bluuebunny/crossref_metadata_2025 huggingface dataset:
Acetycholine activates WNK kinase signaling in endothelial cells to stimulate vasodilation
Authors: Garrud Tessa , Mata-Daboin Alejandro , Collier Daniel , Jaggar Jonathan
Published: May 2023

The endothelium integrates numerous external signals to regulate vascular contractility. Vasodilators such as acetylcholine (ACh) bind to endothelial cell receptors and activate downstream signaling pathways which act on arterial smooth muscle cells to induce vasorelaxation. The physiological functions of anion channels, such as chloride Cl-channels, in endothelial cells is poorly understood. TMEM16A is a calcium (Ca2+)-activated Cl- channel expressed by endothelial cells. WNK kinase is a Cl--sensitive kinase present in endothelial cells that activates the downstream substrate kinases SPS-1 related proline/alanine rich kinase (SPAK) and oxidative-stress responsive kinase-1 (OSR1). The goal of this study was to examine the functional significance of TMEM16A channels and the WNK kinase pathway in endothelial cell-mediated vasodilation in resistance-size mesenteric arteries. Tamoxifen-inducible, endothelial cell-specific TMEM16A knockout (TMEM16A ecKO) mice and their controls (TMEM16Afl/fl) were used for experiments. ACh rapidly increased SPAK and OSR1 phosphorylation in TMEM16Afl/fl arteries and this effect was blocked by WNK463, a specific WNK kinase inhibitor. ACh did not increase SPAK or OSR1 phosphorylation, nor was there any effect of WNK463, in TMEM16A ecKO arteries. ACh increased the frequency and spatial spread of intracellular Ca2+ signals in endothelial cells of en face TMEM16Afl/fl arteries. The ACh-induced increase in Ca2+ signal frequency and spread in endothelial cells was inhibited by WNK 463, HC067047, a TRPV4 channel inhibitor, or TMEM16A knockout. WNK463, HC067047 or TMEM16A knockout also attenuated ACh-induced vasodilation in pressurized arteries. In contrast, WNK463 did not alter ACh-induced vasodilation in TMEM16A ecKO arteries. Taken together, our data suggest that ACh stimulates Ca2+ signaling in endothelial cells and vasodilation via WNK kinase-, TRPV4- and TMEM16A-dependent mechanism.

Funding from NIH R01HL155180, NIH R01HL158846 and AHA 830462.

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Postural hyperventilation in POTS is not driven by exaggerated peripheral chemoreceptor activity
Authors: Baker Jacquie , Incognito Anthony , Ranada Shaun , Phillips Aaron , Sheldon Robert , Wilson Richard , Raj Satish
Published: May 2023

BACKGROUND: Postural Orthostatic Tachycardia Syndrome (POTS) is a chronic form of autonomic nervous system dysfunction primarily affecting premenopausal females. POTS patients experience chronic orthostatic symptoms, and a heart rate (HR) increase of ≥30 bpm within 10 min of standing in the absence of orthostatic hypotension (>20/10 mmHg decrease in blood pressure [BP]). On standing, patients can exhibit increased ventilation (VE) with subsequent reductions in end-tidal CO2 (i.e., hypocapnia). Hypocapnia exacerbates tachycardia and symptoms. Previous research has suggested exaggerated peripheral chemoreceptor (pChR) activity drives postural hyperventilation in POTS. OBJECTIVE: We explored exaggerated pChR activity in the pathophysiology of postural hyperventilation in POTS. HYPOTHESIS: We hypothesized that POTS patients have a greater VE reduction during pChR inhibition compared to healthy controls. METHODS: We conducted a randomized crossover trial of hyperoxia vs. normoxia while supine and during head-up tilt (HUT) in 25 POTS patients (F=25; age: 31±8 years) and 12 healthy controls (HC) (F=12; age: 31±6 years; p=0.8). To inhibit pChR, participants breathed 5x30sec bouts of normocapnic hyperoxia (O2: ~400mmHg). Continuous respiratory (VE, ETO2, ETCO2) and hemodynamics (HR, BP, stroke volume, cardiac output, systemic vascular resistance) were measured. Data are reported as mean±SD. Continuous variables were compared using a 3-factor mixed-effects model: [Group: Control vs. Patient] x [Position (repeated measure): supine vs. HUT] x [Gas (repeated measure): ON vs. OFF). RESULTS During normoxic HUT, VE increased relative to supine baseline in both POTS (Supine: 10.3±2.3 L/min, HUT: 15.2±4.2; p<0.001) and HC (Supine: 11.1±2.1 L/min, HUT: 12.9±2.3 L/min; p=0.04). As predicted, the ΔVE in response to HUT was significantly greater in POTS (4.8±3.3 L/min) vs. HC (1.8±1.6 L/min; p=0.001). During HUT, hyperoxia reduced VE in both POTS (p<0.001) and HC (p<0.001). However, the magnitude of the ventilatory inhibition was not different between groups (POTS: -1.9±1.0 L/min; HC: -1.8±1.0 L/min; p=1.0). CONCLUSION: POTS patients have increased postural VE relative to HC. During pChR inhibition with hyperoxia, VE was reduced in both HC and POTS, and the magnitude of ventilatory inhibition in response to hyperoxia was the same in both groups. These findings suggest pChR activity is not exaggerated in POTS patients, and thus is not the primary driver of postural hyperventilation observed in POTS.

