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Ventricular catheterization with a pressure-volume (PV) catheter is the gold-standard method for assessing in vivo cardiac function in animal studies, providing valuable "load-independent" indices of systolic and diastolic heart performance. PV studies are commonly performed to elucidate mechanistic insights into cardiovascular disease using surgical and genetically engineered rat and mouse models, but there is considerable heterogeneity in how these studies are performed. Wide variation in protocol design, volume calibration, anesthesia, manipulation of cardiac loading conditions, how load-independent indices of cardiac function are derived, as well as in data analysis and reporting is constraining reliability and reproducibility in the field. The purpose of this manuscript is to combine our collective expertise in performing open- and closed-chest left and right ventricle PV studies in rodents to provide consensus guidelines on how to perform, analyze, and interpret these studies using either conductance or admittance PV catheters. We first review recent methodological reporting in rodent PV studies in this journal and discuss important details required to improve reproducibility within and across PV studies. We then recommend steps to obtain high-quality PV data, from volume calibration to choice of anesthetic agent and acquiring load-independent indices of both systolic and diastolic function. We also consider between- and within-animal variation and recommend how to perform data analysis and visualization. We hope that this consensus paper guides those performing PV studies in rodents and helps align the field with best practices in surgical/analytical methodologies and reporting, facilitating better reliability and reproducibility in the PV field. [ABSTRACT FROM AUTHOR]

The rat, Rattus norvegicus, has provided an important model for investigation of a range of characteristics of biomedical importance. Here we survey the origins of this species, its introduction into laboratory research, and the emergence of genetic and genomic methods that utilize this model organism. Genomic studies have yielded important progress and provided new insight into several biologically important traits. However, some studies have been impeded by the lack of a complete and accurate reference genome for this species. New sequencing and genome assembly methods applied to the rat have resulted in a new reference genome assembly, GRCr8, which is a near telomere-to-telomere assembly of high base-level accuracy that incorporates several elements not captured in prior assemblies. As genome assembly methods continue to advance and production costs become a less significant obstacle, genome assemblies for multiple inbred rat strains are emerging. These assemblies will allow a rat pangenome assembly to be constructed that captures all the genetic variations in strains selected for their utility in research and will overcome reference bias, a limitation associated with reliance on a single reference assembly. By this means, the full utility of this model organism to genomic studies will begin to be revealed. [ABSTRACT FROM AUTHOR]

Perceptual decision-making involves multiple cognitive processes, including accumulation of sensory evidence, planning, and executing a motor action. How these processes are intertwined is unclear; some models assume that decision-related processes precede motor execution, whereas others propose that movements reflecting ongoing decision processes occur before commitment to a choice. Here we combine two complementary methods to study the relationship between decision processes and the movements leading up to a choice. The first is a free-response pulse-based evidence accumulation task, in which stimuli continue until choice is reported, and the second is a motion-based drift diffusion model (mDDM), in which movement variables from video pose estimation constrain decision parameters on a trial-by-trial basis. We find that the mDDM provides a better fit to rats' decisions in the free-response accumulation task than traditional drift diffusion models. Interestingly, on each trial we observed a period, before choice, that was characterized by head immobility. The length of this period was positively correlated with the rats' decision bounds, and stimuli presented during this period had the greatest impact on choice. Together these results support a model in which internal decision dynamics are reflected in movements and demonstrate that inclusion of movement parameters improves the performance of diffusion-to-bound decision models. NEW & NOTEWORTHY: In this study we combine a novel pulse-based evidence accumulation task with a newly developed motion-based drift diffusion model (mDDM). In this model, we incorporate movement parameters derived from high-resolution video data to estimate parameters of the model on a trial-by-trial basis. We find that this new model is an improved description of animal choice behavior. [ABSTRACT FROM AUTHOR]

Rhythmic electrical events, termed slow waves, govern the timing and amplitude of phasic contractions of the gastric musculature. Extracellular multielectrode measurement of gastric slow waves can be a biomarker for phenotypes of motility dysfunction. However, a gastric slow-wave conduction pathway for the rat, a common animal model, is unestablished. In this study, the validity of extracellular recording was demonstrated in vitro with simultaneous intracellular and extracellular recordings and by pharmacological inhibition of slow waves. The conduction pathway was determined by in vivo extracellular recordings while considering the effect of motion. Slow-wave characteristics [means (SD)] varied regionally having higher amplitude in the antrum than the distal corpus [1.03 (0.12) mV vs. 0.75 (0.31) mV; n ¼ 7; P ¼ 0.025 paired t test] and faster propagation near the greater curvature than the lesser curvature [1.00 (0.14) mm·s-1 vs. 0.74 (0.14) mm·s-1 ; n ¼ 9 GC, 7 LC; P ¼ 0.003 unpaired t test]. Notably, in some subjects, separate wavefronts propagated near the lesser and greater curvatures with a loosely coupled region occurring in the area near the distal corpus midline at the interface of the two wavefronts. This region had either the greater or lesser curvature wavefront propagating through it in a time-varying manner. The conduction pattern suggests that slow waves in the rat stomach form annular wavefronts in the antrum and not the corpus. This study has implications for interpretation of the relationship between slow waves, the interstitial cells of Cajal network structure, smooth muscles, and gastric motility. NEW & NOTEWORTHY Mapping of rat gastric slow waves showed regional variations in their organization. In some subjects, separate wavefronts propagated near the lesser and greater curvatures with a loosely coupled region near the midline, between the wavefronts, having a varying slow-wave origin. Furthermore, simultaneous intracellular and extracellular recordings were concordant and independent of movement artifacts, indicating that extracellular recordings can be interpreted in terms of their intracellular counterparts when intracellular recording is not possible [ABSTRACT FROM AUTHOR]

Although the cause of interstitial cystitis/painful bladder syndrome (IC/PBS) remains unknown, autoimmune involvement has been strongly suggested to be a contributing factor. To elucidate the pathophysiology of IC/PBS, we characterized the experimental autoimmune cystitis (EAC) in rats. Adult female Sprague-Dawley rats were divided into the EAC and control groups. The EAC rats were generated by administrating a homogenate of donor rat bladder tissue as a bladder antigen. The characteristics of the two groups were determined by evaluating pain behavior and conducting cystometry, histopathology, and molecular analyses. The EAC rats showed: 1) a decreased paw withdrawal threshold, 2) a reduced intercontraction interval on cystometry, 3) the irregular surfaces of the umbrella cells of epithelium throughout the bladder wall, 4) accumulation of stress granules in the bladder and vascular endothelium, 5)the increased expression of genes related to inflammation and ischemia at the mRNA and protein levels, 6) a significantly increased paw withdrawal threshold with pain treatment, and 7) the induction of glomerulation of the bladder wall, epithelium denudation, and lymphocyte infiltration in the interstitium by bladder distension. These results suggest that the EAC rats showed pain and frequent urination with the overexpression of inflammatory chemokines, reflecting clinical IC/BPS, and the bladder epithelium and vascular endothelium may be the primary sites of IC/BPS, and bladder injury, such as bladder distension, can cause progression from BPS to IC with Hunner lesions. NEW & NOTEWORTHY: The experimental autoimmune cystitis model rats showed pain and frequent urination with the overexpression of inflammatory chemokines, reflecting clinical interstitial cystitis/painful bladder syndrome (IC/PBS), and the bladder epithelium and vascular endothelium may be the primary sites of IC/BPS, and bladder injury, such as bladder distension, can cause progression from BPS to IC with Hunner lesions. [ABSTRACT FROM AUTHOR]

Hypertension affects approximately one in two United States adults and sex plays an important role in the pathogenesis of hypertension. The Na+-Cl− cotransporter (NCC), regulated by a kinase network including with-no-lysine kinase (WNK)1 and WNK4, STE20/SPS1-related proline alanine-rich kinase (SPAK), and oxidative stress response 1 (OxSR1), is critical to Na+ reabsorption and blood pressure regulation. Dietary salt differentially modulates NCC in salt-sensitive and salt-resistant rats, in part by modulation of WNK/SPAK/OxSR1 signaling. In this study, we tested the hypothesis that sex-dependent differences in NCC regulation contribute to the development of the salt sensitivity of blood pressure using male and female Sprague-Dawley (SD), Dahl salt-resistant (DSR), and Dahl salt-sensitive (DSS) rats. In normotensive salt-resistant SD and DSR rats, a high-salt diet evoked significant decreases in NCC activity, expression, and phosphorylation. In males, these changes were associated with no change in WNK1 expression, a decrease in WNK4 levels, and suppression of SPAK/OxSR1 expression and phosphorylation. In contrast, in females, there was decreased NCC activity associated with suppression of SPAK/OxSR1 expression and phosphorylation. In hypertensive DSS rats, the ability of females to suppress NCC (in opposition to males) via a SPAK/OxSR1 mechanism likely contributes to their lower magnitude of salt-sensitive hypertension. Collectively, our findings support the existence of sex differences in male versus female rats with NCC regulation during dietary salt intake involving suppression of WNK4 expression in male rats only and the involvement of SPAK/OxSR1 signaling in both males and females. NEW & NOTEWORTHY: NCC regulation is sex dependent. In normotensive male and female Sprague-Dawley and Dahl salt-resistant rats, which exhibit dietary Na+-evoked NCC suppression, male rats exhibit decreased WNK4 expression and decreased SPAK and OxSR1 levels, whereas female rats only suppress SPAK and OxSR1. In hypertensive Dahl salt-sensitive rats, the ability of females to suppress NCC (in opposition to males) via a SPAK/OxSR1 mechanism likely contributes to their lower magnitude of salt-sensitive hypertension. [ABSTRACT FROM AUTHOR]

Adult rats exposed to hyperoxia (>95% O2) die from respiratory failure in 60–72 h. However, rats preconditioned with >95% O2 for 48 h followed by 24 h in room air acquire tolerance of hyperoxia (H-T), whereas rats preconditioned with 60% O2 for 7 days become more susceptible (H-S). Our objective was to evaluate lung tissue mitochondrial bioenergetics in H-T and H-S rats. Bioenergetics was assessed in mitochondria isolated from lung tissue of H-T, H-S, and control rats. Expressions of complexes involved in oxidative phosphorylation (OxPhos) were measured in lung tissue homogenate. Pulmonary endothelial filtration coefficient (Kf) and tissue mitochondrial membrane potential (Δψm) were evaluated in isolated perfused lungs (IPLs). Results show that ADP-induced state 3 OxPhos capacity (Vmax) decreased in H-S mitochondria but increased in H-T. Δψm repolarization time following ADP-stimulated depolarization increased in H-S mitochondria. Complex I expression decreased in H-T (38%) and H-S (43%) lung homogenate, whereas complex V expression increased (70%) in H-T lung homogenate. Δψm is unchanged in H-S and H-T lungs, but complex II has a larger contribution to Δψm in H-S than H-T lungs. Kf increased in H-S, but not in H-T lungs. For H-T, increased complex V expression and Vmax counter the effect of the decrease in complex I expression on Δψm. A larger complex II contribution to Δψm along with decreased Vmax and increased Kf could make H-S rats more hyperoxia susceptible. Results are clinically relevant since ventilation with ≥60% O2 is often required for extended periods in patients with acute respiratory distress syndrome (ARDS). NEW & NOTEWORTHY: We assessed lung tissue mitochondrial bioenergetics in rats with tolerance (H-T) or susceptibility (H-S) to hyperoxia-induced ARDS. Results from studies in isolated mitochondria, tissue homogenate, and isolated perfused lungs show that mitochondrial bioenergetics are differentially altered in H-T and H-S lungs suggesting a potential role for mitochondrial bioenergetics in hyperoxia-induced ARDS. Results are clinically relevant since hyperoxia exposure is a primary therapy for patients with ARDS, and differential sensitivity to hyperoxia surely occurs in humans. [ABSTRACT FROM AUTHOR]

