Differential roles of VPS and RAAS in water homeostasis and a risk for kidney dysfunction in rats undergoing rapid fasting/dehydration with regular exercise

Figures

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  Figure 1. Effects of the rapid restriction of food and water intake on body weight, kidney weight, urine volume, urine osmolality, and blood biomarkers of rats with and without regular exercise. (a) Body weight of rats at the end of the study period. (b) Weight of kidney obtained from rats at the end of the study period. (c) The ratio of kidney weight to body weight. (d) Volume of urine samples collected during the last 12 hr before sacrifice. (e) Osmolality of urine samples obtained in (d). The osmolality was calculated from the freezing point depression. (f) Plasma Na+ concentration. (g, h) Concentrations of blood urea nitrogen (g) and plasma creatinine (h). Data are expressed as mean ± SEM (n = 5/group). *p < .05; **p < .01; ***p < .001; ****p < .0001. NS, not significant; Ex, exercise; RR, rapid restriction

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  Figure 2. Effects of the rapid restriction of food and water intake on vasopressin levels in rats with and without regular exercise. (a) Expression levels of AVP heteronuclear RNA in the hypothalamus. RNA expression levels were analyzed by quantitative RT‐PCR and normalized to those of GAPDH. Fold changes are displayed relative to the control group receiving neither regular exercise nor rapid restriction (1.0). (b) Expression levels of AVP mature mRNA in the hypothalamus. Data are normalized and displayed as described in (a). (c) Circulating levels of vasopressin. The amounts of vasopressin in the plasma were quantified by ELISA. Data are expressed as mean ± SEM (n = 5/group). *p < .05; **p < .01. Ex, exercise; RR, rapid restriction

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  Figure 3. Effects of the rapid restriction of food and water intake on the renal expression levels of water channels in rats with and without regular exercise. (a) Gene expression levels of AQP2 in the kidney medulla. The mRNA levels of AQP2 were obtained by quantitative RT‐PCR and normalized to those of GAPDH. Expression levels are displayed relative to those of the control group receiving neither regular exercise nor rapid restriction (1.0). (b) Protein levels of AQP2 in the kidney medulla. The amounts of the nonglycosylated (29 kDa) and glycosylated form (35 kDa) forms of AQP2 protein were analyzed by Western blot (left panel) and densitometry, and the sum of the two values was compared between the groups (right panel). The image shows representative immunoblots. (c–f) Gene expression levels and protein levels of AQP3 (c and d, respectively) and AQP4 (e and f, respectively) in the kidney medulla. Data are obtained and displayed as described in (a and b). Data are expressed as mean ± SEM (mRNA levels, n = 5/group; protein levels, n = 4/group). *p < .05. NS, not significant; Ex, exercise; RR, rapid restriction

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  Figure 4. Effects of the rapid restriction of food and water intake on the circulating levels of aldosterone in rats with and without regular exercise. The amounts of aldosterone in the plasma were measured by ELISA. Data are expressed as mean ± SEM (n = 5/group). **p < .01. Ex, exercise; RR, rapid restriction

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  Figure 5. Effects of the rapid restriction of food and water intake on the renal expression levels of ion channels in rats with and without regular exercise. (a–d) Gene expression levels of SGK‐1 (a), GILZ (b), CHIF (c), and ENaCα (d) in the kidney medulla. The mRNA levels were analyzed by quantitative RT‐PCR and normalized to those of GAPDH. Fold changes are displayed relative to the control group receiving neither regular exercise nor rapid restriction (1.0). Data are expressed as mean ± SEM (n = 5/group). *p < .05; ***p < .001. NS, not significant; Ex, exercise; RR, rapid restriction

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  Figure 6. Effects of rapid restriction of food and water intake on the levels of inflammation, activation of an apoptotic effector, and levels of tissue injury in the kidney of rats with and without regular exercise. (a–c) Gene expression levels of TNF‐α (a), IL‐1β (b), and IL‐6 (c) in the kidney medulla. The mRNA levels were obtained using quantitative RT‐PCR and normalized to those of GAPDH. Fold changes are displayed relative to the control group receiving neither regular exercise nor rapid restriction (1.0). (d) Protein levels of active caspase‐3 in the kidney medulla. The amounts of active caspase‐3 protein were analyzed via Western blot (left panel) and densitometry (right panel). The image shows representative immunoblots. (e and f) Gene expression levels of Kim‐1 (e) and Ngal (f) in the kidney medulla. Data are expressed as mean ± SEM (mRNA levels, n = 5/group; protein levels, n = 4/group). *p < .05; **p < .01; ***p < .001; ****p < .0001. NS, not significant; Ex, exercise; RR, rapid restriction

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  Figure 7. Effects of the rapid restriction of food and water intake on the pathways for water homeostasis in rats with and without regular exercise. The VPS regulates water homeostasis to the extent of the steady‐state conditions, irrespective of the presence or absence of regular exercise/rapid restriction. Conversely, the RAAS responds to rapid restriction and contributes to water homeostasis to deal with body fluid‐related acute stress. When rapid restriction is loaded in combination with regular exercise, some factors derived from the combination of these two stresses would exhibit deleterious effects on kidney function. VPS, vasopressinergic system; RAAS, renin–angiotensin–aldosterone system