Search

Acute psychosocial stress has been linked to the onset of vaso-occlusive pain crises in patients with sickle cell disease (SCD), however mechanistic insight is lacking. Studies suggest that histamine is a stress-responsive factor and promotes inflammation. In SCD, histamine levels are elevated in association with vaso-occulsive crisis, however, the impact of acute stress is unknown. We hypothesized that acute stress in a humanized SCD mouse model stimulates the histamine pathway, inflammatory mediator release, and a pressor response. Acute stress was induced using cage switch stress (CSS) in male humanized SCD (HbSS) or control (HbAA) mice (n=6-8/group) with blood pressure (BP) monitored by radiotelemetry and mice terminated at baseline (BL, no CSS) and 30 min post-CSS for plasma measurements. Plasma histamine was unchanged in HbAA mice but was elevated in HbSS in response to CSS (nM; HbAA: 90.2±21.3; 104.3±10.2; HbSS: 91.1±13.6; 174.3±19.2*, *pgenotype =0.0001, *pgenotype genotype

Vaso-occlusive crisis often occurs in sickle cell disease (SCD). Since psychosocial stress leads to impaired vascular function and inflammation, we hypothesized that acute psychosocial stress elicits an exaggerated blood pressure response and/or increased circulating inflammatory mediators in a mouse model of SCD. We utilized adult male (16-20 wks) humanized Townes sickle cell (HbSS) and control (HbAA) mice with telemetry (n=8) to monitor changes in systolic blood pressure (SBP), diastolic blood pressure (DBP), spectral analysis (low frequency/high frequency, LF/HF ratio) of blood pressure variability, heart rate (HR), and locomotor activity (LA) in response to cage switch stress (CSS). CSS-induced increases in DBP were significantly attenuated in HbSS versus HbAA (HbSS: 23.3±1.8mmHg, HbAA: 31.6±1.7mmHg, p=0.004), while the increase in SBP was similar in HbSS and HbAA. CSS increased LF/HF in both strains, although HbSS was significantly less than HbAA (CSS HbAA : 3.21±0.2; CSS HbSS : 1.68±0.2, p

Premenopausal women have a lower risk of hypertension compared to age-matched men. Recently, prominent roles have been assigned to P2Y 2 and P2Y 4 purinoceptor subtypes in promoting Na + excretion, implicating dysfunction of these receptors as potential contributors to hypertension. We recently reported that activation of P2Y 2 and P2Y 4 receptors in the renal medulla by UTP promotes Na + excretion in male rats. In intact females, UTP did not stimulate Na + excretion while ovariectomy unmasked UTP-induced natriuresis. These observations led us to hypothesize that intact females have higher basal renal medullary activity of P2Y 2 and P2Y 4 receptors in regulating Na + excretion compared to male and ovariectomized (OVX) rats. To test that, we determined (i) P2Y 2 and P2Y 4 mRNA and protein expression in the inner medulla from male, intact female and OVX Sprague Dawley rats and (ii) the effect of inhibiting medullary purinoceptors (P2 receptors) on Na + excretion in those rats. We found that P2Y 2 and P2Y 4 mRNA expression was higher in the inner medulla from females compared to males (1.00±0.09 vs. 0.70±0.05 and 1.00±0.22 vs. 0.29±0.05, respectively, P2 and P2Y 4 mRNA expression were eliminated by ovariectomy (0.60±0.06 and 0.29±0.04, respectively, p2 receptor, compared to males. In anesthetized rats, medullary P2 receptor inhibition by suramin (P2 receptor antagonist, 750 μg/kg/min) significantly attenuated Na + excretion in intact females (0.4±0.1 vs. 0.9±0.2 μmol/min, P2 ) increases the expression of P2Y 2 and P2Y 4 receptors, we subjected cultured mouse inner medullary collecting duct cells (mIMCD3) to different concentrations of E 2 (0, 10, 100 and 1000 nM). We found that E 2 dose-dependently increased the expression of P2Y 2 and P2Y 4 mRNA in mIMCD3. These data suggest that females have enhanced P2Y 2 and P2Y 4 -dependent regulation of Na + excretion in the renal medulla, compared to male and OVX rats, at least partially via an E 2 -dependent mechanism. This pathway may contribute to facilitated renal Na + handling in premenopausal females.

Circadian rhythms in physiologic functions are driven, at the molecular level, by a group of transcription factors that oscillate over a 24 hour period, collectively termed the molecular clock. Within the kidney, it has been shown that the molecular clock directly influences transcription of Na + transporters and channels, including ENaC. ENaC is regulated by endothelin-1 (ET-1), via ET B receptor activation, in response to high salt intake. Thus, we hypothesized that increases in dietary sodium regulate the renal molecular clock (which in turn would facilitate Na+ homeostasis) through an ET B dependent mechanism. To address this question, we examined the effect of high salt (HS) intake on renal clock gene ( Bmal1, Cry1, Per1, Per2 ) expression. Control and ET B receptor deficient (ET B def) rats (a model of elevated renal ENaC) were placed on either HS or normal salt (NS) for two weeks and euthanized every 4 hours beginning at Zeitgeber Time 0 (Lights on). In the inner medulla, HS causes a phase delay in Bmal1 (Fig 1A) expression in control but not ET B def rats (Fig 1B). In addition, HS suppressed the expression of Cry1 , and Per2 during the respective acrophase in both control and ET B def rats (Fig 1C-1F) with no significant effect on Per1 . In contrast, no significant difference in the expression of Bmal1, Cry1, Per2, or Per1 (Fig 1I-1P) was found in response to HS in the renal cortex of either control or ET B def. These data indicate that HS feeding desynchronizes the molecular clock within the kidney and provides evidence that peripheral clocks are regulated in a cell type specific manner, even within the same organ.

Our lab has recently shown that ETB deficient (ETB def) rats have a time of day dependent impairment in their ability to excrete a Na+ load. These observations suggest an interaction between renal ETB receptors and circadian mechanisms that regulate renal tubular Na+ transport and excretion. Given that knockout of the circadian clock gene Bmal1 reduces blood pressure in mice, we hypothesized that a high salt intake impairs the clock mechanism in the renal inner medulla in an ETB dependent manner. Transgenic control (Tg con) or ETB def rats were fed normal (NS, 0.8% NaCl) or high (HS, 4% NaCl) salt for two weeks. In one group, rats were euthanized every 4 hours beginning at zeitgeber time 0 (lights on) for tissue collection (and subsequent assessment of circadian clock genes), while in a second group of rats urine was collected in 12-hour intervals (active vs. inactive). Consistent with our hypothesis, we observed that HS abolished the normal oscillation in Bmal1 expression in the renal inner medulla of Tg con rats, and effect not observed in ETB def rats. Interestingly, renal production of ET-1, was significantly higher during the active period vs. inactive period in both NS (3.6±1.1 vs. 0.8±0.2 pg/12hr respectively) and HS (9.2±4.1 vs. 1.6±0.3 pg/12hr respectively) fed Tg con rats. There was no time-of-day-dependent difference in ET-1 excretion in ETB def rats on NS (6.6±2.2 vs. 4.6±1.7 pg/12hr respectively), although this pattern was restored in ETB def rats fed HS (2.2±1.0 vs. 9.2±2.5 pg/12hr inactive vs. active). Taken together, these data indicate that an increase in renal ET-1/ETB activation in response to HS modulates inner medullary clock gene expression to promote renal Na+ excretion.