Nitric oxide (NO) inhalation improves pulmonary hemodynamics in participants with pulmonary arterial hypertension (PAH). Although it can reduce pulmonary vascular resistance (PVR) in PAH, its impact on the dynamic mechanics of pulmonary arteries and its potential difference between control and participants with PAH remain unclear. PA impedance provides a comprehensive description of PA mechanics. With an arterial model, PA impedance can be parameterized into peripheral pulmonary resistance (Rp), arterial compliance (Cp), characteristic impedance of the proximal arteries (Zc), and transmission time from the main PA to the reflection site. This study investigated the effects of inhaled NO on PA impedance and its associated parameters in control and monocrotaline-induced pulmonary arterial hypertension (MCT-PAH) male rats (6/group). Measurements were obtained at baseline and during NO inhalation at 40 and 80 ppm. In both groups, NO inhalation decreased PVR and increased the left atrial pressure. Notably, its impact on PA impedance was frequency dependent, as revealed by reduced PA impedance modulus in the low-frequency range below 10 Hz, with little effect on the high-frequency range. Furthermore, NO inhalation attenuated Rp, increased Cp, and prolonged transmission time without affecting Zc. It reduced Rp more pronouncedly in MCT-PAH rats, whereas it increased Cp and delayed transmission time more effectively in control rats. In conclusion, the therapeutic effects of inhaled NO on PA impedance were frequency dependent and may differ between the control and MCT-PAH groups, suggesting that the effect on the mechanics differs depending on the pathological state. NEW & NOTEWORTHY: Nitric oxide inhalation decreased pulmonary arterial impedance in the low-frequency range (<10 Hz) with little impact on the high-frequency range. It reduced peripheral pulmonary resistance more pronouncedly in pulmonary hypertension rats, whereas it increased arterial compliance and transmission time in control rats. Its effect on the mechanics of the pulmonary arteries may differ depending on the pathological status. [ABSTRACT FROM AUTHOR]

Diets varying in macronutrient composition, energy density, and/or palatability may cause differences in outcome of bariatric surgery. In the present study, rats feeding a healthy low-fat (LF) diet or an obesogenic high-fat/sucrose diet (HF/S) were either subjected to Roux-en-Y gastric bypass surgery (RYGB) or sham surgery, and weight loss trajectories and various energy balance parameters were assessed. Before RYGB, rats eating an HF/S (n = 14) diet increased body weight relative to rats eating an LF diet (n = 20; P < 0.01). After RYGB, absolute weight loss was larger in HF/S (n = 6) relative to LF feeding (n = 6) rats, and this was associated with reduced cumulative energy intake (EI; P < 0.05) and increased locomotor activity (LA; P < 0.05-0.001), finally leading to similar levels of reduced body fat content in HF/S and LF rats 3 wk after surgery. Regression analysis revealed that variation in RYGB-induced body weight loss was best explained by models including 1) postoperative cumulative EI and preoperative body weight (R2=0.87) and 2) postoperative cumulative EI and diet (R2=0.79), each without significant contribution of LA. Particularly rats on the LF diet became transiently more hypothermic and circadianally arrhythmic following RYGB (i.e., indicators of surgery-associated malaise) than HF/S feeding rats. Our data suggest that relative to feeding an LF diet, continued feeding an HF/S diet does not negatively impact recovery from RYGB surgery, yet it promotes RYGB-induced weight loss. The RYGB-induced weight loss is primarily explained by reduced cumulative EI and higher preoperative body weight, leading to comparably low levels of body fat content in HF/S and LF feeding rats. NEW & NOTEWORTHY Relative to feeding an LF diet, continued feeding an HF/S diet does not negatively impact recovery from RYGB surgery in rats. Relative to feeding an LF diet, continued feeding an HF/S diet promotes RYGB-induced weight loss. The RYGB-induced weight loss is primarily explained by reduced cumulative EI and higher preoperative body weight, leading to comparably low levels of body fat content in HF/S and LF feeding rats. [ABSTRACT FROM AUTHOR]

We investigated the role played by ATP-sensitive purinergic 2 (P2) receptors in evoking the pressor response to treadmill exercise in male and female rats with and without femoral arteries that were ligated for -72 h to induce simulated peripheral artery disease (PAD). We hypothesized that PPADS (P2 receptor antagonist, 10 mg iv) would reduce the pressor response to 4 min of treadmill exercise (15 m·min-1, 1-incline) and steady-state exercise plasma norepinephrine (NE) values in male and female rats, and that the magnitude of effect of PPADS would be greater in rats with simulated PAD ("ligated") than in sham-operated rats. In males, PPADS significantly reduced the difference between steady-state exercise and baseline mean arterial pressure (DMAP) response to treadmill exercise in sham (n = 8; pre-PPADS: 12 ± 2, post-PPADS: 1 ± 5 mmHg; P = 0.037) and ligated (n = 4; pre-PPADS: 20 ± 2, post-PPADS: 11 ± 3 mmHg; P = 0.028) rats with a similar magnitude of effect observed between groups (P = 0.720). In females, PPADS had no effect on the DMAP response to treadmill exercise in sham (n = 6; pre-PPADS: 9 ± 2, post-PPADS: 7 ± 2 mmHg; P = 0.448) or ligated (n = 6; pre-PPADS: 15 ± 2, post-PPADS: 16 ± 3 mmHg; P = 0.684) rats. When NE values were grouped by sex independent of ligation/sham status, PPADS significantly reduced plasma NE in male (P = 0.016) and female (P = 0.027) rats. The data indicate that P2 receptors contribute to the sympathetic response to exercise in both male and female rats but that the sympathoexcitatory role for P2 receptors translates into an obligatory role in the blood pressure response to exercise in male but not in female rats. NEW & NOTEWORTHY Here, we demonstrate that purinergic 2 (P2) receptors contribute significantly to the blood pressure response to treadmill exercise in male rats both with and without simulated PAD induced by femoral artery ligation. We found no role for P2 receptors in the blood pressure response to treadmill exercise in female rats, thus revealing clear sex differences in P2 receptor-mediated blood pressure control during exercise. [ABSTRACT FROM AUTHOR]

Sodium-glucose cotransporter-2 inhibitors (SGLT2i) reduce blood pressure (BP) in patients with hypertension, yet the precise molecular mechanisms remain elusive. SGLT2i inhibits proximal tubule (PT) NHE3-mediated sodium reabsorption in normotensive rodents, yet no hypotensive effect is observed under this scenario. This study examined the effect of empagliflozin (EMPA) on renal tubular sodium transport in normotensive and spontaneously hypertensive rats (SHRs). It also tested the hypothesis that EMPA-mediated PT NHE3 inhibition in normotensive rats is associated with upregulation of distal nephron apical sodium transporters. EMPA administration for 14 days reduced BP in 12-wk-old SHRs but not in age-matched Wistar rats. PT NHE3 activity was inhibited by EMPA treatment in both Wistar and SHRs. In Wistar rats, EMPA increased NCC activity, mRNA expression, protein abundance, and phosphorylation levels, but not in SHRs. SHRs showed higher NKCC2 activity and an abundance of cleaved ENaC α and γ subunits compared with Wistar rats, none of which were affected by EMPA. Another set of male Wistar rats was treated with EMPA, the NCC inhibitor hydrochlorothiazide (HCTZ), and EMPA combined with HCTZ or vehicle for 14 days. In these rats, BP reduction was observed only with combined EMPA and HCTZ treatment, not with either drug alone. These findings suggest that NCC upregulation counteracts EMPA-mediated inhibition of PT NHE3 in male normotensive rats, maintaining their baseline BP. Moreover, the reduction of NHE3 activity without further upregulation of major apical sodium transporters beyond the PT may contribute to the BP-lowering effect of SGLT2i in experimental models and patients with hypertension. NEW & NOTEWORTHY: This study suggests that reduced NHE3-mediated sodium reabsorption in the renal proximal tubule may account, at least in part, for the BP-lowering effect of SGLT2 inhibitors in the setting of hypertension. It also demonstrates that chronic treatment with SGLT2 inhibitors upregulates NCC activity, phosphorylation, and expression in the distal tubule of normotensive but not hypertensive rats. SGLT2 inhibitor-mediated upregulation of NCC seems crucial to counteract proximal tubule natriuresis in subjects with normal BP. [ABSTRACT FROM AUTHOR]

One exercise session can increase subsequent insulin-stimulated glucose uptake (ISGU) by skeletal muscle from rodents and humans of both sexes. We recently found that concurrent mutation of three key sites to prevent their phosphorylation (Ser588, Thr642, and Ser704) on Akt substrate of 160 kDa (AS160; also known as TBC1D4) reduced the magnitude of the enhancement of postexercise ISGU (PEX-ISGU) by muscle from male, but not female rats. However, we did not test the role of individual phosphorylation sites on PEX-ISGU. Accordingly, our current aim was to test whether AS160 Ser704 phosphorylation (pSer704) is required for elevated PEX-ISGU by muscle. AS160-knockout (AS160-KO) rats (female and male) were studied when either in sedentary or 3 h after acute exercise. Adeno-associated virus (AAV) vectors were used to enable muscle expression of wild-type AS160 (AAV-WT-AS160) or AS160 mutated Ser704 to alanine to prevent phosphorylation (AAV-1P-AS160). Paired epitrochlearis muscles from each rat were injected with AAV-WT-AS160 or AAV-1P-AS160. We discovered that regardless of sex 1) AS160 abundance in AS160-KO rats was similar in paired muscles expressing WT-AS160 versus 1P-AS160; 2) muscles from exercised versus sedentary rats had greater ISGU, and PEX-ISGU was slightly greater for muscles expressing 1P-AS160 versus contralateral muscles expressing WT-AS160; and 3) pAS160Thr642 was lower in muscles expressing 1P-AS160 versus paired muscles expressing WT-AS160. These results indicate that pAS160Ser704 was not essential for elevated PEX-ISGU by skeletal muscle from rats of either sex. Furthermore, elimination of the postexercise increase in pAS160Thr642 did not lessen the postexercise effect on ISGU. NEW & NOTEWORTHY: The current study evaluated the role of Akt substrate of 160 kDa (AS160) phosphorylation on Ser704 in increased insulin-stimulated glucose uptake by skeletal muscle after exercise. Adeno-associated virus vectors were engineered to express either wild-type-AS160 or AS160 mutated so that it could not be phosphorylated on Ser704 in paired muscles from AS160-knockout rats. The results demonstrated that AS160 phosphorylation on Ser704 was not essential for exercise-induced elevation in insulin-stimulated glucose uptake by rats of either sex. [ABSTRACT FROM AUTHOR]