This work was supported by Standing up to POTS.

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Role of scleraxis in angiotensin II-induced cardiac fibrosis
Authors: Chattopadhyaya Sikta , Nagalingam Raghu Sundaresan , Ledingham D. Allison , Czubryt Michael P.
Published: May 2023

Cardiac fibrosis involves the activation and conversion of fibroblasts to myofibroblasts in response to pathophysiological stresses, resulting in structural and functional remodeling of the heart leading to heart failure and death. Although it affects millions of people worldwide, currently no medications are available against cardiac fibrosis. Similar to the pro-fibrotic growth factor TGF-β, angiotensin II (AngII) contributes to cardiac fibrosis, however AngII inhibitors are not used against fibrosis despite growing evidence of efficacy. AngII induces the nuclear translocation of Myocardin Related Transcription Factor-A (MRTF-A) which acts with Serum Response Factor (SRF) to play a major role in the conversion of fibroblasts to myofibroblasts, similar to the transcription factor scleraxis. Scleraxis has been shown by our lab to induce fibrosis by transactivating various pro-fibrotic gene promoters. Here we investigated whether AngII works through an MRTF-A/SRF/scleraxis pathway to induce cardiac fibrosis. AngII delivery (1mg/kg/day for 28 days) through osmotic mini-pumps in C57Bl/6 mice significantly increased blood pressure (systolic, diastolic, and mean) and cardiac hypertrophy compared to saline control group. In the AngII group, cardiac fibrosis was observed with enhanced mRNA expression of pro-fibrotic genes, Col1a1, Col3a1, periostin, and EDA-fibronectin. Interestingly there was a significant increase in scleraxis as well, suggesting its involvement in AngII-mediated fibrosis. Our in vitro studies of rat cardiac fibroblasts treated with AngII showed a significant increase in scleraxis mRNA and protein expression, and scleraxis knockdown significantly attenuated AngII-induced collagen expression, confirming a requirement for scleraxis in the AngII-mediated fibrosis pathway. NIH3T3 cells transfected with MRTF-A and SRF together showed a high level of induction of scleraxis expression. Luciferase assays in both COS7 and NIH3T3 cells showed that MRTF-A and SRF act together to regulate scleraxis expression by binding to and transactivating the scleraxis promoter, indicating that AngII activates a MRTF-A/SRF/scleraxis pathway for inducing cardiac fibrosis, therefore targeting scleraxis may be an effective anti-fibrotic strategy.

Canadian Institutes of Health Research Project Grant

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Brachial vascular function after acute aerobic exercise in older adults with and without type 2 Diabetes mellitus
Authors: Marôco João , Arrais Inês , Silvestre Tiago , Pinto Marco , Laranjo Sérgio , Magalhães João , Santa-Clara Helena , Fernhall Bo , Melo Xavier
Published: May 2023

Brachial vascular function after acute aerobic exercise in older adults with and without type 2 diabetes Purpose: The acute exercise model provides clinical insights regarding vascular dysfunction in adults with type 2 Diabetes Mellitus (T2DM), as it uncovers abnormalities not normally present at rest. However, whether vascular responses to acute exercise in adults with T2DM are intensity-dependent, and whether response patterns differ with aging and disease, remains unknown. Thus, we examined brachial artery flow-mediated dilation (FMD) and flow-mediated slowing (FMS) of pulse wave velocity (PWV), 10 and 60-min after an acute bout of high-intensity interval exercise (HIIE) and moderate-intensity continuous exercise (MICE) in middle-aged and older men with and without T2DM compared to young men without T2DM. Hypothesis: HIIE would induce a larger decrease in FMD immediately after exercise cessation compared to MICE in older men with and without T2DM. Methods: Twelve males with (57-84 yr) and without (57-76 yr) T2DM, and 12 healthy young adults (20-40 yr), completed an isocaloric acute exercise bout of HIIE, MICE, and a non-exercise condition in randomized order. FMD and FMS were evaluated and offline analyzed by the same researcher following standard guidelines. Central and peripheral arterial stiffness were measured via applanation tonometry estimates of carotid-femoral PWV (cfPWV) and carotid dorsalis pedis PWV (cdPWV), respectively. Statistical inferences were performed using linear mixed models. Results: %FMD was reduced (d= - 5.94, 95% CI: -10.50 to -1.38 %, p=0.002), whereas %FMS was increased (d= 4.55, 95% CI: 0.62 to 8.48%, p=0.01), 10-min after HIIE only in older adults with T2DM, returning to resting values 60-min into recovery. Conversely, %FMD was increased (d= 5.33, 95% CI: 0.76 to 9.89%, p=0.009) 10-min after MICE only in older adults with T2DM, while cfPWV and cdPWV remained unchanged following HIIE and MICE in all groups. There were no group differences at rest in brachial %FMD and %FMS, while cfPWV and cdPWV were higher in older adults with T2DM compared to age-matched controls (cfPWV: d= 1.94, 95% CI: 0.44 to 3.44 m. s-1, p=0.009). Conclusions: We found evidence of intensity- and disease-dependent vascular responses to acute aerobic exercise. These findings suggest that high-intensity acute exercise transiently impairs vascular function in older adults with T2DM, which may constitute an important trigger for exercise-based long-term improvements in vascular function.

