Threshold Depolarization Research Articles

Polar Liquid Cloud Base Detection Algorithms for High Spectral Resolution or Micropulse Lidar Data

AbstractLiquid layers in clouds affect their microphysical processes, as well as the surface energy budget. Studies focusing on these and other areas of research are often in need of skillful estimation of liquid‐bearing cloud layer boundaries. The bases of these layers are predominantly determined by ground‐based lidar instruments. Most studies requiring liquid cloud base height (LCBH) information use either fixed lidar parameter (depolarization and/or backscatter cross section) thresholds or cloud base height data products that do not distinguish between ice and liquid, all of which might introduce inconsistencies and errors in the resolved LCBH. In this paper, two explicit LCBH detection algorithms are presented. The first algorithm uses the high spectral resolution lidar (HSRL) data. Examination of this algorithm in multiple cases and scenarios during numerous days and first‐order comparison with microwave‐radiometer data show satisfactory results. The second algorithm incorporates widely available micropulse lidar (MPL) data for the LCBH detection. A 1‐year long comparison of data gathered at Barrow, Alaska, and McMurdo Station, Antarctica, which includes other cloud base detection methodologies (ceilometer, MPL value‐added product cloud base height, and LCBHs detected using a fixed MPL depolarization threshold), emphasizes the merits of the presented MPL algorithm. Examination of several unusual LCBH configurations suggests that the current practice of operating lidar at a tilting angle of 4° off zenith may not be sufficient to avoid specular reflection from oriented ice crystals. Data collected at Madison Wisconsin are used to show that specular reflection may impact measurements even at 4°.

Plasma membrane depolarization and permeabilization due to electric pulses in cell lines of different excitability

In electroporation-based medical treatments, excitable tissues are treated, either intentionally (irreversible electroporation of brain cancer, gene electrotransfer or ablation of the heart muscle, gene electrotransfer of skeletal muscles), or unintentionally (excitable tissues near the target area). We investigated how excitable and non-excitable cells respond to electric pulses, and if electroporation could be an effective treatment of the tumours of the central nervous system. For three non-excitable and one excitable cell line, we determined a strength-duration curve for a single pulse of 10ns–10ms. The threshold for depolarization decreased with longer pulses and was higher for excitable cells. We modelled the response with the Lapicque curve and the Hodgkin-Huxley model. At 1μs a plateau of excitability was reached which could explain why high-frequency irreversible electroporation (H-FIRE) electroporates but does not excite cells. We exposed cells to standard electrochemotherapy parameters (8×100μs pulses, 1Hz, different voltages). Cells behaved similarly which indicates that electroporation most probably occurs at the level of lipid bilayer, independently of the voltage-gated channels. These results could be used for optimization of electric pulses to achieve maximal permeabilization and minimal excitation/pain sensation. In the future, it should be established whether the in vitro depolarization correlates to nerve/muscle stimulation and pain sensation in vivo.

O99 Muscle velocity recovery cycles in neurogenic muscles

Muscle Velocity Recovery Cycles (MVRC) enable assessment of muscle membrane properties. MVRC has been studied in some myopathies, however, the method has never been tested in neurogenic muscles. We aimed in this study to examine whether MVRC differ in neurogenic and normal muscles and how MVRC variables correlate to the incidence of spontaneous activity (SA) in electromyography (EMG). Twenty-five patients with confirmed peroneal neuropathy (twenty) or L5 root lesion (five) were prospectively included. In addition to EMG, MVRC was done in anterior tibial muscle using Qtrac-software. MVRC parameters comprised muscle relative refractory period (MRRP) and early, late and residual supernormality after 1, 2 or 5 conditioning stimuli. Results were compared to 16 age- and sex-matched healthy subjects (HS). There was significant correlation between the incidence of SA and MRRP (r = 0.54) and early and late supernormality most pronounced in early supernormality after 5 conditioning stimulation (r = 0.64) (p < 0.001). The same MVRC parameters differed between patients and HS (t-test, p < 0.05). In contrast, residual supernormality did not differ between groups and there was no correlation to SA. Significantly prolonged MRRP and pronounced slowing in propagation velocity in early supernormal period suggest lower depolarization threshold in the denervated fibers. This is supported by significant correlation to incidence of SA in EMG. MVRC may be a potential diagnostic tool for electrodiagnosis of neuromuscular disorders and may provide information in understanding muscle pathophysiology. Future studies are required with larger patient groups with comparison of different nerve and muscle disorders.