Blood flow regulation within the microvasculature reflects a complex interaction of regulatory mechanisms and varies spatially and temporally according to conditions such as metabolism, growth, injury, and disease. Understanding the role of microvascular flow distributions across conditions is of interest to investigators spanning multiple disciplines; however, data collection within networks can be labor-intensive and challenging due to limited resolution. To overcome these experimental challenges, computational network models that can accurately simulate vascular behavior are highly beneficial. Constrained constructive optimization (CCO) is a commonly used algorithm for vascular simulation, particularly well known for its adaptability toward vascular modeling across tissues. The present work demonstrates an implementation of CCO aimed to simulate a branching arteriolar microvasculature in healthy skeletal muscle, validated against literature including comprehensive rat gluteus maximus vasculature datasets, and reviews a list of user-specified adjustable model parameters to understand how their variability affects the simulated networks. Network geometric properties, including mean element diameters, lengths, and numbers of bifurcations per order, Horton's law ratios, and fractal dimension, demonstrate good validation once model parameters are adjusted to experimental data. This model successfully demonstrates hemodynamic properties such as Murray's law and the network Fahraeus effect. Application of centrifugal and Strahler ordering schemes results in divergent descriptions of identical simulated networks. This work introduces a novel CCO-based model focused on generating branching skeletal muscle microvascular arteriolar networks based on adjustable model parameters, thus making it a valuable tool for investigations into skeletal muscle microvascular structure and tissue perfusion. NEW & NOTEWORTHY: The present work introduces a CCO-based algorithm for generating branching arteriolar networks, with adjustable model parameters to enable modeling in varying skeletal muscle tissues. The geometric and hemodynamic parameters of the generated networks have been comprehensively validated using experimental data collected previously in-house and from literature. This is one of few validated CCO-based models to specialize in skeletal muscle microvasculature and acts as a beneficial tool for investigating the microvasculature for hypothesis testing and validation. [ABSTRACT FROM AUTHOR]

A significant number of pregnancies occur at advanced maternal age (>35 yr), which is a risk factor for pregnancy complications. Healthy pregnancies require massive hemodynamic adaptations, including an increased blood volume and cardiac output. There is growing evidence that these cardiovascular adaptations are impaired with age, however, little is known about maternal cardiac function with advanced age. We hypothesized that cardiac adaptations to pregnancy are impaired with advanced maternal age. Younger (4 mo; ~early reproductive maturity in humans) and aged (9 mo; ~35 yr in humans) pregnant Sprague-Dawley rats were assessed and compared with age-matched nonpregnant controls. Two-dimensional echocardiographic images were obtained (ultrasound biomicroscopy; under anesthesia) on gestational day 19 (term = 22 days) and compared with age-matched nonpregnant rats (n = 7-9/group). Left ventricular structure and function were assessed using short-axis images and transmitral Doppler signals. During systole, left ventricular anterior wall thickness increased with age in the nonpregnant rats, but there was no age-related difference between the pregnant groups. There were no significant pregnancy-associated differences in left ventricular wall thickness. Calculated left ventricular mass increased with age in nonpregnant rats and increased with pregnancy only in young rats. Compared with young pregnant rats, the aortic ejection time of aged pregnant rats was greater and Tei index was lower. Overall, the greater aortic ejection time and lower Tei index with age in pregnant rats suggest mildly altered cardiac adaptations to pregnancy with advanced maternal age, which may contribute to adverse outcomes in advanced maternal age pregnancies. NEW & NOTEWORTHY We demonstrated that even before the age of reproductive senescence, rats show signs of age-related alterations in cardiac structure that suggests increased cardiac work. Our data also demonstrate, using an in vivo echocardiographic approach, that advanced maternal age in a rat model is associated with altered cardiac function and structure relative to younger pregnant controls. [ABSTRACT FROM AUTHOR]

Recent evidence indicates that sex-based differences in cardiovascular disease (CVD) begin early in life, particularly when associated with risk factors such as a sedentary lifestyle. CVD is associated with elevated sympathetic nerve activity (SNA), quantified as increased SNA burst activity in humans. Whether burst characteristics are influenced by sex or sedentary conditions at younger ages is unknown. The purpose of our study is to compare SNA bursts in active and sedentary female and male rats at ages including prepuberty and young adulthood. We hypothesized that burst characteristics and blood pressure are higher under sedentary conditions and lower in female rats compared with males. We analyzed splanchnic SNA (SpSNA) recordings from Inactin-anesthetized male and female rats at 4-, 8-, and 16-wk of age. Physically active and sedentary rats were each housed in separate, environmentally controlled chambers where physically active rats had free access to an in-cage running wheel. Sympathetic bursts were obtained by rectifying and integrating the raw SpSNA signal. Burst frequency, burst height, and burst width were calculated using the Peak Parameters extension in LabChart. Our results showed that sedentary conditions produced a greater burst width in 8- and 16-wk-old rats compared with 4-wk-old rats in both males and females (P < 0.001 for both). Burst frequency and incidence were both higher in 16-wk-old males compared with 16-wk-old females (P < 0.001 for both). Our results suggest that there are sedentary lifestyle- and sex-related mechanisms that impact sympathetic regulation of blood pressure at ages that range from prepuberty into young adulthood. NEW & NOTEWORTHY: The mechanisms of decreased incidence of cardiovascular disease (CVD) in reproductive-age women compared with age-matched men are unknown. The strong association between elevated sympathetic activity and CVD led us to characterize splanchnic sympathetic bursts in female and male rats. Prepubescent males and females exhibited narrower sympathetic bursts, whereas young adult males had higher resting burst frequency compared with age-matched females. Sex-based regulation of sympathetic activity suggests a need for sex-dependent therapeutic strategies to combat CVD. [ABSTRACT FROM AUTHOR]

Cyclooxygenase (COX) products of arachidonic acid metabolism, specifically prostaglandins, play a role in evoking and transmitting the exercise pressor reflex in health and disease. Individuals with type 2 diabetes mellitus (T2DM) have an exaggerated exercise pressor reflex; however, the mechanisms for this exaggerated reflex are not fully understood. We aimed to determine the role played by COX products in the exaggerated exercise pressor reflex in T2DM rats. The exercise pressor reflex was evoked by static muscle contraction in unanesthetized, decerebrate, male, adult University of California Davis (UCD)-T2DM (n = 8) and healthy Sprague-Dawley (n = 8) rats. Changes (Δ) in peak mean arterial pressure (MAP) and heart rate (HR) during muscle contraction were compared before and after intra-arterial injection of indomethacin (1 mg/kg) into the contracting hindlimb. Data are presented as means ± SD. Inhibition of COX activity attenuated the exaggerated peak MAP (Before: Δ32 ± 13 mmHg and After: Δ18 ± 8 mmHg; P = 0.004) and blood pressor index (BPi) (Before: Δ683 ± 324 mmHg·s and After: Δ361 ± 222 mmHg·s; P = 0.006), but not HR (Before: Δ23 ± 8 beats/min and After Δ19 ± 10 beats/min; P = 0.452) responses to muscle contraction in T2DM rats. In healthy rats, COX activity inhibition did not affect MAP, HR, or BPi responses to muscle contraction. Inhibition of COX activity significantly reduced local production of prostaglandin E2 in T2DM and healthy rats. We conclude that peripheral inhibition of COX activity attenuates the pressor response to muscle contraction in T2DM rats, suggesting that COX products partially contribute to the exaggerated exercise pressor reflex in those with T2DM. NEW & NOTEWORTHY: We compared the pressor and cardioaccelerator responses to static muscle contraction before and after inhibition of cyclooxygenase (COX) activity within the contracting hindlimb in decerebrate, unanesthetized type 2 diabetic mellitus (T2DM) and healthy rats. The pressor responses to muscle contraction were attenuated after peripheral inhibition of COX activity in T2DM but not in healthy rats. We concluded that COX products partially contribute to the exaggerated pressor reflex in those with T2DM. [ABSTRACT FROM AUTHOR]

The external carotid artery (ECA) plays a major role in supplying blood to the head and neck. Although impeded blood flow in the ECA is expected to affect orofacial functions, few studies have shown how blood flow obstruction in the ECA contributes to impairment of these functions, including chewing and swallowing. This study was performed to investigate the effects of ECA ligation (ECAL) on immediate and long-term changes in masticatory and swallowing functions as well as the jaw-opening reflex evoked in the digastric muscle. The experiments were carried out using male Sprague-Dawley rats. In the acute experiment, the digastric reflex evoked by low-threshold electrical stimulation of the inferior alveolar nerve and the swallow reflex, identified by digastric and thyrohyoid electromyographic (EMG) bursts, were compared between before and 1 h after ECAL. The chronic experiment was conducted on freely moving rats. EMGs of the masseter, digastric, and thyrohyoid muscles were chronically recorded. The long-term effects of ECAL on behavior and muscle histology were compared between rats with an intact ECA and rats with ECAL. In the acute experiment, the peak amplitude of the digastric reflex on the ECAL side was significantly decreased 1 h after ECAL. In the chronic experiment, although most parameters of the masticatory and swallowing EMGs were not significantly different between the groups, the results suggest wide variation of the effect of ECAL on the muscles. Blood supply compensation from collaterals of the internal carotid artery may be permanent in some animals. [ABSTRACT FROM AUTHOR]

Although body fluid volume control by the kidneys may be classified as a long-term arterial pressure (AP) control system, it does not necessarily follow that the urine flow (UF) response to changes in AP is slow. We quantified the dynamic characteristics of the UF response to short-term AP changes by changing mean AP between 60 mmHg and 100 mmHg every 10 s according to a binary white noise sequence in anesthetized rats (n = 8 animals). In a baro-on trial (the carotid sinus baroreflex was enabled), the UF response represented the combined synergistic effects of pressure diuresis (PD) and neurally mediated antidiuresis (NMA). In a baro-fix trial (the carotid sinus pressure was fixed at 100 mmHg), the UF response mainly reflected the effect of PD. The UF step response was quantified using the sum of two exponential decay functions. The fast and slow components had time constants of 6.5 ± 3.6 s and 102 ± 85 s (means ± SD), respectively, in the baro-on trial. Although the gain of the fast component did not differ between the two trials (0.49 ± 0.21 vs. 0.66 ± 0.22 lL·min-1·kg-1·mmHg-1), the gain of the slow component was greater in the baro-on than in the baro-fix trial (0.51 ± 0.14 vs. 0.09 ± 0.39 lL·min-1·kg-1·mmHg-1, P = 0.023). The magnitude of NMA relative to PD was calculated to be 32.2 ± 29.8%. In conclusion, NMA contributed to the slow component, and its magnitude was approximately one-third of that of the effect of PD. [ABSTRACT FROM AUTHOR]

Although Gaussian white noise (GWN) inputs offer a theoretical framework for identifying higher-order nonlinearity, an actual application to the data of the neural arc of the carotid sinus baroreflex did not succeed in fully predicting the well-known sigmoidal nonlinearity. In the present study, we assumed that the neural arc can be approximated by a cascade of a linear dynamic (LD) component and a nonlinear static (NS) component. We analyzed the data obtained using GWN inputs with a mean of 120 mmHg and standard deviations (SDs) of 10, 20, and 30 mmHg for 15 min each in anesthetized rats (n = 7). We first estimated the linear transfer function from carotid sinus pressure to sympathetic nerve activity (SNA) and then plotted the measured SNA against the linearly predicted SNA. The predicted and measured data pairs exhibited an inverse sigmoidal distribution when grouped into 10 bins based on the size of the linearly predicted SNA. The sigmoidal nonlinearity estimated via the LD-NS model showed a midpoint pressure (104.1 ± 4.4 mmHg for SD of 30 mmHg) lower than that estimated by a conventional stepwise input (135.8 ± 3.9 mmHg, P < 0.001). This suggests that the NS component is more likely to reflect the nonlinearity observed during pulsatile inputs that are physiological to baroreceptors. Furthermore, the LD-NS model yielded higher R2 values compared with the linear model and the previously suggested second-order Uryson model in the testing dataset. [ABSTRACT FROM AUTHOR]

