Electrical Field Stimulation Research Articles

Improvement in Persistent Post-Concussive Symptoms After Treatment with Neuromodulation: A Case Study

AbstractUp to 30% of youth with concussion experience persistent post-concussive symptoms (PPCS) impacting cognitive, social, mental, and physical health. Limited evidence-based therapies exist to treat PPCS. We present a case of an adolescent female with multiple sports-related concussions with persistent symptoms, including headache, nausea, dizziness, brain fog, and anxiety. After 4 weeks of treatment with an auricular percutaneous electrical nerve field stimulation (PENFS) device, she had near-complete resolution of her symptoms, allowing a return to her previous level of academic achievement, as well as physical activity. Emerging evidence suggests that various forms of neuromodulation, such as repetitive transcranial magnetic stimulation (rTMS) and transcranial direct current stimulation (tDCS), can benefit patients with post-concussive symptoms. This adolescent female’s improvement with PENFS contributes to this expanding field.

Conditional deletion of IP3R1 by Islet1-Cre in mice reveals a critical role of IP3R1 in interstitial cells of Cajal in regulating GI motility.

Inositol 1,4,5-trisphosphate receptor type 1 (IP3R1) has been proposed to play a physiological role in regulating gastrointestinal (GI) motility, but the underlying cell-dependent mechanism remains unclear. Here, we utilized cell-specific IP3R1 deletion strategies to address this question in mice. Conditional IP3R1 knockout mice using Wnt1-Cre, Islet1-Cre mice, and smMHC-CreEGFP were generated. Cell lineage tracing was performed to determine where gene deletion occurred in the GI tract. Whole-gut transit assay and isometric tension recording were used to assess GI function in vivo and in vitro. In the mouse GI tract, Islet1-Cre targeted smooth muscle cells (SMCs) and interstitial cells of Cajal (ICCs), but not enteric neurons. IP3R1 deletion by Islet1-Cre (isR1KO) caused a phenotype of intestinal pseudo-obstruction (IPO), evidenced by prolonged whole-gut transit time, enlarged GI tract, abdominal distention, and early lethality. IP3R1 deletion by Islet1-Cre not only reduced the frequency of spontaneous contractions but also decreased the contractile responses to the muscarinic agonist carbachol (CCh) and electrical field stimulation (EFS) in colonic circular muscles. By contrast, smMHC-CreEGFP only targeted SMCs in the mouse GI tract. Although IP3R1 deletion by smMHC-CreEGFP (smR1KO)also reduced the contractile responses to CCh and EFS in colonic circular muscles, the frequency of spontaneous contractions was less affected, and neither global GI abnormalities nor early lethality was found in smR1KO mice. IP3R1 deletion in both ICCs and SMCs but not in SMCs alone causes an IPO phenotype, suggesting that IP3R1 in ICCs plays an essential role in regulating GI motility in vivo.

Effect of Atomoxetine on Mouse Isolated Vas Deferens Contractility

Objective: Atomoxetine (ATX), a selective noradrenaline re-uptake inhibitor, is a preferred drug with sufficient efficacy and favorable safety profile for the treatment of attention-deficit hyperactivity disorder. Ejaculatory dysfunctions have been reported in the patients receiving ATX as sexual side effect, of which underlying mechanisms are largely unknown. The present study aimed to investigate the effect of ATX on mouse isolated vas deferens (VD) contractility as a potential mechanism of ATX-induced ejaculatory dysfunction. Material and Methods: Isolated organ bath studies were performed on prostatic parts of VD obtained from adult male Balb/c mice. The effect of ATX (10-6, 10-5, 3x10-5 and 10-4 M) on KCl (80 mM), phenylephrine (PhE, 3x10-4 M), adenosine 5’-triphosphate (ATP, 10-2 M) and electrical field stimulation (EFS; 100 V, 64 Hz)-induced contractions of VD strips were evaluated in concentration dependent manner. Results: ATX at 10-6 and 10-5 did not alter the contractile responses (p>0.05), however, higher concentrations of ATX (3x10-5 or 10-4 M) significantly inhibited the KCl, PhE, ATP and EFS-induced contractions of VD strips (p<0.05). Conclusion: The present study demonstrated for the first time that ATX decreased the contractile responses of mouse isolated VD concentration-dependently. Our results suggest that ejaculatory dysfunction might be related to the inhibitory effect on ATX on VD.

