Innervation of adipocytes is limited in mouse perivascular adipose tissue.

Perivascular adipose tissue (PVAT) regulates vascular tone by releasing anticontractile factors. These anticontractile factors are driven by processes downstream of adipocyte stimulation by norepinephrine; however, whether norepinephrine originates from neural innervation or other sources is unknown. The goal of this study was to test the hypothesis that neurons innervating PVAT provide the adrenergic drive to stimulate adipocytes in aortic and mesenteric perivascular adipose tissue (aPVAT and mPVAT), and white adipose tissue (WAT). Healthy male and female mice (8-13 wk) were used in all experiments. Expression of genes associated with synaptic transmission were quantified by qPCR and adipocyte activity in response to neurotransmitters and neuron depolarization was assessed in AdipoqCre+;GCaMP5g-tdTf/WT mice. Immunostaining, tissue clearing, and transgenic reporter lines were used to assess anatomical relationships between nerves and adipocytes. Although synaptic transmission component genes are expressed in adipose tissues (aPVAT, mPVAT, and WAT), strong nerve stimulation with electrical field stimulation does not significantly trigger calcium responses in adipocytes. However, norepinephrine consistently elicits strong calcium responses in adipocytes from all adipose tissues studied. Bethanechol induces minimal adipocyte responses. Imaging neural innervation using various techniques reveals that nerve fibers primarily run alongside blood vessels and rarely branch into the adipose tissue. Although nerve fibers are associated with blood vessels in adipose tissue, they demonstrate limited anatomical and functional interactions with adjacent adipocytes, challenging the concept of classical innervation. These findings dispute the significant involvement of neural input in regulating PVAT adipocyte function and emphasize alternative mechanisms governing adrenergic-driven anticontractile functions of PVAT.NEW & NOTEWORTHY This study challenges prevailing views on neural innervation in perivascular adipose tissue (PVAT) and its role in adrenergic-driven anticontractile effects on vasculature. Contrary to existing paradigms, limited anatomical and functional connections were found between PVAT nerve fibers and adipocytes, underscoring the importance of exploring alternative mechanistic pathways. Understanding the mechanisms involved in PVAT's anticontractile effects is critical for developing potential therapeutic interventions against dysregulated vascular tone, hypertension, and cardiovascular disease.

A cell atlas of thoracic aortic perivascular adipose tissue: a focus on mechanotransducers.

Perivascular adipose tissue (PVAT) is increasingly recognized for its function in mechanotransduction. However, major gaps remain in our understanding of the cells present in PVAT, as well as how different cells contribute to mechanotransduction. We hypothesized that snRNA-seq would reveal the expression of mechanotransducers, and test one (PIEZO1) to illustrate the expression and functional agreement between single-nuclei RNA sequencing (snRNA-seq) and physiological measurements. To contrast two brown tissues, subscapular brown adipose tissue (BAT) was also examined. We used snRNA-seq of the thoracic aorta PVAT (taPVAT) and BAT from male Dahl salt-sensitive (Dahl SS) rats to investigate cell-specific expression mechanotransducers. Localization and function of the mechanostransducer PIEZO1 were further examined using immunohistochemistry (IHC) and RNAscope, as well as pharmacological antagonism. Approximately 30,000 nuclei from taPVAT and BAT each were characterized by snRNA-seq, identifying eight major cell types expected and one unexpected (nuclei with oligodendrocyte marker genes). Cell-specific differential gene expression analysis between taPVAT and BAT identified up to 511 genes (adipocytes) with many (≥20%) being unique to individual cell types. Piezo1 was the most highly, widely expressed mechanotransducer. The presence of PIEZO1 in the PVAT but not the adventitia was confirmed by RNAscope and IHC in male and female rats. Importantly, antagonism of PIEZO1 by GsMTX4 impaired the PVAT's ability to hold tension. Collectively, the cell compositions of taPVAT and BAT are highly similar, and PIEZO1 is likely a mechanotransducer in taPVAT.NEW & NOTEWORTHY This study describes the atlas of cells in the thoracic aorta perivascular adipose tissue (taPVAT) of the Dahl-SS rat, an important hypertension model. We show that mechanotransducers are widely expressed in these cells. Moreover, PIEZO1 expression is shown to be restricted to the taPVAT and is functionally implicated in stress relaxation. These data will serve as the foundation for future studies investigating the role of taPVAT in this model of hypertensive disease.

