Elsevier

Brain Research Bulletin

Volume 69, Issue 6, 31 May 2006, Pages 669-679
Brain Research Bulletin

Calcineurin inhibitors cause an acceleration of the neurological phenotype in a mouse transgenic for the human Huntington's disease mutation

https://doi.org/10.1016/j.brainresbull.2006.03.013Get rights and content

Abstract

Calcineurin (CaN) is a Ca2+- and calmodulin-dependent protein serine-threonine phosphatase that is thought to play an important role in the neuronal response to changes in the intracellular Ca2+ concentration. CaN has been implicated in numerous physiological processes including learning and memory. Decreases in CaN expression are thought to be responsible for some of the pathological features seen in brain ischemia, Down's syndrome and Alzheimer's disease. In this study, we examined the possibility of CaN playing a role in the progressive neurological phenotype of the R6/2 mouse of Huntington's disease. We studied the effects of the CaN inhibitors cyclosporin A and FK506 on the progressive neurological phenotype in the R6/2 mouse. We found that an immunosuppressive dose of both drugs dramatically accelerated the main features of the neurological phenotype in R6/2 mice. This was unlikely to be due solely to the immunosuppressive action of these drugs, since treatment with cyclophosphamide, an immunosuppressant drug with a mechanism of action that is not mediated via CaN, did not have deleterious effects on the R6/2 mouse. If anything, cyclophosphamide improved the neurological symptoms in the R6/2 mice. Together, our data suggest a central role for CaN in the deleterious phenotype of the R6/2 mouse. Treatments aimed at preventing the loss of CaN or stimulating its function may be beneficial in the treatment of HD.

Introduction

Huntington's disease (HD) is a genetic neurodegenerative disorder that typically develops in the fourth or fifth decade of life and progresses relentlessly until death. It is characterized by movement disorders, personality changes and cognitive impairment [for references, see 3]. The profound atrophy seen in the striatum of HD patients is caused by the selective loss of medium spiny γ-amino-butyric acid (GABA) containing neurons. However, neither the mechanisms underlying the selective neuronal degeneration in HD, nor the process by which the HD mutation causes the neurological symptoms in HD are clearly understood.

The Ca2+- and calmodulin-dependent protein serine-threonine phosphatase 2B, calcineurin (CaN) is highly enriched in neural tissue, particularly in striatum and hippocampus [31], [32], [56]. CaN is a heterodimer made up of a 58–64 kDa catalytic subunit (CaNA) and a 19 kDa regulatory subunit (CaNB) [see 50]. CaN has a number of important functions in the brain, including neurotransmitter signal transduction, long-term memory formation, excitotoxicity, regulation of intracellular Ca2+, transcriptional regulation and apoptosis [24], [44], [50], [59] (Table 1). CaN is also thought to be involved in several other neurodegenerative conditions. For example, a decrease in CaN expression has been reported after ischemic damage [43], [62], and over-expression of the CaN intracellular constitutive inhibitor Down's syndrome critical region 1 (DSCR1)-Adapt 78 protein has been postulated to be causative of certain features of the pathogenesis of Down's syndrome [11], [15], [19] and Alzheimer's disease [16], [26], [38]. Additionally, CaN over-activation plays a role in brain ageing [1], [17].

The aim of this study was to examine the role of CaN in the neurological phenotype in the R6/2 transgenic mouse model of HD. The R6/2 mouse [41] carries exon one of the HD gene with a pathologically expanded CAG repeat. This produces a rapidly progressing disease in mice, with cognitive deficits starting as early as 3–4 weeks [39] and motor deficits appearing at around 5–6 weeks [10] and premature death at around 4–5 months of age. The behavioural phenotype of the R6/2 mice was examined following treatment with two inhibitors of CaN, FK506 and cyclosporin A (CsA). The behaviour of the mice was compared to that seen after treatment with another immunosuppressive drug, cyclophosphamide (CyPho). A number of different parameters were measured (survival, body weights, diabetes onset, and motor performance), to assess the effect of CaN inhibition on the progressive neurological phenotype of R6/2 mice.

Section snippets

Animals

Mice were taken from a colony of R6/2 mice established in the Department of Pharmacology, University of Cambridge and maintained by backcrossing onto CBA × C57BL/6 F1 × CBA mice. Mice were housed in cages of mixed genotype with 12 h light and dark cycles in a temperature and humidity-controlled room, with ad libitum access to food and water. Mice were assigned randomly to the different groups of mice and experiments carried out blind to genotype until the mice developed an overt phenotype.

Genotyping

Genotypes

Survival

None of the WT mice died in the course of the experiment (Fig. 1a). As expected, saline-treated R6/2 mice died prematurely at around 4 months of age (mean age at death = 100.8 ± 4.5 days) (Fig. 1b). However, the survival of the R6/2 mice in both of the CaN inhibitor treatment groups was significantly reduced (Fig. 1b). Mean age at death was 76.9 ± 3.7 days for FK506-treated mice (p < 0.001 compared to saline-treated mice) and 84.5 ± 4.4 days for CsA-treated mice (p < 0.05, compared to saline-treated mice).