Findings from Condensed-State Physics on Branched-Chain Amino Acids Apply to Voltage-Sensitive Ion Channels

Dramatic effects on channel function by mutations of branched to unbranched residues demonstrate the importance of branched-chain amino acids (BCAAs)—I, L and V—in voltage-sensitive ion channels (VSICs): L to A mutations in Shaker K channel S4 segments shift conductance-voltage curves.1 Substitution of unbranched for branched residues in fragments of Nav channels alters their voltage response.2 Mutations of L to A in S4 segments of Nav1.4 channels alters steady-state activation curves and slows inactivation.3 Voltage-sensor motion in Ci-VSP channels is tuned over two orders of magnitude or shifted 120 mV by I126 of S1.4 In a Kv channel, I237 controls the voltage dependence of charge movement, shifting the Q-V curve toward more positive voltages; I287 controls the energy barrier underlying activation.5 In 1986, Yoshino et al. investigated whether sidechains of amino acids engender ferroelectric properties in molecules with aromatic rings and chiral centers. Three of them did, dramatically: the BCAA sidechains. One compound with an isoleucine sidechain, 4'-3M2CPOOB, exhibited a ferroelectric phase with extremely large spontaneous polarization and dielectric permittivity ε over 3000.6, 7 Applying these condensed-state-physics findings to VSICs suggests that the high-electric-field resting channel is ferroelectric, with a high ε, so that the electrostatic repulsions between the positively charged arginine and lysine residues of S4 are weak. Threshold depolarization, according to the Channel Activation by Electrostatic Repulsion (CAbER) hypothesis,8 converts the channel to a nonpolar phase with much lower ε, causing these repulsions to greatly increase, which expands the S4 segments. CAbER proposes that this observed outward S4 motion drives the channel into a stochastically ion-conducting, active, conformation. 1. Lopez GA et al. (1991) Neuron 7:327-336. 2. Helluin O et al. (2001) IEEE Trans Diel Elect Insul 8:637-643. 3. Bendahhou S et al. (2007) BBA 1768:1440-1447. 4. Lacroix JJ, Bezanilla F (2012) Biophys J 103: L23-L25. 5. Lacroix JJ et al. (2014) PNAS 111(19):E1950-E1959. 6. Yoshino K et al. (1986) Japan J App Phys 25:L416-L418. 7. Yoshino K, Sakurai T (1991) In: Goodby JW et al., Ferroelectric Liquid Crystals, Gordon and Breach. 8. Leuchtag HR (2008) Voltage-Sensitive Ion Channels, Springer; (2016) Biophys J 110(3) 277a; http://doi.org/10.13140/RG.2.1.3360.2328.

What's Wrong with the Hodgkin-Huxley Model? An Exercise in Critical Thinking

Critical thinking is fundamental to science: let us examine an assumption of the Hodgkin-Huxley model of excitable membranes. The circuit elements of the HH model are a capacitance C and parallel ionic conductances g in series with their electromotive forces. HH assumes that C is constant while the gs vary with voltage and time. This assumption, adapted to voltage-sensitive ion channels, places the entire burden of explaining channel function on the gs. While the assumption of constant C—and its dimensionless coefficient, dielectric permittivity ε—is a good approximation for a membrane patch, its application at the macromolecular nanoscale is dubious, considering the ubiquitous presence of branched-chain amino acids—isoleucine, leucine and valine—in the channels. The sidechains of these amino acids (and no others) strongly affect ε in certain molecules with structures resembling channel segments. These ferroelectric liquid crystals, such as 4'-3M2CPOOB, exhibit ε values as high as 3000 in their ferroelectric state. Changes in channel ε affect all electrostatic interactions, including the mutual repulsions between the positively charged residues in S4 segments, key components of voltage sensors. The Channel Activation by Electrostatic Repulsion (CAbER) hypothesis assumes the resting channel, at a high electric field, is in a polar, far-from-equilibrium state with high ε, and that threshold depolarization converts it to a nonpolar state with much lower ε. The consequently increased Coulomb repulsion between arginines and lysines explains the observed outward motion of the S4 segments, which then drives the opening of the pore domain to stochastic ion currents. Does critical thinking support constant-C or CAbER? Problem: Two proton charges on a helix of radius 0.80 nm are 0.45 nm apart axially, with an angular separation of 300°. What is their repulsion energy (kJ/mol) when ε is 500? When it is 4? Further reading: HR Leuchtag, Voltage-Sensitive Ion Channels, Springer, 2008.

