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The progression of chronic kidney disease (CKD) occurs faster in males (M); however, the underlying mechanisms remain poorly understood. A potential contributing factor to sex differences in CKD progression is differences in glomerular capillary morphology. Glomerular capillaries with larger radii or longer lengths may predispose capillaries to barotrauma-induced injury. The goal of the present study was to assess glomerular capillary morphology in M vs. female (F) rats with intact kidneys and after uninephrectomy (UNX). We hypothesized that glomerular capillary radii (RCAP) and length (LCAP) would be greater in M rats.This study utilized rats (8-11-week old, Charles River) with intact kidneys (n=4 M, n=4 F) and UNX (n=4 M, n=4 F) provided a 0.4% NaCl diet and water ad libitum. Kidneys were perfusion-fixed with 2% paraformaldehyde and 2.5% glutaraldehyde in phosphate buffered saline (pH=7.4). The left kidney was excised and a 3 mm transverse section through the midline of the kidney was selected for further processing. Multiple 1 mm3 cubes were randomly excised from the left, middle, and right regions of the outer cortex and embedded in EPONTM. The 1 mm3 cubes were sectioned (1 μm), placed on slides, and stained with toluidine blue. Four glomeruli from each region were randomly selected for stereological analysis using STEPanizer (Tschanz SA et al. J. Microsc. 243(1):47-59, 2011). Values of RCAP and LCAP were derived using standard stereological formulas (Nyengaard JR J. Am. Soc. Nephrol. 10(5):1100-1123, 1999). Effects of sex and UNX on body weight (BW), kidney weight (KW), glomerular volume (VG), RCAP, and LCAP were assessed with a 2-way ANOVA with Sidak post hoc analysis. All data are mean±SE and PG (0.65± 0.02 vs. 0.62±0.04 x 106 μm3), RCAP (2.8±0.1 vs. 2.8±0.1 μm), or LCAP (7.3±0.3 vs. 7.1±0.1 mm). In F rats with UNX vs. intact kidneys, BW was not significantly different; however, KW, VG, RCAP, and LCAP were 1.1-1.6-fold greater (PG were 2-3.5-fold greater (PCAP and LCAP were 1.1-1.5-fold greater (PG (1.3±0.1 vs. 1.0±0.01 x 106 μm3), and LCAP (11.0±0.3 vs. 9.7±0.3 mm) as compared to F rats with UNX. RCAP was similar in M and F rats with UNX (3.3±0.1 vs. 3.1±0.1 μm). These data indicate that M rats exhibit greater compensatory increases in LCAP following UNX as compared to F rats. The greater LCAP in M may result in reduced podocyte density and contribute, in part, to their greater susceptibility to hypertensive injury and CKD progression as compared to F rats. NIH (HL150467) and East Tennessee State University This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

There is limited data on sex differences in renal hemodynamics and autoregulation (AR). We assessed renal vascular resistance (RVR) and renal AR in conscious male (M) and female (F) rats with intact kidneys, uninephrectomy (UNX), and UNX administered deoxycorticosterone acetate (DOCA)+salt. The susceptibility to hypertension-induced renal damage (HIRD) was also assessed in rats with UNX administered DOCA+salt. We hypothesized that M rats exhibit a lower RVR, weaker renal AR responses, and an increased susceptibility to HIRD.Sprague-Dawley rats (8-11-week-old) administered a 0.4% NaCl chow diet and water ad libitum were used. Blood pressure (BP, DSI) and renal blood flow (RBF, Transonic) were assessed in conscious rats with intact kidneys (n=12M,13F) and in rats with UNX (n=7M,8F) before and during 1 week of DOCA (3.3 mg/day s.c.) + salt (1% NaCl in drinking water). RVR was calculated as BP/RBF. Renal AR responses (fractional Δ in RBF / fractional Δ in BP) at 2.5 seconds and 20 seconds following spontaneous BP fluctuations ≥ ±5 mmHg were calculated as recently described (PMID 31792155). A 2-way ANOVA with Sidak post hoc analysis was used to assess differences among groups. Data are mean±SE, and P NIH (HL154067) and East Tennessee State University This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Tissue electrolyte measurements from 5/6 nephrectomized rats treated with either 2 weeks of 0.1M NaHCO3 or equimolar NaCl.

