Changes In Electrophysiological Properties Research Articles

Generation of a Human Induced Pluripotent Stem Cell Line Expressing a Magnetic Resonance Imaging Reporter Gene.

The objective of the current study is to develop a new method for tracking transplanted human induced pluripotent stem cells-derived cardiomyocytes (hiPSC-CMs) using magnetic resonance imaging (MRI). The CRISPR/dCas9 activation system is employed to overexpress ferritin heavy chain (FHC) in hiPSC-CMs. The mRNA and protein expression of FHC in hiPSC and hiPSC-CMs significantly increased after transfection. Iron chloride does not affect the cell viability in a concentration range from 0 to 2000 µm. hiPSCs overexpressing FHC (hiPSC- FHCOE) and hiPSC-CMs overexpressing FHC (hiPSC-CM-FHCOE) significantly enhanced cellular uptake of iron chloride but with no changes in electrophysiological properties compared to hiPSC-CM-Control. Furthermore, hiPSC-CM-FHCOE presented robust contrast and lower T2* values, signifying their potential as highly effective candidates for cardiac MRI. Next, hiPSC-CM-FHCOE is injected into mouse hearts and after 3 days of transplantation, MR images are obtained. hiPSC-CM-FHCOE cells exhibited clear signals in the hearts with lower T2* and rapid signal decay. Collectively, data from this proof-of-concept study demonstrated that endogenous labeling with FHC in hiPSC-CMs can be a potent strategy for enhancing the accuracy of cardiac MRI. This technology represents a significant step forward in tracking the transplanted hiPSC-CMs in the hearts of live animals.

Electrophysiological properties of layer 2/3 pyramidal neurons in the primary visual cortex of a retinitis pigmentosa mouse model (rd10).

Retinal degeneration is one of the main causes of visual impairment and blindness. One group of retinal degenerative diseases, leading to the loss of photoreceptors, is collectively termed retinitis pigmentosa. In this group of diseases, the remaining retina is largely spared from initial cell death making retinal ganglion cells an interesting target for vision restoration methods. However, it is unknown how downstream brain areas, in particular the visual cortex, are affected by the progression of blindness. Visual deprivation studies have shown dramatic changes in the electrophysiological properties of visual cortex neurons, but changes on a cellular level in retinitis pigmentosa have not been investigated yet. Therefore, we used the rd10 mouse model to perform patch-clamp recordings of pyramidal neurons in layer 2/3 of the primary visual cortex to screen for potential changes in electrophysiological properties resulting from retinal degeneration. Compared to wild-type C57BL/6 mice, we only found an increase in intrinsic excitability around the time point of maximal retinal degeneration. In addition, we saw an increase in the current amplitude of spontaneous putative inhibitory events after a longer progression of retinal degeneration. However, we did not observe a long-lasting shift in excitability after prolonged retinal degeneration. Together, our results provide evidence of an intact visual cortex with promising potential for future therapeutic strategies to restore vision.

Developmental changes in electrophysiological properties of auditory cortical neurons in the Cntnap2 knockout rat.

Disruptions in the CNTNAP2 gene are known to cause language impairments and symptoms associated with autism spectrum disorder (ASD). Importantly, knocking out this gene in rodents results in ASD-like symptoms that include auditory processing deficits. This study used in vitro patch-clamp electrophysiology to examine developmental alterations in auditory cortex pyramidal neurons of Cntnap2-/- rats, hypothesizing that CNTNAP2 is essential for maintaining intrinsic neuronal properties and synaptic wiring in the developing auditory cortex. Whole-cell patch-clamp recordings were conducted in wildtype and Cntnap2-/- littermates at 3 postnatal age ranges (P8-12, P18-21, and P70-90). Consistent changes across age were seen in all measures of intrinsic membrane properties and spontaneous synaptic input. Intrinsic cell properties such as action potential half widths, rheobase, and action-potential firing frequencies were different between wildtype and Cntnap2-/- rats predominantly during the juvenile stage (P18-21), whereas adult Cntnap2-/- rats showed higher frequencies of spontaneous and mini excitatory post-synaptic currents (sEPSC; mEPSC), with lower sEPSC amplitudes. These results indicate that intrinsic cell properties are altered in Cntnap2-/- during the juvenile age, leading to a hyperexcitable phenotype during this stage of synaptic remodeling and refinement. While intrinsic properties eventually normalize by reaching adulthood, changes in synaptic input, potentially caused by the differences in intrinsic membrane properties, seem to manifest in the adult age and are presumably responsible for the hyperreactive behavioral phenotype. In conjunction with a previous study, the present results also indicate a large influence of breeding scheme, i.e., pre- or postnatal environment, on the impact of Cntnap2 on cellular physiology.