Hyperphosphorylated tau causes reduced hippocampal CA1 excitability by relocating the axon initial segment

Hyperphosphorylated tau has a critical role in tauopathies such as Alzheimer’s disease and frontotemporal dementia, impairing neuronal function and eventually leading to neurodegeneration. A critical role for tau is supported by studies in transgenic mouse models that express the P301L tau mutation found in cases of familial frontotemporal dementia, with the accumulation of hyperphosphorylated tau in the hippocampus causing reductions in hippocampal long-term potentiation and impairments in spatial learning and memory. However, what has remained unexplored is the role of hyperphosphorylated tau in reducing neuronal excitability. Here, we show in two complementary P301L tau transgenic mouse models that hyperphosphorylated tau induces a more depolarized threshold for action potential initiation and reduces firing in hippocampal CA1 neurons, which was rescued by the suppression of transgenic tau. Furthermore, using mutagenesis and primary hippocampal neuronal cultures, we reveal that this reduction in neuronal excitability results from the relocation of the axon initial segment (AIS) down the axon in a tau phosphorylation-dependent manner. We also demonstrate that this effect is microtubule-dependent. In addition, pharmacological stabilization was found to prevent both the structural and functional deficits caused by tau hyperphosphorylation. Finally, we demonstrate that the AIS of neurons from tau transgenic mice is further down the axon, which correlates with a reduction in excitability. We therefore propose that a reduction in hippocampal excitability due to a tau-mediated distal relocalization of the AIS contributes to the hippocampal dysfunction observed in tauopathies.

Two regulatory RNA elements affect TisB-dependent depolarization and persister formation.

Bacterial survival strategies involve phenotypic diversity which is generated by regulatory factors and noisy expression of effector proteins. The question of how bacteria exploit regulatory RNAs to make decisions between phenotypes is central to a general understanding of these universal regulators. We investigated the TisB/IstR-1 toxin-antitoxin system of Escherichia coli to appreciate the role of the RNA antitoxin IstR-1 in TisB-dependent depolarization of the inner membrane and persister formation. Persisters are phenotypic variants that have become transiently drug-tolerant by arresting growth. The RNA antitoxin IstR-1 sets a threshold for TisB-dependent depolarization under DNA-damaging conditions, resulting in two sub-populations: polarized and depolarized cells. Furthermore, our data indicate that an inhibitory 5' UTR structure in the tisB mRNA serves as a regulatory RNA element that delays TisB translation to avoid inappropriate depolarization when DNA damage is low. Investigation of the persister sub-population further revealed that both regulatory RNA elements affect persister levels as well as persistence time. This work provides an intriguing example of how bacteria exploit regulatory RNAs to control phenotypic heterogeneity.

Sensitized signalling between L-type Ca2+ channels and ryanodine receptors in the absence or inhibition of FKBP12.6 in cardiomyocytes.

The heart contraction is controlled by the Ca2+-induced Ca2+ release (CICR) between L-type Ca2+ channels and ryanodine receptors (RyRs). The FK506-binding protein FKBP12.6 binds to RyR subunits, but its role in stabilizing RyR function has been debated for long. Recent reports of high-resolution RyR structure show that the HD2 domain that binds to the SPRY2 domain of neighbouring subunit in FKBP-bound RyR1 is detached and invisible in FKBP-null RyR2. The present study was to test the consequence of FKBP12.6 absence on the in situ activation of RyR2. Using whole-cell patch-clamp combined with confocal imaging, we applied a near threshold depolarization to activate a very small fraction of LCCs, which in turn activated RyR Ca2+ sparks stochastically. FKBP12.6-knockout and FK506/rapamycin treatments increased spark frequency and LCC-RyR coupling fidelity without altering LCC open probability. Neither FK506 nor rapamycin further altered LCC-RyR coupling fidelity in FKBP12.6-knockout cells. In loose-seal patch-clamp experiments, the LCC-RyR signalling kinetics, indexed by the delay for a LCC sparklet to trigger a RyR spark, was accelerated after FKBP12.6 knockout and FK506/rapamycin treatments. These results demonstrated that RyRs became more sensitive to Ca2+ triggers without FKBP12.6. Isoproterenol (1 μM) further accelerated the LCC-RyR signalling in FKBP12.6-knockout cells. The synergistic sensitization of RyRs by catecholaminergic signalling and FKBP12.6 dysfunction destabilized the CICR system, leading to chaotic Ca2+ waves and ventricular arrhythmias. FKBP12.6 keeps the RyRs from over-sensitization, stabilizes the potentially regenerative CICR system, and thus may suppress the life-threatening arrhythmogenesis.

