Glomerular filtration rate in dogs and cats

R Heiene, Oslo, Norway and HP Lefebvre, Toulouse, France

Introduction

Glomerular filtration rate (GFR) is considered the single most useful and sensitive test of renal function.¹ Any decrease in GFR generally means that kidney disease is occurring or progressing. Assessment of GFR is therefore pivotal for evaluating severity and progression of renal diseases, especially chronic kidney diseases (CKD). In small animal medicine, the staging system proposed by the International Renal Interest Society (IRIS) is currently based on the concentration of creatinine in blood, but the IRIS board considers that, in the future, GFR measurement could become the major criterion for staging, as it is in humans.² Several methods for measuring GFR have now been validated in dogs and cats, including plasma clearance techniques using non-radioactive markers such as creatinine and iohexol, that are easier to use than urinary clearance methods. These plasma clearance methods make determination of GFR practical in both clinical and research settings.³ Here we review some recent developments and current challenges regarding plasma clearance methods to assess GFR.

Blood sampling procedure and strategy

Plasma clearance methods require repeated blood samples over a period of several hours following intravenous administration of a suitable marker. Clearance (Cl) is then calculated using the formula Cl = GFR = dose/AUC, where AUC is the area under the plasma disappearance curve of the marker.

Fig 1a: Example of a plasma disappearance curve, where the elimination is monoexponential after an initial "distribution phase" of the marker

The greater the number of blood samples and the longer the period of collection, the more accurate will be the estimation of the AUC, and thus of the GFR. Sampling should continue ideally until all marker is excreted, but testing is easier if the number of samples and the sampling period are limited. Much research has focussed on determining the optimal number of samples and sampling times to provide suitable limited-sample strategies.

Repeated collection of 5-mL blood samples in cats or miniature dogs may be difficult and lead to excessive blood loss. The total volume of blood collected can be reduced if the assay requires very small volumes of plasma or serum. For example, 0.2 mL of blood is sufficient for blood creatinine assays using an enzymatic method. Paediatric devices based on a capillary system have been proposed as alternatives to vacuum tubes for sampling for these tests.⁴ Potential advantages include absence of vein collapse, limited blood withdrawal and improved safety.

The timing of blood sampling should be aimed to minimize the proportion of the AUC that is extrapolated.¹ For this, the last sampling is especially critical, because it determines the percentage of extrapolated area, which is calculated from the slope of the elimination phase and the last measured concentration. A slight error in calculating the slope may also induce an error in calculating clearance. Ideally, the extrapolated area should be < 20% of the total AUC. The lower the renal function, the slower the elimination of the marker.

Fig1b: Example of how plasma disappearance curves have large or small areas under the curve (AUC) if the marker is slowly (red) or rapidly (green) excreted - reflecting low or high renal function.

Unfortunately, in clinical practice the GFR of the patient is unknown and the optimum final blood sampling time is therefore uncertain. For most markers this is a problem only if the GFR is very low, in which case blood creatinine is increased. Late sampling can then be planned for.

Creatinine has 2–3 times longer elimination half life than most other markers due to a larger volume of distribution. Blood sampling at 1 and 10 h post-administration provides a relatively accurate estimation of the AUC, with a maximum error of –14% compared to the AUC determined from an 11-point kinetic profile. However, sampling at inappropriate times can lead to errors exceeding 400%.⁶ It may be an option to keep the dog in the clinic overnight for a 24 hour sample, if the creatinine in the samples can be analyzed directly and it is observed that blood creatinine remains high during the day of sampling.

Correction formulae derived from those used in human medicine (e.g. Brøchner-Mortensen formula) have been proposed and applied to dogs and cats to estimate the actual GFR value from the clearance determined with a limited sampling strategy.⁵⁻¹⁰

Repeated GFR measurements

A potential problem with repeated GFR measurements in patients is within-individual and between-day variability (i.e. the reproducibility of measurement). If a dog is tested on two separate occasions, what minimum difference between the two GFR findings is needed to exceed physiological and analytical variability, and therefore be interpretable as clinically relevant? Studies using healthy or diseased animals have provided between-day coefficients of variation generally <20% for most markers; 11–14 considered to be acceptable.

Repeated clearance measurements in a given patient should be performed using the same technique to avoid misinterpretation due to methodological differences. Differences of 10–20% are often found in clearance of various markers when performed in the same animals and at the same laboratory.⁸,¹⁵⁻¹⁸

  • Among dietary factors which can affect renal function, protein intake is the most important, as a protein rich meal may increase GFR. Hydration status is also crucial because dehydration - even if subclinical - will reduce GFR. Thus, animals should be well hydrated but fasted overnight before the test.