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Low Type 3 Deiodinase (DIO3) activity Post-Myocardial Infarction leads to a Greater Incidence of Heart Failure in Female Mice
Authors: Teixeira Silvania , Bethea Maigen , Cavasin Maria , Knaub Leslie , Hernandez Arturo , Scalzo Rebecca , Sandoval Darleen
Published: May 2023

Objective: Studies in male mice show that myocardial infarction (MI) leads to the upregulation of type 3 deiodinase (DIO3), the enzyme that inactivates thyroid hormones, which has been regarded as a predictor of poor outcomes after MI. Interestingly, a recent human study has shown a sex-specific regulation of DIO3 after an ischemic event, with women presenting a significant downregulation of this gene. Therefore, this study aims to investigate the biological importance of DIO3 activity on MI recovery in female mice. Methods: Female mice expressing a tamoxifen-inducible Cre-recombinase under the control of the cardiomyocyte-specific Myh6 promoter and carrying a floxed selenocysteine insertion sequence in the DIO3 gene were used to reduce heart DIO3 activity. Animals were treated with tamoxifen (1.5 mg, once daily, for 3 days). MI was induced via ligation of the left anterior descending coronary artery 4 weeks after tamoxifen treatment. Ejection fraction (EF) was measured at baseline, 2 weeks, and 4 weeks after MI induction using echocardiography. Animals were sacrificed 4 weeks post-MI, and heart mitochondrial respiration was measured using Oxygraph-2K. Results: Our data demonstrated that 4 weeks post-MI, females with low DIO3 heart activity show low EF compared to wild-type control mice. We also observed that the cardiac mitochondria of females with low DIO3 activity after MI had lower lipid-linked respiration compared to wild-type control mice. Discussion/Significance of Impact: Impaired cardiomyocyte mitochondrial function has been linked to a reduced ability to recover from MI. We found that lowering cardiac DIO3 activity in female mice caused a reduction in heart function and in cardiomyocyte mitochondrial respiration, suggesting these animals have a reduced ability to recover from MI. The data have important translational implications and clinical significance given that it challenges the current knowledge about DIO3 function after MI and will contribute to explaining why women face a 20% greater risk than men of developing heart failure after MI. Furthermore, it highlights the importance of DIO3 on female heart function since the human DIO3 gene is transcriptionally responsive to estrogen, and MI numbers are relatively low in premenopausal women but significantly increase after menopause.

CTSA Grant UL1 TR002535, KL2 TR002534, & TL1 TR002533

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
ROS scavenger TEMPOL mitigates TNF alpha induced ER stress in hASM cells
Authors: Perez-Franco Rosana , Yap Jane , Blandon-Delgado Leslly , Panigrahy Nisha , Dasgupta Debanjali , Delmotte Philippe , Sieck Gary
Published: May 2023

The impact of airway inflammation is mediated by pro-inflammatory cytokines such as tumor necrosis factor alpha (TNFα), which underlies several respiratory diseases, including COVID-19, asthma, chronic bronchitis, and chronic obstructive pulmonary disease (COPD). In human airway smooth muscle (hASM) cells, exposure to TNFα leads to an accumulation of unfolded proteins in the ER, which triggers a homeostatic ER stress response activating a signaling cascade to restore normal function. Previously, we showed that TNFα selectively activates the inositol-requiring enzyme 1α (IRE1α)/X-box binding protein 1 (XBP1) ER stress pathway in hASM cells. Most likely, the protein unfolding in hASM cells caused by TNFα results from an increase in reactive oxygen species (ROS) formation. We hypothesize that ER stress induced by TNFα is mitigated by the ROS scavenger TEMPOL. Primary hASM cells were isolated from bronchiolar tissue samples obtained from 5 patients (both sexes included) who had no history of respiratory diseases. hASM cells were serum deprived for 48 h before treatment with 1) TNFα (20 ng/ml) for 12 h; 2) TEMPOL (500 μM) for 30 min followed by TNFα (20 ng/ml) for 12 h; and 3) untreated control for 12 h. At the end of the treatment period, hASM cells were processed for determination of protein expression through Western blotting. Activation of the ER stress response was assessed by phosphorylation of IRE1α and splicing of XBP1. Confirming our previous results, we found that 12 h TNFα exposure induced an ER stress response in hASM cells, evidence by phosphorylation of IRE1α and XBP1 splicing. Treating hASM cells with TEMPOL before exposure to TNFα mitigated the ER stress response. These results support our hypothesis and indicate that ROS scavenging prevents TNFα induced protein unfolding and triggering of the IRE1α/XBP1 ER stress pathway.

