A method for accurate measurement of GFR in conscious, spontaneously voiding rats

A method for accurate measurement of GFR in conscious, spontaneously voiding rats. Renal function measurement by clearance methods relies on accurately timed urine collection. In small experimental animals, renal function measurement is usually performed under anesthesia and/or with the application of bladder catheters to ensure accurate urine collection. To avoid both anesthesia and the need for bladder catheters we developed a method to measure glomerular filtration rate (GFR) in spontaneously voiding conscious rats. GFR was measured as the urinary clearance of constantly infused 125I-iothalamate. To correct for incomplete bladder emptying, urinary clearance of 125I-iothalamate was multiplied by the ratio of plasma and urinary clearance of simultaneously infused 131I-hippuran, a correction method that has been previously validated in humans. Reproducibility of the technique was evaluated by analysis of the results of four consecutive clearance periods during the day (intra-assay variation) in a group of 17 rats and of two consecutive clearance periods on two or three separate days in a group of 20 rats (inter-assay variation), all with normal renal function. Application of the correction method reduced the intra-assay coefficient of variation (mean ± SD) from 37.4 ± 14.3 to 5.4 ± 2.3% (P < 0.05). The mean inter-assay coefficient of variation fell slightly from 23.4 ± 10.3 to 11.0 ± 7.2% (P < 0.10). In rats with moderately impaired renal function (N = 8) the intra-assay variation fell from 27.9 ± 20.7 to 2.7 ± 1.6% (P < 0.05). Our data show that this correction method is a useful technique to assess renal function in conscious, spontaneously voiding rats

Author index for volume 286

Susceptibility to renal injury varies among individuals. Previously, we found that individual endothelial function of healthy renal arteries in vitro predicted severity of renal damage after 5/6 nephrectomy. Here we hypothesized that individual differences in endothelial function in vitro and renal perfusion in vivo predict the severity of renal damage in a model of adriamycin-induced nephropathy. In three separate studies, the following baseline parameters were measured in healthy male Wistar rats: (1) acetylcholine (ACh)-induced relaxation in small renal arteries in vitro (n = 16) and the contribution of prostaglandins, nitric oxide (NO) and endothelium-dependent hyperpolarizing factor (EDHF) to the relaxation; (2) glomerular filtration rate (GFR) and effective renal plasma flow (ERPF) in spontaneously voiding rats in vivo (n = 16) and (3) the acute effect of the NO-synthase inhibitor N(G)-nitro-l-arginine methyl ester (L-NAME, n = 12) on renal blood flow (RBF) as compared to vehicle infusion (n = 9). Following these measurements, adriamycin (1.75 mg/kg i.v.) was injected and subsequent renal damage after 6 weeks was related to the baseline parameters. Total ACh-induced (r = 0.51, P <0.05) and EDHF-mediated relaxation (r = 0.68, P <0.05), as well as ERPF (r = 0.66, P <0.01), positively correlated with the severity of proteinuria 6 weeks after injection. In contrast, pronounced baseline NO-mediated dilation was associated with lower proteinuria (r = 0.71, P <0.01). Nevertheless, an acute L-NAME infusion, strongly reducing RBF by 22 +/- 8%, during adriamycin administration provided protection against the development of proteinuria. Individual animals with pronounced baseline endothelial dilatory ability measured in vitro and high ERPF in vivo are vulnerable to renal damage after the adriamycin injection. Acute inhibition of NO during adriamycin administration, resulting in a decrease of RBF, protects against renal injury, probably by limiting the delivery of the drug to the kidney. Therefore, interindividual variability in renal haemodynamics might be crucially involved in susceptibility to nephrotoxic renal damag

Early, but not late therapy with a vasopressin V1a-antagonist ameliorates the development of renal damage after 5/6 nephrectomy

INTRODUCTION: Vasopressin, mainly through the V1a-receptor, is thought to be a major player in the maintenance of hyperfiltration. Its inhibition could therefore lead to a decrease in progression of chronic renal failure. To this end, the effect of the vasopressin V1a-receptor-selective antagonist, YM218, was studied on proteinuria and focal glomerulosclerosis in early and late intervention after 5/6 nephrectomy in rats, and compared with an angiotensin-converting enzyme inhibitor (ACE-I). MATERIALS AND METHODS: After 5/6 nephrectomy, early intervention was performed between week 2 and 10 thereafter with the V1a-receptor-selective antagonist (VRA, 10 mg/kg/day, n=10), enalapril (ACE-I, 10 mg/kg/day, n=9), or vehicle (n=8). Late intervention was performed in another group between week 6 and 12 with VRA (10 mg/kg/day, n=7), lisinopril (ACE-I, 5 mg/kg/day, n=7), or vehicle (n=7). RESULTS: In early intervention, proteinuria and focal glomerulosclerosis were significantly decreased by VRA compared to vehicle (44+7% and 59+8% respectively). ACE-I significantly decreased proteinuria (67+7%) and a trend towards a decrease in focal glomerulosclerosis was observed (30+18%). In late intervention, VRA did not decrease proteinuria and focal glomerulosclerosis compared to vehicle (21+20% and 0%, respectively), ACE-I significantly lowered proteinuria (92+2%) and a focal glomerulosclerosis (69+1%) lowering trend was observed. CONCLUSION: These results indicate that VRA may protect against early progression of renal injury after 5/6 nephrectomy, whereas its effectiveness seems limited in established renal damage

Myocardial infarction does not further impair renal damage in 5/6 nephrectomized rats