Perceptual decision-making is a dynamic cognitive process and is shaped by many factors, including behavioral state, reward contingency, and sensory environment. To understand the extent to which adaptive behavior in decision-making is dependent on pupil-linked arousal, we trained head-fixed rats to perform perceptual decision-making tasks and systematically manipulated the probability of Go and No-go stimuli while simultaneously measuring their pupil size in the tasks. Our data demonstrated that the animals adaptively modified their behavior in response to the changes in the sensory environment. The response probability to both Go and No-go stimuli decreased as the probability of the Go stimulus being presented decreased. Analyses within the signal detection theory framework showed that while the animals’ perceptual sensitivity was invariant, their decision criterion increased as the probability of the Go stimulus decreased. Simulation results indicated that the adaptive increase in the decision criterion will increase possible water rewards during the task. Moreover, the adaptive decision-making is dependent on pupil-linked arousal as the increase in the decision criterion was the largest during low pupil-linked arousal periods. Taken together, our results demonstrated that the rats were able to adjust their decision-making to maximize rewards in the tasks, and that adaptive behavior in perceptual decision-making is dependent on pupil-linked arousal. NEW & NOTEWORTHY Perceptual decision-making is a dynamic cognitive process and is shaped by many factors. However, the extent to which changes in sensory environment result in adaptive decision-making remains poorly understood. Our data provided new experimental evidence demonstrating that the rats were able to adaptively modify their decision criterion to maximize water reward in response to changes in the statistics of the sensory environment. Furthermore, the adaptive decision-making is dependent on pupil-linked arousal. [ABSTRACT FROM AUTHOR]

High versus low aerobic capacity significantly impacts the risk for metabolic diseases. Rats selectively bred for high or low intrinsic aerobic capacity differently modify hepatic bile acid metabolism in response to high-fat diets (HFDs). Here we tested if a bile acid sequestrant would alter hepatic and whole body metabolism differently in rats with high and low aerobic capacity fed a 1-wk HFD. Male rats (8 mo of age) that were artificially selected to be high (HCR) and low-capacity runners (LCR) with divergent intrinsic aerobic capacities were transitioned from a low-fat diet (LFD, 10% fat) to an HFD (45% fat) with or without a bile acid sequestrant (BA-Seq, 2% cholestyramine resin) for 7 days while maintained in an indirect calorimetry system. HFD þ BA-Seq increased fecal excretion of lipids and bile acids and prevented weight and fat mass gain in both strains. Interestingly, HCR rats had increased adaptability to enhance fecal bile acid and lipid loss, resulting in more significant energy loss than their LCR counterpart. In addition, BA-Seq induced a greater expression of hepatic CYP7A1 gene expression, the rate-limiting enzyme of bile acid synthesis in HCR rats both on HFD and HFD þ BA-Seq diets. HCR displayed a more significant reduction of RQ in response to HFD than LCR, but HFD þ BA-Seq lowered RQ in both groups compared with HFD alone, demonstrating a pronounced impact on metabolic flexibility. In conclusion, BA-Seq provides uniform metabolic benefits for metabolic flexibility and adiposity, but rats with higher aerobic capacity display adaptability for hepatic bile acid metabolism. [ABSTRACT FROM AUTHOR]

The study aimed to investigate the bladder and urethral activity and nitric oxide (NO)-related molecular changes in aging rats. Rats were divided into two groups: Group Y (young rats; 12 wk) and Group A (aging rats; 15 mo). A 24-h voiding assay was performed, and the urodynamic parameters were evaluated using awake cystometry (CMG) and urethral perfusion pressure (UPP) recordings under urethane anesthesia. The mRNA expression levels of NO-, ischemia-, and inflammation-related markers in urethra and bladder tissues and cGMP levels in the urethra were assessed. Body weight was significantly higher in Group A than in Group Y. Voiding assay results (24 h) were insignificant. In the CMG, the number of non-voiding contractions per voiding cycle and post-void residual volume were significantly higher in Group A than in Group Y; voiding efficiency was significantly lower in Group A than in Group Y. In the UPP recordings, the urethral pressure reduction and high-frequency oscillation (HFO) amplitude were significantly lower in Group A than in Group Y. The mRNA expression levels of Hif-1α, Vegf-a, and Tgf-β1 in the bladder were significantly higher in Group A than in Group Y. The mRNA expression levels of Nos1 and Prkg1 and the cGMP concentrations in the urethra were significantly lower in Group A than in Group Y. Aging rats can be useful models for studying the natural progression of age-related lower urinary tract dysfunctions, for which impaired NO-mediated transmitter function is likely to be an important mechanism. NEW & NOTEWORTHY Aging rats can be useful models for studying the natural progression of age-related lower urinary tract dysfunctions, for which impaired nitric oxide-mediated transmitter function is likely to be an important mechanism. [ABSTRACT FROM AUTHOR]

Chronic infusion of subpressor level of angiotensin II (ANG II) increases the abundance of Na+ transporters along the distal nephron, balanced by suppression of Na+ transporters along the proximal tubule and medullary thick ascending limb (defined as “proximal nephron”), which impacts K+ handling along the entire renal tubule. The objective of this study was to quantitatively assess the impact of chronic ANG II on the renal handling of Na+ and K+ in female rats, using a computational model of the female rat renal tubule. Our results indicate that the downregulation of proximal nephron Na+ reabsorption (TNa), which occurs in response to ANG II-triggered hypertension, involves changes in both transporter abundance and trafficking. Our model suggests that substantial (∼30%) downregulation of active NHE3 in proximal tubule (PT) microvilli is needed to reestablish the Na+ balance at 2 wk of ANG II infusion. The 35% decrease in SGLT2, a known NHE3 regulator, may contribute to this downregulation. Both depression of proximal nephron TNa and stimulation of distal ENaC raise urinary K+ excretion in ANG II-treated females, while K+ loss is slightly mitigated by cortical NKCC2 and NCC upregulation. Our model predicts that K+ excretion may be more significantly limited during ANG II infusion by ROMK inhibition in the distal nephron and/or KCC3 upregulation in the PT, which remain open questions for experimental validation. In summary, our analysis indicates that ANG II hypertension triggers a series of events from distal TNa stimulation followed by compensatory reduction in proximal nephron TNa and accompanying adjustments to limit excessive K+ secretion. NEW & NOTEWORTHY We used a computational model of the renal tubule to assess the impact of 2-wk angiotensin II (ANG II) infusion on the handling of Na+ and K+ in female rats. ANG II strongly stimulates distal Na+ reabsorption and K+ secretion. Simulations indicate that substantial downregulation of proximal tubule NHE3 is needed to reestablish Na+ balance at 2 wk. Proximal adaptations challenge K+ homeostasis, and regulation of distal NCC and specific K+ channels likely limit urinary K+ losses. [ABSTRACT FROM AUTHOR]

Hypothermia develops during systemic anaphylaxis in rodents. The aim of this study was to elucidate the mechanism for the hypothermia by assessing the roles of locomotor activity, tail heat dissipation, heat production in the brown adipose tissue (BAT) activity, and chemical mediators during ovalbumin-induced anaphylactic hypotension in awake rats. We measured the core body temperature (Tcore) and mean blood pressure (MBP), along with the surface temperature of the interscapular region (TiScap), an indirect measure of BAT activity, and the tail (Ttail). During anaphylaxis, MBP decreased to the nadir of 53 ± 2 mmHg at 8 min with recovery toward baseline. Tcore began to decrease at 7.5 min with the nadir of 36.1 ± 0.2°C at 30 min from the baseline of 38.0 ± 0.1°C. TiScap also significantly decreased, but its onset was preceded by that of Tcore. Ttail decreased after antigen, suggesting the absence of increased heat dissipation from the tail. The physical activity, as evaluated by moved distances, did not decrease until 20 min after antigen, followed by a progressive decrease. Reduced movement using a restraint maneuver not only reduced Tcore in nonsensitized rats but also augmented the anaphylactic hypothermia in the early phase (1.5-18 min) in sensitized rats. Combined antagonism against platelet-activating factor (PAF) and histamine H1 receptors abolished antigen-induced hypotension but only attenuated hypothermia. In conclusion, decreased locomotor activity, but not tail heat dissipation or decreased BAT activity, may at least in part contribute to this hypothermia. PAF and histamine are involved mainly in hypotension but only partly in hypothermia during rat anaphylaxis. NEW & NOTEWORTHY Anaphylactic shock is a life-threatening systemic hypotension. Hypothermia is observed during systemic anaphylaxis of rats. We determined the mechanism as follows: decreased locomotor activity, but not tail heat dissipation or decreased BAT activity, may at least in part contribute to this hypothermia. PAF and histamine are involved mainly in hypotension, but only partly in hypothermia during rat anaphylaxis. [ABSTRACT FROM AUTHOR]

Intracellular Ca2+ concentration ([Ca2+]i) is considered important in the regulation of skeletal muscle mass. This study tested the hypothesis that chronic repeated cooling and/or caffeine ingestion would acutely increase [Ca2+]i and hypertrophy muscles potentially in a fiber-type-dependent manner. Control rats and those fed caffeine were subjected to repeated bidiurnal treatments of percutaneous icing, under anesthesia, to reduce the muscle temperature below ~5°C. The predominantly fast-twitch tibialis anterior (TA) and slow-twitch soleus (SOL) muscles were evaluated after 28 days of intervention. The [Ca2+]i elevating response to icing was enhanced by caffeine loading only in the SOL muscle, with the response present across a significantly higher temperature range than in the TA muscle under caffeine-loading conditions. In both the TA and SOL muscles, myofiber cross-sectional area (CSA) was decreased by chronic caffeine treatment (mean reductions of 10.5% and 20.4%, respectively). However, in the TA, but not the SOL, CSA was restored by icing (+15.4 ± 4.3% vs. noniced, P < 0.01). In the SOL, but not TA, icing + caffeine increased myofiber number (20.5 ± 6.7%, P < 0.05) and satellite cell density (2.5 ± 0.3-fold) in cross sections. These contrasting muscle responses to cooling and caffeine may reflect fiber-type-specific [Ca2+]i responses and/or differential responses to elevated [Ca2+]i. [ABSTRACT FROM AUTHOR]