R01HL 157984 (GCS)

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Cyclooxygenase inhibition attenuates peripheral venous distension reflex in healthy humans
Authors: Hamaoka Takuto , Leuenberger Urs , Kronfli Anthony , Gao Zhaohui , Braha Cheryl , Luck Jonathan , Dalton Paul , Sinoway Lawrence , Cui Jian
Published: May 2023

Background: Venous distension in limbs evokes a pressor reflex (venous distension reflex, VDR). Group III and IV nerves, innervating peripheral veins, are suggested as the afferent arm of the VDR. Prostaglandins stimulate/sensitize group III/IV nerves. The purpose of this study was to evaluate the effect of cyclooxygenase (COX) blockade on muscle sympathetic nerve activity (MSNA) and blood pressure (BP) responses to venous distension. Hypothesis: We hypothesized that inhibition of prostaglandin synthesis by local COX blockade would attenuate the MSNA and BP responses to peripheral venous distension. Methods: Nineteen healthy volunteers (age, 27 ± 5 years) participated in the study with a randomized, double-blind, placebo-controlled, and crossover study design with two separate visits. To induce venous distension, a volume of solution (saline alone or 9 mg ketorolac tromethamine in saline) was infused into the vein in the antecubital fossa of an arterially occluded forearm. During the procedure, beat-by-beat heart rate (HR), BP, and MSNA were recorded simultaneously. The vein size at the site with the catheter for the infusion was measured with ultrasound. This study is associated with a registered clinical trial (NCT03513770). Data and a summary of the results: In both visits, the venous distension procedure significantly increased the vein size, and the increase in vein size was not different between the visits (P > 0.05). A significant decrease (P < 0.001) in TXB2 level (an index for prostaglandin synthesis) by the procedure was observed only in the ketorolac visit. In both visits, the venous distension procedure significantly increased BP, HR, and MSNA (all, P < 0.05). The increase in mean arterial pressure (MAP) and MSNA in the ketorolac visit was significantly lower than in the control visit (ΔMAP, 7.0 ± 6.2 vs. 13.8 ± 7.7 mmHg; ΔMSNA, 6.0 ± 7.1 vs. 14.8 ± 7.7 bursts/minute; both, P < 0.05). Discussion and conclusion: The presented data show that COX blockade attenuates the responses in MSNA and BP to peripheral venous distension. The results suggest that COX products play a key role in evoking afferent activation responsible for the VDR. Vein size was measured with ultrasonography only at the catheter site. Some members of COX products have vasoactive effects. Future studies evaluating the effect of COX blockade on vein size in more detail using other approaches (e.g., magnetic resonance imaging) are warranted to verify the detailed mechanism of the attenuated VDR by the COX blockade.

This study was supported by National Institutes of Health Grants R01 HL144781 (to J.C. and L.I.S).

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Effects of lower limb heating on the ankle-brachial index
Authors: Cui Jian , Aziz Faisal , Hamaoka Takuto , Blaha Cheryl , Luck Jonathan , Dalton Paul , Leuenberger Urs
Published: May 2023

Background: Peripheral artery disease (PAD) is an atherosclerotic disease with luminal narrowing of blood vessels, leading to an obstruction of blood flow. It most commonly affects blood vessels of the lower extremities. The ankle-brachial index (ABI, the ratio of ankle pressure over brachial pressure) is a primary index in the diagnosis (ABI < 0.9) of PAD. However, resting ABI may not be sensitive enough to detect mild/moderate PAD in some patients. The ABI after a 5 -min exercise stress test is currently used to confirm a diagnosis of PAD in patients with classical symptoms of PAD, but with normal resting ABIs. However, not all of the patients can tolerate the treadmill test. The purpose of this study was to examine the effects of lower limb heating on the ABI measurement, since lower limb heating can also induce lower limb vasodilation as walking does. Hypothesis: We hypothesized that lower limb heating would specifically decrease ABI in PAD patients. Methods: In 10 PAD patients and 12 healthy control subjects, resting ABI was measured at first. Then, both feet and lower legs were immersed in warm water (~ 40 ºC) for 30 min. Thereafter, ABI was measured again. This study is associated with a registered clinical trial (NCT03900832). Data and a summary of the results: Lower limb heating did not significantly alter the hemodynamic variables in both groups. Lower limb heating significantly decreased the ABI of both legs in PAD patients (0.75 ± 0.03 to 0.67 ± 0.04, P = 0.003), and did not alter the ABI in healthy controls (1.01 ± 0.01 to 1.03 ± 0.01, P = 0.103). Discussion and Conclusion: The presented data show that lower limb heating decreases ABI in PAD patients, while it does not alter ABI in healthy individuals. The results suggest that lower limb heating can increase the sensitivity of the ABI measurement in PAD patients. Many patients with PAD can have comorbidities that are either absolute or relative contraindications to exercise treadmill testing. Moreover, many patients with intermittent claudication may not tolerate exercise treadmill testing. We speculate that the lower limb heating approach can be applied in these patients. Further studies on the mechanisms are warranted.

This study was supported by National Institutes of Health Grants R01 HL141198 and R01 HL164571.