Variable Electrostatic Repulsion Model of Ion-Channel Activation Vs. Gated Pore with “Neutralized” S4 segments

The assumption of a gated pore in a voltage-sensitive ion channel (VSIC) [Hille, Ion Channels of Excitable Membranes, 2001] treats the channel's nanoworld as the macroworld with shrunken dimensions. It suggests deterministically voltage-dependent opening and closing of ion channels, contrary to the observed stochastic single-channel currents. Another anomaly is neglect of the mutual electrostatic repulsions between the positively charged arg and lys residues in the S4 segments, on grounds of a supposed neutralization by the fewer negatively charged residues in neighboring segments and/or an assumed transmembrane water column [Bezanilla 2008 Neuron 60:456-468; Börjesson, Elinder 2008 Cell Biochem Biophys 52:149-174]. An interdisciplinary model of the way membrane depolarization activates ion channels considers ferroelectric properties [Liu et al. 2012 Phys Rev Lett 108:078103] of resting VSICs [Leuchtag, Voltage-Sensitive Ion Channels, 2008]. In ferroelectric liquid crystals (FLCs) containing branched-chain amino acid (BCAA) (ile, leu, val) sidechains, the dielectric permittivity ε in the ferroelectric phase reaches values over 3000 [Yoshino et al. 1986 Japan J App Phys 25:L416-L418]. Assuming the VSIC with its ubiquitous BCAAs to be a FLC implies that a threshold depolarization changes the channel's phase from ferroelectric to nonpolar, sharply reducing its mean ε. This greatly increases the repulsive Coulomb forces within the S4s, expanding their α-helical pitch, consistent with observed outward S4 movements [Posson, Selvin 2008 Neuron 59:98-109]. The present model postulates that S4 expansion drives an allosteric conformational transition that allows permeant ions to hop along the widened pore domain and percolate [Zallen, The Physics of Amorphous Solids 1983] across the membrane. Observed effects of BCAA replacement with unbranched residues in sodium channels confirm model predictions [Helluin et al. 2001 IEEE Trans Diel Elect Insul 8:637-643; Bendahhou et al. 2007 Biochim Biophys Acta 1768:1440-1447].

Investigation in the Role of Lamin A (LMNA) Deficiency in Heart Block and Dilated Cardiomyopathy (DCM) Using Human Induced Pluripotent Stem Cell Derived Cardiomyocytes

Investigation in the Role of Lamin A (LMNA) Deficiency in Heart Block and Dilated Cardiomyopathy (DCM) Using Human Induced Pluripotent Stem Cell Derived Cardiomyocytes

Subthreshold High-Frequency Electrical Field Stimulation Induces VEGF Expression in Cardiomyocytes.

Subthreshold electrical stimulation (SES) has been shown to induce an improvement of angiogenesis in ischemic and nonischemic skeletal muscles, mediated by increased VEGF expression. VEGF plays a key role in physiological and pathological angiogenesis. Cardiomyocytes possess the ability to synthesize and secrete VEGF. Thus, we thought to investigate the effect of SES on VEGF regulation in cultured neonatal rat ventricular myocytes (NRVMs), in the aim to reveal new techniques for therapeutic angiogenesis in ischemic heart disease. Cell cultures of NRVMs were electrically stimulated with field strengths below the myocyte depolarization threshold (0.5 V/cm with 1 ms bipolar impulse duration). Frequencies ranging from 5 Hz up to 25, 50, and 99 Hz were applied over a period of 48 h. The expression of VEGF and its receptor KDR was determined with Western blot and ELISA. To reveal the biological activity of the secreted VEGF amount, cultured human coronary artery endothelial cells (HCAECs) were treated with the cell culture supernatant of NRVMs exposed to SES. A dominant effect of SES was observed at 25 Hz. Within this particular frequency the VEGF protein amount in the cytoplasm as well as in the cell culture supernatant increased significantly. In parallel, the protein expression of the KDR receptor decreased in a significant manner. Moreover, cell culture supernatant of NRVMs exposed to SES augmented the growth of HCAECs. Cardiomyocytes respond to SES with an increase in biologically active VEGF expression that promotes cell proliferation of HCAECs. This mechanism may provide new approaches to develop therapeutic angiogenesis in the ischemic heart.

PKD1 Protein Is Involved in Reactive Oxygen Species-mediated Mitochondrial Depolarization in Cooperation with Protein Kinase Cδ (PKCδ)

In this study, we used gene targeting in mice to identify the in vivo functions of PKD1. In addition to phenotypically characterizing the resulting knock-out animals, we also used mouse embryonic fibroblasts to investigate the associated signaling pathways in detail. This study is the first to use genetic deletion to reveal that PKD1 is a key regulator involved in determining the threshold of mitochondrial depolarization that leads to the production of reactive oxygen species. In addition, we also provide clear evidence that PKCδ is upstream of PKD1 in this process and acts as the activating kinase of PKD1. Therefore, our in vivo data indicate that PKD1 functions not only in the context of aging but also during nutrient deprivation, which occurs during specific phases of tumor growth.