Changes in electrophysiological properties of pyramidal neuron in motor cortex during the postnatal early development of mice

To investigate the electrophysiological properties of pyramidal neurons in mouse motor cortex during the early postnatal development. Thirty-six mice were randomly divided into postnatal 1-, 2-, 3-Week and 1-, 2-,3-Month groups (n=6). Membrane properties, action potentials (AP) and spontaneous excitatory postsynaptic currents (sEPSCs) of motor cortex pyramidal neurons were recorded to evaluate the changes in the intrinsic electrophysilogical characteristics by using whole cell patch clamp. Pyramidal neurons and interneurons were distinguished according to the AP firing patterns. Comparing with interneurons, pyramidal neurons exhibited regular spiking (RS) with smaller frequency. During the period of postnatal 1 Week-3 Months, some of the intrinsic membrane properties of motor cortex pyramidal neurons changed. Compared to the 1-Week mice, the resting membrane potential (RMP) of 2-Week decreased significantly (P＜0.01), and the membrane input resistance (Rin) of 1-Month got a hyperpolarization (P＜0.01), and they showed no significant change in the next period, while the membrane capacitance (Cm) showed no significant changes during the whole postnatal development. The AP dynamic properties changed significantly during this period. Compared to the 1-Week mice, the absolute value of the AP threshold and the AP amplitude of the 3-Week increased significantly (P＜0.01), while the spike half width of the 2-Week decreased substantially (P＜0.05), and they showed no significant change in the next period. The sEPSCs frequency and amplitude of 1- Month increased significantly compared to the 1-Week mice(P＜0.01), while during the period of next 1 Month-3 Months, the amplitude and frequency showed no significant change. These results suggest that the motor cortex pyramidal neurons have time-specific eletrophysilogical properties during the postnatal development. The electrophysiological properties can be used as a functional index to detect the degree of neurons maturity, and as a marker to distinguish the pyramidal neurons and interneurons.

Atrial AMP-activated protein kinase is critical for prevention of dysregulation of electrical excitability and atrial fibrillation.

Metabolic stress is an important cause of pathological atrial remodeling and atrial fibrillation. AMPK is a ubiquitous master metabolic regulator, yet its biological function in the atria is poorly understood in both health and disease. We investigated the impact of atrium-selective cardiac AMPK deletion on electrophysiological and structural remodeling in mice. Loss of atrial AMPK expression caused atrial changes in electrophysiological properties and atrial ectopic activity prior to the onset of spontaneous atrial fibrillation. Concomitant transcriptional downregulation of connexins and atrial ion channel subunits manifested with delayed left atrial activation and repolarization. The early molecular and electrophysiological abnormalities preceded left atrial structural remodeling and interstitial fibrosis. AMPK inactivation induced downregulation of transcription factors (Mef2c and Pitx2c) linked to connexin and ion channel transcriptional reprogramming. Thus, AMPK plays an essential homeostatic role in atria, protecting against adverse remodeling potentially by regulating key transcription factors that control the expression of atrial ion channels and gap junction proteins.

Kinetic Alterations in Resurgent Sodium Currents of Mutant Nav1.4 Channel in Two Patients Affected by Paramyotonia Congenita

Simple SummaryParamyotonia congenita (PMC) is a rare muscle disorder that causes myotonia and weakness of facial and limb muscles. The electromyography in PMC shows continual spontaneous, high-frequency spike potentials in skeletal muscles. Genetic mutations in the Nav1.4 channel that cause hyperexcitability of muscle fibers are responsible for PMC. However, the genotype–phenotype relationship is highly diversified, and the molecular pathology remains unclear. Here, we investigated the electrophysiology in the Nav1.4 channel with mutations, p.V781I and p.A1737T, which were found in two Taiwanese patients. We identified the distinct changes in gating mechanisms altered by mutations which may underlie the clinical phenotype.Paramyotonia congenita (PMC) is a rare skeletal muscle disorder characterized by muscle stiffness upon repetitive exercise and cold exposure. PMC was reported to be caused by dominant mutations in the SCN4A gene encoding the α subunit of the Nav1.4 channel. Recently, we identified two missense mutations of the SCN4A gene, p.V781I and p.A1737T, in two PMC families. To evaluate the changes in electrophysiological properties caused by the mutations, both mutant and wild-type (WT) SCN4A genes were expressed in CHO-K1 and HEK-293T cells. Then, whole-cell patch-clamp recording was employed to study the altered gating of mutant channels. The activation curve of transient current showed a hyperpolarizing shift in both mutant Nav1.4 channels as compared to the WT channel, whereas there was a depolarizing shift in the fast inactivation curve. These changes confer to an increase in window current in the mutant channels. Further investigations demonstrated that the mutated channel proteins generate significantly larger resurgent currents as compared to the WT channel and take longer to attain the peak of resurgent current than the WT channel. In conclusion, the current study demonstrates that p.V781I and p.A1737T mutations in the Nav1.4 channel increase both the sustained and the resurgent Na+ current, leading to membrane hyperexcitability with a lower firing threshold, which may influence the clinical phenotype.

