Myogenic Response Research Articles

Potentiation of active force by cyclic strain in sheep carotid arterial smooth muscle.

The ability to generate force in large arteries is known to be augmented by cyclic strain that mimics the mechanically dynamic in vivo environment associated with blood pressure fluctuation experienced by these arteries. Cyclic strain does not induce a contractile response, like that observed in the myogenic response seen in small arteries, but prompts a substantial increase in the response to electrical stimulation. We coined this phenomenon "force potentiation." Because protein kinase C (PKC) and rho-kinase (ROCK) are known to play a role in increasing contractility of arterial smooth muscle by inhibition of myosin light chain phosphatase, and integrin-link kinase (ILK) is crucial in mechanotransduction, we examined how inhibition of these kinases affected force potentiation in sheep carotid artery. We found that phosphorylation of the regulatory myosin light chain was enhanced by cyclic strain, but the enhancement was observed only in activated, not in relaxed muscle. Inhibition of ROCK diminished force potentiation and active isometric force, likely due to the disinhibition of myosin light chain phosphatase. Inhibition of PKC abolished force potentiation without an effect on active force, suggesting a more exclusive role of PKC (compared with ROCK) in mediating force potentiation. Inhibition of ILK had a similar effect as PKC inhibition, suggesting that ILK may be an upstream kinase for PKC activation by mechanical stimuli. Taken together, the findings suggest that ILK, PKC, and ROCK are important kinases in the signal transduction pathway that mediate the effect of mechanical strain on force potentiation.NEW & NOTEWORTHY When subjected to mechanical strain, smooth muscle from large arteries has the ability to increase its force generation (force potentiation), which could be important in autoregulation of blood pressure. This phenomenon, however, does not involve a myogenic response, such as the one seen in small arteries and arterioles. Our work shows the involvement of ILK, PKC, and ROCK in the signal transduction pathway that mediates the force-potentiating effect of mechanical strain in large arteries.

Autoregulatory mission impossible: when afferent arterioles lose contractility

The myogenic response of afferent arterioles is a key autoregulatory mechanism that protects the glomeruli from barotrauma. Afferent arteriolar smooth muscle cells contract to increased intraluminal pressure through mechanosensitive cation channels and interactions between integrin and extracellular matrix that trigger calcium-dependent actomyosin contraction. The study by Feng etal. provides evidence supporting the concept that increased matrix metalloproteinase 9 in kidney microvessels of Dahl salt-sensitive rats interferes with integrin-matrix binding and promotes phenotypic transformation of afferent arterioles, causing loss of myogenic constriction and hypertensive nephropathy.

Characterisation of Lipoma-Preferred Partner as a Novel Mechanotransducer in Vascular Smooth Muscle Cells.

In arteries and arterioles, a chronic increase in blood pressure raises wall tension. This continuous biomechanical strain causes a change in gene expression in vascular smooth muscle cells (VSMCs) that may lead to pathological changes. Here we have characterised the functional properties of lipoma-preferred partner (LPP), a Lin11-Isl1-Mec3 (LIM)-domain protein, which is most closely related to the mechanotransducer zyxin but selectively expressed by smooth muscle cells, including VSMCs in adult mice. VSMCs isolated from the aorta of LPP knockout (LPP-KO) mice displayed a higher rate of proliferation than their wildtype (WT) counterparts, and when cultured as three-dimensional spheroids, they revealed a higher expression of the proliferation marker Ki 67 and showed greater invasion into a collagen gel. Accordingly, the gelatinase activity was increased in LPP-KO but not WT spheroids. The LPP-KO spheroids adhering to the collagen gel responded with decreased contraction to potassium chloride. The relaxation response to caffeine and norepinephrine was also smaller in the LPP-KO spheroids than in their WT counterparts. The overexpression of zyxin in LPP-KO VSMCs resulted in a reversal to a more quiescent differentiated phenotype. In native VSMCs, i.e., in isolated perfused segments of the mesenteric artery (MA), the contractile responses of LPP-KO segments to potassium chloride, phenylephrine or endothelin-1 did not vary from those in isolated perfused WT segments. In contrast, the myogenic response of LPP-KO MA segments was significantly attenuated while zyxin-deficient MA segments displayed a normal myogenic response. We propose that LPP, which we found to be expressed solely in the medial layer of different arteries from adult mice, may play an important role in controlling the quiescent contractile phenotype of VSMCs.

Glomerular microcirculation: Implications for diabetes, preeclampsia, and kidney injury.