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Chemical chaperone 4-PBA mitigates TNF alpha mediated ER stress in hASM cells
Authors: Yap Jane , Blandon Delgado Leslly , Perez-Franco Rosana , Panigrahy Nisha , Dasgupta Debanjali , Delmotte Philippe , Sieck Gary
Published: May 2023

Airway inflammation and pro-inflammatory cytokines such as tumor necrosis factor alpha (TNFα) underlie the pathophysiology of respiratory diseases, including asthma. Previously, we showed that TNFα selectively activates the inositol-requiring enzyme 1α (IRE1α)/X-box binding protein 1 spliced (XBP1s) ER stress pathway in human airway smooth muscle (hASM) cells. The ER stress pathway is activated by the accumulation of unfolded proteins in the ER. Accordingly, chemical chaperones such as 4-phenylbutyrate (4-PBA) may ameliorate ER stress activation. In the present study, we hypothesize that 4-PBA mitigates TNFα induced ER stress in hASM cells. hASM cells were isolated from bronchiolar tissue obtained from 5 donors with no history of current smoking or respiratory diseases. hASM cells were serum-deprived for 48 h, and phenotype confirmed by expression of smooth muscle actin and fusiform morphology. hASM cells from the same donor were then separated into 4 12-h treatment groups: 1) TNFα (20 ng/mL), 2) TNFα + 4-PBA (30 min pretreatment), and 3) untreated control. Protein expression and phosphorylation of IRE1α and XBP1s were determined by Western blot. Gene expression of XBP1s was analyzed by PCR. We found that TNFα induced IRE1α phosphorylation and XBP1 splicing, which were significantly reduced by pretreatment with 4-PBA. These results support our hypothesis and suggest that, in the presence of chemical chaperone 4-PBA, TNFα induced ER stress is significantly decreased.

R01HL 157984 (GCS)

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Spinal adenosine 2A receptor inhibition restores phrenic long-term facilitation during mild systemic inflammation
Authors: Marciante Alexandria , Mitchell Gordon
Published: May 2023

Inflammation undermines several forms of neuroplasticity, including phrenic long-term facilitation (pLTF), a form of respiratory motor plasticity elicited by moderate acute intermittent hypoxia (mAIH; 3, 5-min episodes, 5 min intervals; arterial PO2 ~ 40–50 mmHg). Mild inflammation elicited by a low dose of the TLR-4 receptor agonist lipopolysaccharide (LPS; 100 mg/kg, i.p.) abolishes mAIH-induced pLTF by a mechanism that requires spinal p38 MAP kinase and protein phosphatase activity. Neuroinflammation activates microglia and other cells in the CNS, elevating extracellular levels of adenosine—a purine nucleoside known to suppress mAIH-induced pLTF. We hypothesized basal adenosine levels in ventral spinal C3-C5 segments are increased in adult male Sprague-Dawley rats 24 hours post-LPS administration, thereby undermining mAIH-induced pLTF via adenosine 2A receptor (A2A) activation. Thus, we predicted that cervical intrathecal administration of the A2A receptor inhibitor MSX-3 (10 μM, 12 μL) would rescue mAIH-induced pLTF in rats pre-treated with LPS. As predicted, LPS significantly increased basal ventral C3-C5 adenosine levels (p=0.010 vs controls; n=7 per group). In control rats (saline injection, i.p.), MSX-3 enhanced pLTF versus vehicle controls (LPS: 110 ± 16% baseline; controls: 53 ± 6%; p=0.002; each n=6). As expected, mAIH-induced pLTF was abolished in LPS-treated rats (4 ± 6% baseline; n=6). MSX-3 restored mAIH-induced pLTF to levels equivalent to MSX-3 treated control rats (120 ± 14%; p<0.001; n=6; versus LPS controls with MSX-3: p=0.539). Thus, mild LPS-induced inflammation abolishes mAIH-induced pLTF by a mechanism associated with increased spinal adenosine levels and A2A receptor activation. As we move mAIH forward as a therapeutic modality to improve breathing and non-respiratory movements in people with spinal cord injury or ALS, pretreatment with A2A receptor inhibitors may offset the undermining effects of systemic inflammation associated with these severe neuromuscular disorders.

NIH HL147554, HL148030, T32HL134621 (ABM) and the UF McKnight Brain Institute

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Acute inhibition of autophagy impairs maximal diaphragm force generation in old mice
Authors: Saldarriaga Carlos , Alaout Mayar , Khurram Obaid , Gransee Heather , Sieck Gary , Mantilla Carlos
Published: May 2023