Perivascular Adipose Tissue Remodels Only after Elevation of Blood Pressure in the Dahl SS Rat Fed a High-Fat Diet.

INTRODUCTION: Tunica media extracellular matrix (ECM) remodeling is well understood to occur in response to elevated blood pressure, unlike the remodeling of other tunicas. We hypothesize that perivascular adipose tissue (PVAT) is responsive to hypertension and remodels as a protective measure. METHODS: The adventitia and PVAT of the thoracic aorta were used in measuring ECM genes from 5 pairs of Dahl SS male rats on 8 or 24 weeks of feeding from weaning on a control (10% Kcal fat) or high-fat (HF; 60%) diet. A PCR array of ECM genes was performed with cDNA from adventitia and PVAT after 8 and 24 weeks. A gene regulatory network of the differentially expressed genes (DEGs) (HF 2-fold > con) was created using Cytoscape. RESULTS: After 8 weeks, 29 adventitia but 0 PVAT DEGs were found. By contrast, at 24 weeks, PVAT possessed 47 DEGs while adventitia had 3. Top DEGs at 8 weeks in adventitia were thrombospondin 1 and collagen 8a1. At 24 weeks, thrombospondin 1 was also a top DEG in PVAT. The transcription factor Adarb1 was identified as a regulator of DEGs in 8-week adventitia and 24-week PVAT. CONCLUSION: These data support that PVAT responds biologically once blood pressure is elevated.

ß-arrestin biased signaling is not involved in the hypotensive actions of 5-HT7 receptor stimulation: use of Serodolin.

The 5-hydroxytryptamine 7 receptor (5-HT7) is necessary for 5-HT to cause a concentration-dependent vascular relaxation and hypotension. 5-HT7 is recognized as having biased signaling, transduced through either Gs or ß -arrestin. It is unknown whether 5-HT7 signals in a biased manner to cause vasorelaxation/hypotension. We used the recently described ß-arrestin selective 5-HT7 receptor agonist serodolin to test the hypothesis that 5-HT7 activation does not cause vascular relaxation or hypotension via the ß -arrestin pathway. Isolated abdominal aorta (no functional 5-HT7) and vena cava (functional 5-HT7) from male Sprague Dawley rats were used in isometric contractility studies. Serodolin (1 nM - 10 µM) did not change baseline tone of isolated tissues and did not relax the endothelin-1 (ET-1)-contracted vena cava or aorta. In the aorta, serodolin acted as a 5-HT2A receptor antagonist, evidenced by a rightward shift in 5-HT-induced concentration response curve [pEC50 5-HT [M]: Veh = 5.2 +/- 0.15; Ser (100 nM) = 4.49 +/- 0.08; p < 0.05]. In the vena cava, serodolin acted as a 5-HT7 receptor antagonist, shifting the concentration response curve to 5-HT left and upward (%10 µM NE contraction; Veh = 3.2 +/- 1.7; Ser (10 nM) = 58 +/- 11; p < 0.05) and blocking relaxation of pre-contracted tissue to the 5-HT1A/7 agonist 5-carboxamidotryptamine. In anesthetized rats, 5-HT or serodolin was infused at 5, 25 and 75 µg/kg/min, iv. Though 5-HT caused concentration-dependent depressor responses, serodolin caused an insignificant small depressor responses at all three infusion rates. With the final dose of serodolin on board, 5-HT was unable to reduce blood pressure. Collectively the data indicate that serodolin functions as a 5-HT7 antagonist with additional 5-HT2A blocking properties. 5-HT7 activation does not cause vascular relaxation or hypotension via the ß -arrestin pathway.

Integrins play a role in stress relaxation of perivascular adipose tissue.