Impairments in insulin sensitivity can occur in patients with chronic kidney disease (CKD). Correction of metabolic acidosis has been associated with improved insulin sensitivity in CKD, suggesting that metabolic acidosis may directly promote insulin resistance. Despite this, the effect of acid or alkali loading on insulin sensitivity in a rodent model of CKD (remnant kidney) has not been directly investigated. Such studies could better define the relationship between blood pH and insulin sensitivity. We hypothesized that in remnant kidney rats, acid or alkali loading would promote loss of pH homeostasis and consequently decrease insulin sensitivity. To test this hypothesis, we determined the impact of alkali (2 wk) or acid (5–7 days) loading on plasma electrolytes, acid-base balance, and insulin sensitivity in either sham control rats, 2/3 nephrectomized rats, or 5/6 nephrectomized rats. Rats with 5/6 nephrectomy had the greatest response to insulin followed by rats with 2/3 nephrectomy and sham control rats. We found that treatment with 0.1 M sodium bicarbonate solution in drinking water had no effect on insulin sensitivity. Acid loading with 0.1 M ammonium chloride resulted in significant reductions in pH and plasma bicarbonate. However, acidosis did not significantly impair insulin sensitivity. Similar effects were observed in Zucker obese rats with 5/6 nephrectomy. The effect of renal mass reduction on insulin sensitivity could not be explained by reduced insulin clearance or increased plasma insulin levels. We found that renal mass reduction alone increases sensitivity to exogenous insulin in rats and that this is not acutely reversed by the development of acidosis. NEW & NOTEWORTHY Impairments in insulin sensitivity can occur in patients with chronic kidney disease, and previous work has suggested that metabolic acidosis may be the underlying cause. Our study investigated the effect of acid or alkali loading on insulin sensitivity in a rodent model of chronic kidney disease. We found that renal mass reduction increases the blood glucose response to insulin and that this is not acutely reversed by the development of acidosis.

Donepezil is a centrally acting acetylcholinesterase (AChE) inhibitor with therapeutic potential in inflammatory diseases; however, the underlying autonomic and cholinergic mechanisms remain unclear. Here, we assessed effects of donepezil on mean arterial pressure (MAP), heart rate (HR), HR variability, and body temperature in conscious adult male C57BL/6 mice to investigate the autonomic pathways involved. Central versus peripheral cholinergic effects of donepezil were assessed using pharmacological approaches including comparison with the peripherally acting AChE inhibitor, neostigmine. Drug treatments included donepezil (2.5 or 5 mg/kg sc), neostigmine methyl sulfate (80 or 240 μg/kg ip), atropine sulfate (5 mg/kg ip), atropine methyl bromide (5 mg/kg ip), or saline. Donepezil, at 2.5 and 5 mg/kg, decreased HR by 36 ± 4% and 44 ± 3% compared with saline ( n = 10, P < 0.001). Donepezil, at 2.5 and 5 mg/kg, decreased temperature by 13 ± 2% and 22 ± 2% compared with saline ( n = 6, P < 0.001). Modest ( P < 0.001) increases in MAP were observed with donepezil after peak bradycardia occurred. Atropine sulfate and atropine methyl bromide blocked bradycardic responses to donepezil, but only atropine sulfate attenuated hypothermia. The pressor response to donepezil was similar in mice coadministered atropine sulfate; however, coadministration of atropine methyl bromide potentiated the increase in MAP. Neostigmine did not alter HR or temperature, but did result in early increases in MAP. Despite the marked bradycardia, donepezil did not increase normalized high-frequency HR variability. We conclude that donepezil causes marked bradycardia and hypothermia in conscious mice via the activation of muscarinic receptors while concurrently increasing MAP via autonomic and cholinergic pathways that remain to be elucidated.