This review outlines the features of tandem regulation of glomerular microcirculation by autoregulatory mechanisms and intraglomerular redistribution of blood flow. Multiple points of cooperation exist between autoregulatory and distributional mechanisms. Mutual interactions between myogenic and tubuloglomerular feedback (TGF) mechanisms regulating the inflow are briefly discussed. In addition to this, TGF operation involving purinergic, autocoid, and NO signaling affects, however, not only afferent arteriolar tone, but mesangial cell tone as well. The latter reversibly reconfigures the distribution of blood flow between the shorter and longer pathways in the glomerular tuft. I advance a hypothesis that blood flow in these pathways spontaneously alternates, and mesangial cell tonicity serves as a rheostatic shift between them. Furthermore, humoral messengers from macula densa cells, themselves dependent on myogenic mechanisms, fine-tune the secretion of renin and, subsequently, the local, intrarenal generation of angiotensin II, which, in turn, provides additional vasomotor signaling to glomerular capillaries through changing the tone of mesangial cells. This complex regulatory network may partially explain the phenomenon of renal functional reserve, as well as suggest implications for changes in renal function during pregnancy, early diabetes mellitus, and acute kidney injury.

Abstract 083: Neurovascular And Cognitive Dysfunction In Middle-Aged Mice With High Blood Pressure Variability

Blood pressure variability (BPV) is a novel risk factor for cognitive decline. This study tested the hypothesis that BPV disrupts cellular function and communication at the neurovascular complex (vascular, glia, and neuron) contributing to BPV-induced neurovascular dysfunction. A main goal of the study was to establish a murine BPV model and examine how this risk factor impacts key functional outputs of the neurovascular complex. Middle-aged C57BL6 mice were implanted a cranial window and injected GCaMP viral vectors to monitor Ca 2+ activity in cortical neurons or astrocytes. DSI telemetry was used to monitor cardiovascular variables simultaneously ( in vivo ) with arteriole diameter and astrocyte/neuronal Ca 2+ responses via two-photon imaging. In addition, mice were implanted a programmable pump and infused 6-8 pulses/day of saline (Control) or angiotensin II (Ang II, 18 μg/day) for 20-25 days. Ang II infusions evoked transient BP increases (Δ 23±2 mmHg, mean arterial pressure (MAP, P=0.0001, n=14)), and increased BPV (P=0.04, n=12) measured by the average real variability. The 24-hour MAP was not different compared to Controls, allowing for the investigation of BPV-evoked changes in neurovascular function in the absence of hypertension. BPV mice displayed cognitive decline (NOR, P=0.006, n=10). Myogenic responses were lost in parenchymal arterioles of BPV mice but preserved in Controls (P<0.01 n=6). In addition to impaired vascular function, during Ang II-induced BP increases, astrocytes of BPV mice exhibited greater Ca 2+ event frequency (P<0.0001, n=4-6), but of lesser amplitude and reduced area (P=0.02, n=4-6). Furthermore, the neurovascular coupling response, evoked via whisker stimulation, was blunted in mice with high BPV (P<0.0001, n=6-10). Relevant to inflammation, microglial cells from BPV mice had elevated Iba1 (P=0.03, n=4-5) and CD68+ (P=0.003), an inflammatory marker. These findings show that BPV, in the absence of hypertension, result in a loss of cerebral autoregulation, a shift in spontaneous astrocyte Ca 2+ dynamics during blood pressure fluctuations, and increased microglia reactivity. Future mechanistic studies are needed to determine how BPV contributes to cognitive decline.

Abstract P235: The Absence Of Inhibitory G Protein Regulation By Rgs2 Increases Basal Phosphodiesterase Activity To Decrease Forskolin-stimulated Camp Generation And Enhances Myogenic Response In Uterine Arteries Of Non-pregnant Mice

