Our understanding of
human endocrine pancreas function has been greatly enhanced by comparative studies on teleost fish. Perhaps the most important work on pancreas function utilizing teleosts was performed by J.J.R. Macleod,1
who used the large, isolated principal islets of angler fish and sculpin to definitively establish that the islets secrete insulin, with no contribution from acinar tissue. This pioneering study confirmed the work of Banting and Best,2
who extracted insulin from the pancreas of dogs, and showed that injecting it into pancreatectomized dogs alleviated diabetes. For their discovery of insulin, Banting and Macleod shared the Nobel Prize in physiology or medicine in 1923. Since then, pancreas function has been studied in a wide variety of fish, and the literature contains a wealth of comparative data for fish blood glucose levels. Recently, zebrafish (Danio rerio
) blood glucose studies have been added to this list.3–5
The zebrafish is a small, freshwater teleost that has become a popular and informative developmental and genetic model. Zebrafish pancreas specification and morphogenesis have been well studied (reviewed in Refs.6,7
), making this organism attractive for further development into a physiological model of pancreas function. The zebrafish islet, like that of other teleosts, is organized similarly to the human islet and consists of α-, β-, δ-, and
-cells, with pancreatic polypeptide antibody-reactive cells also reported.8
Additionally, several zebrafish models have been established for studying β-cell or islet regeneration.5,9–12
These studies demonstrate the ability of the zebrafish islet to regenerate following isletectomy or β-cell-specific ablation. Studies such as these have the potential to significantly enhance diabetes-related research. However, we currently lack tools for evaluating zebrafish islet function.
An important parameter for evaluating islet function is blood glucose level, which is measured, for example, to determine whether the islet is secreting a sufficient amount of insulin. Adult zebrafish are small (typically <4
cm in length), which makes them an attractive model for many applications, but which presents a challenge for measuring blood glucose. Existing methods used for other animals need to be scaled-down or redesigned. Fish biologists have traditionally used the glucose oxidase assay, which is also used for the clinical assay of human blood. However, these studies have utilized larger fish for which it is relatively easy to sample blood, and to collect adequate sample volumes. For zebrafish, performing a traditional glucose assay on individuals presents problems. To collect the required sample volume, blood from multiple individuals must be combined, which is less than ideal, or the blood from an individual must be diluted, which is only feasible if the particular assay is sensitive enough.
As an alternative to either of these potential sampling methods, we tested two hand-held meters used by diabetic humans for measuring blood glucose. The meters have several advantages over traditional laboratory methods, including speed and portability: glucose can be measured within seconds of blood collection, on site. More important, the hand-held meters require only a microliter or less of blood, giving this method a distinct advantage over traditional laboratory methods. Here we present the details of tests of both meters. We found that both Meter A, which uses the glucose oxidase method, and Meter B, which uses pyrroloquinoline quinone (PQQ) glucose dehydrogenase, were consistent with results from a laboratory glucose oxidase assay. We also present methods for applying anesthesia, for testing fasted and postprandial glucose levels, and for performing an intraperitoneal glucose tolerance test (IP-GTT). These methods will enable zebrafish researchers to study pancreatic function, and will also open the way to studies of metabolic disorders and diseases that perturb glucose homeostasis. As glucose homeostasis involves multiple organs and cell types, including the liver, skeletal muscle, and adipose tissue, these new methodologies will have very broad application.