Age-dependent pattern of cerebellar susceptibility to bilirubin neurotoxicity in vivo in mice.

Giulia Bortolussi,Giulia Viviani,Tamara Bittolo,Gabriele Baj,Andrés F Muro,Simone Vodret

doi:10.1242/dmm.016535

Abstract

Neonatal jaundice is caused by high levels of unconjugated bilirubin. It is usually a temporary condition caused by delayed induction of UGT1A1, which conjugates bilirubin in the liver. To reduce bilirubin levels, affected babies are exposed to phototherapy (PT), which converts toxic bilirubin into water-soluble photoisomers that are readily excreted out. However, in some cases uncontrolled hyperbilirubinemia leads to neurotoxicity. To study the mechanisms of bilirubin-induced neurological damage (BIND) in vivo, we generated a mouse model lacking the Ugt1a1 protein and, consequently, mutant mice developed jaundice as early as 36 hours after birth. The mutation was transferred into two genetic backgrounds (C57BL/6 and FVB/NJ). We exposed mutant mice to PT for different periods and analyzed the resulting phenotypes from the molecular, histological and behavioral points of view. Severity of BIND was associated with genetic background, with 50% survival of C57BL/6‑Ugt1−/− mutant mice at postnatal day 5 (P5), and of FVB/NJ-Ugt1−/− mice at P11. Life-long exposure to PT prevented cerebellar architecture alterations and rescued neuronal damage in FVB/NJ-Ugt1−/− but not in C57BL/6-Ugt1−/− mice. Survival of FVB/NJ-Ugt1−/− mice was directly related to the extent of PT treatment. PT treatment of FVB/NJ-Ugt1−/− mice from P0 to P8 did not prevent bilirubin-induced reduction in dendritic arborization and spine density of Purkinje cells. Moreover, PT treatment from P8 to P20 did not rescue BIND accumulated up to P8. However, PT treatment administered in the time-window P0–P15 was sufficient to obtain full rescue of cerebellar damage and motor impairment in FVB/NJ-Ugt1−/− mice. The possibility to modulate the severity of the phenotype by PT makes FVB/NJ-Ugt1−/− mice an excellent and versatile model to study bilirubin neurotoxicity, the role of modifier genes, alternative therapies and cerebellar development during high bilirubin conditions.

Highlights

Using phototherapy at different times after birth, the authors identified the critical window of neuronal susceptibility to bilirubin toxicity and showed that the neurotoxic effects of bilirubin depend on the developmental stage of the cerebellum
We showed that susceptibility to bilirubin damage and, survival of mutant animals is strain-specific, with a significant increase in the survival of FVB/NJ-Ugt1−/− compared with C57BL/6-Ugt1−/− mice, the cerebellum being the most affected organ in both mutant strains
Phenotype severity is associated with the genetic background We have previously reported the generation of a mouse model of neonatal severe hyperbilirubinemia bearing a targeted mutation in the Ugt1 gene, which resembles the human Crigler-Najjar syndrome type I (CNSI) from both the genetic and phenotypic points of view (Bortolussi et al, 2012)

Summary

Introduction

Neonatal jaundice occurs in more than 60% of normal newborns during their first week of life (Bhutani and Johnson, 2009a; Bhutani and Johnson, 2009b; Karen et al, 2009) as a result of excessive unconjugated bilirubin (UCB) formation and transient inability of. Up to 35% of newborns with kernicterus die, and some of the survivors suffer from permanent neurological sequelae (Kaplan et al, 2011)

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Conclusion