G protein regulation by regulators of G protein signaling (RGS) proteins is crucial to maintaining normal vascular tone for promoting uterine vascular bed perfusion. However, the specific regulatory mechanisms involved are incompletely understood. Previously, we showed that loss of regulation of Gi/o and Gq/11 by RGS2 contributes to enhanced myogenic tone and decreased uterine blood flow in non-pregnant mice. Similarly, RGS5, closely related to RGS2, was found to fine-tune G protein signaling and modulate GPCR-evoked vascular reactivity. RGS2 and RGS5 are co-expressed in the uterine vascular bed. However, it is unclear whether their regulatory activities are coordinated in fine-tuning G protein signaling to modulate vascular tone. Here, we tested whether the combined activity of RGS2 and 5 reduces myogenic tone by negatively regulating Gi/o signaling to promote cAMP-dependent uterine artery relaxation. We used commercially available kits to measure forskolin-stimulated cAMP generation and basal phosphodiesterase (PDE) activity in freshly isolated uterine arteries from non-pregnant wild type, Rgs5 KO and Rgs2 KO mice. Basal tone and myogenic response to increasing intraluminal pressure were assessed using ex vivo pressure myography. We found that the absence of RGS2 but not RGS5 decreased forskolin-stimulated cAMP generation. Conversely, basal PDE activity was augmented in both Rgs2 KO and Rgs5 KO arteries, though the increase was less robust in Rgs5 KO arteries. Basal tone and myogenic response were reduced in Rgs5 KO arteries compared to WT or Rgs2 KO arteries in the absence or presence of nitric oxide synthase blockade with L-NAME. Activating Gi/o with dopamine increased myogenic constriction in all genotypes but less robustly in Rgs5 KO arteries, while pre-incubation of the vessels with exogenous cAMP or PDE inhibition with IBMX normalized maximal myogenic response to similar levels in all groups. Collectively, these findings indicate that altered Gi/o regulation due to RGS2 absence enhances myogenic tone by promoting PDE-mediated inhibition of cAMP-dependent vasodilation. While both RGS2 and RGS5 negatively regulate this novel Gi/o-PDE-cAMP signaling pathway, RGS5 dampens the inhibitory efficacy of RGS2 towards Gi/o in uterine arteries.

Reduced tubuloglomerular feedback activity and absence of its synchronization in a connexin40 knockout rat.

Introduction: Tubuloglomerular feedback (TGF) is the negative feedback component of renal blood flow (RBF) autoregulation. Neighbouring nephrons often exhibit spontaneous TGF oscillation and synchronization mediated by endothelial communication, largely via connexin40 (Cx40). Methods: We had a knockout (KO) rat made that lacks Cx40. One base pair was altered to create a stop codon in exon 1 of Gja5, the gene that encodes Cx40 (the strain is WKY-Gja55em1Mcwi). Blood pressure (BP)-RBF transfer functions probed RBF dynamics and laser speckle imaging interrogated the dynamics of multiple efferent arterioles that reach the surface (star vessels). Results: The distribution of wild type (WT), heterozygote, and KO pups at weaning approximated the Mendelian ratio of 1:2:1; growth did not differ among the three strains. The KO rats were hypertensive. BP-RBF transfer functions showed low gain of the myogenic mechanism and a smaller TGF resonance peak in KO than in WT rats. Laser speckle imaging showed that myogenic mechanism had higher frequency in KO than in WT rats, but similar maximum spectral power. In contrast, the TGF frequency was similar while peak power of its oscillation was much smaller in KO than in WT rats. In WT rats, plots of instantaneous TGF phase revealed BP-independent TGF synchronization among star vessels. The synchronization could be both prolonged and widespread. In KO rats TGF synchronization was not seen, although BP transients could elicit short-lived TGF entrainment. Discussion: Despite the reduced TGF spectral power in KO rats, there was sufficient TGF gain to induce oscillations and therefore enough gain to be effective locally. We conclude that failure to synchronize is dependent, at least in part, on impaired conducted vasomotor responses.

Myogenic and cortical evoked potentials vary as a function of stimulus pulse geometry delivered in the subthalamic nucleus of Parkinson's disease patients.

The therapeutic efficacy of deep brain stimulation (DBS) of the subthalamic nucleus (STN) for Parkinson's disease (PD) may be limited for some patients by the presence of stimulation-related side effects. Such effects are most often attributed to electrical current spread beyond the target region. Prior computational modeling studies have suggested that changing the degree of asymmetry of the individual phases of the biphasic, stimulus pulse may allow for more selective activation of neural elements in the target region. To the extent that different neural elements contribute to the therapeutic vs. side-effect inducing effects of DBS, such improved selectivity may provide a new parameter for optimizing DBS to increase the therapeutic window. We investigated the effect of six different pulse geometries on cortical and myogenic evoked potentials in eight patients with PD whose leads were temporarily externalized following STN DBS implant surgery. DBS-cortical evoked potentials were quantified using peak to peak measurements and wavelets and myogenic potentials were quantified using RMS. We found that the slope of the recruitment curves differed significantly as a function of pulse geometry for both the cortical- and myogenic responses. Notably, this effect was observed most frequently when stimulation was delivered using a monopolar, as opposed to a bipolar, configuration. Manipulating pulse geometry results in differential physiological effects at both the cortical and neuromuscular level. Exploiting these differences may help to expand DBS' therapeutic window and support the potential for incorporating pulse geometry as an additional parameter for optimizing therapeutic benefit.

