notably increased the rate and extent of behavioral recovery without altering infarct size. Conclusions: These data provide the first evidence that enhanced GABA A -mediated synaptic activity during the recovery phase improves stroke outcome. These data identify modulation of phasic GABA signaling as a novel therapeutic strategy for stroke, indicate zolpidem as a potential drug to improve recovery, and underscore the necessity to distinguish the role of tonic and phasic GABA inhibition in stroke recovery.

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Ion channels in spinal cord astrocytes in vitro. I. Transient expression of high levels of Na+ and K+ channels

Journal of Neurophysiology ◽

10.1152/jn.1992.68.4.985 ◽

1992 ◽

Vol 68 (4) ◽

pp. 985-1000 ◽

Cited By ~ 88

Author(s):

H. Sontheimer ◽

J. A. Black ◽

B. R. Ransom ◽

S. G. Waxman

Keyword(s):

Spinal Cord ◽

Membrane Potentials ◽

Resting Potential ◽

K Channel ◽

Na Channels ◽

Patch Clamp Recording ◽

K Channels ◽

Na Currents ◽

Channel Expression

1. Na+ and K+ channel expression was studied in cultured astrocytes derived from P--0 rat spinal cord using whole cell patch-clamp recording techniques. Two subtypes of astrocytes, pancake and stellate, were differentiated morphologically. Both astrocyte types showed Na+ channels and up to three forms of K+ channels at certain stages of in vitro development. 2. Both astrocyte types showed pronounced K+ currents immediately after plating. Stellate but not pancake astrocytes additionally showed tetrodotoxin (TTX)-sensitive inward Na+ currents, which displayed properties similar to neuronal Na+ currents. 3. Within 4-5 days in vitro (DIV), pancake astrocytes lost K(+)-current expression almost completely, but acquired Na+ currents in high densities (estimated channel density approximately 2-8 channels/microns2). Na+ channel expression in these astrocytes is approximately 10- to 100-fold higher than previously reported for glial cells. Concomitant with the loss of K+ channels, pancake astrocytes showed significantly depolarized membrane potentials (-28.1 +/- 15.4 mV, mean +/- SD), compared with stellate astrocytes (-62.5 +/- 11.9 mV, mean +/- SD). 4. Pancake astrocytes were capable of generating action-potential (AP)-like responses under current clamp, when clamp potential was more negative than resting potential. Both depolarizing and hyperpolarizing current injections elicited overshooting responses, provided that cells were current clamped to membrane potentials more negative than -70 mV. Anode-break spikes were evoked by large hyperpolarizations (less than -150 mV). AP-like responses in these hyperpolarized astrocytes showed a time course similar to neuronal APs under conditions of low K+ conductance. 5. In stellate astrocytes, AP-like responses were not observed, because the K+ conductance always exceeded Na+ conductance by at least a factor of 3. Thus stellate spinal cord astrocyte membranes are stabilized close to EK as previously reported for hippocampal astrocytes. 6. It is concluded that spinal cord pancake astrocytes are capable of synthesizing Na+ channels at densities that can, under some conditions, support electrogenesis. In vivo, however, AP-like responses are unlikely to occur because the cells' resting potential is too depolarized to allow current activation. Thus the absence of electrogenesis in astrocytes may be explained by two mechanisms: 1) a low Na-to-K conductance ratio, as in stellate spinal cord astrocytes and in other previously studied astrocyte preparations; or, 2) as described in detail in the companion paper, a mismatch between the h infinity curve and resting potential, which results in Na+ current inactivation in spinal cord pancake astrocytes.

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Culture-induced alterations in alveolar type II cell Na+ conductance

AJP Cell Physiology ◽

10.1152/ajpcell.1993.265.3.c630 ◽

1993 ◽

Vol 265 (3) ◽

pp. C630-C640 ◽

Cited By ~ 31

Author(s):

G. Yue ◽

P. Hu ◽

Y. Oh ◽

T. Jilling ◽

R. L. Shoemaker ◽

...

Keyword(s):

Alveolar Type ◽

Type Ii ◽

Na Transport ◽

Western Blots ◽

Adult Rat ◽

Antigenic Sites ◽

Cytoplasmic Proteins ◽

Na Currents ◽

Type Ii Cell ◽

Molecular Masses

Changes in Na+ transport in rat alveolar type II (ATII) cells during culture were quantified and related to alterations in spatial distribution of proteins antigenically related to amiloride-sensitive Na+ channels. Adult rat ATII cells were cultured for periods ranging from 24 to 96 h. When patch clamped in the whole cell mode, both freshly isolated and cultured ATII cells exhibited outwardly rectified Na+ currents. At 0 and 24 h in culture, these currents were equally inhibited by amiloride, benzamil, and 5-(N-ethyl-N-isopropyl)-2',4'-amiloride (inhibitory constant approximately 1 microM). These conductive pathways were equally permeable to Na+ and K+. Immunocytochemical localization at 0 or 24 h in culture revealed the presence of plasma membrane antigenic sites; after 48 h, the appearance of intracellular antigenic sites increased significantly. A single band of molecular mass 135 kDa in membrane proteins of freshly isolated ATII cells was recognized in Western blots; at 48 h in culture, two lower bands with molecular masses of 75 and 65 kDa were detected in either membrane or cytoplasmic proteins. Photolabeling with 2'-methoxy-5'-nitrobenzamil showed that the 135-, 75-, and 65-kDa bands contained amiloride-binding sites. These results suggest the presence of low amiloride affinity conductive pathways in freshly isolated and cultured ATII cells. Culturing ATII cells resulted in internalization and possible breakdown of these pathways and decreased Na+ transport.

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Colonic inflammation increases Na+ currents in bladder sensory neurons

Neuroreport ◽

10.1097/00001756-200412030-00008 ◽

2004 ◽

Vol 15 (17) ◽

pp. 2601-2605 ◽

Cited By ~ 47

Author(s):

Anna P. Malykhina ◽

Chao Qin ◽

Robert D. Foreman ◽

Hamid I. Akbarali

Keyword(s):

Sensory Neurons ◽

Colonic Inflammation ◽

Na Currents

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Modulation of respiration during brain hypoxia

Journal of Applied Physiology ◽

10.1152/jappl.1990.68.2.441 ◽

1990 ◽

Vol 68 (2) ◽

pp. 441-451 ◽

Cited By ~ 200

Author(s):

J. A. Neubauer ◽

J. E. Melton ◽

N. H. Edelman

Keyword(s):

Animal Models ◽

Respiratory Depression ◽

Cellular Metabolism ◽

Oxygen Availability ◽

Cellular Systems ◽

Respiratory Neuron ◽

Brain Hypoxia ◽

Cellular Basis ◽

Neuronal Inhibition ◽

Limited Oxygen

This review is a summary of the effects of brain hypoxia on respiration with a particular emphasis on those studies relevant to understanding the cellular basis of these effects. Special attention is given to mechanisms that may be responsible for the respiratory depression that appears to be the primary sequela of brain hypoxia in animal models. Although a variety of potential mechanisms for hypoxic respiratory depression are considered, emphasis is placed on changes in the neuromodulator constituency of the respiratory neuron microenvironment during hypoxia as the primary cause of this phenomenon. Hypoxia is accompanied by a net increase in neuronal inhibition due to both decreased excitatory and increased inhibitory neuromodulator levels. A survey of hypoxia-tolerant cellular systems and organisms suggests that hypoxic respiratory depression may be a manifestation of the depression of cellular metabolism, which appears to be a major adaptation to limited oxygen availability in these systems.

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