Perivascular adipose tissue (PVAT) is known for being anti-contractile in healthy tissues. We discovered a new function of PVAT, the ability to stress relax and maintain a tone in response to a stretch. This is of note because stress relaxation has been attributed to smooth muscle, of which PVAT has none that is organized in a functional layer. We test the hypothesis the interactions of integrins with collagen play a role in stress relaxation. Our model is the thoracic aorta of the male Dahl SS rat. The PVAT and aorta were physically separated for most assays. Results from single nuclei RNA sequencing (snRNAseq) experiments, histochemistry and isometric contractility were also used. Masson Trichrome staining made evident the expression of collagen in PVAT. From snRNA seq experiments of the PVAT, mRNA for multiple collagen and integrin isoforms were detected: the α1 and ß1 integrin were most highly expressed. Pharmacological inhibition of integrin/collagen interaction was effected by the specific α1ß1 distintegrin obtustatin or general integrin inhibitor RGD peptide. RGD peptide but not obtustatin increased the stress relaxation. Cell-cell communication inference identified integrins αv and α5, two major RGD motif containing isoforms, as potential signaling partners of collagens. Collectively, these findings validate that stress relaxation can occur in a non-smooth muscle tissue, doing so in part through integrin-collagen interactions that may not include α1ß1 heterodimers. The importance of this lies in considering PVAT as a vascular layer that possesses mechanical functions.

International Union of Basic and Clinical Pharmacology. CX. Classification of Receptors for 5-hydroxytryptamine; Pharmacology and Function.

5-HT receptors expressed throughout the human body are targets for established therapeutics and various drugs in development. Their diversity of structure and function reflects the important role 5-HT receptors play in physiologic and pathophysiological processes. The present review offers a framework for the official receptor nomenclature and a detailed understanding of each of the 14 5-HT receptor subtypes, their roles in the systems of the body, and, where appropriate, the (potential) utility of therapeutics targeting these receptors. SIGNIFICANCE STATEMENT: This review provides a comprehensive account of the classification and function of 5-hydroxytryptamine receptors, including how they are targeted for therapeutic benefit.

Chemerin is resident to vascular tunicas and contributes to vascular tone.

The adipokine chemerin may support blood pressure, evidenced by a fall in mean arterial pressure after whole body antisense oligonucleotide (ASO)-mediated knockdown of chemerin protein in rat models of normal and elevated blood pressure. Although the liver is the greatest contributor of circulating chemerin, liver-specific ASOs that abolished hepatic-derived chemerin did not change blood pressure. Thus, other sites must produce the chemerin that supports blood pressure. We hypothesize that the vasculature is a source of chemerin independent of the liver that supports arterial tone. RNAScope, PCR, Western blot analyses, ASOs, isometric contractility, and radiotelemetry were used in the Dahl salt-sensitive (SS) rat (male and female) on a normal diet. Retinoic acid receptor responder 2 (Rarres2) mRNA was detected in the smooth muscle, adventitia, and perivascular adipose tissue of the thoracic aorta. Chemerin protein was detected immunohistochemically in the endothelium, smooth muscle cells, adventitia, and perivascular adipose tissue. Chemerin colocalized with the vascular smooth muscle marker α-actin and the adipocyte marker perilipin. Importantly, chemerin protein in the thoracic aorta was not reduced when liver-derived chemerin was abolished by a liver-specific ASO against chemerin. Chemerin protein was similarly absent in arteries from a newly created global chemerin knockout in Dahl SS rats. Inhibition of the receptor Chemerin1 by the receptor antagonist CCX832 resulted in the loss of vascular tone that supports potential contributions of chemerin by both perivascular adipose tissue and the media. These data suggest that vessel-derived chemerin may support vascular tone locally through constitutive activation of Chemerin1. This posits chemerin as a potential therapeutic target in blood pressure regulation.NEW & NOTEWORTHY Vascular tunicas synthesizing chemerin is a new finding. Vascular chemerin is independent of hepatic-derived chemerin. Vasculature from both males and females have resident chemerin. Chemerin1 receptor activity supports vascular tone.

5-HT7 receptors mediate dilation of rat cremaster muscle arterioles in vivo.

