Glucagon revisited: Coordinated actions on the liver and kidney.

Glucagon secretion is stimulated by a low plasma glucose concentration. By activating glycogenolysis and gluconeogenesis in the liver, glucagon contributes to maintain a normal glycemia. Glucagon secretion is also stimulated by the intake of proteins, and glucagon contributes to amino acid metabolism and nitrogen excretion. Amino acids are used for gluconeogenesis and ureagenesis, two metabolic pathways that are closely associated. Intriguingly, cyclic AMP, the second messenger of glucagon action in the liver, is released into the bloodstream becoming an extracellular messenger. These effects depend not only on glucagon itself but on the actual glucagon/insulin ratio because insulin counteracts glucagon action on the liver. This review revisits the role of glucagon in nitrogen metabolism and in disposal of nitrogen wastes. This role involves coordinated actions of glucagon on the liver and kidney. Glucagon influences the transport of fluid and solutes in the distal tubule and collecting duct, and extracellular cAMP influences proximal tubule reabsorption. These combined effects increase the fractional excretion of urea, sodium, potassium and phosphates. Moreover, the simultaneous actions of glucagon and extracellular cAMP are responsible, at least in part, for the protein-induced rise in glomerular filtration rate that contributes to a more efficient excretion of protein-derived end products.

Improved protocols for the study of urinary electrolyte excretion and blood pressure in rodents: use of gel food and stepwise changes in diet composition.

Many experimental protocols in rodents require the comparison of groups that are fed different diets. Changes in dietary electrolyte and/or fat content can influence food intake, which can potentially introduce bias or confound the results. Unpalatable diets slow growth or cause weight loss, which is exacerbated by housing the animals in individual metabolic cages or by surgery. For balance studies in mice, small changes in body weight and food intake and low urinary flow can amplify these challenges. Powder food can be administered as gel with the addition of a desired amount of water, electrolytes, drugs (if any), and a small amount of agar. We describe here how the use of gel food to vary water, Na, K, and fat content can reduce weight loss and improve reproducibility of intake, urinary excretion, and blood pressure in rodents. In addition, mild food restriction reduces the interindividual variability and intergroup differences in food intake and associated variables, thus improving the statistical power of an experiment. Finally, we also demonstrate the advantages of using gel food for weight-based drug dosing. These protocols can improve the accuracy and reproducibility of experimental data where dietary manipulations are needed and are especially advisable in rodent studies related to water balance, obesity, and blood pressure.

Hydration and Chronic Kidney Disease Progression: A Critical Review of the Evidence.

We performed a comprehensive literature review to examine evidence on the effects of hydration on the kidney. By reducing vasopressin secretion, increasing water intake may have a beneficial effect on renal function in patients with all forms of chronic kidney disease (CKD) and in those at risk of CKD. This potential benefit may be greater when the kidney is still able to concentrate urine (high fluid intake is contraindicated in dialysis-dependent patients). Increasing water intake is a well-accepted method for preventing renal calculi, and current evidence suggests that recurrent dehydration and heat stress from extreme occupational conditions is the most probable cause of an ongoing CKD epidemic in Mesoamerica. In polycystic kidney disease (PKD), increased water intake has been shown to slow renal cyst growth in animals via direct vasopressin suppression, and pharmacologic blockade of renal vasopressin-V2 receptors has been shown to slow cyst growth in patients. However, larger clinical trials are needed to determine if supplemental water can safely slow the loss of kidney function in PKD patients.

Improvement of skin wound healing in diabetic mice by kinin B2 receptor blockade.