Noninvasive determination of the severity of parenchymal injury in acute kidney injury remains challenging. Edema is an early pathological process following injury, which may correlate with changes in kidney volume. The goal of the present study was to test the hypothesis that “increases in kidney volume measured in vivo using ultrasound correlate with the degree of renal parenchymal injury.” Ischemia-reperfusion (IR) of varying length was used to produce graded tissue injury. We first determined 1) whether regional kidney volume in rats varied with the severity (0, 15, 30, and 45 min) of warm bilateral IR and 2) whether this correlated with tubular injury score. We then determined whether these changes could be measured in vivo using three-dimensional ultrasound. Finally, we evaluated cumulative changes in kidney volume up to 14 days post-IR in rats to determine whether changes in renal volume were predictive of latent tubular injury following recovery of filtration. Experiments concluded that noninvasive ultrasound measurements of change in kidney volume over 2 wk are predictive of tubular injury following IR even in animals in which plasma creatinine was not elevated. We conclude that ultrasound measurements of volume are a sensitive, noninvasive marker of tissue injury in rats and that the use of three-dimensional ultrasound measurements may provide useful information regarding the timing, severity, and recovery from renal tissue injury in experimental studies.

Brown-Norway chromosome 1 mitigates the upregulation of pro-inflammatory pathways in mTAL cells and subsequent age-related CKD in Dahl SS/JrHsdMcwi Rats

Background Abnormal renal hemodynamic responses to salt‐loading are thought to contribute to salt‐sensitive (SS) hypertension. However, this is based largely on studies in anesthetized animals, and little data are available in conscious SS and salt‐resistant rats. Methods and Results We assessed arterial blood pressure, renal function, and renal blood flow during administration of a 0.4% NaCl and a high‐salt (4.0% NaCl) diet in conscious, chronically instrumented 10‐ to 14‐week‐old Dahl SS and consomic SS rats in which chromosome 1 from the salt‐resistant Brown‐Norway strain was introgressed into the genome of the SS strain (SS.BN1). Three weeks of high salt intake significantly increased blood pressure (20%) and exacerbated renal injury in SS rats. In contrast, the increase in blood pressure (5%) was similarly attenuated in Brown‐Norway and SS.BN1 rats, and both strains were completely protected against renal injury. In SS.BN1 rats, 1 week of high salt intake was associated with a significant decrease in renal vascular resistance (−8%) and increase in renal blood flow (15%). In contrast, renal vascular resistance failed to decrease, and renal blood flow remained unchanged in SS rats during high salt intake. Finally, urinary sodium excretion and glomerular filtration rate were similar between SS and SS.BN1 rats during 0.4% NaCl and high salt intake. Conclusions Our data support the concept that renal vasodysfunction contributes to blood pressure salt sensitivity in Dahl SS rats, and that genes on rat chromosome 1 play a major role in modulating renal hemodynamic responses to salt loading and salt‐induced hypertension.

Acute kidney injury (AKI) is a major complication in hospitalized patients and is associated with elevated mortality rates. Numerous recent studies indicate that AKI also significantly increases the risk of chronic kidney disease (CKD), end-stage renal disease (ESRD), hypertension, cardiovascular disease, and mortality in those patients who survive AKI. Moreover, the risk of ESRD and mortality after AKI is substantially higher in patients with preexisting CKD. However, the underlying mechanisms by which AKI and CKD interact to promote ESRD remain poorly understood. The recently developed models that superimpose AKI on rodents with preexisting CKD have provided new insights into the pathogenic mechanisms mediating the deleterious interactions between AKI and CKD. These studies show that preexisting CKD impairs recovery from AKI and promotes the development of mechanisms of CKD progression. Specifically, preexisting CKD exacerbates microvascular rarefaction, failed tubular redifferentiation, disruption of cell cycle regulation, hypertension, and proteinuria after AKI. The purpose of this review is to discuss the potential mechanisms by which microvascular rarefaction and hypertension contribute to impaired recovery from AKI and the subsequent progression of renal disease in preexisting CKD states.