OBJECTIVE: Serotonin (5-HT) infusion in vivo causes hypotension and a fall in total peripheral resistance. However, the vascular segment and the receptors that mediate this response remain in question. We hypothesized that 5-HT7 receptors mediate arteriolar dilation to 5-HT in skeletal muscle microcirculation. METHODS: Cremaster muscles of isoflurane-anesthetized male Sprague-Dawley rats were prepared for in vivo microscopy of third- and fourth-order arterioles and superfused with physiological salt solution at 34°C. Quantitative real-time PCR (RT-PCR) was applied to pooled samples of first- to third-order cremaster arterioles (2-4 rats/sample) to evaluate 5-HT7 receptor expression. RESULTS: Topical 5-HT (1-10 nmols) or the 5-HT1/7 receptor agonist, 5-carboxamidotryptamine (10-30 nM), dilated third- and fourth-order arterioles, responses that were abolished by 1 µM SB269970, a selective 5-HT7 receptor antagonist. In contrast, dilation induced by the muscarinic agonist, methacholine (100 nmols) was not inhibited by SB269970. Serotonin (10 nmols) failed to dilate cremaster arterioles in 5-HT7 receptor knockout rats whereas arterioles in wild-type litter mates dilated to 1 nmol 5-HT, a response blocked by 1 µM SB269970. Quantitative RT-PCR revealed that cremaster arterioles expressed mRNA for 5-HT7 receptors. CONCLUSIONS: 5-HT7 receptors mediate dilation of small arterioles in skeletal muscle and likely contribute to 5-HT-induced hypotension, in vivo.

Higher OAK (Oral Anticoagulation Knowledge) score at baseline associated with better TTR (Time in Therapeutic Range) in patients taking warfarin.

A lack in patient knowledge of warfarin therapy is associated with poor adherence. This knowledge gap may result in a lower INR Time in Therapeutic Range (TTR). To investigate association between patient anticoagulation knowledge and warfarin control. Michigan Anticoagulation Quality Improvement Initiative (MAQI2) is a Blue Cross Blue Shield of Michigan sponsored consortium of six anticoagulation management services. Patients prescribed warfarin at two MAQI2 sites completed a voluntary Oral Anticoagulation Knowledge (OAK) questionnaire at warfarin initiation and 6-month follow-up. The results of 20 OAK questions and TTRs (excluding 1st month post-initiation) were compared using chi-square tests, t-tests and multivariate analysis adjusting for SAMe-TT2R2 and days on warfarin. Of 1836 surveys distributed at warfarin initiation, 481 (26.2%) patients completed the baseline questionnaire (within 1 month post-initiation): mean OAK score: 14.6 ± 3.4. Of those, 147 (30.6%) completed 6-month follow-up surveys (OAK: 12.7 ± 5.8). Patients with TTR ≥ 70% at baseline scored higher on OAK tests than patients with TTR < 70% in unadjusted analyses (15.1 ± 3.2 v. 14.2 ± 3.5, p = 0.003) and adjusted analysis (p = 0.020). There was no unadjusted or adjusted difference in OAK scores at 6-month follow-up between patients with TTR ≥ 70% and TTR < 70%. For patients who completed baseline and follow-up surveys, there was a decrease of 2.4 points in OAK score between baseline and 6-month follow up (p < 0.001). Higher baseline, but not follow-up, OAK score is associated with better warfarin control and average OAK scores decreased between baseline and follow-up. Further studies are needed to determine what type of patient education may improve patient knowledge retention and warfarin control.

The moderating and mediating role of telepresence and cognitive change in cognitive behaviour therapy delivered via videoconference.

In this study, we combined the results of two controlled trials and examined the relationships between working alliance, telepresence, cognitive change and treatment outcome. Sixty-five participants with a primary diagnosis of generalized anxiety disorder (GAD) or panic disorder with agoraphobia (PDA) received cognitive behaviour therapy delivered via videoconference. Participants completed measures of working alliance and telepresence after three psychotherapy sessions. They also completed measures of treatment outcome and dysfunctional beliefs (cognitive change) specific to PDA and GAD at pretreatment and posttreatment. Results revealed that telepresence at the fifth session moderated the relationship between the working alliance at the first and fifth sessions. As telepresence increased, its impact on the working alliance diminished. Cognitive change mediated the relationship between the working alliance at the fifth session and treatment outcome.

Aortic stiffness is lower when PVAT is included: a novel ex vivo mechanics study.

