Oral Presentation 5th International Symposium on Phaeochromocytoma and Paraganglioma 2017

Zebrafish mutants in the von Hippel-Lindau tumor suppressor display a normetanephrine dominant phenotype (#11)

Peter MT Deen 1 , Selma Waaijers 1 2 , Antonius E van Herwaarden 2 , Rachel Giles 3 , Henri JLM Timmers 4
  1. Physiology, Radboud Univ. Med. Center, Nijmegen, Netherlands
  2. Laboratory Medicine, Radboud Univ. Nijmegen Med Center, Nijmegen, Gelderland, The Netherlands
  3. Nephrology and hypertension, University Medical Center Utrecht, Utrecht, The Netherlands
  4. Internal Medicine, Radboud Univ. Nijmegen Med Center, Nijmegen, The Netherlands

Introduction: Zebrafish vhl mutants develop key aspects of the human disease condition, including activation of the hypoxia-inducible factor (HIF) signaling pathway, polycythemia, excessive neovascularization, macular edema, and pronephric abnormalities.1 Whether the human VHL-related pheochromocytoma/paraganglioma (PPGL) phenotype is replicated in vhl mutant zebrafish has not been investigated. Our aim was to develop a method to measure catecholamines and metanephrines in zebrafish and to investigate the presence of features of PPGL.

 Methods: liquid chromatography/mass spectrometry was used to measure metanephrines in zebrafish of lysates of larvae and in (urine containing) medium they were swimming in. Concentrations of metanephrines were compared between vhl-/- and wild-type (wt) zebrafish.

 Results: Protocols for optimal sample preparation and (nor)metanephrine assays were established for both fish lysates and swimming medium. In lysates of 30 5-days-post-fertilization wt zebrafish larvae, the mean levels of metanephrine and normetanephrine were 0.010±0.001 (SEM and 0.036±0.002 pmol/larva, respectively. In 30 vhl-/- zebrafish larvae, normetanephrine levels were 2-fold higher (0.056±0.01; P<0,01), whereas metanephrine levels were >2 (0.004±0.001; P<0,001). In the swimming water, metanephrins were undetectable, while in medium of vhl-/- larvae the normetanephrine levels were 2,5-fold increased () as compared to wt (1.093±0.115 versus 0.429±0.049 pmol/larva; P<0,01). No obvious tumors of the fish’s ‘adrenal’ gland were observed.

 Conclusion: We have established a method for the quantification of (nor)metanephrines in zebrafish and Vhl knock-out larvae display a normetanephrine dominant phenotype despite absence of obvious PPGL-like tumors. As such, vhl zebrafish represent a unique in vivo model for studying human VHL genotype–phenotype correlations, including (features of) PPGL.


  1. Van Rooijen E, et al. Zebrafish mutants in the von Hippel-Lindau tumor suppressor display a hypoxic response and recapitulate key aspects of Chuvash polycythemia. Blood. 2009 Jun 18;113(25):6449