Spinal cord injury-mediated changes in electrophysiological properties of rat gastric nodose ganglion neurons

In preclinical rodent models, spinal cord injury (SCI) manifests as gastric vagal afferent dysfunction both acutely and chronically. However, the mechanism that underlies this dysfunction remains unknown. In the current study, we examined the effect of SCI on gastric nodose ganglia (NG) neuron excitability and on voltage-gated Na+ (NaV) channels expression and function in rats after an acute (i.e. 3-days) and chronic (i.e. 3-weeks) period. Rats randomly received either T3-SCI or sham control surgery 3-days or 3-weeks prior to experimentation as well as injections of 3% DiI solution into the stomach to identify gastric NG neurons. Single cell qRT-PCR was performed on acutely dissociated DiI-labeled NG neurons to measure NaV1.7, NaV1.8 and NaV1.9 expression levels. The results indicate that all 3 channel subtypes decreased. Current- and voltage-clamp whole-cell patch-clamp recordings were performed on acutely dissociated DiI-labeled NG neurons to measure active and passive properties of C- and A-fibers as well as the biophysical characteristics of NaV1.8 channels in gastric NG neurons. Acute and chronic SCI did not demonstrate deleterious effects on either passive properties of dissociated gastric NG neurons or biophysical properties of NaV1.8. These findings suggest that although NaV gene expression levels change following SCI, NaV1.8 function is not altered. The disruption throughout the entirety of the vagal afferent neuron has yet to be investigated.

Rutaecarpine targets hERG channels and participates in regulating electrophysiological properties leading to ventricular arrhythmia.

Drug‐mediated or medical condition‐mediated disruption of hERG function accounts for the main cause of acquired long‐QT syndrome (acLQTs), which predisposes affected individuals to ventricular arrhythmias (VA) and sudden death. Many Chinese herbal medicines, especially alkaloids, have risks of arrhythmia in clinical application. The characterized mechanisms behind this adverse effect are frequently associated with inhibition of cardiac hERG channels. The present study aimed to assess the potent effect of Rutaecarpine (Rut) on hERG channels. hERG‐HEK293 cell was applied for evaluating the effect of Rut on hERG channels and the underlying mechanism. hERG current (IhERG) was measured by patch‐clamp technique. Protein levels were analysed by Western blot, and the phosphorylation of Sp1 was determined by immunoprecipitation. Optical mapping and programmed electrical stimulation were used to evaluate cardiac electrophysiological activities, such as APD, QT/QTc, occurrence of arrhythmia, phase singularities (PSs), and dominant frequency (DF). Our results demonstrated that Rut reduced the IhERG by binding to F656 and Y652 amino acid residues of hERG channel instantaneously, subsequently accelerating the channel inactivation, and being trapped in the channel. The level of hERG channels was reduced by incubating with Rut for 24 hours, and Sp1 in nucleus was inhibited simultaneously. Mechanismly, Rut reduced threonine (Thr)/ tyrosine (Tyr) phosphorylation of Sp1 through PI3K/Akt pathway to regulate hERG channels expression. Cell‐based model unables to fully reveal the pathological process of arrhythmia. In vivo study, we found that Rut prolonged QT/QTc intervals and increased induction rate of ventricular fibrillation (VF) in guinea pig heart after being dosed Rut for 2 weeks. The critical reasons led to increased incidence of arrhythmias eventually were prolonged APD90 and APD50 and the increase of DF, numbers of PSs, incidence of early after‐depolarizations (EADs). Collectively, the results of this study suggest that Rut could reduce the IhERG by binding to hERG channels through F656 and Y652 instantaneously. While, the PI3K/Akt/Sp1 axis may play an essential role in the regulation of hERG channels, from the perspective of the long‐term effects of Rut (incubating for 24 hours). Importantly, the changes of electrophysiological properties by Rut were the main cause of VA.

A single injection of periostin decreases cardiac voltage-gated Na+ channel in rat ventricles.