Postnatal development of extracellular matrix and vascular function in small arteries of the rat.

Introduction: Vascular extracellular matrix (ECM) is dominated by elastic fibers (elastin with fibrillin-rich microfibrils) and collagens. Current understanding of ECM protein development largely comes from studies of conduit vessels (e.g., aorta) while resistance vessel data are sparse. With an emphasis on elastin, we examined whether changes in postnatal expression of arteriolar wall ECM would correlate with development of local vasoregulatory mechanisms such as the myogenic response and endothelium-dependent dilation. Methods: Rat cerebral and mesenteric arteries were isolated at ages 3, 7, 11, 14, 19days, 2months, and 2years. Using qPCR mRNA expression patterns were examined for elastin, collagen types I, II, III, IV, fibrillin-1, and -2, lysyl oxidase (LOX), and transglutaminase 2. Results: Elastin, LOX and fibrillar collagens I and III mRNA peaked at day 11-14 in both vasculatures before declining at later time-points. 3D confocal imaging for elastin showed continuous remodeling in the adventitia and the internal elastic lamina for both cerebral and mesenteric vessels. Myogenic responsiveness in cannulated cerebral arteries was detectable at day 3 with constriction shifted to higher intraluminal pressures by day 19. Myogenic responsiveness of mesenteric vessels appeared fully developed by day 3. Functional studies were performed to investigate developmental changes in endothelial-dependent dilation. Endothelial-dependent dilation to acetylcholine was less at day 3 compared to day 19 and at day 3 lacked an endothelial-derived hyperpolarizing factor component that was evident at day 19. Conclusion: Collectively, in the rat small artery structural remodeling and aspects of functional control continue to develop in the immediate postnatal period.

Kv2.1 Channels Prevent Vasomotion and Safeguard Myogenic Reactivity in Rat Small Superior Cerebellar Arteries.

Vascular smooth muscle voltage-gated potassium (Kv) channels have been proposed to contribute to myogenic autoregulation. Surprisingly, in initial experiments, we observed that the Kv2 channel inhibitor stromatoxin induced vasomotion without affecting myogenic tone. Thus, we tested the hypothesis that Kv2 channels contribute to myogenic autoregulation by fine-tuning the myogenic response. Expression of Kv2 channel mRNA was determined using real-time PCR and 'multiplex' single-cell RT-PCR. Potassium currents were measured using the patch-clamp technique. Contractile responses of intact arteries were studied using isobaric myography. Expression of Kv2.1 but not Kv2.2 channels was detected in intact rat superior cerebellar arteries and in single smooth muscle cells. Stromatoxin, a high-affinity inhibitor of Kv2 channels, reduced smooth muscle Kv currents by 61% at saturating concentrations (EC50 36 nmol/L). Further, stromatoxin (10-100 nmol/L) induced pronounced vasomotion in 48% of the vessels studied. In vessels not exhibiting vasomotion, stromatoxin did not affect myogenic reactivity. Notably, in vessels exhibiting stromatoxin-induced vasomotion, pressure increases evoked two effects: First, they facilitated the occurrence of random vasodilations and/or vasoconstrictions, disturbing the myogenic response (24% of the vessels). Second, they modified the vasomotion by decreasing its amplitude and increasing its frequency, thereby destabilizing myogenic tone (76% of the vessels). Our study demonstrates that (i) Kv2.1 channels are the predominantly expressed Kv channels in smooth muscle cells of rat superior cerebellar arteries, and (ii) Kv2.1 channels provide a novel type of negative feedback mechanism in myogenic autoregulation by preventing vasomotion and thereby safeguarding the myogenic response.

CP55940-induced vasorelaxation is endothelial-dependent and mediated by the CB1R through NOS, COX and EDHF pathways in porcine cerebral arteries.