High K+ supplementation is correlated with a lower risk of the composite of death, major cardiovascular events, and ameliorated blood pressure, but the exact mechanisms have not been established. Inwardly rectifying K+ (Kir) channels expressed in the basolateral membrane of the distal nephron play an essential role in maintaining electrolyte homeostasis. Mutations in this channel family have been shown to result in strong disturbances in electrolyte homeostasis, among other symptoms. Kir7.1 is a member of the ATP-regulated subfamily of Kir channels. However, its role in renal ion transport and its effect on blood pressure have yet to be established. Our results indicate the localization of Kir7.1 to the basolateral membrane of aldosterone-sensitive distal nephron cells. To examine the physiological implications of Kir7.1, we generated a knockout of Kir7.1 (Kcnj13) in Dahl salt-sensitive (SS) rats and deployed chronic infusion of a specific Kir7.1 inhibitor, ML418, in the wild-type Dahl SS strain. Knockout of Kcnj13 (Kcnj13-/-) resulted in embryonic lethality. Heterozygous Kcnj13þ/- rats revealed an increase in K+ excretion on a normal-salt diet but did not exhibit a difference in blood pressure development or plasma electrolytes after 3 wk of a high-salt diet. Wild-type Dahl SS rats exhibited increased renal Kir7.1 expression when dietary K+ was increased. K+ supplementation also demonstrated that Kcnj13þ/- rats excreted more K+ on normal salt. The development of hypertension was not different when rats were challenged with high salt for 3 wk, although Kcnj13þ/- rats excrete less Na+. Interestingly, chronic infusion of ML418 significantly increased Na+ and Cl- excretion after 14 days of high salt but did not alter salt-induced hypertension development. Here, we found that reduction of Kir7.1 function, either through genetic ablation or pharmacological inhibition, can influence renal electrolyte excretion but not to a sufficient degree to impact the development of SS hypertension. [ABSTRACT FROM AUTHOR]

Recently, we have reported that the early progression of proteinuria in the obese Dahl salt-sensitive (SS) leptin receptor mutant (SSLepRmutant) strain was associated with increased renal macrophage infiltration before puberty. Macrophages can be divided into two distinct phenotypes: M1 (proinflammatory) and M2 (anti-inflammatory). Moreover, previous studies have demonstrated that interleukin (IL)-25 converts resting macrophages and M1 into M2. Therefore, the present study examined whether treatment with IL-25 would reduce the early progression of renal injury in SSLepRmutant rats by increasing renal M2. We also investigated the impact of IL-25 on M2 subtypes: M2a (wound healing/anti-inflammatory), M2b (immune mediated/proinflammatory), M2c (regulatory/anti-inflammatory), and M2d (tumor associated/proangiogenic). Four-wk-old SS and SSLepRmutant rats were treated with either control (IgG) or IL-25 (1 lg/day ip every other day) for 4 wk. The kidneys from SSLepRmutant rats displayed progressive proteinuria and renal histopathology versus SS rats. IL-25 treatment had no effect on these parameters in SS rats. However, in the SSLepRmutant strain, proteinuria was markedly reduced after IL-25 treatment. Chronic treatment with IL-25 significantly decreased glomerular and tubular injury and renal fibrosis in the SSLepRmutant strain. Although the administration of IL-25 did not change total renal macrophage infiltration in both SS and SSLepRmutant rats, IL-25 increased M2a by >50% and reduced M1 by 60% in the kidneys of SSLepRmutant rats. Overall, these data indicate that IL-25 reduces the early progression of renal injury in SSLepRmutant rats by inducing M2a and suppressing M1 and suggest that IL-25 may be a therapeutic target for renal disease associated with obesity. [ABSTRACT FROM AUTHOR]

Heart failure with preserved ejection fraction (HFpEF) is characterized by obesity, hypertension, diabetes mellitus, and chronic kidney disease. Obese ZSF1 rats, a model of HFpEF, exhibit multiple such comorbidities that can disturb cardiac function. Little attention has been paid to how these comorbidities affect renal disease in ZSF1 rats. HFpEF is found predominantly in women in whom obesity and hypertension are particularly prevalent. Therefore, we characterized the renal phenotype in female and male lean and obese ZSF1 rats and investigated additional effects of worsened hypertension on disease severity. Systolic blood pressure and renal function were assessed biweekly from 12 to 26 wk. From 19 wk, rats were implanted with either a deoxycorticosterone acetate pellet and fed a high-salt diet (DS) or a placebo pellet and fed a normal-salt diet. At 26 wk of age, terminal glomerular filtration rate was assessed via inulin clearance under isoflurane. Renal sections were processed for histological analysis. Lean and obese ZSF1 rats, both female and male, were mildly hypertensive (systolic blood pressure: 140-150 mmHg). All obese ZSF1 rats showed HFpEF. In female normoglycemic ZSF1 rats, obesity associated with mild proteinuria, decreased glomerular filtration rate, and glomerular hypertrophy. DS-worsened hypertension enhanced proteinuria and triggered glomerulosclerosis. Male obese ZSF1 rats were hyperglycemic and showed proteinuria, glomerular hypertrophy and sclerosis, and tubulointerstitial damage. DS-worsened hypertension aggravated this phenotype in male ZSF1 rats. In conclusion, female obese ZSF1 rats develop mild renal dysfunction and DS-worsened hypertension compromises renal function and structure in normoglycemic female obese ZSF1 rats as in hyperglycemic male obese ZSF1 rats. [ABSTRACT FROM AUTHOR]

Hereditary pulmonary veno-occlusive disease (hPVOD) is a severe form of autosomal recessive pulmonary hypertension and is due to biallelic loss of function of the EIF2AK4 gene (alias GCN2) coding for GCN2. GCN2 is a stress kinase that belongs to the integrated stress response pathway (ISR). Three rat lines carrying biallelic Gcn2 mutation were generated and found phenotypically normal and did not spontaneously develop a PVOD-related disease. We submitted these rats to amino acid deprivation to document the molecular and cellular response of the lungs and to identify phenotypic changes that could be involved in PVOD pathophysiology. Gcn2-/- rat lungs were analyzed under basal conditions and 3 days after a single administration of PEG-asparaginase (ASNase). Lung mRNAs were analyzed by RNAseq and single-cell RNAseq (scRNA-seq), flow cytometry, tissue imaging, and Western blots. The ISR was not activated after ASNase treatment in Gcn2-/- rat lungs, and apoptosis was increased. Several proinflammatory and innate immunity genes were overexpressed, and inflammatory cells infiltration was also observed in the perivascular area. Under basal conditions, scRNA-seq analysis of Gcn2-/- rat lungs revealed increases in two T-cell populations, a LAG3þ T-cell population and a proliferative T-cell population. Following ASNase administration, we observed an increase in calprotectin expression involved in TLR pathway activation and neutrophil infiltration. In conclusion, under basal and asparagine and glutamine deprivation induced by asparaginase administration, Gcn2-/- rats display molecular and cellular signatures in the lungs that may indicate a role for Gcn2 in immune homeostasis and provide further clues to the mechanisms of hPVOD development. [ABSTRACT FROM AUTHOR]

This article discusses the challenges of studying pulmonary hypertension (PH) in animal models, particularly in mice. PH is characterized by elevated pulmonary arterial pressure and pulmonary vascular resistance, and understanding its pathophysiological mechanisms is difficult due to the diverse clinical conditions associated with the disease. The article highlights the use of combined stimuli to induce experimental PH, similar to the "two-hit" model in humans, where two insults are required to induce clinical disease. The authors attempted to develop a new model of severe and persistent PH in mice by combining surgical pneumonectomy with different stimuli, but the results were essentially negative. The study emphasizes the resilience of mice to PH stimuli compared to rats and humans, but also acknowledges the progress made in the field of PH research through murine studies. The authors suggest considering the addition of multiple concurrent stimuli in future studies to better mimic the complexity of human disease. The article concludes by commending the authors for publishing a well-performed negative result, which adds to the scientific literature and helps other researchers avoid unnecessary efforts in creating the same model. [Extracted from the article]

The utility of rodents for research related to atrial fibrillation (AF) is growing exponentially. However, the obtained arrhythmic waveforms are often mixed with ventricular signals and the ability to analyze regularity and complexity of such events is limited. Recently, we introduced an implantable quadripolar electrode adapted for advanced atrial electrophysiology in ambulatory rats. Notably, we have found that the implantation itself leads to progressive atrial remodeling, presumably because of mechanical loading of the atria. In the present study, we developed an algorithm to clean the atrial signals from ventricular mixing and thereafter quantify the AF substrate in an objective manner based on waveform complexity. Rats were sequentially examined 1-, 4-, and 8-wk postelectrode implantation using a standard AF triggering protocol. Preburst ventricular mixing was sampled and automatically subtracted based on QRS detection in the ECG. Thereafter, the "pure" atrial signals were analyzed by Lempel-Ziv complexity algorithm and a complexity ratio (CR) was defined for each signal by normalizing the postburst to the preburst values. Receiver operating characteristic (ROC) curve analysis indicated an optimal CR cutoff of 1.236 that detected irregular arrhythmic events with high sensitivity (94.5%), specificity (93.1%), and area under the curve (AUC) (0.96, 95% confidence interval, 0.945-0.976). Automated and unbiased analysis indicated a gradual increase in signal complexity over time with augmentation of high frequencies in power spectrum analysis. Our findings indicate that CR algorithm detects irregularity in a highly efficient manner and can also detect the atrial remodeling induced by electrode implantation. Thus, CR analysis can strongly facilitate standardized AF research in rodents. [ABSTRACT FROM AUTHOR]

Stimulation of the mesencephalic locomotor region elicits exaggerated sympathetic nerve and pressor responses in spontaneously hypertensive rats (SHR) as compared with normotensive Wistar-Kyoto rats (WKY). This suggests that central command or its influence on vasomotor centers is augmented in hypertension. The decerebrate animal model possesses an ability to evoke intermittent bouts of spontaneously occurring motor activity (SpMA) and generates cardiovascular responses associated with the SpMA. It remains unknown whether the changes in sympathetic nerve activity and hemodynamics during SpMA are altered by hypertension. To test the hypothesis that the responses in renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) during SpMA are exaggerated with hypertension, this study aimed to compare the responses in decerebrate, paralyzed SHR, WKY, and normotensive Sprague-Dawley (SD) rats. In all strains, an abrupt increase in RSNA occurred in synchronization with tibial motor discharge (an index of motor activity) and was followed by rises in MAP and heart rate. The centrally evoked increase in RSNA and MAP during SpMA was much greater (306 ± 110%) in SHR than WKY (187 ± 146%) and SD (165 ± 44%). Although resting baroreflex-mediated changes in RSNA were not different across strains, mechanically or pharmacologically induced elevations in MAP attenuated or abolished the RSNA increase during SpMA in WKY and SD but had no effect in SHR. It is likely that the exaggerated sympathetic nerve and pressor responses during SpMA in SHR are induced along a central command pathway independent of the arterial baroreflex and/or result from central command-induced inhibition of the baroreflex. [ABSTRACT FROM AUTHOR]

Inhalation of noxious irritants activates nociceptive sensory afferent nerves innervating the airways, inducing reflex regulation of autonomic networks and the modulation of respiratory drive and cardiovascular (CV) parameters such as heart rate and blood pressure. In healthy mammals, irritant-evoked pulmonary-cardiac reflexes cause parasympathetic-mediated bradycardia. However, in spontaneously hypertensive (SH) rats, irritant inhalation also increases sympathetic drive to the heart. This remodeled pulmonary-cardiac reflex may contribute to cardiovascular risk caused by inhalation of air pollutants/irritants in susceptible individuals with cardiovascular disease (CVD). Previous studies have shown that the cooling mimic L-menthol, an agonist for the cold-sensitive transient receptor potential melastatin 8 (TRPM8), can alleviate nasal inflammatory symptoms and respiratory reflexes evoked by irritants. Here, we investigated the impact of inhalation of TRPM8 agonists L-menthol and WS-12 on pulmonary-cardiac reflexes evoked by inhalation of the irritant allyl isothiocyanate (AITC) using radiotelemetry. L-Menthol, but not its inactive analog d-menthol, significantly reduced the AITC-evoked reflex tachycardia and premature ventricular contractions (PVCs) in SH rats but had no effect on the AITC-evoked bradycardia in either SH or normotensive Wistar–Kyoto (WKY) rats. WS-12 reduced AITC-evoked tachycardia and PVCs in SH rats, but this more potent TRPM8 agonist also reduced AITC-evoked bradycardia. L-Menthol had no effect on heart rate when given alone, whereas WS-12 evoked a minor bradycardia in WKY rats. We conclude that stimulation of TRPM8-expressing afferents within the airways reduces irritant-evoked pulmonary-cardiac reflexes, especially the aberrant reflex tachyarrhythmia in SH rats. Airway menthol treatment may be an effective therapy for reducing pollution-associated CV exacerbations. [ABSTRACT FROM AUTHOR]