Perivascular adipose tissue (PVAT) is increasingly recognized as an essential layer of the functional vasculature, being responsible for producing vasoactive substances and assisting arterial stress relaxation. Here, we test the hypothesis that PVAT reduces aortic stiffness. Our model was the thoracic aorta of the male Sprague-Dawley rat. Uniaxial mechanical tests for three groups of tissue were performed: aorta with PVAT attached (+PVAT) or removed (-PVAT), and isolated PVAT (PVAT only). The output of the mechanical test is reported in the form of a Cauchy stress-stretch curve. This work presents a novel, physiologically relevant approach to measure mechanical stiffness ex vivo in isolated PVAT. Low-stress stiffness (E0), high-stress stiffness (E1), and the stress corresponding to a stretch of 1.2 (σ1.2) were measured as metrics of distensibility. The low-stress stiffness was largest in the -PVAT samples and smallest in PVAT only samples. Both the high-stress stiffness and the stress at 1.2 stretch were significantly higher in -PVAT samples when compared with +PVAT samples. Taken together, these results suggest that -PVAT samples are stiffer (less distensible) both at low stress (not significant) as well as at high stress (significant) when compared with +PVAT samples. These conclusions are supported by the results of the continuum mechanics material model that we also used to interpret the same experimental data. Thus, tissue stiffness is significantly lower when considering PVAT as part of the aortic wall. As such, PVAT should be considered as a target for improving vascular function in diseases with elevated aortic stiffness, including hypertension.NEW & NOTEWORTHY We introduce a novel and physiologically relevant way of measuring perivascular adipose tissue (PVAT) mechanical stiffness which shows that PVAT's low, yet measurable, stiffness is linearly correlated with the amount of collagen fibers present within the tissue. Including PVAT in the measurement of the aortic wall's mechanical behavior is important, and it significantly affects the resulting metrics by decreasing aortic stiffness.

Identification of Piezo1 channels in perivascular adipose tissue (PVAT) and their potential role in vascular function.

The vasculature constantly experiences distension/pressure exerted by blood flow and responds to maintain homeostasis. We hypothesized that activation of the stretch sensitive, non-selective cation channel Piezo1 would directly increase vascular contraction in a way that might be modified by perivascular adipose tissue (PVAT). The presence and function of Piezo1 was investigated by RT-PCR, immunohistochemistry, and isolated tissue bath contractility. Superior and mesenteric resistance arteries, aortae, and their PVATs from male Sprague Dawley rats were used. Piezo1 mRNA was detected in aortic vessels, aortic PVAT, mesenteric vessels, and mesenteric PVAT. Both adipocytes and stromal vascular fraction of mesenteric PVAT expressed Piezo1 mRNA. In PVAT, expression of Piezo1 mRNA was greater in magnitude than that of Piezo2, transient receptor potential cation channel, subfamily V, member 4 (TRPV4), anoctamin 1, calcium activated chloride channel (TMEM16), and Pannexin1 (Panx1). Piezo1 protein was present in endothelium and PVAT of rat aortic and in PVAT of mesenteric artery. The Piezo1 agonists Yoda1 and Jedi2 (1 nM - 10 µM) did not stimulate aortic contraction [max < 10% phenylephrine (PE) 10 µM contraction] or relaxation in tissues + or -PVAT. Depolarizing the aorta by modestly elevated extracellular K+ did not unmask aortic contraction to Yoda1 (max <10% PE 10 µM contraction). Finally, the Piezo1 antagonist Dooku1 did not modify PE-induced aorta contraction + or -PVAT. Surprisingly, Dooku1 directly caused aortic contraction in the absence (Dooku1 =26 ± 11; Vehicle = 11 ± 11%PE contraction) but not in the presence of PVAT (Dooku1 = 2 ± 1; Vehicle = 8 ± 5% PE contraction). Thus, Piezo1 is present and functional in the isolated rat aorta but does not serve direct vascular contraction with or without PVAT. We reaffirmed the isolated mouse aorta relaxation to Yoda1, indicating a species difference in Piezo1 activity between mouse and rat.

Is the 5-hydroxytryptamine 7 Receptor Constitutively Active in the Vasculature? A Study in Veins/Vein.