Using swine as an experimental model, we chose to examine whether the cannabinoid receptors, e.g. CB1R and the CB2R, could affect vasomotor tone in isolated pial arteries. It was hypothesized that the CB1R would specifically mediate cerebral artery vasorelaxation in an endothelial-dependent manner. First-order pial arteries were isolated from female Landrace pigs (age = 2 months; N = 27) for wire and pressure myography. Arteries were pre-contracted with a thromboxane A2 analogue (U-46619) and vasorelaxation in response to the CB1R and CB2R receptor mixed agonist CP55940 was examined in the following conditions: 1) untreated; 2) inhibition of the CB1R (AM251); or 3) inhibition of the CB2R receptor (AM630). The data revealed that CP55940 elicits a CB1R-dependent relaxation in pial arteries. CB1R expression was confirmed using immunoblot and immunohistochemical analyses. Subsequently, the role of different endothelial-dependent pathways in the CB1R-mediated vasorelaxation was examined using: 1) denudation (removal of the endothelium); 2) inhibition of cyclooxygenase (COX; Naproxen); 3) inhibition of nitric oxide synthase (NOS; L-NAME); 4) combined inhibition of COX + NOS. The data revealed CB1R-mediated vasorelaxation was endothelial-dependent, with contributions from COX-derived prostaglandins, NO, and endothelium-dependent hyperpolarizing factor (EDHF). Pressurized arteries underwent myogenic curves (20-100 mmHg) under the following conditions: 1) untreated; 2) inhibition of the CB1R. The data revealed CB1R inhibition increased basal myogenic tone, but not myogenic reactivity. As the vascular responses were assessed in isolated pial arteries, this work reveals that the CB1R modulates cerebrovascular tone independently of changes in brain metabolism.

The isolated effects of local cold application on proteolytic and myogenic signaling

Post-exercise cooling studies reveal inhibitory effects on markers of skeletal muscle growth. However, the isolated effect of local cold application has not been adequately addressed. It is unclear if the local cold or the combination of local cold and exercise is driving negatively altered skeletal muscle gene expression. The purpose was to determine the effects of a 4 h local cold application to the vastus lateralis on the myogenic and proteolytic response. Participants (n = 12, 27 ± 6 years, 179 ± 9 cm, 82.8 ± 13.0 kg, 18.4 ± 7.1 %BF) rested with a thermal wrap placed on each leg with either circulating cold fluid (10 °C, COLD) or no fluid circulation (room temperature, RT). Muscle samples were collected to quantify mRNA (RT-qPCR) and proteins (Western Blot) associated with myogenesis and proteolysis. Temperatures in COLD were lower than RT at the skin (13.2 ± 1.0 °C vs. 34.8 ± 0.9 °C; p < 0.001) and intramuscularly (20.5 ± 1.3 °C vs. 35.6 ± 0.8 °C, p < 0.001). Myogenic-related mRNA, MYO-G and MYO-D1, were lower in COLD (p = 0.001, p < 0.001, respectively) whereas myogenic-mRNA, MYF6, was greater in COLD (p = 0.002). No other myogenic associated genes were different between COLD and RT (MSTN, p = 0.643; MEF2a, p = 0.424; MYF5, p = 0.523; RPS3, p = 0.589; RPL3-L, p = 0.688). Proteolytic-related mRNA was higher in COLD (FOXO3a, p < 0.001; Atrogin-1, p = 0.049; MURF-1, p < 0.001). The phosphorylation:total protein ratio for the translational repressor of muscle mass, 4E-BP1Thr37/46, was lower in COLD (p = 0.043), with no differences in mTORser2448 (p = 0.509) or p70S6K1Thr389 (p = 0.579). Isolated local cooling over 4 h exhibits inhibited myogenic and higher proteolytic skeletal muscle molecular response.

Renoprotective effects of GLP-1 receptor agonists and SGLT-2 inhibitors-is hemodynamics the key point?

Two novel treatments for diabetic kidney disease have emerged after decades with little progression. Both agents were developed for improved glycemic control in patients with type-2 diabetes. However, large clinical trials showed renoprotective effects beyond their ability to lower plasma glucose levels, body weight, and blood pressure. How this renal protection occurs is unknown. We will discuss their physiological effects, with special focus on the renal effects. We discuss how these drugs affect the function of the diabetic and nondiabetic kidneys to elucidate mechanisms by which the renoprotection could arise. Diabetic kidney disease affects the glomerular capillaries, which are usually protected by the renal autoregulatory mechanisms, the myogenic response, and the tubuloglomerular feedback mechanism. Animal models with reduced renal autoregulatory capacity develop chronic kidney disease. Despite different cellular targets, both drugs are suspected to affect renal hemodynamics through changes in the renal autoregulatory mechanisms. The glucagon-like peptide-1 receptor agonists (GLP-1RAs) exert a direct vasodilatory effect on the afferent arteriole (AA) positioned just before the glomerulus. Paradoxically, this effect is expected to increase glomerular capillary pressure, causing glomerular injury. In contrast, the sodium-glucose transporter-2 inhibitors (SGLT2i) are believed to activate the tubuloglomerular feedback mechanism to elicit vasoconstriction of the afferent arteriole. Because of their opposing effects on the renal afferent arterioles, it appears unlikely that their renoprotective effects can be explained by common effects of renal hemodynamics, but both drugs appear to add protection to the kidney beyond what can be obtained with classical treatment targeted at lowering blood glucose levels and blood pressure.