Background. Recent observational studies show that reduced renal function is an independent risk factor for the development of cardiovascular disease. Previously, we reported that myocardial infarction (MI) indeed enhanced mild renal function decline in rats after unilateral nephrectomy (NX) and that RAAS intervention inhibited this decline. The effects of an MI on pre-existing sever renal function loss and the effects of RAAS intervention interrupting this hypothesized cardiorenal interaction are however unknown and clinically even more relevant. Methods. Male Wistar rats underwent MI, sham MI, 5/6NX, or 5/6NX and MI. Six weeks later, the NX rats were treated with an angiotensin-converting enzyme inhibitor (ACEi) or vehicle for 6 weeks. Results. An MI did not significantly induce more proteinuria (303 +/- 46 versus 265 +/- 24 mg/24 h) and glomerulosclerosis (40 +/- 11 versus 28 +/- 4 arbitrary units) in 5/6NX+MI compared to 5/6NX, and ACEi therapy was equally effective in reducing renal damage in these groups. In the 5/6NX+MI group, decreased renal blood flow and creatinine clearance were observed compared to 5/6NX (2.2 +/- 0.6 versus 3.6 +/- 0.4 ml/min/kg and 2.1 +/- 0.3 versus 2.9 +/- 0.3 ml/min/kg), which both increased after ACEi to levels comparable found in the group that underwent 5/6NX alone. Conclusions. MI does not further deteriorate structural renal damage induced by 5/6NX compared with 5/6NX alone. Furthermore, renal haemodynamic impairment occurs after MI, which can be improved applying ACEi therapy. Therefore, we conclude that treatment with ACEi should be optimized in patients with chronic kidney disease after MI to improve renal function

Therapeutic resistance to angiotensin converting enzyme (ACE) inhibition is related to pharmacodynamic and -kinetic factors in 5/6 nephrectomized rats

Proteinuria plays a pathogenic role in the development of end stage renal disease. Angiotensin converting enzyme (ACE) inhibitors lower proteinuria and are renoprotective. However, large inter-individual variation in antiproteinuric response to ACE inhibitors exists. In this study, we explored the mechanism of therapeutic resistance to an ACE inhibitor in the rat 5/6 nephrectomy model. At week 6 after 5/6 nephrectomy, treatment with lisinopril was initiated for 6 weeks. Proteinuria and blood pressure were evaluated weekly. At the end of the experiment, rats were divided into tertiles according to their antiproteinuric response: (1) responders (n=9), (2) intermediate responders (n=8) and (3) non-responders to ACE inhibitor therapy (n=9). At the start of treatment, proteinuria had progressively increased to 154 (95% confidence interval [Cl]: 123-185) mg/24 It in the entire cohort, with comparable proteinuria and blood pressure in all groups. Following treatment with ACE inhibitor, proteinuria was significantly lower in the responders (68, Cl: 46-89 mg/24 h) compared to the non-responders (25 1, CI: 83-420) mg/24 h). Similarly, blood pressure was reduced in the responders, but unaffected in the non-responders. At autopsy, renal ACE activity and renal ACE expression were significantly lower in the responders compared to the non-responders. Although lisinopril intake was comparable in all animals, urinary drug excretion was increased in the non-responders, demonstrating increased drug clearance. Average urinary lisinopril excretion was correlated with antiproteinuric response (R-2=0.32, P=0.003). In conclusion, both pharmacodynamic and -kinetic factors account for the non-response to lisinopril. Whether these can be overcome simply by increasing drug dosage in non-responders should be investigated. (c) 2007 Elsevier B.V All rights reserved

Renal damage after myocardial infarction is prevented by renin-angiotensin-aldosterone-system intervention

Recently, it was shown that myocardial infarction aggravates preexistent mild renal damage that is elicited by unilateral nephrectomy in rats. The mechanism behind this cardiorenal interaction likely involves the renin-angiotensin-aldosterone-system and/or vasoactive peptides that are metabolized by neutral endopeptidase (NEP). The renoprotective effect of angiotensin-converting enzyme inhibition (ACEi) as well as combined ACE/NEP inhibition with a vasopeptidase inhibitor (VPI) was investigated in the same model to clarify the underlying mechanism. At week 17 after sequential induction of unilateral nephrectomy and myocardial infarction, treatment with lisinopril (ACEi), AVE7688 (VPI), or vehicle was initiated for 6 wk. Proteinuria and systolic BP (SBP) were evaluated weekly. Renal damage was assessed primarily by proteinuria, interstitial a-smooth muscle actin (alpha-SMA) staining, and the incidence of focal glomerulosclerosis (FGS). At start of treatment, proteinuria had increased progressively to 167 +/- 20 mg/d in the entire cohort (n = 42). Both ACEi and VPI provided a similar reduction in proteinuria, a-SMA, and FGS compared with vehicle at week 23 (proteinuria 76 +/- 6 versus 77 +/- 4%; a-SMA 60 6 versus 77 +/- 3%; FGS 52 +/- 14 versus 61 +/- 10%). Similar reductions in systolic BP were observed in both ACEi- and VPI-treated groups (33 +/- 3 and 37 +/- 2%, respectively). Compared with ACEi, VPI-treated rats displayed a significantly larger reduction of plasma (41 +/- 5 versus 61 +/- 4%) and renal (53 +/- 6 versus 74 4%) ACE activity. It is concluded that both ACEi and VPI intervention prevent renal damage in a rat model of cardiorenal interaction. VPI treatment seemed to provide no additional renoprotection compared with sole ACEi after 6 wk of treatment in this model, despite a more pronounced ACE-inhibiting effect of VPI