Semaglutide improves contractile function in isolated human atrium

Abstract Background Recently, the results of the STEP-HFpEF trial were announced, in which high-dose treatment with the antidiabetic glucagon-like-peptide 1 agonist semaglutide resulted in a significant improvement of HFpEF-associated symptoms and reduction of NT-proBNP levels. Interestingly, treatment effects were independent of initial body weight and left-ventricular ejection fraction, sparking interest regarding the precise mechanism of action. Purpose We tested the effects of acute semaglutide treatment in isolated human right-atrial trabeculae. Methods Trabeculae were prepared from right atrial appendage biopsies from 32 patients undergoing elective coronary artery bypass grafting and/or valve surgery. Regular contractions were elicited by electrical field stimulation (1Hz, 5V/50ms, 37°C). Paired experiments were performed simultaneously per patient with a control group and one exposed to semaglutide (100 or 300nM). Developed tension was analysed at steady state after 30 min. Arrhythmias were defined as deviations from baseline not arising from electrical stimulation. L-type calcium channel blockade was achieved with nifedipine (2µM) in some experiments. Rapid cooling contraction was elicited to assess sarcoplasmic reticulum Ca2+ content (RCC1) by rapidly superfusing the trabeculae with chilled solution (2°C). After a short rewarming period of 10s to 37°C, RCC was repeated to assess the Na+/Ca2+ exchanger function (RCC2). Statistical analysis was conducted using paired t-test or Wilcoxon test as appropriate (for 2 groups) and one-way ANOVA with test for trend (for >2 groups). Results Exposure to semaglutide increased developed tension by over 3-fold in human atrial trabeculae in a dose-dependent manner: from 3.14±1.14 for control to 5.43±2.35mN/mm2 for 100nM semaglutide (p=0.03 vs. control) to 12.5±3.53 for 300nM semaglutide (p=0.004 vs. control, mean±SEM, fig. A-D, p=0.005 for linear trend). L-type calcium channel blockade with nifedipine blunted developed tension overall but did not abolish the effect of semaglutide (fig. E, p=0.02). In contrast, the developed tensions upon RCC1 and RCC2 were increased by semaglutide (300nM) compared to control, while the ratio of RCC2:RCC1 was elevated by semaglutide, indicating both increased SR Ca2+ reuptake and less external Ca2+ removal in the presence of semaglutide (fig. H-J). Relaxation time to 50% of baseline was unaltered (not shown), while relaxation time to 90% of baseline was mildly prolonged by semaglutide at 300 nM (fig. F). No increase in the rate of arrhythmias was observed by semaglutide treatment (fig. G). Conclusion Acute exposure of human atrial trabeculae to semaglutide results in a strong positive inotropic effect in a dose-dependent manner without increasing the propensity for arrhythmias. This effect is most likely due to increased SR Ca2+ reuptake. Treatment of heart failure patients with high-dose semaglutide can improve atrial function, which may ameliorate symptoms.

Circadian rhythm disruption modulates enteric neural precursor cells differentiation leading to gastrointestinal motility dysfunction via the NR1D1/NF-κB axis

ObjectivesCircadian rhythm disruption (CRD) is implicated with numerous gastrointestinal motility diseases, with the enteric nervous system (ENS) taking main responsibility for the coordination of gastrointestinal motility. The purpose of this study is to explore the role of circadian rhythms in ENS remodeling and to further elucidate the underlying mechanisms.MethodsFirst, we established a jet-lagged mice model by advancing the light/dark phase shift by six hours every three days for eight weeks. Subsequent changes in gastrointestinal motility and the ENS were then assessed. Additionally, a triple-transgenic mouse strain (Nestin-creERT2 × Ngfr-DreERT2: DTRGFP) was utilized to track the effects of CRD on the differentiation of enteric neural precursor cells (ENPCs). RNA sequencing was also performed to elucidate the underlying mechanism.ResultsCompared to the control group, CRD significantly accelerated gastrointestinal motility, evidenced by faster intestinal peristalsis (P < 0.01), increased fecal output (P < 0.01), and elevated fecal water content (P < 0.05), as well as enhanced electrical field stimulation induced contractions (P < 0.05). These effects were associated with an increase in the number of glial cells and nitrergic neurons in the colonic myenteric plexus. Additionally, ENPCs in the colon showed a heightened differentiation into glial cells and nitrergic neurons. Notably, the NR1D1/nuclear factor-kappaB (NF-κB) axis played a crucial role in the CRD-mediated changes in ENPCs differentiation. Supplementation with NR1D1 agonist or NF-κB antagonist was able to restore gastrointestinal motility and normalize the ENS in jet-lagged mice.ConclusionsCRD regulates the differentiation of ENPCs through the NR1D1/NF-κB axis, resulting in dysfunction of the ENS and impaired gastrointestinal motility in mice.