Impaired skin wound healing is a major medical problem in diabetic subjects. Kinins exert a number of vascular and other actions limiting organ damage in ischaemia or diabetes, but their role in skin injury is unknown. We investigated, through pharmacological manipulation of bradykinin B1 and B2 receptors (B1R and B2R respectively), the role of kinins in wound healing in non-diabetic and diabetic mice. Using two mouse models of diabetes (streptozotocin-induced and db/db mice) and non-diabetic mice, we assessed the effect of kinin receptor activation or inhibition by subtype-selective pharmacological agonists (B1R and B2R) and antagonist (B2R) on healing of experimental skin wounds. We also studied effects of agonists and antagonist on keratinocytes and fibroblasts in vitro. Levels of Bdkrb1 (encoding B1R) and Bdkrb2 (encoding B2R) mRNAs increased 1-2-fold in healthy and wounded diabetic skin compared with in non-diabetic skin. Diabetes delayed wound healing. The B1R agonist had no effect on wound healing. In contrast, the B2R agonist impaired wound repair in both non-diabetic and diabetic mice, inducing skin disorganization and epidermis thickening. In vitro, B2R activation unbalanced fibroblast/keratinocyte proliferation and increased keratinocyte migration. These effects were abolished by co-administration of B2R antagonist. Interestingly, in the two mouse models of diabetes, the B2R antagonist administered alone normalized wound healing. This effect was associated with the induction of Ccl2 (encoding monocyte chemoattractant protein 1)/Tnf (encoding tumour necrosis factor α) mRNAs. Thus stimulation of kinin B2 receptor impairs skin wound healing in mice. B2R activation occurs in the diabetic skin and delays wound healing. B2R blockade improves skin wound healing in diabetic mice and is a potential therapeutic approach to diabetic ulcers.

Protein- and diabetes-induced glomerular hyperfiltration: role of glucagon, vasopressin, and urea.

A single protein-rich meal (or an infusion of amino acids) is known to increase the glomerular filtration rate (GFR) for a few hours, a phenomenon known as "hyperfiltration." It is important to understand the factors that initiate this upregulation because it becomes maladaptive in the long term. Several mediators and paracrine factors have been shown to participate in this upregulation, but they are not directly triggered by protein intake. Here, we explain how a rise in glucagon and in vasopressin secretion, directly induced by protein ingestion, might be the initial factors triggering the hepatic and renal events leading to an increase in the GFR. Their effects include metabolic actions in the liver and stimulation of sodium chloride reabsorption in the thick ascending limb. Glucagon is not only a glucoregulatory hormone. It is also important for the excretion of nitrogen end products by stimulating both urea synthesis in the liver (along with gluconeogenesis from amino acids) and urea excretion by the kidney. Vasopressin allows the concentration of nitrogenous end products (urea, ammonia, etc.) and other protein-associated wastes in a hyperosmotic urine, thus allowing a very significant water economy characteristic of all terrestrial mammals. No hyperfiltration occurs in the absence of one or the other hormone. Experimental results suggest that the combined actions of these two hormones, along with the complex intrarenal handling of urea, lead to alter the composition of the tubular fluid at the macula densa and to reduce the intensity of the signal activating the tubuloglomerular feedback control of GFR, thus allowing GFR to raise. Altogether, glucagon, vasopressin, and urea contribute to set up the best compromise between efficient urea excretion and water economy.

Cardioprotective effect of VEGF and venom VEGF-like protein in acute myocardial ischemia in mice: effect on mitochondrial function.

Coronary endothelial dysfunction is involved in cardiac ischemia-reperfusion (IR) injury. Vascular endothelial growth factor (VEGF) activates endothelial cells and exerts cardioprotective effects in isolated hearts. The recently discovered viper venom protein called increasing capillary permeability protein (ICPP) exerts VEGF-like effects in endothelial cells. We examined whether VEGF or ICPP can influence IR outcome in vivo in mice. Dosages of VEGF and ICPP were determined by preliminary blood pressure study. In IR, both the proteins administered intravenously at reperfusion reduced infarct size (IS) by 57% for VEGF and 52% for ICPP (P < 0.01). Pretreatment with a selective VEGFR2 receptor antagonist abolished the reduction in IS. VEGF and ICPP induced ERK phosphorylation in the myocardium. IR triggered mitochondrial pore opening and impaired mitochondrial respiratory function. These effects of IR were prevented by VEGF or ICPP, which increased mitochondrial calcium retention capacity by 37% compared with saline (P < 0.05) and improved mitochondrial respiratory function (by 71% and 65%, respectively for state 3, and 51% and 38% for state 4, P < 0.01 for VEGF). Thus, intravenous administration of VEGF or ICPP at reperfusion largely reduces IS in IR, through stimulation of VEGFR2 receptors. This effect is mediated, at least in part, by improvement of IR-induced mitochondrial dysfunction.

Synthesis and fragmentation of hyaluronan in renal ischaemia.