ABSTRACT: The 5-hydroxytryptamine 7 (5-HT 7 ) receptor is reported to have considerable constitutive activity when transfected into cells. Constitutive activity-receptor activity in the absence of known agonist-is important for understanding the contributions of a receptor to (patho)physiology. We test the hypothesis that the 5-HT 7 receptor possesses constitutive activity in a physiological situation. Isolated veins from male and female Sprague Dawley rats were used as models for measuring isometric force; the abdominal vena cava possesses a functional 5-HT 7 receptor that mediates relaxation, whereas the small mesenteric vein does not. Compounds reported to act as inverse agonists were investigated for their ability to cause contraction (moving a constitutively active relaxant receptor to an inactive state, removing relaxation). Compared with a vehicle control, clozapine, risperidone, ketanserin, and SB269970 caused no contraction in the isolated male abdominal vena cava. By contrast, methiothepin caused a concentration-dependent contraction of the male but not female abdominal vena cava, although with low potency (-log EC 50 [M] = 5.50 ± 0.45) and efficacy (∼12% of contraction to endothelin-1). Methiothepin-induced contraction was not reduced by the 5-HT 7 receptor antagonist (SB269970, 1 µM, not active in the vena cava). These same compounds showed little to no effect in the isolated mesenteric vein. We conclude that the 5-HT 7 receptor in the isolated veins of the Sprague Dawley rat does not possess constitutive activity. We raise the question of the physiological relevance of constitutive activity of this receptor important to such diverse physiological functions as sleep, circadian rhythm, temperature, and blood pressure regulation.

Retrospective Assessment of Patient and Catheter Characteristics Associated With Malpositioned Central Venous Catheters in Pediatric Patients.

OBJECTIVES: The primary objective was to determine the prevalence and characteristics associated with malpositioned temporary, nontunneled central venous catheters (CVCs) placed via the internal jugular (IJ) and subclavian (SC) veins in pediatric patients. DESIGN: Single-center retrospective cohort study. SETTING: Quaternary academic PICU. PATIENTS: Children greater than 1 month to less than 18 years who had a CVC placed between January 2014 and December 2018. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: The primary outcome was the CVC tip position located on the first postprocedural radiograph. CVC tip was defined as follows: "recommended" (tip location between the carina and two vertebral bodies inferior to the carina), "high" (tip location between one and four vertebral bodies superior to the carina), "low" (tip position three or more vertebral bodies inferior to the carina), and "other" (tip grossly malpositioned). Seven hundred eighty-one CVCs were included: 481 (61.6%) were in "recommended" position, 157 (20.1%) were "high," 131 (16.8%) were "low," and 12 (1.5%) were "other." Multiple multinomial regression (referenced to "recommended" position) showed that left-sided catheters (adjusted odds ratio [aOR], 2.00, 95% CI 1.17-3.40) were associated with "high" CVC tip positions, whereas weight greater than or equal to 40 kg had decreased odds of having a "high" CVC tip compared with the reference (aOR, 0.45; 95% CI, 0.24-0.83). Further, weight category 20-40 kg (aOR, 2.42; 95% CI, 1.38-4.23) and females (aOR, 1.51; 95% CI, 1.01-2.26) were associated with "low" CVC tip positions. There was no difference in rates of central line-associated blood stream infection, venous thromboembolism, or tissue plasminogen activator usage or dose between the CVCs with tips outside and those within the recommended location. CONCLUSIONS: The prevalence of IJ and SC CVC tips outside of the recommended location was high. Left-sided catheters, patient weight, and sex were associated with malposition. Malpositioned catheters were not associated with increased harm.

Male and female high-fat diet-fed Dahl SS rats are largely protected from vascular dysfunctions: PVAT contributions reveal sex differences.

Vascular dysfunctions are observed in the arteries from hypertensive subjects. The establishment of the Dahl salt-sensitive (SS) male and female rat models to develop a reproducible hypertension with high-fat (HF) diet feeding from weaning allows addressing the question of whether HF diet-associated hypertension results in vascular dysfunction similar to that of essential hypertension in both sexes. We hypothesized that dysfunction of three distinct vascular layers, i.e., endothelial, smooth muscle, and perivascular adipose tissue (PVAT), would be present in the aorta from HF diet-fed versus control diet-fed male and female rats. Dahl SS rats were fed a control (10% kcal of fat) or HF (60%) diet from weaning for 24 wk. Male and female Dahl SS rats became equally hypertensive when placed on a HF diet. For male and female rats, the thoracic aorta exhibited medial hypertrophy in HF diet-induced hypertension versus control, but neither displayed a hyperresponsive contraction to the α-adrenergic agonist phenylephrine nor an endothelial cell dysfunction as measured by acetylcholine-induced relaxation. A beneficial PVAT function, support of stress relaxation, was reduced in the male versus female rats fed a HF diet. PVAT in the aorta of males but not in females retained the anticontractile activity. We conclude that this HF model does not display the same vascular dysfunctions observed in essential hypertension. Moreover, both male and female show significantly different vascular dysfunctions in this HF feeding model.NEW & NOTEWORTHY Although the aorta exhibits medial hypertrophy in response to HF diet-induced hypertension, it did not exhibit hyperresponsive contraction to an α-adrenergic agonist nor endothelial cell dysfunction; this was true for both sexes. Unlike other hypertension models, PVAT around aorta from (male) rats on the HF diet retained significant anticontractile activity. PVAT around aorta of the male on a HF diet was modestly more fibrotic and lost the ability to assist in arterial stress relaxation.