  • In growing Beagle puppies, the mean GFR decreased ~40% (from 4.1 to 2.5 mL/min/kg) between ages of 9 and 27 weeks.¹⁹ GFR was 87% higher in 2-month-old Beagle puppies than in 6–9 year-old Beagle dogs.²⁰ These results indicate that a progressive decline in GFR is normal in the growing pup and should not be interpreted as development of CKD. In kittens, the GFR is higher at ages 9 to 19 weeks than in younger or older individuals.²¹

    In adult dogs, aging seemed to have a limited effect, or an effect obscured by other influences, in longitudinal²² and cross-sectional⁵ studies. For cats, plasma clearance of creatinine, but not of iohexol, was ~25% lower at ages 9–12 years than in those 7–12 months old.¹⁴ Another feline study found no correlation between age and plasma clearance of iohexol or creatinine.⁹ Thus, any decline in renal function with aging seems limited in healthy dogs and cats. However, the potential clinical relevance of GFR decline in aged animals needs further study.

  • GFR is lower in large dogs than in small dogs. In healthy adult dogs, negative correlations have been shown between body weight and GFR, assessed by plasma clearance of creatinine (n = 113)²² and of iohexol (n = 118).⁵

    Also in cats a negative linear relationship was evident between body weight and GFR assessed by the plasma clearance of iohexol with creatinine, although the R² value was low (<20%).⁹ In domestic shorthair cats, body weight appears to have a slight effect on blood creatinine concentration (mean ± SD values 142 ± 36 and 161 ± 22 µmol/L for body weights <4 kg and >5 kg, respectively).²³

    The effect of size may initially seem surprising, but it is not unexpected, as a number of species have been shown to have kidney function more closely related to metabolic needs than to body weight.²⁴

  • Breed effects on blood creatinine concentrations were recently reported in dogs²³ and cats.²⁴ But this does not mean that a breed-dependency of GFR necessarily exists, as endogenous production of creatinine in muscle also affects blood creatinine concentrations. A good example of this is the Greyhound breed, where the higher blood creatinine concentrations found have been shown to be due to higher production of creatinine rather than to lower GFR.²⁵,²⁶

    However, breed differences might still influence GFR. In a recent study,²⁷ GFR was lower in German Shepherds than in English Pointers and English Setters (2.5 ± 0.7, vs. 3.5 ± 0.6 and 3.4 ± 0.8 mL/min/kg, respectively). Age was similar in the three breeds but German Shepherds were heavier; thus differences in body weight, a confounding factor in this study, could have contributed to the observed differences in GFR.

Indexation of glomerular filtration rate

Clearance is measured as mL/min. Standardization (indexation; scaling) of GFR values to the body weight allows comparison of the clearance value to reference ranges for healthy animals. GFR in dogs and cats is traditionally expressed in mL/min/kg. A current challenge is to define the most appropriate way to standardize GFR. Body surface area (BSA) has been proposed as the reference for indexing physiological variables and it is routinely used in human medicine. However, it has been recommended that such an indexation for GFR should be abandonned²⁸ and the formulae used to estimate BSA in dogs are probably inaccurate.²⁹ Indexation to extracellular fluid volume (ECFV) has been proposed as an alternative because one of the major roles of the kidney is to regulate body fluid composition.³⁰ Nevertheless, standardization to ECFV did not produce substantial changes in the relationships between GFR estimates and body weight in adult dogs.⁵ Moreover, with indexation to ECFV, differences between puppies and adult dogs were still observed, but were inversed.²⁰ In an earlier canine study, standardization to body weight, BSA or ECFV was shown to produce quite different results and for some dogs altered the clinical interpretation of the GFR value obtained.¹⁰ A similar problem exists in the cat: standardization to BSA resulted in larger between-individual coefficient of variation (36%) than did standardization to body weight (27%) or ECFV (24%).⁹

Standardization of GFR is also of concern in obese human patients, who are clearly different from lean individuals of similar body weights. The higher the weight, the lower the GFR indexed to BSA. Other ways to index GFR have been tested unsuccessfully. An absolute, non-corrected GFR is currently recommended in obese patients.³¹ There are no published data regarding indexation of body weight in obese dogs or cats. In children, variation in body shape or constitution causes difficulties with prediction formulae and simplified approaches.³²

Further research is needed in order to evaluate the best method for standardization of GFR values in dogs and cats. Such research is not simple because there is no "golden standard" to relate to. Ideally, regression analysis should be used on a large representative population of adult dogs (different breeds, body weights, age and sex), evaluating different methods of standardization. A simplified alternative would be to stratify the canine population into body weight categories and define an “average” cut-off for the GFR estimates in each category. The main limitation then would be that the greater the range of body weights for a given category, the more inaccurate the corresponding derived cut-off values. For the time being, it seems logical to standardize to body weight as most publications on reference intervals make use of this approach.

Reference intervals

Defining GFR reference intervals for dogs and cats is a prerequisite for classification of patients as normal or abnormal. Reference values are only published for adult animals; while values are different for puppies and kittens (see above). It is generally considered that the lower cut-off value is between 1.5 and 2.0 mL/min/kg. Considering the effect of physiological variability observed in recent studies, however, it appears that a unique cut-off value is not acceptable for the overall canine population. Stratification is advisable according to body weight, but possibly not to age.