Transdiaphragmatic pressure (Pdi) measurements are used to determine diaphragm force generation across various motor behaviors. Previous studies in the diaphragm muscle found selective age-related changes at fast-twitch fatigable motor units including increased susceptibility to neuromuscular transmission failure and denervation. Accordingly, age-related changes at diaphragm motor units predominantly impair the ability to accomplish higher force behaviors, and these aging effects are evident prior to the onset of muscle atrophy. Recent findings suggest that autophagy plays a critical role in the development of age-related neuromuscular dysfunction; however, the possible contribution of autophagy impairment to the maintenance of diaphragm force generation in old age is unknown. We hypothesized that inhibition of autophagy (using chloroquine) decreases diaphragm force-generating capacity (Pdi) in mice susceptible to age-related neuromuscular dysfunction. Adult male and female C57BL/6 mice at 16-28 months of age were randomized to receive a single intraperitoneal injection of 50 mg/kg of chloroquine or vehicle (n= 9 per treatment group) four hours before Pdi measurements, with the experimenter blinded. Pdi was measured during eupnea (room air breathing), exposure to hypoxia/hypercapnia (10% O2/5% CO2), tracheal occlusion, spontaneous deep breaths (“sighs”) and during maximal activation elicited by bilateral phrenic nerve stimulation. Consistent with our hypothesis, there was an ~25% reduction in maximal Pdi in the chloroquine group (46 ± 12 cm H2O) compared to the vehicle control (61 ± 16 cm H2O; p < 0.05). There was no effect of chloroquine on Pdi during eupnea, hypoxia/hypercapnia, tracheal occlusion, or sigh behaviors. Acute inhibition of autophagy results in reduced maximal Pdi, consistent with that evident in old mice. Thus, we propose that autophagy impairment may be an important contributor to the reduced diaphragm force generation associated with aging. These findings help us further understand the mechanisms regulating age-related neuromuscular dysfunction and may serve as a foundation to develop novel treatments to mitigate aging effects.

Support: NIH R01 AG057052.

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Age-associated impairments in exercise tolerance and critical power are related to vascular endothelial dysfunction
Authors: Gifford Jayson , Dorff Abigail
Published: May 2023

Background: Aging results in lower exercise tolerance, manifested as decreased Critical Power (CP). Aging is also associated with reduced physical activity, decreased muscle mass, and impaired vascular endothelial function, all of which may contribute to the age-related decrease in CP. Purpose: The purpose of this study was to determine if the age-related decrease in CP occurs independently of changes in physical activity and muscle mass and if it is related to impaired vascular endothelial function. Methods: 10 Old (63.1 ± 2.5 years, 5 female and 5 male) and 10 Young (24.4 ± 4.0 years, 5 female and 5 male) physically active volunteers enrolled in this study. Physical activity was measured with accelerometry. Leg muscle mass was quantified with dual x-ray absorptiometry (DEXA). CP during single leg knee extension exercise was determined over the course of 4 visits with 3-5 constant-power tests to task failure. On the fifth visit, vascular function of the leg was assessed with the passive leg movement (PLM) hyperemia. Results: Young and Old subjects did not differ in daily step count (Old = 13001.1 ± 2464.0 vs Young = 13527.0 ± 3213.8 steps, P = 0.735) or in leg lean mass (P = 0.901). The Old subjects had a lower mass-specific CP (Old = 3.20 ± 0.94 vs Young = 4.60 ± 0.87 W/kg, P = 0.004). The Old subjects also had significantly lower vascular function assessed by the peak hyperemic response to PLM, even after being normalized for leg-lean mass (Old = 79.4 ± 38.3 vs Young = 128.8 ± 34.9 ml/min/kg, P = 0.010). CP was related to PLM hyperemia (R= 0.63; P = 0.004). Importantly, simultaneously controlling for physical activity (steps per day) and for leg lean mass with partial correlation analysis, actually strengthened the relationship between CP and vascular function assessed by PLM (R = 0.69; P = 0.003). Conclusion: The age-related decline in CP is not merely the consequence of low physical activity or reduced muscle mass. Reductions in the vascular endothelial function within the exercising muscle are strongly related to the observed decline in CP. Future research should determine if interventions known to improve vascular function can ameliorate exercise tolerance in Old adults.

BYU College Undergraduate Research Award

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Endothelial cell TMEM16A channels regulate arterial contractility and blood pressure
Authors: Mata-Daboin Alejandro , Garrud Tessa , Fernandez-Pena Carlos , Peixoto-Neves Dieniffer , Bernardelli Angelica , Leo M Dennis , Jaggar Jonathan
Published: May 2023

Endothelial cells (ECs) are electrically coupled to arterial smooth muscle cells in the vascular wall and can modulate their contractility through direct control of membrane potential and via the production of several vasoactive substances. ECs express several cation channels which are known to regulate arterial contractility. In contrast, physiological functions of anion channels in ECs are unclear. ECs express TMEM16A, a Ca2+-activated Cl-channel. To examine physiological functions of TMEM16A channels in ECs, we generated an inducible, EC-specific TMEM16A knockout (TMEM16A ecKO) mouse. An increase in intracellular Ca2+ concentration ([Ca2+]i) or the application of acetylcholine (ACh) activated Cl‑ currents in fresh-isolated mesenteric artery ECs of control ( TMEM16Afl/fl) mice that were inhibited by tannic acid or benzbromarone, which are TMEM16A channel blockers. In contrast, an increase in [Ca2+]i or ACh did not activate Cl‑ currents in TMEM16A ecKO ECs. HC067047, a TRPV4 channel blocker, inhibited TMEM16A current activation by ACh in TMEM16Afl/fl ECs. GSK101, a TRPV4 channel activator, stimulated TMEM16A currents in TMEM16Afl/fl ECs, but not in TMEM16A ecKO ECs. Super resolution single-molecule localization microscopy demonstrated that surface clusters of TMEM16A and TRPV4 channels locate in nanoscale spatial proximity in the plasma membrane of ECs. ACh produced membrane hyperpolarization in pressurized TMEM16Afl/fl arteries that was attenuated in TMEM16A ecKO arteries. Vasodilation to ACh was smaller in pressurized TMEM16A ecKO arteries than in TMEM16Afl/fl arteries. TMEM16A knockout in ECs increased blood pressure in mice. These data indicate that ACh activates TRPV4-coupled TMEM16A channels in ECs to induce membrane hyperpolarization, vasodilation and a reduction in blood pressure.