Abstract 216: The Cerebral Blood Flow Threshold for Membrane Repolarization is 3 Times Higher Than That for Membrane Depolarization and is Lowered by Intraischemic Hypothermia

Introduction: It has been reported that cerebral blood flow (CBF) supplied by chest compression (20-40% of control levels) exceeds the CBF threshold for membrane depolarization (20%). However, the CBF threshold for membrane repolarization has not been studied. It is unknown whether chest compression can restore membrane potential. In the present study, we compared the CBF threshold for depolarization with that for repolarization. In addition, the effects of brain hypothermia on the repolarization threshold were evaluated. Methods: Rats (10 in each group) were anesthetized with isoflurane. CBF was measured by placing a laser-Doppler flow probe placed adjacent to a direct current (DC) potential electrode in the parietal cerebral cortex. Following bilateral occlusion of common carotid artery, CBF was continuously decreased by exsanguination at a speed of 5% of the baseline level every 2 min until a sudden negative DC shift was observed. After 5 or 10 min of ischemic depolarization, the CBF was restored at the same rate by returning blood until a positive DC shift was observed. In the hypothermia groups, brain temperature was decreased to 31°C by initiating nasopharyngeal cooling after depolarization. Result: The threshold for repolarization (46.5 ± 12%) was significantly higher than that for depolarization (19.2 ± 5%, p &lt; 0.001) after 5 min of depolarization time, and further increased to 61.5 ± 14% (p &lt; 0.05) after 10 min of depolarization time. Brain hypothermia significantly decreased the threshold for repolarization (5 min of ischemic depolarization: 33.8 ± 10%, p &lt; 0.05 compared with normal temperature, 10 min: 36.6 ± 6%, p &lt; 0.05). Conclusion: Thresholds for repolarization were significantly higher than those for depolarization and are likely to exceed the levels supplied by chest compression. Induced brain hypothermia during ischemia lowered CBF thresholds for repolarization and may help protect the brain during CPR.

S17-3. Clinical significance of ictal direct current shifts: Is it made by glia?

To assess clinical usefulness of ictal direct current (DC) shifts and also high frequency oscillations (HFO) for delineating epileptogenic zone in human partial epilepsy, we analyzed subdurally recorded EEG in 16 patients, and correlated the findings with pathology. 4, 9, 1 and 2 patients had the diagnosis of glioma, focal cortical dysplasia, low grade neuroepithelial tumor and hippocampal sclerosis, respectively. As the results, ictal DC shifts occurred in 75%, and ictal HFO in 44% of patients. 2 patients with glioma accompanied with focal cortical dysplasia showed ictal DC shifts and ictal HFO with good reproducibility. Ictal DC shifts occurred earlier than or as early as ictal HFO. Both of ictal DC shifts and HFO or either occurred in more restricted areas than conventional ictal changes. We conclude that both ictal DC shifts and HFO finally complementarily contributed to delineate core epileptogenic zone. Earlier occurrence of ictal DC shifts than HFO suggests an active role of glia in seizure generation. A tendency of ictal DC shifts and HFO to occur more in glioma with focal cortical dysplasia may reflect decreased threshold for abnormal depolarization of pyramidal neurons due to glial dysfunction.

Differential distribution of voltage-gated channels in myelinated and unmyelinated baroreceptor afferents

Voltage gated ion channels (VGC) make possible the frequency coding of arterial pressure and the neurotransmission of this information along myelinated and unmyelinated fiber pathways. Although many of the same VGC isoforms are expressed in both fiber types, it is the relative expression of each that defines the unique discharge properties of myelinated A-type and unmyelinated C-type baroreceptors. For example, the fast inward Na⁺ current is a major determinant of the action potential threshold and the regenerative transmembrane current needed to sustain repetitive discharge. In A-type baroreceptors the TTX-sensitive Na(v)1.7 VGC contributes to the whole cell Na⁺ current. Na(v)1.7 is expressed at a lower density in C-type neurons and in conjunction with TTX-insensitive Na(v)1.8 and Na(v)1.9 VGC. As a result, action potentials of A-type neurons have firing thresholds that are 15-20 mV more negative and upstroke velocities that are 5-10 times faster than unmyelinated C-type neurons. A more depolarized threshold in conjunction with a broader complement of non-inactivating K(V) VGC subtypes produces C-type action potentials that are 3-4 times longer in duration than A-type neurons and at markedly lower levels of cell excitability. Unmyelinated baroreceptors also express KCa1.1 which provides approximately 25% of the total outward K⁺ current. KCa1.1 plays a critically important role in shaping the action potential profile of C-type neurons and strongly impacts neuronal excitability. A-type neurons do not functionally express the KCa1.1 channel despite having a whole cell Ca(V) current quite similar to that of C-type neurons. As a result, A-type neurons do not have the frequency-dependent braking forces of KCa1.1. Lack of a KCa current and only a limited complement of non-inactivating K(V) VGC in addition to a hyperpolarization activated HCN1 current that is nearly 10 times larger than in C-type neurons leads to elevated levels of discharge in A-type neurons, a hallmark of myelinated baroreceptors. Interestingly, HCN2 and HCN4 expression levels are comparable in both fiber types. Collectively, such apportion of VGC constrains the neural coding of myelinated A-type baroreceptors to low threshold, high frequency, high fidelity discharge but with a limited capacity for neuromodulation of afferent bandwidth. Unmyelinated C-type baroreceptors require greater depolarizing forces for spike initiation and have a low frequency discharge profile that is often poorly correlated with the physiological stimulus. But the complement of VGC in C-type neurons provides far greater capacity for neuromodulation of cell excitability than can be obtained from A-type baroreceptors.