Guidelines for the measurement of vascular function and structure in isolated arteries and veins.

The measurement of vascular function in isolated vessels has revealed important insights into the structural, functional, and biomechanical features of the normal and diseased cardiovascular system and has provided a molecular understanding of the cells that constitutes arteries and veins and their interaction. Further, this approach has allowed the discovery of vital pharmacological treatments for cardiovascular diseases. However, the expansion of the vascular physiology field has also brought new concerns over scientific rigor and reproducibility. Therefore, it is appropriate to set guidelines for the best practices of evaluating vascular function in isolated vessels. These guidelines are a comprehensive document detailing the best practices and pitfalls for the assessment of function in large and small arteries and veins. Herein, we bring together experts in the field of vascular physiology with the purpose of developing guidelines for evaluating ex vivo vascular function. By using this document, vascular physiologists will have consistency among methodological approaches, producing more reliable and reproducible results.

Vascular reactivity stimulated by TMA and TMAO: Are perivascular adipose tissue and endothelium involved?

Trimethylamine (TMA), formed by intestinal microbiota, and its Flavin-Monooxygenase 3 (FMO3) product Trimethylamine-N-Oxide (TMAO), are potential modulators of host cardiometabolic phenotypes. High circulating levels of TMAO are associated with increased risk for cardiovascular diseases. We hypothesized that TMA/TMAO could directly change the vascular tone. Perivascular adipose tissue (PVAT) helps to regulate vascular homeostasis and may also possess FMO3. Thoracic aorta with(+) or without(-) PVAT, also + or - the endothelium (E), of male Sprague Dawley rats were isolated for measurement of isometric tone in response to TMA/TMAO (1nM-0.5 M). Immunohistochemistry (IHC) studies were done to identify the presence of FMO3. TMA and TMAO elicited concentration-dependent arterial contraction. However, at a maximally achievable concentration (0.2 M), contraction stimulated by TMA was of a greater magnitude (141.5 ± 16% of maximum phenylephrine contraction) than that elicited by TMAO (19.1 ± 4.03%) with PVAT and endothelium intact. When PVAT was preserved, TMAO-induced contraction was extensively reduced the presence (19.1 ± 4.03%) versus absence of E (147.2 ± 20.5%), indicating that the endothelium plays a protective role against TMAO-induced contraction. FMO3 enzyme was present in aortic PVAT, but the FMO3 inhibitor methimazole did not affect contraction stimulated by TMA in aorta + PVAT. However, the l-type calcium channel blocker nifedipine reduced TMA-induced contraction by ∼50% compared to the vehicle. Though a high concentration of these compounds was needed to achieve contraction, the findings that TMA-induced contraction was independent of PVAT and E and mediated by nifedipine-sensitive calcium channels suggest metabolite-induced contraction may be physiologically important.

5-HT7 Receptor Restrains 5-HT-induced 5-HT2A Mediated Contraction in the Isolated Abdominal Vena Cava.