Advancing cardiac 3D organoid-on-chip: integrating iPSC-derived MELAS syndrome models to study electromechanical synchronicity and cardiac memory using contactless technology

Abstract Background Mitochondrial DNA mutations affect 1:5000 newborns, posing life-threatening risks, especially to high-energy organs such as the brain, heart, and muscles. Mortality rates can reach 71% when cardiomyopathy is present. MELAS syndrome (Mitochondrial Encephalopathy with Lactic Acidosis and Stroke-like Episodes), primarily affecting protein synthesis (MTTL1 gene, tRNALeu(UUR)), disrupts the electron transport chain complex I and is associated with cardiac conduction defects and hypertrophy. Manifesting symptoms typically appear between ages 3 and adulthood, presenting a complex clinical scenario. Human induced pluripotent stem cells (hiPSCs) offer a unique platform for studying cardiac manifestations of this disease when differentiated into cardiomyocytes. Purpose We adopted a highly adaptable in-vitro model, organized in 3D cardioids, to explore potential impairments in electromechanical coupling of cells. Methods We utilized hiPSCs carrying the MELAS mutation and their respective isogenic controls differentiated into cardiomyocytes, cultured in 3D cardioids. Electromechanical coupling was assessed using laser-poration technology combined with video kinematics evaluation under various stimulation patterns. Results Morphological analysis revealed a statistically significant reduction in the diameter of MELAS cardioids compared to the isogenic 3D structures (368±17.15 μm vs. 527±36.76 μm). In a spontaneous regimen, MELAS samples exhibited higher beating frequency (0.88±0.28 Hz vs. 0.17±0.05 Hz) and contractile speed (99.35±39.11 μm/s vs. 33.46±5.57 μm/s). Additionally, the response to different electric field stimulation protocols varied significantly, with the isogenic cell line showing better adaptation to sinusoidal stimuli than the pathological one, characterized by rectangular monophasic stimuli. Laser-poration technology, combined with MEA chips and video kinematic evaluation, revealed modulation of electromechanical coupling, affecting action potential upstroke and duration, reducing contraction velocity, and altering electromechanical delay. Furthermore, MELAS hiPSCs and primary cardiomyocytes exhibited reduced protein levels of various OXPHOS subunits (SDHB, ATPB, and COX IV) compared to isogenic controls. The mutated cell line also showed increased ROS production in non-stimulated samples compared to stimulated ones (1.3-fold vs. 1.2-fold), with this difference magnifying after 24 hours from stimulation (1.8-fold vs. 1.3-fold). Conclusions This study characterizes MELAS hiPSCs differentiated into cardiac organoids, offering insight into the cardiac aspect of this complex syndrome. Our findings deepen understanding of the syndrome's cardiac impact and response to extracellular stimuli, potentially benefiting affected tissues. This research defines critical physiological and kinematic parameters for MELAS, paving the way for future investigations.

Interaction between fluticasone furoate and umeclidinium in passively sensitized isolated human airways

Asthma management often includes inhaled corticosteroids (ICSs), with additional controllers like long-acting muscarinic antagonists (LAMAs) for severe cases. The primary goal of this study was to investigate the pharmacological interaction between various concentrations of fluticasone furoate (FF) and umeclidinium (UME) in isolated human airways to determine the nature of their interaction, whether synergistic or additive. Medium bronchi and small airways obtained from patients undergoing lobectomy were passively sensitized to mimic asthmatic conditions. The effects of FF and UME, alone and in combination, on airway relaxation were evaluated using histamine-induced contraction and electrical field stimulation. Pharmacological interactions were analyzed using the Bliss Independence theory. Results indicated that FF induced a partial, concentration-dependent relaxation of sensitized airways, while UME induced a larger relaxation in medium bronchi but a weaker effect in small airways. The combination of FF and UME resulted in significantly greater relaxation than either drug alone, demonstrating synergism at high concentrations in medium bronchi but only additive effects in small airways. This study suggests that higher doses of FF might be necessary in a fixed dose combination to achieve optimal synergistic bronchodilation with UME. Future research should focus on clinical trials to confirm these findings and explore the molecular mechanisms underlying these interactions, potentially improving personalized asthma therapy.

Perivascular Adipose Tissue Becomes Pro-Contractile and Remodels in an IL10-/- Colitis Model of Inflammatory Bowel Disease.

Inflammatory Bowel Diseases (IBDs) are associated with aberrant immune function, widespread inflammation, and altered intestinal blood flow. Perivascular adipose tissue (PVAT) surrounding the mesenteric vasculature can modulate vascular function and control the local immune cell population, but its structure and function have never been investigated in IBD. We used an IL10-/- mouse model of colitis that shares features with human IBD to test the hypothesis that IBD is associated with (1) impaired ability of PVAT to dilate mesenteric arteries and (2) changes in PVAT resident adipocyte and immune cell populations. Pressure myography and electrical field stimulation of isolated mesenteric arteries show that PVAT not only loses its anti-contractile effect but becomes pro-contractile in IBD. Quantitative immunohistochemistry and confocal imaging studies found significant adipocyte hyperplasia and increased PVAT leukocytes, particularly macrophages, in IBD. PCR arrays suggest that these changes occur alongside the altered cytokine and chemokine gene expression associated with altered NF-κB signaling. Collectively, these results show that the accumulation of macrophages in PVAT during IBD pathogenesis may lead to local inflammation, which ultimately contributes to increased arterial constriction and decreased intestinal blood flow with IBD.