A convolutional deep neural network is employed to assess renal autoregulation using time series of arterial blood pressure and blood flow rate measurements in conscious rats. The network is trained using representative data samples from rats with intact autoregulation and rats whose autoregulation is impaired by the calcium channel blocker amlodipine. Network performance is evaluated using test data of the types used for training, but also with data from other models for autoregulatory impairment, including different calcium channel blockers and also renal mass reduction. The network is shown to provide effective classification for impairments from calcium channel blockers. However, the assessment of autoregulation when impaired by renal mass reduction was not as clear, evidencing a different signature in the hemodynamic data for that impairment model. When calcium channel blockers were given to those animals, however, the classification again was effective.

Supplementary methods and figures for the manuscript, "Renal Mass Reduction Increases the Response to Exogenous Insulin Independent of Acid-Base Status or Plasma Insulin Levels in Rats".

Unilateral ischemia-reperfusion (UIR) injury leads to progressive renal atrophy and tubulointerstitial fibrosis (TIF) and is commonly used to investigate the pathogenesis of the acute kidney injury-chronic kidney disease transition. Although it is well known that contralateral nephrectomy (CNX), even 2 wk post-UIR injury, can improve recovery, the physiological mechanisms and tubular signaling pathways mediating such improved recovery remain poorly defined. Here, we examined the renal hemodynamic and tubular signaling pathways associated with UIR injury and its reversal by CNX. Male Sprague-Dawley rats underwent left UIR or sham UIR and 2 wk later CNX or sham CNX. Blood pressure, left renal blood flow (RBF), and total glomerular filtration rate were assessed in conscious rats for 3 days before and over 2 wk after CNX or sham CNX. In the presence of a contralateral uninjured kidney, left RBF was lower ( P < 0.05) from 2 to 4 wk following UIR (3.6 ± 0.3 mL/min) versus sham UIR (9.6 ± 0.3 mL/min). Without CNX, extensive renal atrophy, TIF, and tubule dedifferentiation, but minimal pimonidazole and hypoxia-inducible factor-1α positivity in tubules, were present at 4 wk post-UIR injury. Conversely, CNX led ( P < 0.05) to sustained increases in left RBF (6.2 ± 0.6 mL/min) that preceded the increases in glomerular filtration rate. The CNX-induced improvement in renal function was associated with renal hypertrophy, more redifferentiated tubules, less TIF, and robust pimonidazole and hypoxia-inducible factor-1α staining in UIR injured kidneys. Thus, contrary to expectations, indexes of hypoxia are not observed with the extensive TIF at 4 wk post-UIR injury in the absence of CNX but are rather associated with the improved recovery of renal function and structure following CNX.

Supplementary methods and figure for intraperitoneal versus intravenous administration of exogenous insulin for insulin tolerance testing in rats.