Safety Factor for Neuromuscular Transmission Is Reduced in Mild and Moderate Myasthenia Gravis Due to Loss of Both Sodium Channels and Acetylcholine Receptors (S35.006)

Objective: To evaluate the contributions of loss of Na + channels and acteylcholine receptors (AChR) to reduction in safety factor for neuromuscular transmission (SF) in patients with MGFA class IIa and IIIa myasthenia gravis (MG). I hypothesized that loss of endplate Na+ channels would contribute to the reduction of SF in patients with MG. Background Endplate Na + channels improve neuromuscular transmission by reducing the threshold depolarization needed to trigger an action potential (E AP ). In MG, AChRs and Na + channels are lost from the endplate. Na+ channels as well as AChRs were lost from the endplate membrane. Design/Methods: I used standard electrophysiological techniques to compare membrane currents and potentials in intercostal nerve-muscle preparations from 5 MGFA IIIa, 2 MGFA IIa and 7 control adult male subjects. Na + currents recorded using loose patch voltage clamp. Action potential (AP) characteristics measured with a 2 electrode clamp. Endplate potentials (EPPs) studied using μ-conotoxin GIIIb to block muscle Na + channels. The motor nerve was stimulated using a suction electrode. I report the findings for recordings from fast-glycolytic muscle fibers. Results: In MG, EPP was reduced from 40.2 ±1.3 to 30.1±1.3 mV (IIa) and 23.5 ±1.7 mV (IIIa) (p + channel loss increased E AP from −71.9 ±2.2 mV to -66.1 ±2.7 mV (IIa; p AP was the same for MG fibers and controls. The SF for neuromuscular transmission was reduced from 2.98 to 1.58 (IIa) and 1.09 (IIIa). Loss of Na + channels accounted for 53% (IIa) or 41% of the SF reduction. Reduction of EPP produced 47% (IIa) or 59% (IIIa) of the SF decrease. Conclusions: Loss of endplate Na + channels contributes to the reduced SF in MG to a similar extent as does the loss of endplate AChRs. Supported by: Department Veterans Affairs. Disclosure: Dr. Ruff has nothing to disclose.

Childhood ADHD Diagnosis, Behaviors, and Motor Overflow Correlate with Slower Inter-Hemispheric Inhibitory Signaling as Measured with Transcranial Magnetic Stimulation (P07.142)