Quantitative data about the internal lung structure are needed to better understand normal and pathological lung development. Aberrant lung development causes deficits in alveolar and microvascular development; however, the normal temporal relationship between these processes is still not fully understood. We hypothesized that alveolar and capillary development show a differential time pattern. Lungs of rats aged 3, 7, 14, 21 days (d) or 3 mo (n = 8-10 each) were fixed by vascular perfusion and processed for light microscopy. Using design-based stereology number, the surface area and volume of alveoli, septal capillaries, and alveolar septa were quantified. The total number and the total volume of alveoli increased progressively during postnatal development. Interestingly, the numerical density of capillary loops was significantly higher in 14- and 21-d-old rats than before or after this age, causing a duplication of the total number of capillary loops between 1 and 2 wk of age. The mean thickness of alveolar septa started to decline slightly at the age of 14d and more pronounced at later stages. Although the septal epithelial surface area increased in proportion to alveolar number during the first 3 wk, the capillary endothelial surface area grew only slightly compared with the number of capillaries. In conclusion, the number of elements composing the alveolar capillary network expands massively during the first two postnatal weeks and exceeds the formation of alveoli. The thinning of the alveolar septa during further development suggests a reduction of the capillary network during alveolarization. [ABSTRACT FROM AUTHOR]

In people with type 1 diabetes, hypoglycemia can induce cardiac arrhythmias. In rodent experiments, severe hypoglycemia can induce fatal cardiac arrhythmias, especially so in diabetic models. Increased oxidative stress associated with insulin-deficient diabetes was hypothesized to increase susceptibility to severe hypoglycemia-induced fatal cardiac arrhythmias. To test this hypothesis, Sprague-Dawley rats were made insulin deficient with streptozotocin and randomized into two groups: 1) control (n = 22) or 2) vitamin E treated (four doses of α-tocopherol, 400 mg/kg, n = 20). Following 1 week of treatment, rats were either tested for cardiac oxidative stress or underwent a hyperinsulinemic-severe hypoglycemic (10-15 mg/dL) clamp with electrocardiogram recording. As compared with controls, vitamin E-treated rats had threefold less cardiac oxidative stress, sixfold less mortality due to severe hypoglycemia, and sevenfold less incidence of heart block. In summary, vitamin E treatment and the associated reduction of cardiac oxidative stress in diabetic rats reduced severe hypoglycemia-induced fatal cardiac arrhythmias. These results indicate that in the setting of diabetes, pharmacological treatments that reduce oxidative stress may be an effective strategy to reduce the risk of severe hypoglycemia-induced fatal cardiac arrhythmias. [ABSTRACT FROM AUTHOR]

Polycystic ovary syndrome (PCOS) is an extremely common endocrine-metabolic disorder and the main cause of infertility in premenopausal women, thus targeted treatments are sorely needed. Accumulative evidence showed that exogenous supplementation of IL-22 in PCOS mice may be of significant positive effect on insulin resistance (IR), a root causative factor for this condition, but much remained unknown about its mechanism. According to our previous study, troxerutin, a common anticoagulant and thrombolytic agent in clinic, alleviated various PCOS-like phenotypes in dihydrotestosterone (DHT)-treated rat model with unclear mechanism. Here, glucose tolerance tests (GTTs), insulin tolerance tests (ITTs), and homeostatic model assessment of insulin resistance (HOMA-IR) analyses revealed that troxerutin treatment in DHT-treated rats also significantly improved insulin resistance and enhanced serum IL-22 levels, which thereby activated IL-22R1/Janus kinase 1 (JAK1)/signal transducer and activator of transcription-3 (STAT3) signaling pathway in pancreatic islet. This protective effect of troxerutin on insulin resistance improvement was blocked by an inhibitor of p-STAT3, S3I-201. Troxerutin administration to DHT rats decreased the relative abundance of Bifidobacterium and enhanced secondary bile acid profiles, which were positively correlated with serum IL-22 concentration. Conclusively, the present study reported that troxerutin is an endogenous enhancer of IL-22 and the effect of troxerutin on insulin resistance improvement was via IL-22R1/JAK1/STAT3 signaling activation in a DHT-induced PCOS rat model. These insights may be translated into a primary therapeutic agent for PCOS with insulin resistance and hyperandrogenism. [ABSTRACT FROM AUTHOR]

Chronic kidney disease (CKD) is associated with an increased risk of cardiovascular disease (CVD). Despite the entry of sodium glucose cotransporter 2 (SGLT2) inhibitors, CKD persists as a medical challenge. Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibition reduces low-density lipoprotein (LDL)-cholesterol, a major risk factor of CVD. Interestingly, studies indicate that PCSK9 inhibition decreases proteinuria in kidney disease, complementing the reduced CVD risk. This study aimed to validate obese ZSF1 rats as a model for the renoprotective effects of PCSK9 and SGLT2 inhibition using alirocumab and empagliflozin for 15 wk. Obese rats revealed a significant reduction in measured glomerular filtration rate (mGFR) and increased urine albumin/creatinine ratio (UACR) during follow-up compared with lean controls. Alirocumab treatment resulted in a decline in mGFR and increased UACR compared with vehicle-treated obese rats. Immunohistochemistry showed increased fibrosis and inflammation in kidney tissue from obese rats treated with empagliflozin or alirocumab, whereas hepatic cholesterol and triglyceride levels were lowered compared with vehicle-treated obese rats. Although alirocumab lowered circulating free cholesterol levels throughout the treatment period, certain cholesteryl esters were increased at the end of the study, resulting in no overall difference in total cholesterol levels in the alirocumab group. Correspondingly, only a trend toward increased hepatic LDL-receptor levels was observed. In conclusion, these findings suggest that alirocumab treatment aggravates kidney dysfunction in obese ZSF1 rats. Moreover, in contrast to the renoprotective properties of empagliflozin observed in patients with CKD, empagliflozin did not ameliorate kidney disease progression in the obese ZSF1 rat. NEW & NOTEWORTHY New treatments to slow kidney disease progression and reduce cardiovascular disease risk are needed for chronic kidney disease (CKD). We investigated the cholesterol-lowering PCSK9 inhibitor alirocumab as a new treatment for proteinuric CKD and the effect of SGLT2 inhibition using empagliflozin in obese ZSF1 rats. Regarding renoprotection, our findings were contradictory with previous preclinical studies and clinical data, suggesting that different pathophysiological mechanisms may apply to this rat model.

The aim of this study was to test whether combined physical exercise training of moderate intensity executed during the development of monocrotaline (MCT)-induced pulmonary arterial hypertension (PAH) hinders the progression of pulmonary and right heart harmful functional and structural remodeling in rats. Wistar rats were injected with MCT (60 mg/kg) and after 24 h were exposed to a combined exercise training program: aerobic exercise (treadmill running-60 min/day; 60% of maximum running speed); and resistance exercise (vertical ladder climbing-15 climbs; 60% of maximum carrying load), on alternate days, 5 days/wk, for ∼3 wk. After euthanasia, the lung and right ventricle (RV) were excised and processed for histological, single myocyte, and biochemical analyses. Combined exercise increased the tolerance to physical effort (time until fatigue and relative maximum load) and prevented increases in pulmonary artery resistance (acceleration time (TA)/ejection time (TE)] and reductions in RV function [tricuspid annular plane systolic excursion (TAPSE)]. Moreover, in myocytes isolated from the RV, combined exercise preserved contraction amplitude, as well as contraction and relaxation velocities, and inhibited reductions in the amplitude and maximum speeds to peak and to decay of the intracellular Ca 2+ transient. Furthermore, combined exercise avoided RV (RV weight, cardiomyocyte, extracellular matrix, collagen, inflammatory infiltrate, and extracellular matrix) and lung (pulmonary alveoli and alveolar septum) harmful structural remodeling. In addition, combined exercise restricted RV [nitric oxide (NO) and carbonyl protein (CP)] and lung [catalase (CAT), glutathione S-transferase (GST), and NO] oxidative stress. In conclusion, the applied combined exercise regime hinders the progression of pulmonary and right heart functional and structural harmful remodeling in rats with MCT-induced PAH. NEW & NOTEWORTHY This study reveals that combined exercise improves tolerance to physical effort, prevents increases in pulmonary artery resistance, and conserves the right heart function during the progression of pulmonary arterial hypertension. Our analyses show that combined exercise hinders harmful right ventricular and lung structural remodeling and oxidative stress, which reflects in the maintenance of right ventricular myocytes' contractile function by preserving the intracellular calcium cycling. An attenuated progression of the disease impacts positively on its prognosis.

Emerging studies have reported the vital role of histone modification in the dysfunction of pulmonary vascular endothelial cells, which acts as the key reason to drive the hypoxia-induced pulmonary vascular remodeling and pulmonary hypertension (PH). This study aims to investigate the role of a histone 3 lysine 9 (H3K9) methyltransferase, SET domain bifurcated 1 (SETDB1), in hypoxia-induced functional and phenotypical changes of pulmonary vascular endothelial cells. Primarily cultured rat pulmonary microvascular endothelial cells (PMVECs) were used as cell model. Specific knockdown and overexpression strategies were used to systematically determine the molecular regulation and function of SETDB1 in PMVECs. SETDB1 is highly expressed and significantly upregulated in the pulmonary vascular endothelium of lung tissue isolated from SU5416/hypoxia-induced PH (SuHx-PH) rats and also in pulmonary arterial endothelial cells (PAECs) from patients with idiopathic pulmonary arterial hypertension (IPAH), comparing with their respective controls. In primarily cultured rat PMVECs, treatment of hypoxia or CoCl 2 induces significant upregulation of HIF2α, SETDB1, and H3K9me3. Specific knockdown and overexpression strategies indicate that the hypoxia- or CoCl 2 -induced upregulation of SETDB1 is mediated through a HIF2α-dependent mechanism. Knockdown of SETDB1 significantly inhibits the hypoxia- or CoCl 2 -induced apoptosis, senescence, and endothelial to mesenchymal transition (EndoMT) in rat PMVECs. Moreover, treatment of the specific inhibitor of histone methyltransferase, Chaetocin, effectively attenuates the disease pathogenesis of SuHx-PH in rat. Our results suggest that the HIF2α-dependent upregulation of SETDB1 facilitates hypoxia-induced functional and phenotypical changes of PMVECs, potentially contributing to the hypoxia-induced pulmonary vascular remodeling and PH. NEW & NOTEWORTHY Abnormal histone modification plays vital role in pulmonary hypertension (PH). This study reports the regulation and role of a histone 3 lysine 9 (H3K9) methyltransferase, SETDB1, in primarily cultured rat pulmonary microvascular endothelial cells (PMVECs). Hypoxia induces significant upregulation of SETDB1 at both mRNA and protein levels, in a HIF2α-dependent manner. The hypoxic upregulation of SETDB1 leads to significant apoptosis, senescence, and endothelial-to-mesenchymal transition in PMVECs. Treatment of a specific inhibitor of histone methyltransferase, Chaetocin, effectively attenuates the disease pathogenesis of PH rat model induced by SU5416/hypoxia.