Influence of the Electrode Montage to Detect Ocular Vestibular Evoked Myogenic Potentials.

To compare ocular vestibular evoked myogenic potentials (oVEMPs) obtained with three different electrode montages (infra-orbital vs belly-tendon vs chin) in a group of healthy subjects. To assess the electrical activity recorded at the level of the reference electrode in the belly-tendon and chin montages. Prospective study. Tertiary referral center. Twenty-five healthy adult volunteers. Each ear was tested separately via air-conducted sound (500 Hz Narrow Band CE-Chirps at 100 dB nHL) for recording contralateral myogenic responses. Recording conditions were randomized. n1-p1 amplitudes values, interaural amplitude asymmetry ratios (ARs) and response rates. The belly-tendon electrode montage (BTEM) produced larger amplitudes than the chin ( p = 0.008) and the IOEM (infra-orbital electrode montage; p < 0.001). The chin montage displayed larger amplitudes than the IOEM ( p < 0.001). The interaural amplitude asymmetry ratios (ARs) were not affected by the different electrode montages ( p = 0.549). In 100% of cases, oVEMPs were detected bilaterally with the BTEM which is higher than with the chin and the IOEM ( p < 0.001; p = 0.020, respectively). We did not record any VEMP when placing the active electrode on the contralateral internal canthus or the chin and the reference electrode on the dorsum of the hand. The BTEM increased the amplitudes recorded and response rate in healthy subjects. No positive or negative reference contamination was observed with the belly-tendon or chin montages.

In vivo pressure-flow relation of human cutaneous vessels following prolonged iterative exposures to hypergravity.

The study examined intra- and interlimb variations in cutaneous vessel responsiveness to acute and repeated transmural pressure elevations. In 11 healthy men, red blood cell flux was assessed via laser-Doppler flowmetry on both glabrous and nonglabrous skin regions of an arm (finger and forearm) and leg (toe and lower leg), across a wide range of stepwise increasing distending pressures imposed in the vessels of each limb separately. The pressure-flux cutaneous responses were evaluated before and after 5 wk of intermittent (40 min, 3 sessions per week) exposures to hypergravity (∼2.6-3.3 G; G training). Before and after G training, forearm and lower leg blood flux were relatively stable up to ∼210 and ∼240 mmHg distending pressures, respectively; and then they increased two- to threefold (P < 0.001). Finger blood flux dropped promptly (P < 0.001), regardless of the G training (P = 0.64). At ≤120-mmHg distending pressures, toe blood flux enhanced by ∼40% (P ≤ 0.05); the increase was augmented after the G training (P = 0.01). At high distending pressures, toe blood flux dropped by ∼70% in both trials (P < 0.001). The present results demonstrate that circulatory autoregulation is more pronounced in glabrous skin than in nonglabrous skin, and in nonglabrous sites of the leg than in those of the arm. Repetitive high-sustained gravitoinertial stress does not modify the pressure-flow relationship in the dependent skin vessels of the arm nor in the nonglabrous sites of the lower leg. Yet it may partly inhibit the myogenic responsiveness of the toe's glabrous skin.

Hearing impairment in patients with alopecia areata.

