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.

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.

Renal and hypothalamic inflammation in renovascular hypertension: role of afferent renal nerves.

Renal denervation (RDN) is a potential therapy for drug-resistant hypertension. However, whether its effects are mediated by ablation of efferent or afferent renal nerves is not clear. Previous studies have implicated that renal inflammation and the sympathetic nervous system are driven by the activation of afferent and efferent renal nerves. RDN attenuated the renal inflammation and sympathetic activity in some animal models of hypertension. In the 2 kidney,1 clip (2K1C) model of renovascular hypertension, RDN also decreased sympathetic activity; however, mechanisms underlying renal and central inflammation are still unclear. We tested the hypothesis that the mechanisms by which total RDN (TRDN; efferent + afferent) and afferent-specific RDN (ARDN) reduce arterial pressure in 2K1C rats are the same. Male Sprague-Dawley rats were instrumented with telemeters to measure mean arterial pressure (MAP), and after 7 days, a clip was placed on the left renal artery. Rats underwent TRDN, ARDN, or sham surgery of the clipped kidney and MAP was measured for 6 wk. Weekly measurements of water intake (WI), urine output (UO), and urinary copeptin were conducted, and urine was analyzed for cytokines/chemokines. Neurogenic pressor activity (NPA) was assessed at the end of the protocol calculated by the depressor response after intraperitoneal injection of hexamethonium. Rats were euthanized and the hypothalamus and kidneys removed for measurement of cytokine content. MAP, NPA, WI, and urinary copeptin were significantly increased in 2K1C-sham rats, and these responses were abolished by both TRDN and ARDN. 2K1C-sham rats presented with renal and hypothalamic inflammation and these responses were largely mitigated by TRDN and ARDN. We conclude that RDN attenuates 2K1C hypertension primarily by ablation of afferent renal nerves which disrupts bidirectional renal neural-immune pathways.NEW & NOTEWORTHY Hypertension resulting from reduced perfusion of the kidney is dependent on renal sensory nerves, which are linked to inflammation in the kidney and hypothalamus. Afferent renal nerves are required for chronic increases in both water intake and vasopressin release observed following renal artery stenosis. Findings from this study suggest an important role of renal sensory nerves that has previously been underestimated in the pathogenesis of 2K1C hypertension.

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.

Skin Vascular Resistance Decreases during 5-HT-Induced Hypotension in the Rat.

A recognized vasodilator, the infusion of 5-hydroxytryptamine (5-HT, serotonin) decreases blood pressure through the reduction of total peripheral resistance in the rat. It is not clear which vascular beds/tissues are responsible for this fall. We hypothesized that an increase in blood flow within the skin, measured as an elevated temperature (T) in the thermoregulatory tail and paws, enables at least part of 5-HT-induced reduction in blood pressure through active vasodilation. The temperature of thermoregulatory regions of the skin of an anesthetized male, Sprague Dawley rats were measured using a Optris PI640 thermal camera. The blood pressure of the animal and the temperature of each paw and four locations along the tail (TL1-4) were recorded before, during, and after the infusion of 5-HT at a rate of 25 mg/min into a femoral vein. Contrary to our hypothesis, the temperature of the paws and tail was stable before and during 5-HT infusion and actually increased during the 15-min recovery period. This finding suggests that hyperemia of the skin circulation is not necessary for the fall in blood pressure observed with infused 5-HT, but that a reduction in cutaneous vascular resistance plays a part in the fall in total peripheral resistance.

A high fat diet does not stimulate blood pressure dependence on chemerin in the Sprague-Dawley rat.

The adipokine chemerin is a candidate for connecting obesity to hypertension. Study objective: To test the hypothesis that a high fat (HF) diet stimulates dependence on chemerin for blood pressure regulation. Design: Blood pressure in male Sprague Dawley rats fed a control (10 % fat) or HF (60 % fat) diet from weaning was measured using radiotelemetry. Antisense oligonucleotides (ASOs), administered after 17 weeks of feeding, were used to abolish chemerin production. Results: The HF diet did not increase blood pressure (mm Hg; control = 117.0 ± 2.5; HF = 122.0 ± 2.2). An ASO against chemerin (dosed 1×/week, 4 weeks) similarly reduced blood pressure in the control (-14.0 ± 2.7 mmHg) and HF rat (-12.4 ± 2.3). Chemerin mRNA was abolished in the liver and fats (primary producers of chemerin) from rats given the ASO chemerin vs control. Conclusion: A HF diet alone is insufficient to stimulate the dependence of blood pressure in the rat on chemerin.