Microscopic mechanistic study of the penetration distributions for plasma reactive oxygen and nitrogen species based on sialic acid targeting on the cell membrane surface

The ability of cold atmospheric plasmas (CAPs) to produce a wide range of active constituents while maintaining a low or even room temperature of the gas has made it a novel research area of great interest. During plasma action, cancer cell membrane surface components are susceptible to oxidative modification by reactive oxygen and nitrogen species (RONS). In this study, the process of oxidative modification of membrane surface components sialic acid by RONS was investigated based on molecular dynamics simulations, and the penetration mechanism of long-lived particles ONOOH and its homolytic products at the membrane-water interface and the effect of appropriate electric field action were studied. The results showed that cancer cells with high sialic acid expression were less stable than healthy cells. Plasma treatment may promote the ONOOH homolysis process, and its homolysis product OH free radical is more likely to adsorb near sialic acid molecules by hydrogen bonding, resulting in oxidative modification. The interaction force between OH free radical and sialic acid molecules is stronger than ONOOH, which helps to further understand the oxidative modification reaction in membrane environment. At the same time, appropriate electric field stimulation can enhance the depth of penetration of RONS to more effectively treat the pathological state of biological tissues. The study proposes the use of membrane surface sialic acid as a cancer therapeutic target and provides guidance for improving the depth of RONS penetration and maximizing the survival of healthy cells, which contributes to the further clinical translation of plasma biomedicine.

Neurogenic-derived 6-nitrodopamine is the most potent endogenous modulator of the mouse urinary bladder relaxation

6-Nitrodopamine (6-ND) modulates vas deferens, seminal vesicles, and corpus cavernosum contractility; however, its role on the lower urinary tract organs has not been evaluated. Investigations of isolated urinary bladders from wild-type (WT) mice revealed 6-ND release was comparable to that of dopamine and adrenaline, whereas noradrenaline was hardly detected, as assessed by liquid chromatography coupled to tandem mass spectrometry. In vitro, 6-ND induced concentration-dependent relaxations in carbachol pre-contracted bladders with high potency (pEC50: 8.04 ± 0.86), independently of eNOS/sGC activity. Co-incubation of 6-ND (1–10 μM) antagonizes the contractile effects of acetylcholine (p < 0.05). Experiments using nitric oxide synthase (NOS) knockout mice demonstrated that 6-ND release from isolated urinary bladder was significantly reduced by neuronal NOS (nNOS−/−) deletion and abolished by triple NOSs deletion (n/i/eNOS−/−), while no significant changes were observed in endothelial (eNOS−/−) or inducible (iNOS−/−) knockout mice. Incubation with tetrodotoxin resulted in a significant decrease in 6-ND release in bladders obtained from WT, but not in nNOS−/− mice. The bladders from nNOS−/− and n/i/eNOS−/− mice exhibited significantly higher contractile responses to electric field stimulation (EFS), compared to eNOS−/−, iNOS−/−, or WT bladders. The hyperreactivity observed in triple NOS knockouts was reversed by the incubation with bladder mucosal layer obtained from a donor WT mice, but not with the muscular layer. These findings clearly demonstrate 6-ND is the most potent endogenous relaxing agent of urinary bladder, and inhibition of its release is associated with bladder hyperreactivity.

Impact of Pulse Electric Field Stimulation on Negative Air Ion Release Capacity of Snake Plants

To investigate the effects of pulse electric field stimulation on the photosynthetic electron transport chain and negative air ion (NAI) release capacity of snake plants, the chlorophyll content, fluorescence induction kinetics curve (OJIP curve), chlorophyll fluorescence parameters, and NAI release concentration of snake plants kept under identical greenhouse conditions under different pulse electric field stimulations were compared and analyzed. The experimental results show that (1) after pulse electric field stimulation, the chlorophyll content in treatment group T1 (5 kv) and T2 (7 kv) of snake plants increased by 6.30% and 6.70%, respectively, with significant differences observed between the two treatment groups and the control group (CK). (2) In both treatment groups, the OJIP curve exhibited higher values for the inflection point (I) and peak (P) compared to the origin (O) and inflection point (J) values, with the rising trend in the I–P segment being more gentle than that of the O–J segment. Additionally, the J band was above 0, with the peak value in the T2 group being higher than that in the T1 group. (3) The chlorophyll fluorescence parameters showed fluctuating variations. Specifically, Fm, TRo/CSo, ETo/CSo, and DIo/CSo showed ascending trends in the treatment groups. Fv/Fo, Sm, and ABS/RC exhibited descending trends; Fv/Fm, Vj, ETo/RC, and φEo showed relatively minor changes. The PIabs displayed a decreasing trend. The PItotal in the CK was greater than that in the T1 and T2 groups. (4) After 4 h of pulse electric field stimulation, the NAI concentration increased by 87.60% in the T1 group and by 62.09% in the T2 group, compared to the same measurement taken at 3 h. Pulse electric field impacts the photosynthetic electron transport chain of snake plants, thereby influencing their NAI release capacity. This study aims to elucidate the physiological responses of the chloroplasts in snake plants to pulsed electric field stimulation and to lay the foundation for enhancing the plant’s release of negative air ion concentrations through physical and technological means.