Background Alopecia areata is an autoimmune disease that damages hair follicles and follicular melanocytes can be involved in the autoimmune process. Therefore, similar to vitiligo, there may be a relationship between sensorineural hearing loss and alopecia areata. Aims/objectives This study aimed to investigate potential hearing impairments in patients with alopecia areata. Methods A total of 42 subjects with alopecia areata and 42 healthy individuals enrolled in this cross-sectional study. The hearing was evaluated by vestibular evoked myogenic potential, otoacoustic emission and pure tone audiometry tests in the patients and control subjects. Results A normal otoacoustic emission was reported in 59.5% and 100% of subjects with alopecia areata and the controls, respectively (P = 0.02). Higher speech recognition thresholds (P = 0.02) and speech discrimination scores were reported more in subjects with alopecia areata than in controls (P < 0.001); however, the most comfortable level of speech was not significantly different between the groups (P = 0.06). The greatest increase in the hearing threshold was recorded at a frequency of 8000 Hz, while at frequencies of 500 and 1000 Hz, the patients and controls did not significantly differ (P > 0.05). About 6 (14.3%) and 2 (4.8%) of patients with unilateral and bilateral involvement, respectively, demonstrated no vestibular evoked myogenic potential response in the alopecia areata group. The patients and controls did not significantly differ in terms of amplitudes of the vestibular evoked myogenic potential test (P = 0.097). Limitation Small sample size and qualitative measurement of otoacoustic emission were limitations of our study. Conclusion Hearing loss was more common in alopecia areata patients than in healthy individuals. Follicular melanocytes may be involved in the alopecia areata inflammatory process, and destroying melanocytes may impact hearing function in the inner ear. However, there was no significant relationship between the duration and severity of alopecia areata and hearing loss.

TRPC6 deficiency impairs middle cerebral artery myogenic response and brain spatial memory

Transient receptor potential channel 6 (TRPC6) is a nonselective cation channel belonging to the TRP ion channel family and expressed in vascular smooth muscle cells. TRPC6 regulates calcium influx in response to vascular wall mechanical stretch. A previous study reported that TRPC6 is critical for the myogenic response of the middle cerebral artery (MCA). Impaired MCA myogenic response is implicated in cerebral blood flow dysfunction and may promote cognitive deficits. In this study, we investigated whether TRPC6 contributes to the MCA myogenic response at pressure range (from 40 to 180 mmHg) in 12-week-old male wild-type (WT) and whole-body TRPC6 knock-out (TRPC6 KO) mice (n=5-8). Cognitive function (spatial learning and reference memory) was examined at 22 weeks of age using the Morris water maze test. Myogenic response in MCA was similar in WT and TRPC6 KO mice when perfusion pressure was increased from 40 to 80 mmHg. At higher pressures (&gt;120 mmHg), however, TRPC6 KO mice displayed significantly reduced myogenic response compared to WT mice (3 % less of myogenic tone in TRPC6 KO compared to WT) which was further exacerbated when perfusion pressure was increased to 180 mmHg (9 % less of myogenic tone in TRPC6 KO compared to WT). There were no significant differences in wall thickness, wall-to-lumen ratio, vascular distensibility, incremental distensibility, or elastic modulus curves in MCA between WT and TRPC6 KO mice. Compared to WT mice, TRPC6 KO mice displayed similar spatial learning curves after 4 days of training but showed significantly impaired reference memory (14±1 % vs 29±4 % in the target quadrant) and reduced frequency (1.0±0.3 vs 2.1±0.4 in the target area crossing) compared to WT. These findings suggest that TRPC6 deficiency impairs myogenic response at higher perfusion pressures and spatial memory, even at a young age. (AG050049 (F.F), AG057842 (F.F), AG079336 (F.F), NIDDK R00DK113280, R01DK121411, NIGMS P20GM104357 and NIGMS U54GM115428) 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.

Abstract 604: Distinct Fibroadipogenic Progenitor Populations And Trajectories Characterize Regenerative And Non-regenerative Responses In The Ischemic Limb

Introduction: Skeletal muscle possesses the capacity for self-repair. This ability is impaired in chronic limb threatening ischemia (CLTI) patients. Myopathy in CLTI patients is associated with adverse clinical outcomes. Skeletal muscle regeneration requires multicellular interactions between muscle progenitor cells and other cell types. Fibroadipogenic progentiors (FAPs) represent a support cell type in the skeletal muscle niche; they modulate inflammatory and myogenic responses via the secretion of a paracrine factors. Hence, FAPs are an attractive therapeutic target for myopathies. We sought to characterize the temporal dynamics and transcriptional heterogeneity of FAPs in failed and successful skeletal muscle regeneration in the ischemic limb. Methods: We analyzed the FAP regenerative response in BALB/c (non-regenerative) and C57BL/6 (regenerative) mice undergoing hindlimb ischemia using single-cell RNA sequencing spanning 4 distinct time points. We also performed RNA velocity analysis to determine the cellular dynamics of FAP sub-populations during the repair process. Results: We identified 10 FAP populations in the ischemic limb, including specific regenerative and non-regenerative FAP sub-clusters. Gene ontology (GO) enrichment analysis demonstrated distinct transcriptional programs between regenerative and non-regenerative FAPs. Regenerative FAPs were enriched in extracellular matrix and angiogenesis pathways, while non-regenerative FAPs were enriched in chemotactic pathways. RNA velocity analysis demonstrated divergent FAP cellular trajectories. Conclusions: Our study maps the dynamic FAP response in the ischemic limb at single-cell resolution. We identify distinct transcriptional programs and cellular trajectories that are associated with failed and successful skeletal muscle repair. This provides a resource to investigate FAP mediated mechanisms of skeletal muscle regeneration in response to limb ischemia.