Changes in electrophysiological properties, such as ion channel expression and activity, are closely related to arrhythmogenesis during heart failure (HF). However, a causative factor for the electrical remodeling in HF has not been determined. Periostin (POSTN), a matricellular protein, is increased in heart tissues of patients with HF. In the present study, we investigated whether a single injection of POSTN affects the electrophysiological properties in rat ventricles. After male Wistar rats were intravenously injected with recombinant rat POSTN (64 µg/kg, 24 hr), electrocardiogram (ECG) was recorded. Whole-cell patch clamp was performed to measure action potential (AP) and Na+ current (INa) in isolated ventricular myocytes. Protein expression of cardiac voltage-gated Na+ channel (NaV1.5) in isolated ventricles was examined by Western blotting. In ECG, POSTN-injection significantly increased RS height. POSTN-injection significantly delayed time to peak in AP and decreased INa in the isolated ventricular myocytes. POSTN-injection decreased NaV1.5 expression in the isolated ventricles. It was confirmed that POSTN (1 µg/ml, 24 hr) decreased INa and NaV1.5 protein expression in neonatal rat ventricular myocytes. This study for the first time demonstrated that a single injection of POSTN in rats decreased INa by suppressing NaV1.5 expression in the ventricular myocytes, which was accompanied by a prolongation of time to peak in AP and an increase of RS height in ECG.

The role of β-adrenergic system remodeling in human heart failure: A mechanistic investigation

β-adrenergic receptor antagonists (β-blockers) are extensively used to improve cardiac performance in heart failure (HF), but the electrical improvements with these clinical treatments are not fully understood. The aim of this study was to analyze the electrophysiological effects of β-adrenergic system remodeling in heart failure with reduced ejection fraction and the underlying mechanisms. We used a combined mathematical model that integrated β-adrenergic signaling with electrophysiology and calcium cycling in human ventricular myocytes. HF remodeling, both in the electrophysiological and signaling systems, was introduced to quantitatively analyze changes in electrophysiological properties due to the stimulation of β-adrenergic receptors in failing myocytes. We found that the inotropic effect of β-adrenergic stimulation was reduced in HF due to the altered Ca2+ dynamics resulting from the combination of structural, electrophysiological and signaling remodeling. Isolated cells showed proarrhythmic risk after sympathetic stimulation because early afterdepolarizations appeared, and the vulnerability was greater in failing myocytes. When analyzing coupled cells, β-adrenergic stimulation reduced transmural repolarization gradients between endocardium and epicardium in normal tissue, but was less effective at reducing these gradients after HF remodeling. The comparison of the selective activation of β-adrenergic isoforms revealed that the response to β2-adrenergic receptors stimulation was blunted in HF while β1-adrenergic receptors downstream effectors regulated most of the changes observed after sympathetic stimulation. In conclusion, this study was able to reproduce an altered β-adrenergic activity on failing myocytes and to explain the mechanisms involved. The derived predictions could help in the treatment of HF and guide in the design of future experiments.

Ranolazine-Mediated Attenuation of Mechanoelectric Feedback in Atrial Myocyte Monolayers.

BackgroundMechanical stretch increases Na+ inflow into myocytes, related to mechanisms including stretch-activated channels or Na+/H+ exchanger activation, involving Ca2+ increase that leads to changes in electrophysiological properties favoring arrhythmia induction. Ranolazine is an antianginal drug with confirmed beneficial effects against cardiac arrhythmias associated with the augmentation of INaL current and Ca2+ overload.ObjectiveThis study investigates the effects of mechanical stretch on activation patterns in atrial cell monolayers and its pharmacological response to ranolazine.MethodsConfluent HL-1 cells were cultured in silicone membrane plates and were stretched to 110% of original length. The characteristics of in vitro fibrillation (dominant frequency, regularity index, density of phase singularities, rotor meandering, and rotor curvature) were analyzed using optical mapping in order to study the mechanoelectric response to stretch under control conditions and ranolazine action.ResultsHL-1 cell stretch increased fibrillatory dominant frequency (3.65 ± 0.69 vs. 4.35 ± 0.74 Hz, p < 0.01) and activation complexity (1.97 ± 0.45 vs. 2.66 ± 0.58 PS/cm2, p < 0.01) under control conditions. These effects were related to stretch-induced changes affecting the reentrant patterns, comprising a decrease in rotor meandering (0.72 ± 0.12 vs. 0.62 ± 0.12 cm/s, p < 0.001) and an increase in wavefront curvature (4.90 ± 0.42 vs. 5.68 ± 0.40 rad/cm, p < 0.001). Ranolazine reduced stretch-induced effects, attenuating the activation rate increment (12.8% vs. 19.7%, p < 0.01) and maintaining activation complexity—both parameters being lower during stretch than under control conditions. Moreover, under baseline conditions, ranolazine slowed and regularized the activation patterns (3.04 ± 0.61 vs. 3.65 ± 0.69 Hz, p < 0.01).ConclusionRanolazine attenuates the modifications of activation patterns induced by mechanical stretch in atrial myocyte monolayers.