Tetrabenazine, a vesicular monoamine transporter 2 inhibitor, inhibits vesicular storage capacity and release of monoamine transmitters in mouse brain tissue

AbstractBackground and PurposeTetrabenazine (TBZ), used for treating hyperkinetic disorders, inhibits vesicular monoamine transporter‐2 (VMAT‐2), which sequesters monoamines into vesicles for exocytosis. However, our knowledge of the effect of TBZ on monoaminergic transmission is limited. Herein, we provide neurochemical evidence regarding the effect of VMAT‐2 inhibition on vesicular neurotransmitter release from the prefrontal cortex (PFC) and striatum (STR) (brain regions involved in characteristic TBZ treatment side effects). The interaction between TBZ and MDMA was also assessed regarding motor behaviour in mice.Experimental ApproachVesicular storage capacity and release of [3H]‐noradrenaline ([3H]‐NA), [3H]‐dopamine ([3H]‐DA), [3H]‐serotonin ([3H]‐5‐HT), and [3H]‐acetylcholine ([3H]‐ACh) was studied in mouse PFC and STR ex vivo slice preparations using electrical field stimulation. Additionally, locomotor activity was assessed in vehicle‐treated mice and compared with that of MDMA, TBZ, and co‐administered animals (n = 6) using the LABORAS system.Key ResultsTBZ lowered the storage capacity and inhibited the vesicular release of [3H]‐NA and [3H]‐DA from the PFC, and [3H]‐DA and [3H]‐5‐HT from the STR in a concentration‐dependent manner. Unlike vesamicol (vesicular ACh uptake inhibitor), TBZ failed to inhibit the vesicular release of [3H]‐ACh from the PFC. When the vesicular storage of the investigated monoamines was inhibited by TBZ in the PFC and STR, MDMA induced the release of transmitters through transporter reversal; MDMA dose dependently increased locomotor activity in vivo.Conclusion and ImplicationsOur observations provide neurochemical evidence explaining the mechanism of VMAT‐2 inhibitors in the brain and support the involvement of dopaminergic and noradrenergic transmission in hyperkinetic movement disorders.

Functional-hydrogel-based electronic-skin patch for accelerated healing and monitoring of skin wounds

Conductive hydrogels feature reasonable electrical performance as well as tissue-like mechanical softness, thus positioning them as promising material candidates for soft bio-integrated electronics. Despite recent advances in materials and their processing technologies, however, facile patterning and monolithic integration of functional hydrogels (e.g., conductive, low-impedance, adhesive, and insulative hydrogels) for all-hydrogel-based soft bioelectronics still poses significant challenges. Here, we report material design, fabrication, and integration strategies for an electronic-skin (e-skin) patch based on functional hydrogels. The e-skin patch was fabricated by using photolithography-compatible functional hydrogels, such as poly(2-hydroxyethyl acrylate) (PHEA) hydrogel (substrate), Ag flake hydrogel (interconnection; conductivity: ∼571.43 S/cm), poly(3,4-ethylenedioxythiophene:polystyrene) (PEDOT:PSS) hydrogel (working electrode; impedance: ∼69.84 Ω @ 1 Hz), polydopamine (PDA) hydrogel (tissue adhesive; shear strength: ∼725.1 kPa), and poly(vinyl alcohol) (PVA) hydrogel (encapsulation). The properties of these functional hydrogels closely resemble those of human tissues in terms of water content and Young's modulus, enabling stable tissue-device interfacing in dynamically changing physiological environments. We demonstrated the efficacy of the e-skin patch through its application to accelerated healing and monitoring of skin wounds in mouse models − efficient fibroblast migration, proliferation, and differentiation promoted by electric field (EF) stimulation and iontophoretic drug delivery, and monitoring of the accelerated healing process through impedance mapping. The all-hydrogel-based e-skin patch is expected to create new opportunities for various clinically-relevant tissue interfacing applications.