National Institutes of Health

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
H2O2-REST signaling pathway is involved in cardiac vagal dysfunction and myocardial infarction-evoked ventricular arrhythmia in type 2 diabetes
Authors: Zhang Dongze , Hu Wenfeng , Tu Huiyin , Wadman Michael , Li Yulong
Published: May 2023

Myocardial infarction (MI)-induced ventricular arrhythmia is the most common cause of death in patients with type 2 diabetes mellitus (T2DM). Patients with T2DM are two to four times more likely to die from MI than non-diabetic patients. Withdrawal of cardiac vagal activity is an important trigger for MI-induced ventricular arrhythmias in T2DM. Our recent study found that T2DM-elevated hydrogen peroxide (H2O2) levels in cardiac vagal postganglionic (CVP) neurons contribute to the withdrawal of cardiac vagal activity and ventricular arrhythmogenesis through inhibition of N-type Ca2+ channels (Cav2.2) in CVP neurons. The repressor element 1-silencing transcription factor (REST) is crucial to inhibit the expression of Cav2.2-α. In general, mature neuronal cells do not express the REST in the physiological condition. Here, we tested whether H2O2 induces REST expression in CVP neurons, which successively inhibits the expression of Cav2.2 in CVP neurons, causes cardiac vagal dysfunction, and triggers MI-evoked ventricular arrhythmias in T2DM rats. Our in vitro experiments demonstrated that protein expression of REST is high, whereas Cav2.2-α expression is low in undifferentiated NG108-15 cells. Differentiation of NG108-15 cells for twenty-one days significantly reduced REST expression and increased Cav2.2-α expression. Additionally, treatment with exogenous H2O2 (1 μM) in differentiated NG108-15 cells markedly increased REST expression and decreased Cav2.2-α expression. Similarly, a high level of REST protein and a low level of Cav2.2-α protein were confirmed in CVP neurons from T2DM rats than those from age-matched sham rats. In vivo transfection of REST shRNA (2 μl, 1×107 TU/ml) into the CVP neurons significantly decreased REST expression and increased Cav2.2-α expression in CVP neurons from T2DM rats. REST shRNA also improved cardiac vagal function, measured by heart rate variability, and reduced the cumulative duration of MI-evoked ventricular arrhythmias in conscious rats. These data suggested that H2O2-REST signaling pathway could be a key factor to mediate cardiac vagal dysfunction and affect the occurrence of ventricular arrhythmias in T2DM.

This study was supported by the NIH-NHLBI (R01HL144146 to YLL).

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Role of microRNA miR-122-5p in post-transcriptional regulation of the mitochondrial thiamin pyrophosphate transporter (MTPPT; SLC25A19) in pancreatic acinar cells (PACs)
Authors: Ramamoorthy Kalidas , Sabui Subrata , Isaiah Manzon Kameron , Appakalai N Balamurugan , Said Hamid M
Published: May 2023

Thiamin (vitamin B1; also known as the “energy vitamin”) plays a critical role in normal energy metabolism and ATP production. Like all other human/mammalian cells, pancreatic acinar cells (PACs) cannot synthesize thiamin, and thus, must obtain the vitamin from circulation. Internalized free thiamin is then converted to TPP in the cytoplasm and transported into mitochondria for utilization via a carrier-mediated process that is mediated by the MTPPT (product of the SLC25A19 gene). Previous studies from our laboratory have characterized different aspects of the mitochondrial TPP uptake process in PACs; we have also delineated aspects of the MTPPT transcriptional regulation. To date, however, there has been nothing known about post-transcriptional regulation of MTPPT expression and function in PACs. We began to address these issues and, in this study, we examined the potential role of miRNAs in this regulation. First, we subjected the 458 bp long hMTPPT-3' untranslated region (UTR) to different in silico analysis (namely: miRDB, Targetscan, and DIANA-microTCDS), which identified multiple putative miRNA binding sites (including sites for miR-122-5p) in this region. These putative miRNA binding sites were found to be conserved in the human and rat MTPPTs. We then examined the effect of transfecting pmirGLO-hMTPPT-3'UTR into PAC (rat-derived AR42J cells) and observed a significant reduction in luciferase activity compared with cells transfected with pmirGLO empty vector. Focusing on the putative miR-122-5p (since it is expressed in normal human pancreatic cells and that its level is induced in pancreatitis), we found that transfecting AR42J cells as well as human native primary PACs (obtained from normal human organ donors) with a mimic of miR-122-5p to lead to a significant inhibition in level of MTPPT mRNA and protein expression as well as in TPP uptake by isolated mitochondria. On the other hand, inhibiting the miR-122-5p with the use of miR-122-5p inhibitor was found to lead to an increase in MTPPT mRNA and protein expression as well as in TPP uptake by isolated mitochondria. Additionally, truncating the region that carries the putative miR-122-5p binding site(s) in the 3'UTR of MTPPT as well as mutating those sites were found to lead to an abrogation in the inhibitory effect of this miRNA on luciferase activity in PACs. In conclusion, our results demonstrate, for the first time, that expression and function of the MTPPT are subject to post-transcriptional regulation by miR-122-5p.