Effect of remote ischemic preconditioning on electrophysiological parameters in nonvalvular paroxysmal atrial fibrillation: The RIPPAF Randomized Clinical Trial

Atrial fibrillation (AF) remains the most relevant arrhythmia with a prevalence of 2%. The treatment options are either highly invasive and cost-intensive or limited by potential side effects or insufficient efficacy. However, no direct means of prevention that could reduce the burden of AF have been tested. The purpose of this study was to determine whether remote ischemic preconditioning (RIPC) has an impact on inducibility and sustainability of AF. A total of 146 patients with paroxysmal AF undergoing electrophysiology study were randomized to receive either RIPC, performed by short episodes of forearm ischemia, or sham intervention (clinicaltrials.gov identifier: NCT02779660). Effective refractory periods, conduction times, velocities, and conduction delays measured were analyzed by pacing from the coronary sinus (CS). End points of the study were the inducibility and sustainability of AF after prespecified rapid pacing sequences. RIPC significantly reduces the inducibility (odds ratio 0.35; 95% confidence interval 0.17-0.71; P = .003) and sustainability (odds ratio 0.36; 95% confidence interval 0.16-0.81; P=.01) of AF. Furthermore, it decreased dispersion of atrial refractory periods (16.0 ± 14.0 ms vs 22.7 ± 19.0 ms; P = .021) as well as atrial conduction delays (49.2 ± 19.6 ms vs 56.2 ± 22.5 ms; P =.049 for proximal CS and 42.4 ± 16.6 ms vs 49.8 ± 22.2 ms; P = .029 for distal CS). In the whole cohort, longer atrial conduction delay (57.6 ± 22.2 ms vs 50.0 ± 20.5 ms; P = .044) and slower conduction velocity (1.74 ± 0.3 mm/ms vs 1.93 ± 0.5 mm/ms; P =.006) were associated with inducibility of AF whereas a wider dispersion of effective refractory periods (25.9 ± 18.3 ms vs 15.7 ± 11.6 ms; P = .028) maintained AF episodes. RIPC reduces the inducibility and sustainability of AF, which is possibly mediated by changes in electrophysiological properties of the atria. It may be used as a simple noninvasive procedure to reduce AF burden.

Abrupt Change in Electrophysiological Properties Begins From Postnatal Day 7 Before Hearing Onset in the Developing Mice Auditory Cortical Layer II/III Neurons.

ObjectivesIn the developing auditory cortex, maturation of electrophysiological properties and cell types before and after hearing onset has been reported previously. However, the exact timing of firing pattern change has not been reported. In this study, firing pattern change was investigated from postnatal day 3 (P3) to P12 in auditory cortical layer II/III neurons to investigate whether firing pattern changes dramatically after a specific point during development.MethodsICR mice pups aged from P3 to P12 were sacrificed to obtain 300-mm-thick brain slices containing the primary auditory cortex. From cortical layer II/III neurons, the patterns of action potential firing generated by current injection were examined using whole cell current clamp technique and the characteristics of Na+ currents involved in action potential firing were investigated using whole cell voltage clamp technique.ResultsFrom P3 to P6, most cells did not show action potential firing (29 of 46 cells), and some cells responding to current injection showed a single action potential at the initial depolarizing current step (17 of 46 cells). This firing pattern changes from P7. From P7 to P9, cells begin to show regular spiking to current injection. The spiking frequency increased after P10. In studying Na+ current with whole cell voltage clamp, Na+ current densities increased gradually (32.0±2.0 pA/pF [P3–P6, n=7], 51.2±2.0 pA/pF [P7–P9, n=13], and 69.5±3.7 pA/pF [P10–P12, n=13]) in low external [Na+] condition. Na+ current recovery was accelerated and inactivation curves shifted to hyperpolarization with age.ConclusionAs regular spiking cells were observed from P7 but never from P3 to P6, P7 might be regarded as an important milestone in the development of auditory cortical layer II/III neurons. This change might mainly result from the increase in Na+ current density.