ABSTRACT: Although discovered as a vasoconstrictor, 5-hydroxytryptamine (5-HT, serotonin) infused into man and rodent reduces blood pressure. This occurs primarily through activation of 5-HT7 receptors and, at least in part, venodilation. Vascular mechanisms by which this could occur include direct receptor activation leading to vasodilation and/or suppression of contractile 5-HT receptor activation. This study tests the hypothesis that the 5-HT7 receptor restrains activation of the 5-HT2A receptor. A subhypothesis is whether agonist-induced activation-independent of constitutive activity-of the 5-HT7 receptor is necessary for this restraint. The isolated abdominal aorta and vena cava from the normal male Sprague-Dawley rat was our model. Studies used real-time PCR and a pharmacological approach in the isolated tissue bath for measurement of isometric tone. Although 5-HT2A receptor mRNA expression in both aorta and vena cava was significantly larger than that of the 5-HT7 receptor mRNA, the 5-HT7/5-HT2A receptor mRNA ratio was greater in the vena cava (0.30) than in the aorta (0.067). 5-HT7 receptor antagonism by SB266970 and DR 4458 increased maximum contraction to 5-HT in the isolated vein by over 50% versus control. The 5-HT2A receptor agonists TCB-2 and NBOH were more potent in the aorta compared with 5-HT but less efficacious, serving as partial agonists. By contrast, these same three agonists caused no contraction in the vena cava isolated from the same rats up to 10 µM agonist. Antagonism of the 5-HT7 receptor by SB269970 did not increase either the potency or efficacy of TCB-2 or NBOH. These data support that the 5-HT7 receptor itself needs to be stimulated to reduce contraction and suggest there is little constitutive activity of the 5-HT7 receptor in the isolate abdominal vena cava.

Diagnostic Accuracy of an Esophageal Screening Protocol Interpreted by the Speech-Language Pathologist.

Oropharyngeal and esophageal dysphagia may occur simultaneously. However, symptoms are often evaluated separately. Few standardized, multi-texture esophageal screening protocols exist as an addition to the modified barium swallow study (MBSS). Given the gap in MBSS evaluation standards, providers may be lacking information needed to fully assess the swallowing process and create appropriate dysphagia management plans. The aim was to assess the diagnostic accuracy of a standardized esophageal screening protocol performed by an SLP compared to formal reference esophageal examinations. A cross-sectional analytic study was performed. Consecutively referred patients who underwent same-day consultation with the SLP and a gastroenterologist were included. MBSS with a standardized esophageal screen was performed. Same-day formal esophageal testing was completed and included timed barium emptying study or high-resolution manometry. Summary diagnostic accuracy measures were calculated. Seventy-three patients matched the inclusion criteria. Median age was 62.5 years (25-87), 55% were female. Sensitivity of the esophageal screen for the detection of esophageal abnormality was 83.7% (95% CI 70-91.9%); specificity was 73.7% (95% CI 55.6-85.8%). The positive likelihood ratio was 3.14 (95% CI 1.71-5.77), whereas the negative likelihood ratio was 0.22 (95% CI 0.11-0.45). Positive and negative predictive values were 82% and 76%, respectively. Use of a systematic, multi-texture esophageal screen protocol interpreted by SLPs accurately identifies multiphase dysphagia and should be considered in addition to standard MBSS testing. Inclusion of a cursory esophageal view may more adequately assess dysphagia symptoms and help to promote multidisciplinary care.

Transglutaminases Are Active in Perivascular Adipose Tissue.

Transglutaminases (TGs) are crosslinking enzymes best known for their vascular remodeling in hypertension. They require calcium to form an isopeptide bond, connecting a glutamine to a protein bound lysine residue or a free amine donor such as norepinephrine (NE) or serotonin (5-HT). We discovered that perivascular adipose tissue (PVAT) contains significant amounts of these amines, making PVAT an ideal model to test interactions of amines and TGs. We hypothesized that transglutaminases are active in PVAT. Real time RT-PCR determined that Sprague Dawley rat aortic, superior mesenteric artery (SMA), and mesenteric resistance vessel (MR) PVATs express TG2 and blood coagulation Factor-XIII (FXIII) mRNA. Consistent with this, immunohistochemical analyses support that these PVATs all express TG2 and FXIII protein. The activity of TG2 and FXIII was investigated in tissue sections using substrate peptides that label active TGs when in a catalyzing calcium solution. Both TG2 and FXIII were active in rat aortic PVAT, SMAPVAT, and MRPVAT. Western blot analysis determined that the known TG inhibitor cystamine reduced incorporation of experimentally added amine donor 5-(biotinamido)pentylamine (BAP) into MRPVAT. Finally, experimentally added NE competitively inhibited incorporation of BAP into MRPVAT adipocytes. Further studies to determine the identity of amidated proteins will give insight into how these enzymes contribute to functions of PVAT and, ultimately, blood pressure.