In one previous study (113 healthy dogs),³³ animals were divided into 4 body weight categories (Mini, Medium, Maxi and Giant). The corresponding GFR values (mean ± SD) were 3.7 ± 0.5, 3.0 ± 0.5, 2.5 ± 0.4, and 2.4 ± 0.6 mL/min/kg, respectively (Lefevbre HP et al ; unpublished data). If the hypothetical distributions of GFR values were normal, the corresponding lower limits of the reference ranges could be considered to be 2.7, 2.0, 1.7 and 1.3 mL/min/kg. In another study involving 118 adult healthy dogs, the reference ranges in different weight quartiles were 1.54–4.25, 1.29–3.50, 1,2–3.36, and 1.12–3.39 mL/min/kg, respectively, for body weight quartiles of 1.8–12.4, 13.2–25.5, 25.7–31.6, and 32.0–70.3 kg.⁵

These results, obtained with two different markers (creatinine and iohexol), demonstrate that depending on body weight, the lower cut-off value is higher for small dogs,and lower for giant dogs. Thus, choosing a single value of 1.5 mL/min/kg to declare a dog as renal-impaired will be too low for small dogs (giving some false negative results) and too high for giant dogs (some false positive results).

Breed may also affect the reference intervals. For example, tentative reference intervals for English Pointers, English Setters and German Shepherds are 2.3–5.1, 1.8–5.0 and 1.7–3.8 mL/min/kg.²⁷ It seems impractical to try to establish breed-specific reference intervals for all breeds, other than the major ones. In our experience, most breeds tend to follow their weight category with respect to reference values in healthy dogs.

Plasma clearance reference ranges for cats have been proposed recently: 1.0–3.5 mL/min/kg for iohexol and 1.3–3.8 mL/min/kg for creatinine.⁹ From the point of view of allometric relationships,²⁴ it is interesting that the cut-off value for GFR is lower in cats than in dogs of similar body weight.

These preliminary results provide some estimates of reference intervals for veterinary nephrologists, but further investigations in larger canine and feline populations are obviously required.

Perspectives
on GFR measurements in cardiovascular and endocrine diseases

GFR measurements may be interesting to assess renal function in small animal patients with non-primary renal diseases, as shown recently for some cardiovascular and endocrine diseases.

Cardiovascular diseases

CKD was shown recently to be prevalent in humans with cardiovascular disease, wherein a very slight decrease in renal function may dramatically increase the cardiovascular risk.³⁴ Azotaemia is common in dogs with cardiac diseases,³⁵ but the degree of azotaemia is mostly mild to moderate and not related to changes in cardiovascular variables.³⁵ Interestingly, the GFR in 24 dogs with chronic valvular disease was significantly lower in NYHA III-IV (1.7±0.7 mL/min/kg) than in NYHA I-II (3.1±0.8 mL/min/kg). Only 1/15 NYHA dogs in class I-II had GFR <2 mL/min/kg and only 2/9 NYHA class III-IV dogs had GFR >2 mL/min/kg.³⁵ Further investigations are needed to identify the cause of decreased GFR and its clinical relevance to the outcome of cardiac disease.

Endocrine diseases

Interactions exist between endocrine systems and kidney function under both physiological and clinical conditions. One of the best examples in veterinary medicine is probably the effect of hyper- or hypothyroidism on renal function in small animals.³⁹

Hyperthyroidism is a common endocrine disorder in aged cats and a 14–40% prevalence of pre-existing CKD has been reported in affected cats.³⁶ Evaluation of kidney function in a hyperthyroid cat is pivotal as hyperthyroidism can mask and sometimes worsen CKD. CKD became clinically apparent after treatment of hyperthyroidism in about 17–39% of cats.³⁶ A challenge in such patients is to predict the risk of post-treatment azotaemia. Pre-treatment GFR was shown to be predictive of development of post-treatment CKD.³⁷,³⁸ An accurate evaluation of kidney function can be performed from 4 weeks after¹³ 1I-treatment.³⁸,³⁹

GFR was shown to be decreased in experimentally-induced canine hypothyroidism whereas blood creatinine concentrations remained unaltered.⁴⁰ In 14 dogs with naturally occurring hypothyroidism, GFR was <2 mL/min/kg in all, and increased with levothyroxine treatment.⁴¹ The potential effects of renal dysfunction and development of subclinical CKD on the clinical outcome of hypothyroid dogs require further investigation.

Other situations where questions might arise about adequacy of kidney function are in diabetes mellitus or spontaneous hyperadrenocorticism, or during glucocorticosteroid therapy. The usefulness of GFR esitmation is of limited value at present in these situations, because little is know about how they affect GFR, so studies are needed to establish GFR values for these patients.

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