Reduction in Hindquarter Vascular Resistance Supports 5-HT7 Receptor Mediated Hypotension.

The 5-HT7 receptor is the primary mediator of both the acute (

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.

Phenotypic Changes in T Cell and Macrophage Subtypes in Perivascular Adipose Tissues Precede High-Fat Diet-Induced Hypertension.

Perivascular adipose tissue (PVAT) may connect adiposity to hypertension because of its vasoactive functions and proximity to blood vessels. We hypothesized that immune cell changes in PVATs precede the development of high fat diet (HFD)-induced hypertension. Both sexes of Dahl S rat become equally hypertensive when fed a HFD. Further, both sexes would have similar immune cell composition in PVATs with the development and progression of hypertension. Male and female Dahl S rats were fed a regular (10% calories from fat; CD) diet or a HFD (60%) from weaning. PVATs from around the thoracic aorta (APVAT) and small mesenteric vessels (MRPVAT) were harvested at 10 weeks (pre-hypertensive), 17 weeks (onset), or 24 (hypertensive) weeks on diet. RNA-sequencing in MRPVAT at 24 weeks indicated sex-differences with HFD (>CD) and diet-differences in males (>females). The top 2 out of 7 immune processes with the maximum number of differentially expressed genes (DEGs) were associated with immune effector processes and leukocyte activation. Macrophages and T cells (and their activation status), neutrophils, mast, B and NK cells were measured by flow cytometry. Sex-specific changes in the number of CD4 memory T cells (males > females) and M2-like macrophages (females > males) in PVATs occur with a HFD before hypertension developed. Sex-differences became more prominent with the development and progression of hypertension, driven by the diet (HFD > CD). These findings suggest that though the magnitudes of increased blood pressure were equivalent in both sexes, the associated phenotypic changes in the immune subsets within the PVATs were different in the male vs. the female with the development and progression of hypertension.

Renal Denervation and Celiac Ganglionectomy Decrease Mean Arterial Pressure Similarly in Genetically Hypertensive Schlager (BPH/2J) Mice.

Renal denervation (RDNX) lowers mean arterial pressure (MAP) in patients with resistant hypertension. Less well studied is the effect of celiac ganglionectomy (CGX), a procedure which involves the removal of the nerves innervating the splanchnic vascular bed. We hypothesized that RDNX and CGX would both lower MAP in genetically hypertensive Schlager (BPH/2J) mice through a reduction in sympathetic tone. Telemeters were implanted into the femoral artery in mice to monitor MAP before and after RDNX (n=5), CGX (n=6), or SHAM (n=6). MAP, systolic blood pressure, diastolic blood pressure, and heart rate were recorded for 14 days postoperatively. The MAP response to hexamethonium (10 mg/kg, IP) was measured on control day 3 and postoperative day 10 as a measure of global neurogenic pressor activity. The efficacy of denervation was assessed by measurement of tissue norepinephrine. Control MAP was similar among the 3 groups before surgical treatments (≈130 mm Hg). On postoperative day 14, MAP was significantly lower in RDNX (-11±2 mm Hg) and CGX (-11±1 mm Hg) groups compared with their predenervation values. This was not the case in SHAM mice (-5±3 mm Hg). The depressor response to hexamethonium in the RDNX group was significantly smaller on postoperative day 10 (-10±5 mm Hg) compared with baseline control (-25±10 mm Hg). This was not the case in mice in the SHAM (day 10; -28±5 mm Hg) or CGX (day 10; -34±7 mm Hg) group. In conclusion, both renal and splanchnic nerves contribute to hypertension in BPH/2J mice, but likely through different mechanisms.

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.