Effects of Cold on the Adrenoreactivity of the Rat Superior Mesenteric Artery

In experiments on segments of the rat superior mesenteric artery (SMA) under isometric conditions, the effect of cold on the reactivity of this artery, estimated by the magnitude of the contractile reaction of its segments to noradrenaline (NA) at a concentration of 0.01–10.0 µM, as well as to electrical field stimulation (EFS) of perivascular nerves with a frequency of 3, 10 and 40 Hz in the absence and presence of NA in the medium and at a temperature of 37 oC or 25 oC was studied. Cooling was found not to change the tone of the SMA caused by NA at all the concentrations used. In the absence and presence of NA at a low concentration (0.01–0.10 µM), cooling leads to a significant decrease in the neurogenic response of the SMA at all frequencies of EFS, while in the presence of NA at a high concentration (1–10 µM), cooling has no statistically significant effect on this response. In the presence of NA at a low concentration, cooling at low EFS frequencies insignificantly, and at high frequency significantly reduces the potentiating effect of NA on neurogenic contraction of SMA. NA at a high concentration under normal temperature conditions, reduces the neurogenic contraction at all EFS frequencies, while under cooling conditions at low frequencies it has a potentiating effect, and at high EFS frequency it has a depressing effect. The results obtained show that the depressing effect of cold on the neurogenic constriction of the rat SMA, observed at low and disappearing at high concentrations of NA, as well as on the NA-evoked potentiation of the neurogenic vasoconstriction, is not associated with a decrease in the contractile effect of NA, which is the main neurotransmitter in this artery. These effects can contribute to the redistribution of blood from the surface deep into the body, thus reducing heat loss and improve thermoregulation.

Abstract P221: Yoda2, a Novel Piezo1 Agonist, Induces Relaxation in Mouse Vascular and Corpus Cavernosum Tissues

Introduction: Piezo1, a mechanosensitive nonselective cation channel, is expressed in endothelial and vascular smooth muscle cells. It plays a critical role in sensing endothelial shear stress, generating nitric oxide (NO), regulating vascular tone, promoting angiogenesis, and controlling blood pressure. Yoda1, a tool compound for Piezo1, has limitations due to its low potency and poor water solubility. To overcome these limitations, a Yoda1 analogue, Yoda2, was developed. Yoda2 features a 4-benzoic acid moiety replacing the pyrazine of Yoda1, thereby enhancing its agonist properties and solubility. We hypothesize that Yoda2 induces relaxation in mouse mesenteric resistance artery (MRA), pudendal artery (PA), and corpus cavernosum (CC), through a NO-dependent mechanism. Methods: Male C57BL/6 mice were euthanized, and their MRA, PA, and CC were isolated and mounted in DMT wire myographs for isometric force measurements. Concentration-response curves to Yoda2 were obtained. Additionally, the contraction of CC induced by electrical field stimulation (EFS) was evaluated in the absence or presence of Yoda2. Data are presented as mean ± S.E.M of 3-4 mice. Non-linear regression curve was performed, and the maximum response (E max ) and pharmacological potency (pEC 50 ) were analyzed using Student's t-test (p&lt;0.05). Results: Yoda2 induced concentration-dependent relaxation in the MRA, PA, and CC, with E max values of 98.31 ± 0.89% for MRA, 95.01 ± 3.75% for PA, and 75.03 ± 15.17% for CC. The role of NO in this pathway was confirmed by the attenuation of Yoda2 effects upon inhibition of NO synthase (NOS) with L-NAME in MRA and PA. Furthermore, Yoda2 (1 µM) decreased the concentration response curves for phenylephrine-induced contraction in PA and MRA. On the other hand, Yoda2 (30 µM) did not change the EFS-induced contraction in CC. Conclusion: This study is the first to demonstrate that Yoda2, a potent and water-soluble Piezo1 channel agonist, induces significant relaxation in the MRA, PA, and CC of mice. Consistent with the effects of Yoda1, the effects of Yoda2 are mediated through NO production.

Abstract P351: Obesity in Early Life Leads to Endothelial and Erectile Dysfunction in Young Adult Rats.