Background Renal autoregulation maintains stable renal function despite BP fluctuations and protects glomerular capillaries from hypertensive injury. However, real-time dynamics of renal autoregulation in conscious animals have not been characterized. Methods To develop novel analytic methods for assessing renal autoregulation, we recorded concurrent BP and renal blood flow in conscious rats, comparing animals with renal autoregulation that was intact versus impaired (from 3/4 nephrectomy), before and after additional impairment (from the calcium channel blocker amlodipine). We calculated autoregulatory indices for adjacent short segments of increasing length (0.5, 1, 2.5, 5, 10, and 20 seconds) that exhibited a mean BP difference of at least 5 mm Hg. Results Autoregulatory restoration of renal blood flow to baseline after BP changes in conscious rats occurs rapidly, in 5-10 seconds. The response is significantly slower in states of impaired renal autoregulation, enhancing glomerular pressure exposure. However, in rats with severe renal autoregulation impairment (3/4 nephrectomy plus amlodipine), renal blood flow in conscious animals (but not anesthetized animals) was still restored to baseline, but took longer (15-20 seconds). Consequently, the ability to maintain overall renal blood flow stability is not compromised in conscious rats with impaired renal autoregulation. Conclusions These novel findings show the feasibility of renal autoregulation assessment in conscious animals with spontaneous BP fluctuations and indicate that transient increases in glomerular pressure may play a greater role in the pathogenesis of hypertensive glomerulosclerosis than previously thought. These data also show that unidentified mechanosensitive mechanisms independent of known renal autoregulation mechanisms and voltage-gated calcium channels can maintain overall renal blood flow and GFR stability despite severely impaired renal autoregulation.

The relative contribution of self-perpetuating versus hemodynamic-induced fibrosis to the progression of chronic kidney disease (CKD) after acute kidney injury (AKI) is unclear. In the present study, male Sprague-Dawley rats underwent right uninephrectomy and were instrumented with a blood pressure radiotelemeter. Two weeks later, separate groups of rats were subjected to 40 minutes renal ischemia–reperfusion or sham surgery and followed up for 4 or 16 weeks to determine the extent to which glomerulosclerosis and tubulointerstitial fibrosis as a result of the AKI–CKD transition (ie, at 4 weeks post AKI) change over time during the progression of CKD (ie, at 16 weeks post AKI). On average, tubulointerstitial fibrosis was ≈3-fold lower ( P P

Altered renal vascular responses to a high salt diet have been proposed to contribute to salt-sensitive (SS) hypertension. The goals of this study were to assess: 1) SS hypertension and renal injury in Dahl SS vs. Brown-Norway (BN) and consomic SS.BN1 rats and 2) renal hemodynamics in conscious SS vs. SS.BN1 rats during consumption of a low and high salt diet. Systolic BP (24 hrs/day via telemetry), proteinuria and renal injury were assessed in 10 week old SS (n=9), BN (n=8) and SS.BN1 (n=8) rats during a 0.4% NaCl diet and for 3 weeks during a 4.0% NaCl diet. On a 0.4% NaCl diet, BP was different (P

The susceptibility to hypertension-induced renal damage (HIRD) varies greatly within the hypertensive population. With the exception of certain genetically susceptible groups such as African-Americans, the vast majority of individuals with essential hypertension (HTN) are quite resistant to the development of significant HIRD. This is because the normal preglomerular renal autoregulatory mechanisms prevent the transmission of systemic HTN to glomerular capillaries protecting them from barotrauma. Significant HIRD only develops when HTN becomes very severe and exceeds this protective capacity. By contrast, these protective autoregulatory mechanisms are impaired in individuals with significant CKD including those with diabetic nephropathy. This impairment allows an enhanced glomerular transmission of even moderate HTN and results in progressive glomerular barotrauma and glomerulosclerosis (GS). Studies in renal mass reduction (RMR) models of CKD have confirmed the quantitative importance of autoregulatory impairment and physical glomerular BP transmission as the predominant determinant of HIRD severity and its rate of progression. Recent data indicate that the loss of NO-mediated efferent vasodilation may independently contribute to the pathogenesis of HIRD in states of endothelial dysfunction. The CKD-associated glomerular capillary hypertrophy may additionally increase HIRD susceptibility due to a reduction in podocyte density and the ability to withstand mechanical stress. Although the renin-angiotensin system (RAS) is widely believed to promote HIRD through BP-independent fibrogenic pathways, most of the benefits of RAS blockade are attributable to BP lowering when BP has been measured adequately. Nevertheless, RAS blockade combined with adequate diuretic therapy offers the most effective strategy to control BP and thereby CKD progression.