Supported by NIH grants AA-018071 & DK-56061, and VA grants 101BX001142 & IK6BX006189

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Amyloid-beta peptide potentiates vascular Cav1.2 channel activity
Authors: Taylor Jade , Martin-Aragon Baudel Miguel , Hell Johannes , Nieves-Cintron Madeline , Navedo Manuel
Published: May 2023

Alzheimer’s disease (AD), the most common form of dementia, affects over 6 million Americans and has devastating emotional and physical consequences for both the patient and surrounding loved ones. Current drug therapies are principally targeted at the main disease-causing peptide in AD, amyloid-beta (Aβ). These drugs reduce symptoms at the onset of the disease but have negligible positive impacts once AD has progressed. Thus, new avenues for therapeutic intervention are imperative. It is currently understood that reduced cerebral blood flow (CBF), due to an increased constriction of cerebral blood vessels, contributes to the pathogenesis of AD. Calcium (Ca2+) influx through voltage-gated L-type calcium CaV1.2 channels is essential for vascular smooth muscle (VSM) contractility, arterial diameter, and CBF. Yet, how Aβ affects CaV1.2 channel activity and vascular function is unclear. Here, we test the hypothesis that Aβ promotes vascular CaV1.2 clustering leading to increased CaV1.2 channel activity and enhanced vascular reactivity. The hypothesis was tested using a multi-scale approach that includes super-resolution microscopy, patch-clamp electrophysiology, and pressure myography using freshly isolated cerebral VSM and arteries from WT mice. Initial experiments found that exposing VSM to 100 nM Aβ for 10 minutes stimulates the redistribution of CaV1.2 into superclusters. This observation was correlated with an increase in CaV1.2 channel activity and the frequency and strength of cooperative events, which are expected to amplify Ca2+ influx and alter vascular contractility. Consistent with this premise, pressurized cerebral arteries exposed to Aβ showed an elevation in myogenic tone. These results suggest that Aβ impacts vascular CaV1.2 spatiotemporal properties to control vascular contractility. Moreover, results may provide new mechanistic insight into a link between impaired CaV1.2 function, VSM dysfunction and AD.

NIH R01HL149127 and R01HL149127S1(to MFN).

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Trait anxiety is associated with blood pressure and muscle sympathetic nerve activity
Authors: Bigalke Jeremy , Durocher John , Greenlund Ian , Keller-Ross Manda , Carter Jason
Published: May 2023

Heightened anxiety is associated with an increased risk for cardiovascular diseases such as coronary heart disease and hypertension. While hyperactivity of the sympathetic nervous system has been suggested to mediate this relationship, direct measures of muscle sympathetic nerve activity (MSNA) in people with elevated anxiety have indicated little, if any, alterations in baseline sympathetic function. However, studies examining anxiety and MSNA have been limited by small sample sizes and lack of detail on anxiety severity, limiting the ability to investigate subtle interactions. The present study assessed associations between trait anxiety, mean arterial pressure (MAP), and MSNA using a large, representative sample of healthy young men and women, with an exploratory assessment of sex-specific interactions. We hypothesized that heightened trait anxiety would be associated with elevated resting MAP and MSNA. Trait anxiety was assessed using the Spielberger Trait Anxiety Inventory (STAI) in 97 healthy adults (52 male, 45 female; 25±1 years; 26±1 kg/m2). In each participant, the average of three seated brachial blood pressures was recorded and used in subsequent analysis. All participants underwent an autonomic function test that simultaneously assessed heart rate (HR), beat-to-beat blood pressure, MSNA burst frequency (BF) and incidence (BI) during 5-10 minutes of supine rest. MSNA recordings were captured in 91 participants. Multiple regression was used to test the relationship between STAI, MAP, MSNA, and HR while accounting for age and sex. Sex-specific interactions were explored with correlations controlling for age. The regression model significantly predicted MAP (R2=.196, P<0.001), BF (R2=.136, P=0.005), and BI (R2=.150, P=0.003), but not HR (R2=.055, P=0.155). STAI scores were independently associated with MAP (β=.207, P=0.029), BF (β=.245, P=0.016), and BI (β=.224, P=0.027). In males, STAI was correlated with MAP (R=.277, P=0.049), BF (R=.363, P=0.009), and BI (R=.346, P=0.013), but not HR (R=.140, P=0.326). In contrast, STAI was not associated with MAP (R=.174, P=0.260), BF (R=.094, P=0.574), BI (R=.069, P=0.681), or HR (R=.226, P=0.141) in females. In a large sample of healthy adults, our results highlight the relationship between the severity of trait anxiety, resting blood pressure, and sympathetic outflow, with a more pronounced relationship in males.

Support: NIH (AA-024892, 1R15HL140596-01, AG064038-01A1, U54GM115371; P20GM103474, 2TL1 TR002318).

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.