Background: Erectile dysfunction (ED) is largely associated with obesity, however, the consequences of early life obesity for ED in young adulthood are unknown. We hypothesize that obesity induced by preweaning overfeeding impairs vascular and erectile function and affects blood pressure in young adult male rats (5-month-old). Methods: At postnatal day (PND) 1, Wistar rats were divided into a normal litter (NL, 5 female + 5 male pups/dam) or small litter (SL, 2 male + 1 female pup/dam) during lactation. After weaning, SL pups had increased body weight. Next, rats from both groups returned to normal size colonies (4-5 rats per cage) and were fed standard chow. At PND160, we evaluated body weight and fat pad deposition, mean arterial pressure (MAP) by tail-cuff, and pudendal artery (PA) and corpus cavernosum (CC) reactivity. Concentration-response curves to phenylephrine (PE) or acetylcholine (ACh) were performed in both preparations. In the CC, contraction and relaxation to electrical field stimulation (EFS) were obtained. Concentration-response curves were analyzed using non-linear regression analysis. RNA sequencing (RNAseq) analyses were performed in both PA and CC. Data were analyzed by Student’s t-test and presented as mean ± S.E.M. Statistical significance was set at p&lt;0.05. Results: At PND160 no differences in body weight were observed in SL (594±14g) compared to NL (559±13g), however, there was an increase in fat deposition in SL (retroperitoneal fat: NL 7.47±0.69g vs SL 11.25±1.23g; perigonadal fat: NL 10.05±0.55g vs SL 13.24±1.16g). MAP was higher in SL (120.5±1.20 mmHg vs 114.9±0.58 NL). In the PA, ACh-induced relaxation was reduced in SL (Emax: 28±6%) vs NL (Emax: 68±5.5%), with no changes in the contraction to PE. In the CC, EFS-induced relaxation was significantly decreased in SL rats compared to NL rats (16 Hz: NL 3.06 ± 0.28% vs SL 2.39 ± 0.53%). RNAseq revealed that, in PA from SL rats, 289 and 224 genes were down and upregulated, respectively, while in the CC from SL rats, 115 genes were downregulated, and 110 genes were upregulated. Among them, hcn2 gene (which encodes HCN2, the hyperpolarization-activated cyclic nucleotide-gated potassium and sodium channel 2) was strongly downregulated in both PA and CC, and this may affect mechanotransduction. Conclusion: Our data shows that early-life obesity causes long-lasting vascular and cavernous damage that might be associated with endothelial and erectile dysfunction in young adulthood.

The impact of long-course chemoradiotherapy on the myenteric plexus, neuromuscular functions and responses to prokinetic drugs in the human rectum.

The long-term effects of chemoradiotherapy on human rectum are poorly understood. The aims were to investigate changes in inflammatory status, myenteric neuron numbers/phenotype, neuromuscular functions and prokinetic drug efficacy. Macroscopically normal proximal-to-mid rectum was obtained from 21 patients undergoing surgery for bowel cancer, 98days (range: 63-350) after concurrent capecitabine and pelvic radiotherapy, and 19 patients without chemoradiotherapy. Inflammatory status was measured by H&E, CD45 staining and qPCR. Myenteric neurons were examined by immunohistochemistry. Neuromuscular functions and drug efficacy were studied using exogenous agents and electrical field stimulation (EFS) to activate intrinsic nerves. Inflammation was not detected. Numbers of myenteric ganglia/neurons were unchanged (11.7±2.4 vs. 10.3±2.2 neurons/mm myenteric plexus with/without chemoradiotherapy) as were the numbers of cholinergic/nitrergic neurons. EFS stimulated cholinergic and nitrergic neurons so the contractile response of the muscle was the sum of both but dominated by cholinergic (causing contraction) or less often, nitrergic activity (relaxation), followed, after termination of EFS, by neuronally mediated contraction. Inhibition of nitric oxide synthase (by L-NAME 300μM) more clearly defined EFS-evoked contractions. The 5-HT4 agonist prucalopride 10μM and the cholinesterase inhibitor donepezil 1µM, respectively increased and greatly increased the composite contractile response to EFS (measured as 'area-under-the curve') and the contractions isolated by L-NAME (respectively, by 22±14% and 334±87%; n=11/8). After chemoradiotherapy, nitrergic-mediated muscle relaxations occurred more often during EFS (in 29.8±6.1% preparations vs. 12.6±5.1% without chemoradiotherapy, n=21/18). With L-NAME, the ability of prucalopride to facilitate EFS-evoked contraction was lost and that of donepezil approximately halved (contractions increased by 132±36%; n=8). Several months after chemoradiotherapy, the rectum was not inflamed and myenteric neuron numbers/phenotype unchanged. However, nitrergic activity was increased relative to cholinergic activity, and prokinetic-like drug activity was lost or greatly reduced. Thus, chemoradiotherapy causes long-term changes in neuromuscular functions and markedly reduces the efficacy of drugs for treating constipation.

Unpinning and elimination of spiral waves and turbulence through local optogenetical illumination.