Pressure Regulation of Vascular L-type Calcium Channels

Myogenic tone is dependent upon the activation of L-type Ca2+ channels and the rise in cytosolic [Ca2+]i. While L-type Ca2+ channel activation is often singularly linked to arterial depolarization, sparse evidence suggests that pressure itself may also enhance functional activity, through cooperative gating and trafficking. In this context, this study explored whether intravascular pressure indeed tune the myogenic response by enhancing the L-type Ca2+ channel trafficking and amplifying cooperative gating.Experimentation began in isolated smooth muscle cells (whole cell/single-channel patch clamp; immunohistochemistry; proximity ligation assay) and extended to intact arteries (pressure myography and Ca2+ measurements). Whole-cell electrophysiology revealed the ability of pressure stimuli to increase the L-type Ca2+ current, a response dependent on the intact cytoskeleton and enzymes tied to functional coupling (PKCα, AKAP150, PKA). Cell-attached electrophysiology confirmed whole-cell observations, registering increased coupling among L-type Ca2+ channels when cells were pressurized. Ensuing experiments on isolated arteries subsequently revealed linkage between intravascular pressure and enhanced L-type Ca2+ channel activity, independent of membrane depolarization.In closing, our results suggest there is more to L-type Ca2+ channel regulation than voltage gating and that pressure itself can enhance functional activity. Work continues to precisely define the pressure-sensitive signaling complexes within vascular smooth muscle. Supported by the Canadian Institute for Health Research, National Institute of Health and the Rorabeck Chair in Vascular Biology and Neuroscience. 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.

Death-associated protein kinase 3 regulates the myogenic reactivity of cerebral arteries.

What is the central question of this study? DAPK3 contributes to the Ca2+ -sensitization of vascular smooth muscle contraction: does this protein kinase participate in the myogenic response of cerebral arteries? What is the main finding and its importance? Small molecule inhibitors of DAPK3 effectively block the myogenic responses of cerebral arteries. HS38-dependent changes to vessel constriction occur independent of LC20 phosphorylation, and therefore DAPK3 appears to operate via the actin cytoskeleton. A role for DAPK3 in the myogenic response was not previously reported, and the results support a potential new therapeutic target in the cerebrovascular system. The vascular smooth muscle (VSM) of resistance blood vessels is a target of intrinsic autoregulatory responses to increased intraluminal pressure, the myogenic response. In the brain, the myogenic reactivity of cerebral arteries is critical to homeostatic blood flow regulation. Here we provide the first evidence to link the death-associated protein kinase 3 (DAPK3) to the myogenic response of rat and human cerebral arteries. DAPK3 is a Ser/Thr kinase involved in Ca2+ -sensitization mechanisms of smooth muscle contraction. Ex vivo administration of a specific DAPK3 inhibitor (i.e., HS38) could attenuate vessel constrictions invoked by serotonin as well as intraluminal pressure elevation. The HS38-dependent dilatation was not associated with any change in myosin light chain (LC20) phosphorylation. The results suggest that DAPK3 does not regulate Ca2+ sensitization pathways during the myogenic response of cerebral vessels but rather operates to control the actin cytoskeleton. A slow return of myogenic tone was observed during the sustained ex vivo exposure of cerebral arteries to HS38. Recovery of tone was associated with greater LC20 phosphorylation that suggests intrinsic signalling compensation in response to attenuation of DAPK3 activity. Additional experiments with VSM cells revealed HS38- and siDAPK-dependent effects on the actin cytoskeleton and focal adhesion kinase phosphorylation status. The translational importance of DAPK3 to the human cerebral vasculature was noted, with robust expression of the protein kinase and significant HS38-dependent attenuation of myogenic reactivity found for human pial vessels.