BACKGROUND: The turnover of hyaluronan (HA), especially the production of low-molecular-weight fragments of HA, was examined in a model of unilateral renal ischaemia-reperfusion (IR) in rats. METHODS: HA was extracted from the outer and inner stripe of the outer medulla (OSOM and ISOM) at different times following IR. Its fragmentation was measured using membrane filtration and size-exclusion chromatography. Quantitative reverse transcription-polymerase chain reaction, zymography and immunohistochemistry were used to assess the expression and localization of various forms of HA synthase (HAS) and hyaluronidase (HYAL). Macrophage infiltration was evaluated using immunohistochemistry. RESULTS: HA accumulated at Day 1 mostly as high-molecular-weight (HMW) species with an elution profile similar to a reference 2500 kDa HA and at Day 14 mostly as medium- to low-size fragments. Within 1 day, HAS1 messenger RNA was up-regulated > 50- and 35-fold in OSOM and ISOM, respectively. Thereafter, HAS1 tended to normalize, while HAS2 increased steadily. Both synthetic enzymes were localized around tubules and in the interstitium. Conversely, HYAL1, HYAL2 and global hyaluronidase activity were repressed during the first 24 h. The patterns were identical in the OSOM and ISOM despite markedly different amounts of HA at baseline. There was no obvious correlation between HA deposits and macrophage infiltration. CONCLUSIONS: In the post-ischaemic kidney, HA starts to accumulate at Day 1 mostly as HMW species. Later on, a large proportion becomes degraded into smaller fragments. This pattern is explained by coordinated changes in the expression of HA synthases and hyaluronidases, especially an early induction of HAS1. The current data open the door to timed pharmacological interventions blocking the production of HA fragments.

Differential regulation of angiotensin II receptors during renal injury and compensatory hypertrophy in the rat.

1. The renin-angiotensin system may be involved in the compensatory adaptations occurring after the reduction of renal mass and during the consecutive changes leading to chronic renal failure. We therefore investigated the regulation of angiotensin II receptors in two models of renal hypertrophy in the rat: hypertrophy following uninephrectomy (UNx) or subtotal nephrectomy (STNx). The level of angiotensin type 1 (AT1A-R and AT1B-R) and type 2 (AT2-R) receptor mRNA was quantified by competitive reverse transcription-polymerase chain reaction (RT-PCR) in specific renal zones and the intrarenal distribution of angiotensin II receptors was analysed by immunohistochemistry. 2. In the UNx rats, AT1-R mRNA expression was not modified in the cortex or in the inner stripe of the outer medulla of the residual kidney at any time after the surgery (1, 4 and 12 weeks). In contrast, AT1-R mRNA expression was significantly reduced in these zones in STNx rats (-33% and -40%, respectively). This downregulation was organ-specific, as AT1-R mRNA levels were not modified in the liver. The proportions of AT1-R subtype (AT1A and AT1B) mRNA were unchanged by UNx or STNx. Very low levels of AT2-R mRNA were found in the cortex of all groups. Immunostaining revealed a similar localization of AT1-R in mesangial cells, proximal tubule, basolateral membrane of thick ascending limb, in both models of hypertrophy. AT1-R labelling was also detected in the apical membrane of intercalated cells of cortical collecting ducts. 3. This differential mRNA expression of angiotensin II receptors during compensatory hypertrophy and renal injury suggests that the development of renal hypertrophy is independent of AT1-R and AT2-R gene expression levels.

Effect of salt and water intake on epithelial sodium channel mRNA abundance in the kidney of salt-sensitive Sabra rats.

1. The level of mRNA expression of epithelial sodium channel (ENaC) subunits was studied in a salt-dependent hypertensive rat strain (Sabra). These rats exhibit high vasopressin levels compared with their normotensive counterparts. We also investigated whether this expression is influenced by changes in the sodium intake/aldosterone axis or in the fluid intake/vasopressin axis. 2. A higher expression of beta- and gamma-subunit mRNA was found in salt-sensitive compared with salt-resistant rats on a normal salt diet. A high-sodium diet did not alter mRNA abundance in either substrain. In contrast, water supplementation in salt-sensitive rats fed the high-sodium diet induced a marked reduction in mRNA abundance of beta- and gamma-subunits. 3. The present study provides evidence that beta- and gamma-subunits of ENaC are differently expressed in the kidney of salt-sensitive and salt-resistant Sabra rats and that their abundance is regulated by vasopressin, not by sodium intake. These results are consistent with the hypothesis that increased vasopressin-dependent ENaC expression and activity may contribute to the pathogenesis of hypertension in salt-sensitive Sabra rats.