Intrauterine inflammation from chorioamnionitis (CA) is associated with placental dysfunction and increased risk of bronchopulmonary dysplasia (BPD), the chronic lung disease of prematurity. Antenatal steroid (ANS) treatment improves early respiratory outcomes for premature infants. However, it remains unclear whether ANS improves long-term respiratory outcomes, and whether these effects are mediated through the improvement of placental dysfunction and/or direct impact on the fetal lung. We hypothesized that maternal ANS therapy preserves long-term lung development and impacts placental structural changes and gene expression in experimental CA with features of BPD. Pregnant rat dams were administered either saline (CTL), intra-amniotic (IA) endotoxin (ETX), ETX plus intramuscular (IM) betamethasone (ETX + BETA), or IM BM alone (BETA) on embryonic day 20 (E20). We collected placental tissue at delivery (E22) and infant lung tissue on the day of life (DOL) 14 . In comparison with controls, IA ETX had impaired infant lung growth and function. Maternal BM treatment of ETX-exposed pregnant dams reduced infant total lung resistance by 15.3% ( P < 0.05), improved infant lung compliance by 9.5% ( P < 0.05), preserved alveolar and vascular growth ( P < 0.05), and improved right ventricular hypertrophy (RVH) by 42.4% ( P < 0.05). ETX + BETA pregnancies were also associated with normalization of placental spiral artery modification and altered placental gene expression. These included the upregulation of placental prolactin, which has regulatory effects on pregnancy homeostasis and has been clinically associated with decreased BPD risk. The current study identifies parallel lung and placental changes associated with ANS treatment, providing a foundation for future studies to identify alternate antenatal therapies with more specific efficacy for BPD prevention. NEW & NOTEWORTHY We performed parallel neonatal lung and placental analyses in a preclinical model to characterize the impact of antenatal betamethasone in experimental chorioamnionitis. Antenatal steroids improved long-term respiratory outcomes and were associated with concurrent structural and molecular changes in the placenta. This study establishes an important model system for future analyses to evaluate mechanistic links determining whether the long-term impact of antenatal steroids on lung development may be through alteration of placental function.

Hyperbaric oxygen (HBO) refers to pure oxygen with a pressure greater than 1 atmospheres absolute (ATA), and when the pressure is too high, it can cause convulsive attacks. Adenosine and dopamine have been shown to be closely associated with HBO-induced convulsion seizures, and their receptors exhibited a coexisting relationship of mutual antagonism on the membrane of nerve cells. We explored the influence of adenosine and dopamine interplay on the occurrence of oxygen convulsion. Rats were individually exposed to HBO of 6 ATA and treated with adenosine, dopamine, and their receptor modulators separately and jointly, with the latency of convulsion onset recorded. In addition, after administering adenosine to rats and exposing them to HBO for 30 min, the content of dopamine and its metabolites and the activity of enzymes related to their metabolism were measured. The results revealed that dopamine was effective in resisting convulsion (>60 min vs. 32.53 ± 5.31 min, P = 0.000), and low-dose adenosine partially counteracted its effect (>60 min vs. 28.18 ± 6.24 min, P = 0.002). The combined use of adenosine A1 and dopamine D1 receptor modulators significantly impacted the incidence of convulsion. The activation or inhibition of the A2A receptor had a particularly significant impact on convulsion, whereas modulating the D2 receptor did not affect their effects. The combination of A1 agonist and D2 agonist was highly effective in resisting convulsion (>60 min vs. 32.53 ± 5.31 min, P = 0.000). Exposure to HBO accelerated the metabolism of dopamine to its end products, which may be related to the enhanced activity of monoamine oxidase (MAO). Adenosine can inhibit MAO activity (0.0766 ± 0.0150 U/mg.prot vs. 0.1055 ± 0.0086 U/mg.prot, P = 0.004), maintaining a higher level of dopamine (1.820 ± 0.379 mg/g vs. 0.602 ± 0.087 mg/g, P = 0.000). The study demonstrated that dopamine plays a significant role in oxygen convulsion and adenosine can affect dopamine metabolism. The interaction between them can have a crucial impact on the occurrence of oxygen convulsion. The findings offer a novel perspective for further investigating the mechanism of oxygen convulsion and exploring effective preventive strategies. NEW & NOTEWORTHY The interaction between adenosine and dopamine is critically important in determining the incidence of oxygen convulsion. Simultaneous regulation of both adenosine and dopamine offers a superior approach to counteract oxygen convulsion, achieving a synergistic effect exceeding the sum of their individual impacts. These findings provide new directions and insights for future in-depth and systematic exploration of the pathogenesis of central nervous system oxygen toxicity.

Integration of autonomic and metabolic regulation, including hepatic function, is a critical role played by the brain's hypothalamic region. Specifically, the paraventricular nucleus of the hypothalamus (PVN) regulates autonomic functions related to metabolism, such as hepatic glucose production. Although insulin can act directly on hepatic tissue to inhibit hepatic glucose production, recent evidence implicates that central actions of insulin within PVN also regulate glucose metabolism. However, specific central circuits responsible for insulin signaling with relation to hepatic regulation are poorly understood. As a heterogeneous nucleus essential to controlling parasympathetic motor output with notable expression of insulin receptors, PVN is an appealing target for insulin-dependent modulation of parasympathetic activity. Here, we tested the hypothesis that insulin activates hepatic-related PVN (PVN hepatic ) neurons through a parasympathetic pathway. Using transsynaptic retrograde tracing, labeling within PVN was first identified 24 h after its expression in the dorsal motor nucleus of the vagus (DMV) and 72 h after hepatic injection. Critically, nearly all labeling in medial PVN was abolished after a left vagotomy, indicating that PVN hepatic neurons in this region are part of a central circuit innervating parasympathetic motor neurons. Insulin also significantly increased the firing frequency of PVN hepatic neurons in this subregion. Mechanistically, rapamycin pretreatment inhibited insulin-dependent activation of PVN hepatic neurons. Therefore, central insulin signaling can activate a subset of PVN hepatic neurons that are part of a unique parasympathetic network in control of hepatic function. Taken together, PVN hepatic neurons related to parasympathetic output regulation could serve as a key central network in insulin's ability to control hepatic functions. NEW & NOTEWORTHY Increased peripheral insulin concentrations are known to decrease hepatic glucose production through both direct actions on hepatocytes and central autonomic networks. Despite this understanding, how (and in which brain regions) insulin exerts its action is still obscure. Here, we demonstrate that insulin activates parasympathetic hepatic-related PVN neurons (PVN hepatic ) and that this effect relies on mammalian target of rapamycin (mTOR) signaling, suggesting that insulin modulates hepatic function through autonomic pathways involving insulin receptor intracellular signaling cascades.

Aging is an intricate and gradual process characterized by tissue and cellular dysfunction. Adipose-derived mesenchymal stem cells (ADMSCs) experience a functional decline as part of systemic aging. However, the alterations in ADMSCs across various anatomical sites throughout an individual's lifespan remain unclear. To shed light on these changes, we collected white adipose tissue and brown adipose tissue samples from the epididymis, perirenal, inguinal, and scapular regions of young, adult, and aged rats and subsequently isolated ADMSCs for RNA sequencing. As aging progressed, we observed a reduction in the number of ADMSCs at all anatomical sites. Marker genes of ADMSCs from different sites were identified. Aging triggered notable activation of inflammatory and immune responses while diminishing the ADMSC differentiation capacity and ability to maintain a normal tissue morphology. Furthermore, miR-195-5p and miR-497-3p, which promoted cell senescence and apoptosis while inhibiting proliferation and differentiation, were positively correlated with aging. These findings increase our understanding of ADMSC senescence and underscore the unique physiological changes and functions of ADMSCs across different anatomical sites during aging. NEW & NOTEWORTHY Dynamic changes in mRNAs and miRNAs of ADMSCs during aging are shown. As aging progressed, we observed a reduction in the number of ADMSCs at all anatomical sites. Aging leads to the activation of inflammatory and cellular dysfunction. miR-195-5p and miR-497-3p are positively correlated with aging, which promoted cell senescence and apoptosis while inhibiting proliferation and differentiation. ADMSCs associated with different anatomical sites have site-specific markers.

Iron deficiency (ID) is common during gestation and early infancy and can alter developmental trajectories with lasting consequences on cardiovascular health. Iron plays a critical role in systemic oxygen transport (via hemoglobin) and aerobic respiration (as a component of mitochondrial complexes). Perinatal ID has been shown to cause cardiac dysfunction in neonates, but the mechanisms underlying these changes have not been characterized. Here, we examined the effects of perinatal ID on cardiac mitochondrial function in rats in the early postnatal period. Female rats were fed an iron-restricted or iron-replete diet before and during pregnancy. Offspring hearts were collected postmortem for quantitative shotgun proteomic analysis [postnatal days (PD) 0 and 28] and mitochondrial function was assessed by high-resolution respirometry (at PD 0, 14, and 28). Markers of oxidative stress were measured by fluorescence microscopy and assessment of antioxidant gene expression profiles. Both male and female ID pups had reduced body weight and increased relative heart weights at all time points assessed, despite recovering from anemia by PD28. Proteomics analysis revealed dysregulation of mitochondrial proteins by ID, and these differences were most pronounced in males. In male hearts, ID increased mitochondrial content and decreased normalized mitochondrial respiration through the NADH-pathway, succinate-pathway, and fatty acid oxidation (FAO)-pathway. In conclusion, ID causes changes in cardiac mitochondrial function in neonates, which may reflect inadequate or maladaptive compensation during the transition from intrauterine to extrauterine life. Furthermore, the results presented herein, which were stratified by offspring sex, underscore the need for follow-up studies to directly assess differences in how male and female offspring cope with ID as a perinatal stressor. NEW & NOTEWORTHY Iron deficiency (ID) is the most common nutritional deficiency worldwide and is highly prevalent among pregnant women and young children. ID causes changes in mitochondrial protein expression and function in neonatal hearts, which may contribute to functional impairments. Improving cardiac energy metabolism may represent a novel approach to improve short- and long-term outcomes in infants affected by ID, but sex of the neonate may be an important determinant of treatment efficacy.