Orofacial operant behaviors and electrophysiological properties of trigeminal ganglion neurons following masseter muscle inflammation in rats

Orofacial muscle pain is a significant clinical problem because it affects eating, speaking, and other orofacial functions in patients. However, mechanisms underlying orofacial muscle pain are not fully understood. In the present study we induced orofacial muscle pain by injecting Complete Freund’s Adjuvant (CFA) into masseter muscle of rats and assessed pain by the orofacial operant test. In comparison with the control group, CFA-injected animals (CFA group) showed decreases in operant behaviors, suggesting the presence of orofacial pain. Trigeminal ganglion (TG) neurons innervating masseter muscles were retrograde-labeled with DiI and their electrophysiological properties studied using patch-clamp recordings. About 20% of DiI-labeled TG neurons showed spontaneous action potentials (APs) in the CFA group but none in the control group. AP rheobase levels were significantly lower in DiI-labeled TG neurons of the CFA group than in the control group. Membrane input resistance of DiI-labeled TG neurons was significantly higher in the CFA group than in the control group. Several other membrane parameters were also different between DiI-labeled TG neurons of the CFA and control groups. Voltage-dependent currents were examined and the most significant changes following CFA were background K+ currents, which showed significantly smaller in DiI-labeled TG neurons of CFA group compared to the control group. Collectively, orofacial muscle pain in CFA model is accompanied with changes of electrophysiological properties and background K+ currents in TG neurons that innervate masseter muscles.

Changes in electrophysiological properties of photoreceptors in Periplaneta americana associated with the loss of screening pigment.

Absence of screening pigment in insect compound eyes has been linked to visual dysfunction. We investigated how its loss in a white-eyed mutant (W-E) alters the photoreceptor electrophysiological properties, opsin gene expression, and the behavior of the cockroach, Periplaneta americana. Whole-cell patch-clamp recordings of green-sensitive photoreceptors in W-E cockroaches gave reduced membrane capacitance, absolute sensitivity to light, and light-induced currents. Decreased low-pass filtering increased voltage-bump amplitudes in W-E photoreceptors. Intracellular recordings showed that angular sensitivity of W-E photoreceptors had two distinct components: a large narrow component with the same acceptance angle as wild type, plus a relatively small wide component. Information processing was evaluated using Gaussian white-noise modulated light stimulation. In bright light, W-E photoreceptors demonstrated higher signal gain and signal power than wild-type photoreceptors. Expression levels of the primary UV- and green-sensitive opsins were lower and the secondary green-sensitive opsin significantly higher in W-E than in wild-type retinae. In behavioral experiments, W-E cockroaches were significantly less active in dim green light, consistent with the relatively low light sensitivity of their photoreceptors. Overall, these differences can be related to the loss of screening pigment function and to a compensatory decrease in the rhabdomere size in W-E retinae.

Changes in membrane properties of rat deep cerebellar nuclear projection neurons during acquisition of eyeblink conditioning

Previous studies have shown changes in membrane properties of neurons in rat deep cerebellar nuclei (DCN) as a function of development, but due to technical difficulties in obtaining viable DCN slices from adult animals, it remains unclear whether there are learning-related alterations in the membrane properties of DCN neurons in adult rats. This study was designed to record from identified DCN cells in cerebellar slices from postnatal day 25-26 (P25-26) rats that had a relatively mature sensory nervous system and were able to acquire learning as a result of tone-shock eyeblink conditioning (EBC) and to document resulting changes in electrophysiological properties. After electromyographic electrode implantation at P21 and inoculation with a fluorescent pseudorabies virus (PRV-152) at P22-23, rats received either four sessions of paired delay EBC or unpaired stimulus presentations with a tone conditioned stimulus and a shock unconditioned stimulus or sat in the training chamber without stimulus presentations. Compared with rats given unpaired stimuli or no stimulus presentations, rats given paired EBC showed an increase in conditioned responses across sessions. Whole-cell recordings of both fluorescent and nonfluorescent DCN projection neurons showed that delay EBC induced significant changes in membrane properties of evoked DCN action potentials including a reduced after-hyperpolarization amplitude and shortened latency. Similar findings were obtained in hyperpolarization-induced rebound spikes of DCN neurons. In sum, delay EBC produced significant changes in the membrane properties of juvenile rat DCN projection neurons. These learning-specific changes in DCN excitability have not previously been reported in any species or task.