Potential role of intermittent functioning of baroreflexes in the etiology of hypertension in spontaneously hypertensive rats.

The spontaneously hypertensive rat (SHR) is a genetic model of primary hypertension with an etiology that includes sympathetic overdrive. To elucidate the neurogenic mechanisms underlying the pathophysiology of this model, we analyzed the dynamic baroreflex response to spontaneous fluctuations in arterial pressure in conscious SHRs, as well as in the Wistar-Kyoto (WKY), the Dahl salt-sensitive, the Dahl salt-resistant, and the Sprague-Dawley rat. Observations revealed the existence of long intermittent periods (lasting up to several minutes) of engagement and disengagement of baroreflex control of heart rate. Analysis of these intermittent periods revealed a predictive relationship between increased mean arterial pressure and progressive baroreflex disengagement that was present in the SHR and WKY strains but absent in others. This relationship yielded the hypothesis that a lower proportion of engagement versus disengagement of the baroreflex in SHR compared with WKY contributes to the hypertension (or increased blood pressure) in SHR compared with WKY. Results of experiments using sinoaortic baroreceptor denervation were consistent with the hypothesis that dysfunction of the baroreflex contributes to the etiology of hypertension in the SHR. Thus, this study provides experimental evidence for the roles of the baroreflex in long-term arterial pressure regulation and in the etiology of primary hypertension in this animal model.

Blood pressure changes PVAT function and transcriptome: use of the mid-thoracic aorta coarcted rat.

Perivascular adipose tissue (PVAT) modifies the contractile function of the vessel it surrounds (outside-in signaling). Little work points to the vessel actively affecting its surrounding PVAT. We hypothesized that inside-out arterial signaling to PVAT would be evidenced by the response of PVAT to changes in tangential vascular wall stress. Rats coarcted in the mid-thoracic aorta created PVAT tissues that would exemplify pressure-dependent changes (above vs. below coarctation); a sham rat was used as a control. Radiotelemetry revealed a ∼20 mmHg systolic pressure gradient across the coarctation 4 wk after surgery. Four measures (histochemical, adipocyte progenitor proliferation and differentiation, isometric tone, and bulk mRNA sequencing) were used to compare PVAT above versus below the ligature in sham and coarcted rats. Neither aortic collagen deposition in PVAT nor arterial media/radius ratio above coarctation was increased versus below segments. However, differentiated adipocytes derived from PVAT above the coarctation accumulated substantially less triglycerides versus those below; their relative proliferation rate as adipogenic precursors was not different. Functionally, the ability of PVAT to assist stress relaxation of isolated aorta was reduced in rings above versus below the coarctation. Transcriptomic analyses revealed that the coarctation resulted in more differentially expressed genes (DEGs) between PVAT above versus below when compared with sham samples from the same locations. A majority of DEGs were in PVAT below the coarctation and were enriched in neuronal/synaptic terms. These findings provide initial evidence that signaling from the vascular wall, as stimulated by a pressure change, influences the function and transcriptional profile of its PVAT.NEW & NOTEWORTHY A mid-thoracic aorta coarcted rat was created to generate a stable pressure difference above versus below the coarctation ligature. This study determined that the PVAT around the thoracic aorta exposed to a higher pressure has a significantly reduced ability to assist stress relaxation versus that below the ligature and appears to retain the ability to be anticontractile. At the same time, the PVAT around the thoracic aorta exposed to higher pressure had a reduced adipogenic potential versus that below the ligature. Transcriptomics analyses indicated that PVAT below the coarctation showed the greatest number of DEGs with an increased profile of the synaptic neurotransmitter gene network.

Activation of the 5-HT7 receptor but not nitric oxide synthase is necessary for chronic 5-hydroxytryptamine-induced hypotension.