Objective: The primary objective of this study was to determine whether Transcranial Magnetic Stimulation (TMS) -evoked Ipsilateral Silent Period (ISP) latency, which reflects trans-callosal, inter-hemispheric inhibition, correlates with presence and severity of Attention Deficit/Hyperactivity Disorder (ADHD) in childhood as well with commonly observed delays in motor control. Background Children with ADHD often fail to meet age-norms in development of motor control, particularly timed repetitive and sequential movements, motor overflow, and balance. The neural substrate of this motor delay and particularly motor overflow may include impaired or delayed mechanisms of inter-hemispheric, inhibitory signaling which contribute to both motor and behavioral symptoms. Design/Methods: Case-Control Study. Behavioral ratings, motor skills, and motor cortex physiology were evaluated in 50 ADHD (mean age 10.9 years, 29 male) and 64 typically developing (TD) (mean age 11.1 years, 33 male) children, all right-handed, ages 8-12 years. Motor skills were evaluated with the Physical and Neurological Examination for Soft Signs (PANESS). ISP and other physiological measures were obtained using TMS in left motor cortex. Results: ISP was successfully elicited in 54 (47%) children (23 ADHD, 31 TD, p = NS). Children in whom ISP was unobtainable were younger (p=.03) and had higher resting and active motor depolarization thresholds, but did not differ in other clinical, intellectual, or motor symptoms. In ADHD children, mean ISP latency was increased by 5 msec (12%) (p=.007). Longer ISP latencies (slower trans-hemispheric signaling) correlated with more severe ADHD symptom scores (r=.36, p = .009), particularly hyperactivity (r = .37, p = .007) as well as with worse motor overflow ratings on the PANESS (r = .27, p = .05) and diminished intra-hemispheric, TMS-evoked Short Interval Cortical Inhibition (SICI) (r = .34, p = .02). Conclusions: Slower trans-hemispheric inhibitory signaling reflects both behavioral and motor control problems in children with ADHD. Supported by: R01 MH078160. Disclosure: Dr. Gilbert has received personal compensation in an editorial capacity for Pediatrics Review and Education Program R11. Dr. Gilbert has received research support from Psyadon Pharmaceuticals and Otsuka Pharmaceuticals. Dr. Wu has received research support from Psyadon Pharmaceuticals, Otsuka Pharmaceutical and Genzyme Corporation. Dr. Shahana has nothing to disclose. Dr. Huddleston has nothing to disclose. Dr. Shiels has nothing to disclose. Dr. Mostofsky has nothing to disclose.

Endplate Na + Channels Loss Impairs Neuromuscular Transmission in Myasthenia Gravis

Myasthenia gravis (MG) is caused by acetylcholine receptor (AChR) antibodies inducing complement-medicated destruction of endplate membrane. Endplate Na+ channels enhance the safety factor for neuromuscular transmission by reducing the threshold depolarization needed to trigger an action potential (EAP). I previously demonstrated that MG antibodies do not directly alter the function of Na+ channels and that Na+ channels are lost by complement-mediated loss of endplate membrane. I examined the impact of postsynaptic voltage-gated Na+ channel loss on the SF. Comparing intercostal nerve-muscle preparations from controls and MG patients with moderate (M) or severe (S) MG, I found that endplate AChR loss decreased the endplate potential (EPP) from 40.2 ±1.3 to 30.1±1.3 mV (M) and 23.5 ±1.7 (S) (p < 0.001 for both). Endplate Na+ channel loss increased EAP from −71.9 ±2.2 to −66.1 ±2.7 (M; p < 0.01) and 62.3 ±2.7 mV (S; p < 0.001). In contrast, on extrajunctional membrane (> 0.1 cm from the endplate), EAP was the same for MG fibers and controls. The SF for neuromuscular transmission was reduced from 2.98 to 1.58 (M) and 1.09 (S). If the EPP was the same as control, the SF for MG fibers would have been 2.12 (M) or 1.86 (S). If EAP for MG fibers were the same as control fibers, the SF for MG fibers would have been 2.23 (M) or 1.74(S). In moderate MG, reduced EPP accounted for 47% of the SF reduction and increased EAP 53% of the SF reduction. In severe MG, reduced EPP accounted for 59% of the SF reduction and increased EAP 40% of the SF reduction. Conclusion: loss of endplate Na+ channels in MG contributes to the reduction in SF to a similar extent as does the loss of endplate AChRs.

Voltage-gated divalent currents in descending vasa recta pericytes

Multiple voltage-gated Ca(2+) channel (Ca(V)) subtypes have been reported to participate in control of the juxtamedullary glomerular arterioles of the kidney. Using the patch-clamp technique, we examined whole cell Ca(V) currents of pericytes that contract descending vasa recta (DVR). The dihydropyridine Ca(V) agonist FPL64176 (FPL) stimulated inward Ca(2+) and Ba(2+) currents that activated with threshold depolarizations to -40 mV and maximized between -20 and -10 mV. These currents were blocked by nifedipine (1 μM) and Ni(2+) (100 and 1,000 μM), exhibited slow inactivation, and conducted Ba(2+) > Ca(2+) at a ratio of 2.3:1, consistent with "long-lasting" L-type Ca(V). In FPL, with 1 mM Ca(2+) as charge carrier, Boltzmann fits yielded half-maximal activation potential (V(1/2)) and slope factors of -57.9 mV and 11.0 for inactivation and -33.3 mV and 4.4 for activation. In the absence of FPL stimulation, higher concentrations of divalent charge carriers were needed to measure basal currents. In 10 mM Ba(2+), pericyte Ca(V) currents activated with threshold depolarizations to -30 mV, were blocked by nifedipine, exhibited voltage-dependent block by diltiazem (10 μM), and conducted Ba(2+) > Ca(2+) at a ratio of ∼2:1. In Ca(2+), Boltzmann fits to the data yielded V(1/2) and slope factors of -39.6 mV and 10.0 for inactivation and 2.8 mV and 7.7 for activation. In Ba(2+), V(1/2) and slope factors were -29.2 mV and 9.2 for inactivation and -5.6 mV and 6.1 for activation. Neither calciseptine (10 nM), mibefradil (1 μM), nor ω-agatoxin IVA (20 and 100 nM) blocked basal Ba(2+) currents. Calciseptine (10 nM) and mibefradil (1 μM) also failed to reverse ANG II-induced DVR vasoconstriction, although raising mibefradil concentration to 10 μM was partially effective. We conclude that DVR pericytes predominantly express voltage-gated divalent currents that are carried by L-type channels.