Epidermal growth factor (EGF) has important effects in the renal collecting duct to regulate salt and water transport. To identify elements of EGF-mediated signaling in the rat renal inner medullary collecting duct (IMCD), we carried out phosphoproteomic analysis. Biochemically isolated rat IMCD suspensions were treated with 1 µM of EGF or vehicle for 30 min. We performed comprehensive quantitative phosphoproteomics using tandem mass tag (TMT)-labeling of tryptic peptides followed by protein mass spectrometry. We present a data resource reporting all detected phosphorylation sites and their changes in response to EGF. For a total of 29,881 unique phosphorylation sites, 135 sites were increased and 119 sites were decreased based on stringent statistical analysis. The data are provided to users at https://esbl.nhlbi.nih.gov/Databases/EGF-phospho/. The analysis demonstrated that EGF signals through canonical EGF pathways in the renal IMCD. Analysis of Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways in which EGF-regulated phosphoproteins are over-represented in native rat IMCD cells confirmed mapping to RAF-MEK-extracellular signal-regulated kinase (ERK) signaling but also pointed to a role for EGF in the regulation of protein translation. A large number of phosphoproteins regulated by EGF contained PDZ domains that are key elements of epithelial polarity determination. We also provide a collecting duct EGF-network map as a user-accessible web resource at https://esbl.nhlbi.nih.gov/Databases/EGF-network/. Overall, the phosphoproteomic data presented provide a useful resource for experimental design and modeling of signaling in the renal collecting duct. NEW & NOTEWORTHY EGF negatively regulates transepithelial water and salt transport across the kidney collecting duct. This study identified phosphoproteins affected by EGF stimulation in normal rat collecting ducts, providing insights into global cell signaling mechanisms. Bioinformatic analyses highlighted enhanced canonical ERK signaling alongside a diminished activity in the PI3K-Akt pathway, which is crucial for cell proliferation and survival. This EGF response differs somewhat from prior studies where both pathways were prominently activated.

We tested the hypothesis that attenuation of the circulating insulin level in rats during early life can provide sustained protection against diet-induced obesity and metabolic dysfunction in adulthood. Male Wistar rats received intraperitoneal scopolamine butylbromide (SB) during the first 12 days of suckling, whereas control rats received 0.9% saline injections. The animals were weaned on day 21 and fed a normal chow diet. At 60 days of age, the control and SB groups were fed a normal chow diet (ND; 4.5% fat) or a high-fat diet (HF; 35% fat) until 90 days of age to induce obesity and metabolic dysfunction. Insulin secretion, food intake, and body weight were measured. Pancreatic islet function, autonomic nervous system function, and glucose homeostasis were evaluated at 90 days of age. During lactation, the plasma insulin concentration was significantly lower in the SB groups than in the control group. SB rats also exhibited reduced body weight. The HF diet resulted in obesity, glucose intolerance, insulin resistance, disruption of insulin secretion, and vagal hyperactivity in adult control rats. Remarkably, SB-treated rats fed the HF diet showed attenuated body weight and adiposity and did not develop diet-induced glucose/insulin imbalance. In addition, vagal activity and adequate pancreatic islet insulin secretion were preserved. Offspring exposed to SB during early life are provided with long-lasting protection against obesity and metabolic complications induced by an HF diet. An attenuated circulating insulin level in early life may have far-reaching consequences on metabolic programming. NEW & NOTEWORTHY High insulin levels during early life may lead to the late development of obesity and diabetes. We investigated whether attenuation of insulin levels by using an antimuscarinic agent could prevent diet-induced obesity and diabetes. Rats' early exposure to an antimuscarinic agent reduced insulin levels during the lactation period and promoted protection against obesity and metabolic dysfunctions. Independent of the programming mechanisms, insulin levels during early life may be a defining factor of health or diseases later in life.

As cardiomyocyte loss causes heart failure, inhibition of cardiomyocyte death may be a therapeutic strategy against heart failure. In this study, we have identified defender against cell death 1 (Dad1) as a candidate regulator of cardiomyocyte death, using complementary DNA microarray and siRNA knockdown screening. Dad1 is a subunit of oligosaccharyltransferase (OST) complex that is responsible for protein N-glycosylation; however, its function in cardiomyocytes remains unknown. Importantly, the knockdown of Dad1 using siRNA reduced the viability of neonatal rat cardiomyocytes (NRCMs), accompanied by cleaved caspase3 expression, independent of endoplasmic reticulum stress. Dad1 knockdown impaired cell spreading and reduced myofibrillogenesis in NRCMs, suggesting that Dad1 knockdown induced anoikis, apoptosis by disrupting cell-matrix interactions. Consistently, knockdown of Dad1 impaired N-glycosylation of integrins α5 and β1, accompanied by inactivation of focal adhesion kinase. When cell adhesion was enhanced using adhesamine, fibronectin, or collagen type IV, cardiomyocyte death induced by Dad1 knockdown was reduced. Dad1 knockdown decreased the expression of staurosporine and temperature-sensitive 3 A (Stt3A), a catalytic subunit of OST complex. Interestingly, Stt3A knockdown using Stt3A siRNA reduced the expression of Dad1, indicating that both Dad1 and Stt3A were required for OST stabilization. In conclusion, Dad1 plays an important role in maintaining the expression of mature N-glycosylated integrins and their downstream signaling molecules to suppress cardiomyocyte anoikis. NEW & NOTEWORTHY This study found for the first time that the knockdown of Dad1 induced cardiomyocyte death, accompanied by impairment of myofibrillogenesis and cell spreading. Dad1 regulates the N-glycosylation of integrins in cooperation with Stt3A and preserves cell adhesion activity, promoting cardiomyocyte survival. This is the first demonstration that Dad1 contributes to the maintenance of cardiac homeostasis through the posttranslational modification of integrins, providing a novel insight into the biological significance of OST complex in cardiomyocytes.

Right ventricular (RV) function determines outcomes in RV pressure loading. A better understanding of the time-course and regional distribution of RV remodeling may help optimize targets and timing for therapeutic intervention. We sought to characterize RV remodeling between zero and 6 wk after the initiation of RV pressure loading. Thirty-six rats were randomized to either sham surgery or to pulmonary artery banding (PAB). After echocardiography and conductance catheter studies, groups of rats were euthanized at 1 wk, 3 wk, and 6 wk after sham surgery, or induction of RV pressure loading, for RV histological, RNA, and molecular analysis. A vigorous inflammatory response characterized by increased RV inflammatory cytokines, chemokines, and macrophage markers was observed at 1 wk following PAB. Metabolic changes, transforming growth factor-β (TGF-β)1 canonical signaling, collagenous fibrosis deposition, and apoptosis were already significantly increased by 1 wk after PAB. Genes marking fibroblast activation were upregulated at 1 wk but not at 6 wk post-PAB surgery. Mitochondrial dysfunction was evidenced by increased pyruvate dehydrogenase kinase (PDK) activity and decreased pyruvate dehydrogenase (PDH) phosphorylation significantly at 6-wk post-PAB. These processes preceded the development of overt myocardial hypertrophy and impaired echo parameters of systolic and diastolic function that occurred significantly from 3 wk after PAB. RV myocardial inflammation, metabolic shift, metabolic gene transcription, and profibrotic signaling occur early after initiation of pressure loading when RV pressures are only moderately elevated, before the development of overt myocardial hypertrophy and dysfunction, suggesting that adaptive hypertrophy and maladaptive remodeling occur simultaneously. These results suggest that therapeutic intervention to reduce adverse RV remodeling may be needed earlier and at lower thresholds than currently used. NEW & NOTEWORTHY Exploring the dynamics of right ventricular remodeling: unveiling the intricate interplay between inflammation, metabolic shifts, and fibrotic signaling in response to pressure loading. Through a comprehensive study spanning from initiation to 6 wk post-pressure loading, our research sheds light on the early onset of crucial molecular processes preceding overt hypertrophy and dysfunction. These findings challenge conventional intervention timing, advocating for early, targeted therapeutic strategies to mitigate adverse remodeling in right ventricular pressure loading.

This study was designed to investigate the effects of three hypoabsorptive bariatric surgeries, namely Roux-en-Y gastric bypass (RYGB), biliopancreatic diversion with duodenal switch (BPD-DS), and single anastomosis duodeno-ileal bypass with sleeve gastrectomy (SADI-S), on bile acids (BAs) and to assess whether the changes in BA plasma levels were associated with the metabolic and homeostatic effects of the surgeries. Male Wistar rats, either fed a high- (HF) or a low-fat (LF) diet, were divided into seven groups: RYGB HF, BPD-DS HF, SADI-S HF, sleeve-gastrectomy (SG) HF, sham-operated (Sham) HF, Sham LF, and Sham HF-pair-weighed to BPD-DS (Sham HF-PW). The rats were treated for 56 days. The results demonstrate the ability of RYGB, BPD-DS, and SADI-S to raise plasma levels of BAs, whose elevations were likely due to changes in gene expression of transporters, enzymes, and receptors in the liver and small intestine. This increase, most notably of the secondary BAs (deoxycholic acid, ursodeoxycholic acid, and lithocholic acid), was negatively associated with body weight gain, fat gain, and fasting insulin levels, and positively with plasma peptide tyrosine-tyrosine (PYY). Plasma BAs also correlated positively with the fecal levels of Clostridium , Sutterella , and Enterobacteriaceae and negatively with Clostridiales&#95;f&#95;g&#95;2, Christensenellaceae , Ruminococcaceae&#95;g&#95;2, Oscillibacter , and Oscillospira . In addition, they are associated positively with the short-chain fatty acid (SCFA) levels of propionate, butyrate, isobutyrate, valerate, and isovalerate. Altogether, the present study emphasizes the ability of RYGB, BPD-DS, and SADI-S to induce circulating BA elevations that predict the beneficial consequences of those hypoabsorptive bariatric surgeries on energy and glucose homeostasis and circulating levels of PYY. The present results also reveal close associations between plasma BAs and SCFAs, whose variations following hypoabsorptive surgeries are linked to significant fat losses and metabolic health improvements. NEW & NOTEWORTHY The study emphasizes the ability of RYGB, BPD-DS, and SADI-S to induce elevated circulating bile acids levels and changes in the gene expression of transporters, enzymes and receptors in the liver and small intestine, predicting positive effects on energy and glucose homeostasis as well as PYY levels. The present results also reveal close associations between plasma BAs and SCFAs, whose variations following hypoabsorptive surgeries are also linked to significant fat losses and metabolic health improvements. These findings provide valuable insights into the mechanisms underlying the positive effects of these surgical interventions.

Kratom ( Mitragyna speciosa ), containing the primary alkaloid mitragynine, has emerged as an alternative self-treatment for opioid use disorder. Mitragynine binds numerous receptor types, including opioid receptors, which are known to modulate food consumption. However, the ability of acute mitragynine to modulate food consumption remains unknown. The current study assessed the effects of acute mitragynine or morphine administration on unconditioned food and water intake in 16 Sprague-Dawley rats. Food and water intake changes were monitored in response to morphine, mitragynine (1.78-56 mg/kg ip), saline, or vehicle controls for 12 h, starting at the onset of the dark cycle. Naltrexone pretreatment was used to examine pharmacological specificity. Both morphine and mitragynine demonstrated a biphasic food intake dose-effect, with low doses (5.6 mg/kg) increasing and high doses (56 mg/kg) decreasing food intake. All morphine doses reduced water intake; however, only the highest dose of mitragynine (56 mg/kg) reduced water intake. Naltrexone attenuated both stimulatory and inhibitory effects of morphine on food intake, but only the stimulatory effect of mitragynine. In conclusion, low doses of mitragynine stimulate food intake via opioid-related pathways, while high doses likely recruit other targets. NEW & NOTEWORTHY This study reveals that morphine and the kratom alkaloid mitragynine produce dose-dependent effects on feeding in rats. Low doses stimulate food intake via opioid pathways, while high doses decrease consumption through nonopioid mechanisms. Morphine potently suppresses water intake at all doses, whereas only high doses of mitragynine reduce drinking. These findings provide novel insights into the complex opioid and nonopioid mechanisms underlying the effects of mitragynine on ingestive behaviors.