Phenotypic plasticity in Periplaneta americana photoreceptors

Plasticity is a crucial aspect of neuronal physiology essential for proper development and continuous functional optimization of neurons and neural circuits. Despite extensive studies of different visual systems, little is known about plasticity in mature microvillar photoreceptors. Here we investigate changes in electrophysiological properties and gene expression in photoreceptors of the adult cockroach, Periplaneta americana, after exposure to constant light (CL) or constant dark (CD) for several months. After CL, we observed a decrease in mean whole-cell capacitance, a proxy for cell membrane area, from 362 ± 160 to 157 ± 58 pF, and a decrease in absolute sensitivity. However, after CD, we observed an increase in capacitance to 561 ± 155 pF and an increase in absolute sensitivity. Small changes in the expression of light-sensitive channels and signaling molecules were detected in CD retinas, together with a substantial increase in the expression of the primary green-sensitive opsin (GO1). Accordingly, light-induced currents became larger in CD photoreceptors. Even though normal levels of GO1 expression were retained in CL photoreceptors, light-induced currents became much smaller, suggesting that factors other than opsin are involved. Latency of phototransduction also decreased significantly in CL photoreceptors. Sustained voltage-activated K+ conductance was not significantly different between the experimental groups. The reduced capacitance of CL photoreceptors expanded their bandwidth, increasing the light-driven voltage signal at high frequencies. However, voltage noise was also amplified, probably because of unaltered expression of TRPL channels. Consequently, information transfer rates were lower in CL than in control or CD photoreceptors. These changes in whole-cell capacitance and electrophysiological parameters suggest that structural modifications can occur in the photoreceptors to adapt their function to altered environmental conditions. The opposing patterns of modifications in CL and CD photoreceptors differ profoundly from previous findings in Drosophila melanogaster photoreceptors.

Purinergic plasticity within petrosal neurons in hypertension.

The carotid bodies are peripheral chemoreceptors and contribute to the homeostatic maintenance of arterial levels of O2, CO2, and [H+]. They have attracted much clinical interest recently because of the realization that aberrant signaling in these organs is associated with several pathologies including hypertension. Herein, we describe data suggesting that sympathetic overactivity in neurogenic hypertension is, at least in part, dependent on carotid body tonicity and hyperreflexia that is related to changes in the electrophysiological properties of chemoreceptive petrosal neurons. We present results showing critical roles for both ATP levels in the carotid bodies and expression of P2X3 receptors in petrosal chemoreceptive, but not baroreceptive, terminals in the etiology of carotid body tonicity and hyperreflexia. We discuss mechanisms that may underlie the changes in electrophysiological properties and P2X3 receptor expression in chemoreceptive petrosal neurons, as well as factors affecting ATP release by cells within the carotid bodies. Our findings support the notion of targeting the carotid bodies to reduce sympathetic outflow and arterial pressure, emphasizing the potential clinical importance of modulating purinergic transmission to treat pathologies associated with carotid body dysfunction but, importantly, sparing physiological chemoreflex function.

Nerve growth factor-dependent hyperexcitability of capsaicin-sensitive bladder afferent neurones in mice with spinal cord injury.

What is the central question of this study? Nerve growth factor (NGF) is reportedly a mediator inducing urinary bladder dysfunction. Is NGF directly involved in hyperexcitability of capsaicin-sensitive C-fibre bladder afferent pathways after spinal cord injury (SCI)? What is the main finding and its importance? Neutralization of NGF by anti-NGF antibody treatment reversed the SCI-induced increase in the number of action potentials and the reduction in spike thresholds and A-type K+ current density in mouse capsaicin-sensitive bladder afferent neurones. Thus, NGF plays an important and direct role in hyperexcitability of capsaicin-sensitive C-fibre bladder afferent neurones attributable to the reduction in A-type K+ channel activity in SCI. Nerve growth factor (NGF) has been implicated as an important mediator in the induction of C-fibre bladder afferent hyperexcitability, which contributes to the emergence of neurogenic lower urinary tract dysfunction after spinal cord injury (SCI). In this study, we determined whether NGF immunoneutralization using an anti-NGF antibody (NGF-Ab) normalizes the SCI-induced changes in electrophysiological properties of capsaicin-sensitive C-fibre bladder afferent neurones in female C57BL/6 mice. The spinal cord was transected at the Th8/Th9 level. Two weeks later, continuous administration of NGF-Ab (10μgkg-1 h-1 , s.c. for 2weeks) was started. Bladder afferent neurones were labelled with Fast-Blue (FB), a fluorescent retrograde tracer, injected into the bladder wall 3weeks after SCI. Four weeks after SCI, freshly dissociated L6-S1 dorsal root ganglion neurones were prepared. Whole-cell patch-clamp recordings were then performed in FB-labelled neurones. After recording action potentials or voltage-gated K+ currents, the sensitivity of each neurone to capsaicin was evaluated. In capsaicin-sensitive FB-labelled neurones, SCI significantly reduced the spike threshold and increased the number of action potentials during membrane depolarization for 800ms. These SCI-induced changes were reversed by NGF-Ab. Densities of slow-decaying A-type K+ (KA ) and sustained delayed rectifier-type K+ currents were significantly reduced by SCI. The NGF-Ab treatment reversed the SCI-induced reduction in the KA current density. These results indicate that NGF plays an important role in hyperexcitability of mouse capsaicin-sensitive C-fibre bladder afferent neurones attributable to a reduction in KA channel activity. Thus, NGF-targeting therapies could be effective for treatment of afferent hyperexcitability and neurogenic lower urinary tract dysfunction after SCI.