Statistical Technique for Classification of MUAPs for Neuromuscular disease

Electromyograph (EMG) is useful to know the state of a patient under medical diagnosis and treatment. As the number of neuromuscular patients is increasing, it is not possible to take care of all the neuromuscular patients by carrying out manual investigations under all the conditions. Therefore it is required to design a computer aided expert system which can analyze and interpret the EMG signal. The EMG data acquisition and preprocessing, detection of MUAPs, classification of EMGs into similar groups, feature extraction of these groups and their usage in disease classification and diagnostics are the important stages in computer aided EMG analysis and interpretation. The objective of the present work is to detection of MUAPs, classification of EMGs into similar groups for computer aided analysis and interpretation of EMG signals for disease diagnosis. In the work real time recordings of myopathy, motor neuron disease and normal cases have been considered for MUAP segmentation and classification by statistical technique. EMG signal recorded by the needle electrode has been used. 1. INTRODUCTIONbody is composed of various systems and sub-systems. All human body functions are carried out by coordinated and controlled activities of the subsystems. The structural support is provided by bones while the functional support is provided by neuromuscular system. As such, the human body works under the influence of inputs from external environment and from within the body itself. Electromyography is used for the classification of various neuromuscular disorders .The word is made up of three parts: electro- + - myo- + -graphy. -Myo- is from the Greek mys, meaning muscle and graphy comes from the Greek grapho meaning to write. So electromyography literally is the writing (recording) of muscle electricity. The bioelectric potentials associated with the muscle activity constitute the electromyogram .EMG stands for electromyography. It is the study of muscle electrical signals EMGs can be used to detect abnormal muscle electrical activity that can occur in many diseases and conditions. Electromyogram (EMG) is a test that is used to record the electrical activity of muscles. When muscles are active, they produce an electrical current. This current is usually proportional to the level of the muscle activity. The electric current may be produced due to voluntary or involuntary muscle contraction. The EMG activity of voluntary muscle contractions is related to tension. The functional unit of the muscle contraction is a motor unit, which is comprised of a single alpha motor neuron and all the fibers it enervates. This muscle fiber contracts when the action potentials (impulse) of the motor nerve which supplies it reaches a depolarization threshold. The depolarization generates an electromagnetic field and the potential is measured as a voltage. The depolarization, which spreads along the membrane of the muscle, is a muscle action potential. The motor unit action potential is the spatial and temporal summation of the individual muscle action potentials for all the fibers of a single motor unit. Therefore, the EMG signal is the algebraic summation of the motor unit action potentials within the pick-up area of the electrode being used. The electrical signals produced by the muscles and nerves are analyzed to assess the state of neuromuscular functions in subjects with suspected neuromuscular disorders. The repetitive activation of several individual motor units results in superposed pulse train and constitutes the electromyogam signal. The potentials from individual motor units are characterized by unpredictable shapes depending upon the motor unit structure, electrode shapes and sizes, electrode placement, intervening tissues, and the state of neuromusculature. The analysis the EMG is based on its constitute i.e. motor unit action potential (MUAPs). The motor unit is the smallest functional unit of a muscle, which can be activated voluntarily. It consists of a group of muscle fibers, which are innervated from the same motor nerve. The shape of MUAP reflects the pathological and functional states of the motor unit. With increasing muscle force, the EMG signal shows an increase in the number of activated MUAPs recruited at increasing firing rate, making it difficult for the neurophysiologist to distinguish individual MUAP waveforms. In most of the clinical EMG examinations, it is the shape of the action potential that is analyzed for diagnostics. For this purpose, clinical examinations are carried out with the EMG signal recorded at low contraction level of the muscle, usually such that individual motor unit action potentials are distinguishable. This single motor unit activity is the summation of electrical activity of each muscle fiber within a motor unit. Figure1 shows the steps for the preprocessing and classification of MUAP into similar Classes.