NEW FINDINGS: What is the central question of this study? What mechanisms account for the hypotension observed during chronic elevations in circulating 5-hydroxytryptamine in rats? What is the main finding and its importance? Chronic 5-hydroxytryptamine-induced hypotension requires continued activation of the 5-HT7 receptor subtype but does not require NO, an outcome that resolves previous conflicting results. Therapeutic interruption of the hypotensive actions of 5-HT under pathophysiological conditions can only be achieved through blockade of the 5-HT7 receptor. ABSTRACT: Low dose infusion of 5-hydroxytryptamine (5-HT) to rats causes both an acute and a chronic fall in arterial blood pressure. The 5-HT7 receptor subtype plays a critical part in the observed hypotension. Acute (minutes to hours) 5-HT infusion shows no depressor role for nitric oxide (NO), but 5-HT depressor responses under chronic conditions suggest that NO production may be critical. We test the hypothesis that NO contributes to the chronic, but not the acute, depressor response to 5-HT. We compared the role of NO and 5-HT7 receptors in 5-HT-induced hypotension under acute and chronic conditions in the same animal. Mean arterial pressure and heart rate were measured by radiotelemetry in conscious rats during 5 days of saline or 5-HT (25 µg kg-1  min-1 ; osmotic pump) infusion and for 2 days after infusion was stopped. To quantify the contributions of NO and the 5-HT7 receptor to 5-HT-induced hypotension, the nitric oxide synthase (NOS) inhibitor l-NAME or the selective 5-HT7 receptor antagonist SB-267790 were given at 1, 3 and 5 days of chronic infusion, and 1 day after 5-HT infusion pumps were removed. Nω -Nitro-l-arginine methyl ester (l-NAME) caused a pressor response of the same magnitude in the absence or presence of 5-HT infusion. Conversely, SB-269970 did not affect mean arterial pressure in the absence of 5-HT infusion and reversed the 5-HT-induced depressor response at each time point. Our findings demonstrate that acute and chronic 5-HT-induced hypotension does not require NOS activation but does require continued activation of the 5-HT7 receptor.

Chemerin contributes to in vivo adipogenesis in a location-specific manner.

Since chemerin's identification as an adipokine, it has been associated with a number of human diseases including diabetes and obesity. However, the basic scientific foundation for these clinical determinations is still lacking. Fibroblastic mouse 3T3 cells are unable to develop lipid droplets if chemerin is not present. Thus, we hypothesized that an in vivo rat model chemerin knockout (KO; an advancement from the previously mentioned in vitro cultures) would have limited accumulation of lipid in adipocytes compared to their wild-type (WT) counterparts. Female WT/KO rats (Sprague Dawley background) were fed a low-fat diet starting at 8 weeks of age with weekly body weight and food consumption monitoring. At 25 weeks of age, adipose tissue depots were dissected and flash frozen for PCR analysis or fixed with paraformaldehyde for histology. Over the 17 weeks of experimentation, WT and KO animals did not have differences in total body weight or food consumption but KO animals had a significantly reduced amount of visceral fat compared to WT animals (via microCT at 8 and 25 weeks). Histology of retroperitoneal and mesenteric depots demonstrated a significant leftward shift in adipocyte size in the mesenteric but not the retroperitoneal depot of the KO compared to WT animals. Similarly, in the mesenteric fat of the KO rat, gene expression of adiponectin, fatty acid synthase, perilipin, and leptin were significantly reduced compared to mesenteric fat of WT animals and retroperitoneal fat of both WT and KO animals. Adiponectin was highlighted by a protein-protein interaction network as being important for the physiological effects of chemerin removal. These data are the first, to our knowledge, to demonstrate chemerin's adipokine potential in vivo and identify it as fat depot location-specific.

A New Function for Perivascular Adipose Tissue (PVAT): Assistance of Arterial Stress Relaxation.

In health, PVAT secretes anti-contractile factors that relax the underlying artery. PVAT's contributions to vascular function include more than production of vasoactive substances. We hypothesized that PVAT benefits the artery by assisting the function of stress (-induced) relaxation. Thoracic aorta rings from Sprague Dawley rats were mounted in isolated tissue baths with (+) and without (-) PVAT. A cumulative length tension (0-6 grams) was generated. The tension to which the tissue stress relaxed over 30 minutes was recorded; the tension lost was stress relaxation. The presence of PVAT increased the amount of stress relaxation (final tension in mgs; aortic ring -PVAT = 4578 ± 190; aortic ring + PVAT = 2730 ± 274, p < 0.05). PVAT left attached but not encompassing the aorta provided no benefit in cumulative stress relaxation (aortic ring +/- PVAT = 4122 ± 176; p > 0.05 vs -PVAT). A PVAT ring separated from the aorta demonstrated more profound stress relaxation than did the aortic ring itself. Finally, PVAT-assisted stress relaxation was observed in an artery with white fat (superior mesenteric artery) and in aorta from both male and female of another rat strain, the Dahl S rat. Knowledge of this new PVAT function supports PVAT as an essential player in vascular health.