Spiral waves in cardiac tissue have been identified as a significant factor leading to life-threatening arrhythmias and ventricular fibrillation. Consequently, understanding the mechanisms underlying the dynamics of such waves and exploring strategies for their elimination have garnered substantial interest and emerged as crucial research objectives. Spiral waves often become pinned (trapped) at anatomical obstacles in cardiac tissue, resulting in increased stability and posing challenges for their elimination. The unpinning of spiral waves can be achieved through the application of an external electric field and has been the subject of previous research. Recently, optogenetics has emerged as an alternative method to modulate electrical activity by illumination of cardiac tissue. In this paper, we employ mathematical modeling to investigate the potential of utilizing local illumination to unpin and eliminate spiral waves in cardiac tissue. We also extend this methodology to explore the effects of more complex turbulent excitation patterns. We conduct simulations using low-dimensional (Barkley) and ionic (Fenton-Karma) models of cardiac tissue, incorporating optogenetical channels. Our findings demonstrate that local suprathreshold illumination can successfully unpin spiral waves in 100% of cases. Notably, unlike unpinning by electrical field stimulation, this approach does not necessitate precise timing of stimulus application during a specific phase of rotation. Additionally, we demonstrate that periodic optogenetical stimulation can effectively eliminate both unpinned spiral waves and turbulence by moving them toward the boundary via an antitachycardia pacing mechanism.

Inducible deletion of endothelial cell Efnb2 delays capillary regeneration and attenuates myofibre reinnervation following myotoxin injury in mice.

Acute injury of skeletal muscle disrupts myofibres, microvessels and motor innervation. Myofibre regeneration is well characterized, however its relationship with the regeneration of microvessels and motor nerves is undefined. Endothelial cell (EC) ephrin-B2 (Efnb2) is required for angiogenesis during embryonic development and promotes neurovascular regeneration in the adult. We hypothesized that, following acute injury to skeletal muscle, loss of EC Efnb2 would impair microvascular regeneration and the recovery of neuromuscular junction (NMJ) integrity. Mice (aged 3-6months) were bred for EC-specific conditional knockout (CKO) of Efnb2 following tamoxifen injection with non-injected CKO mice as controls (CON). The gluteus maximus, tibialis anterior or extensor digitorum longus muscle was then injured with local injection of BaCl2. Intravascular staining with wheat germ agglutinin revealed diminished capillary area in the gluteus maximus of CKO vs. CON at 5days post-injury (dpi); both recovered to uninjured (0dpi) level by 10dpi. At 0dpi, tibialis anterior isometric force of CKO was less than CON. At 10dpi, isometric force was reduced by half in both groups. During intermittent contractions (75Hz, 330mss-1, 120s), isometric force fell during indirect (sciatic nerve) stimulation whereas force was maintained during direct (electrical field) stimulation of myofibres. Neuromuscular transmission failure correlated with perturbed presynaptic (terminal Schwann cells) and postsynaptic (nicotinic acetylcholine receptors) NMJ morphology in CKO. Resident satellite cell number on extensor digitorum longus myofibres did not differ between groups. Following acute injury of skeletal muscle, loss of Efnb2 in ECs delays capillary regeneration and attenuates recovery of NMJ structure and function. KEY POINTS: The relationship between microvascular regeneration and motor nerve regeneration following skeletal muscle injury is undefined. Expression of Efnb2 in endothelial cells (ECs) is essential to vascular development and promotes neurovascular regeneration in the adult. To test the hypothesis that EfnB2 in ECs is required for microvascular regeneration and myofibre reinnervation, we induced conditional knockout of Efnb2 in ECs of mice. Acute injury was then induced by BaCl2 injection into gluteus maximus, tibialis anterior or extensor digitorum longus (EDL) muscle. Capillary regeneration was reduced at 5days post-injury (dpi) in gluteus maximus of conditional knockout vs. controls; at 10dpi, neither differed from uninjured. Nerve stimulation revealed neuromuscular transmission failure in tibialis anterior with perturbed neuromuscular junction structure. Resident satellite cell number on EDL myofibres did not differ between groups. Conditional knockout of EC Efnb2 delays capillary regeneration and attenuates recovery of neuromuscular junction structure and function.

Enhanced Energy Storage Properties of Highly Polarized BMT-Based Thin Films through the Multiscale Structure Synergistic Regulation Strategy.

For solving the trade-off relationship of the polarization and breakdown electric field, ferroelectric films with high polarization are playing a critical role in energy storage capacitor applications, especially at moderate/low electric fields. In this work, we propose a multiscale structure (including defect, domain, and grain structures) synergetic optimization strategy to optimize the polarization behavior and energy storage performances of BiMg0.5Ti0.5O3 (BMT) ferroelectric films by introducing Sr0.7La0.2TiO3 (SLT) without compromising the breakdown strength. At a moderate electric field of 2917 kV/cm, a high discharge density of 72.2 J/cm3 has been achieved in 0.9BMT-0.1SLT films, together with good frequency, thermal, and cycle stabilities for energy storage. Importantly, the phase difference Δφ is utilized to quantitatively evaluate the polarization switching mobility of the ferroelectric domain/PNRs at an external electric field stimulation. This research demonstrates that a multiscale structure optimization strategy could effectively regulate the energy storage performance, and ecofriendly BMT-based materials are promising candidates for next-generation energy storage capacitors, especially at moderate/low electric fields.