PD19-10 P38 MITOGEN-ACTIVATED PROTEIN KINASE INHIBITOR REDUCES HYPEREXCITABILITY OF CAPSAICIN SENSITIVE BLADDER AFFERENT NEURONS IN MICE WITH SPINAL CORD INJURY

You have accessJournal of UrologyUrodynamics/Lower Urinary Tract Dysfunction/Female Pelvic Medicine: Basic Research & Pathophysiology I1 Apr 2018PD19-10 P38 MITOGEN-ACTIVATED PROTEIN KINASE INHIBITOR REDUCES HYPEREXCITABILITY OF CAPSAICIN SENSITIVE BLADDER AFFERENT NEURONS IN MICE WITH SPINAL CORD INJURY Takahisa Suzuki, Takahiro Shimizu, Nobutaka Shimizu, Jianshu Ni, Shinsuke Mizoguchi, Eiichiro Takaoka, Hideaki Miyake, Anthony J Kanai, and Naoki Yoshimura Takahisa SuzukiTakahisa Suzuki More articles by this author , Takahiro ShimizuTakahiro Shimizu More articles by this author , Nobutaka ShimizuNobutaka Shimizu More articles by this author , Jianshu NiJianshu Ni More articles by this author , Shinsuke MizoguchiShinsuke Mizoguchi More articles by this author , Eiichiro TakaokaEiichiro Takaoka More articles by this author , Hideaki MiyakeHideaki Miyake More articles by this author , Anthony J KanaiAnthony J Kanai More articles by this author , and Naoki YoshimuraNaoki Yoshimura More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2018.02.1000AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Nerve growth factor (NGF) has been implicated as an important mediator to induce C-fiber bladder afferent hyperexcitability, which contributes to the emergence of neurogenic detrusor overactivity following spinal cord injury (SCI). It is also known that the second messenger signaling pathways activated by NGF utilize p38 Mitogen-Activated Protein Kinase (MAPK). We examined whether p38 MAPK inhibition using SB203580 (p38 MAPK inhibitor) normalizes the SCI-induced changes in electrophysiological properties of capsaicin-sensitive C-fiber bladder afferent neurons in the mouse SCI model. METHODS The spinal cord was transected at the Th8/9 level in 8-9 weeks old female C57BL/6 mice. Two weeks later, SB203580 (1 mg/ml) was injected intrathecally using an osmotic pump at infusion rate of 0.51 µl/hr for 2 weeks. Instead of SB203580, artificial cerebrospinal fluid was injected in separate groups of spinal intact (SI) and SCI mice as controls. Bladder afferent neurons were labeled with axonal transport of Fast Blue (FB), a fluorescent retrograde tracer, injected into the bladder wall 3 weeks after SCI. Dissociated L6 and S1 dorsal root ganglion neurons were prepared 4 weeks after SCI. Whole cell patch clamp recordings were performed in FB-labeled bladder afferent neurons, and the data were compared among control SI, SCI and SCI treated with p38 MAPK inhibitor mice. After recording action potentials (AP) or voltage-gated K+ (Kv) currents, the sensitivity of each neuron to capsaicin was evaluated. RESULTS In capsaicin-sensitive bladder afferent neurons, the threshold for eliciting AP was significantly reduced in control SCI compared to SI mice. Also, SCI increased the number of AP during 800 ms membrane depolarization. These SCI induced hyperexcitability was reversed by p38 MAPK inhibitor. Densities of slow decaying KA and sustained KDR currents evoked by depolarization to 0 mV were significantly reduced after SCI. The treatment with a p38 MAPK inhibitor reversed the SCI-induced reduction in the KA current density, but not in the KDR. CONCLUSIONS In SCI mice, p38 MAPK signaling pathways play an important role in hyperexcitability of capsaicin sensitive C-fiber bladder afferent neurons due to KA current reduction. Thus, p38 MAPK-targeting therapies could be effective for treatment of afferent hyperexcitability in SCI. © 2018FiguresReferencesRelatedDetails Volume 199Issue 4SApril 2018Page: e399-e400 Advertisement Copyright & Permissions© 2018MetricsAuthor Information Takahisa Suzuki More articles by this author Takahiro Shimizu More articles by this author Nobutaka Shimizu More articles by this author Jianshu Ni More articles by this author Shinsuke Mizoguchi More articles by this author Eiichiro Takaoka More articles by this author Hideaki Miyake More articles by this author Anthony J Kanai More articles by this author Naoki Yoshimura More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...