Effects of high-fat diet on sympathetic neurotransmission in mesenteric arteries from Dahl salt-sensitive rat.

Obesity hypertension is driven by sympathetic neurotransmission to the heart and blood vessels. We tested the hypothesis that high-fat diet (HFD)-induced hypertension is driven by sympathetic neurotransmission to mesenteric arteries (MA) in male but not female Dahl salt-sensitive (Dahl ss) rat. Rats were fed a control diet (CD; 10 kcal% from fat) or HFD (60 kcal% from fat) beginning at 3 weeks (wk) of age; measurements were made at 10-, 17- and 24-wk. Body weight increased with HFD, age and sex. Mean arterial pressure (MAP) was higher in HFD versus CD rats from both sexes at 17- and 24-wk. MA constriction measured using pressure myography, and electrical field stimulation (EFS, 0.2-30 Hz) was greater in HFD versus CD in males at 17-wk; this was not due to changes in α2 autoreceptor or norepinephrine transporter (NET) function. Prazosin (α1-AR antagonist) and suramin (P2 receptor antagonist) inhibited neurogenic MA constriction equally in all groups. Arterial reactivity to exogenous norepinephrine (NE; 10-8 - 10-5 M) was lower in HFD versus CD at 10-wk in males. Female MA reactivity to exogenous ATP was lower at 24-weeks compared to earlier time points. HFD did not affect tyrosine hydroxylase (TH) or the vesicular nucleotide transporter (VNUT) nerve density in MA from both sexes. NE content was lower in MA but higher in plasma at 24-wk compared to 10- and 17-wk in both sexes. In conclusion, HFD-induced hypertension is not driven by increased sympathetic neurotransmission to MA in male and female Dahl ss rats.

Brain Prostaglandin D2 Increases Neurogenic Pressor Activity and Mean Arterial Pressure in Angiotensin II-Salt Hypertensive Rats.

This study tested whether brain L-PGDS (lipocalin-type prostaglandin [PG] D synthase), through prostanoid signaling, might increase neurogenic pressor activity and thereby cause hypertension. Sprague Dawley rats on high-salt diet received either vehicle or Ang II (angiotensin II) infusion. On day 4, the developmental stage of hypertension, brains from different sets of control and Ang II-treated rats were collected for measuring L-PGDS expression, PGD2 levels, and DP1R (type 1 PGD2 receptor) expression. In a different set of 14-day Ang II-salt-treated rats, mini-osmotic pumps were used to infuse either a nonselective COX (cyclooxygenase) inhibitor ketorolac, L-PGDS inhibitor AT56, or DP1R inhibitor BWA868C to test the role of brain COX-PGD2-DP1R signaling in Ang II-salt hypertension. The acute depressor response to ganglion blockade with hexamethonium was used to quantify neurogenic pressor activity. During the developmental stage of Ang II-salt hypertension, L-PGDS expression was higher in cerebrospinal fluid, and PGD2 levels were increased in the choroid plexus, cerebrospinal fluid, and the cardioregulatory brain region rostral ventrolateral medulla. DP1R expression was decreased in rostral ventrolateral medulla. Both brain COX inhibition with ketorolac and L-PGDS inhibition with AT56 lowered mean arterial pressure by altering neurogenic pressor activity compared with vehicle controls. Blockade of DP1R with BWA868C, however, increased the magnitude of Ang II-salt hypertension and significantly increased neurogenic pressor activity. In summary, we establish that the development of Ang II-salt hypertension requires increased COX- and L-PGDS-derived PGD2 production in the brain, making L-PGDS a possible target for treating neurogenic hypertension.