Discussion

The role of CaN in the pathogenesis HD has not been investigated previously. However, there is considerable evidence to suggest that impairments in CaN function might play a deleterious role on other neurological conditions, including forebrain ischemia [43], [62], Down's syndrome [11], [15], [19] and Alzheimer's disease [16], [26], [38]. Moreover, CaN knockout mice models show impaired cognitive function [63]. Here we examined the role of CaN in the behavioural pathogenesis of the R6/2 mouse

Acknowledgements

We would like to thank Mrs. Wendy Leavens for technical assistance on the development of this project. This work was funded by CURE HD Initiative of the Hereditary Disease Foundation and HighQ Foundation.

References (64)

  • M. Morioka et al.

    Potential role of calcineurin for brain ischemia and traumatic injury

    Prog. Neurobiol.

    (1999)
  • R.A. Nichols et al.

    Calcineurin-mediated protein dephosphorylation in brain nerve terminals regulates the release of glutamate

    J. Biol. Chem.

    (1994)
  • R. Sinigaglia-Coimbra et al.

    Protective effect of systemic treatment with cyclosporine A after global ischemia in rats

    J. Neurol. Sci.

    (2002)
  • S.H. Snyder et al.

    Neural actions of immunophilin ligands

    TIPS

    (1998)
  • A. Stelzer

    GABAA receptors control the excitability of neuronal populations

    Int. Rev. Neurobiol.

    (1992)
  • H.Y. Wu et al.

    Critical role of calpain-mediated cleavage of calcineurin in excitotoxic neurodegeneration

    J. Biol. Chem.

    (2004)
  • Y. Yamasaki et al.

    Alteration in the immunoreactivity of the calcineurin subunits after ischemic hippocampal damage

    Neuroscience

    (1992)
  • H. Zeng et al.

    Forebrain-specific calcineurin knockout selectively impairs bidirectional synaptic plasticity and working/episodic-like memory

    Cell

    (2001)
  • L. Andreeva et al.

    Cyclophilins and their possible role in the stress response

    Int. J. Exp. Pathol.

    (1999)
  • G.P. Bates et al.

    Huntington's Disease

    (2002)
  • D.J. Begley et al.

    Permeability of the blood–brain barrier to the immunosuppressive cyclic peptide cyclosporin A

    J. Neurochem.

    (1990)
  • J.A. Bibb et al.

    Severe deficiencies in dopamine signaling in presymptomatic Huntington's disease mice

    Proc. Natl. Acad. Sci. U.S.A.

    (2000)
  • M. Braun et al.

    GABAB receptor activation inhibits exocytosis in rat pancreatic beta-cells by G-protein-dependent activation of calcineurin

    Physiology

    (2004)
  • S.E. Browne et al.

    The energetics of Huntington's disease

    Neurochem. Res.

    (2004)
  • S.P. Butcher et al.

    Neuroprotective actions of FK506 in experimental stroke: in vivo evidence against an antiexcitotoxic mechanism

    J. Neurosci.

    (1997)
  • G. Camirand et al.

    Combined immunosuppression of mycophenolate mofetil and FK506 for myoblast transplantation in mdx mice

    Transplantation

    (2001)
  • R.J. Carter et al.

    Characterization of progressive motor deficits in mice transgenic for the human Huntington's disease mutation

    J. Neurosci.

    (1999)
  • K.T. Chang et al.

    The Drosophila homolog of Down's syndrome critical region 1 gene regulates learning: implications for mental retardation

    Proc. Natl. Acad. Sci. U.S.A.

    (2003)
  • O.M. Colvin

    An overview of cyclophosphamide development and clinical applications

    Curr. Pharm. Des.

    (1999)
  • G. Ermak et al.

    DSCR1 (Adapt78)—a Janus gene providing stress protection but causing Alzheimer's disease?

    IUBMB Life

    (2003)
  • T.C. Foster et al.

    Calcineurin links Ca2+ dysregulation with brain aging

    J. Neurosci.

    (2001)
  • H. Friberg et al.

    Cyclosporine A, but not FK 506, protects mitochondria and neurons against hypoglycemic damage and implicates the mitochondrial permeability transition in cell death

    J. Neurosci.

    (1998)
  • Cited by (18)

    • Pathophysiology of Huntington's disease: Time-dependent alterations in synaptic and receptor function

      2011, Neuroscience
      Citation Excerpt :

      One possible explanation for such differences comes from studies indicating that signaling downstream of NMDARs is dramatically altered with disease onset. In mouse models of HD and/or human HD brains, Ca2+ handling is more efficient (Hansson et al., 2001), scaffolding and signaling proteins in the NMDAR signaling complex show altered expression (Luthi-Carter et al., 2003; Jarabek et al., 2004), and some downstream death signaling proteins, such as calcineurin, are downregulated (Goto et al., 1989; Hernández-Espinosa and Morton, 2006; Xifró et al., 2009). These changes could all contribute to resistance to excitotoxicity, but would also be expected to reduce capacity for learning and promote deficits in motor performance.

    • FK506 ameliorates cell death features in Huntington's disease striatal cell models

      2011, Neurochemistry International
      Citation Excerpt :

      Furthermore, mHtt has been reported to interfere with transcription factor activity or may function to disrupt coactivator complexes on susceptible gene promoters (Rosenstock et al., 2010, for review). Interestingly, FK506 was previously reported to modulate transcription through several targets, such as cAMP response element binding protein (CREB) (Dineley et al., 2010), inhibitor-1 and dopamine- and cyclic AMP-regulated phosphoprotein relative molecular mass 32 kDa (DARPP-32), NMDAR and α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptors (AMPAR), inositol-triphosphate receptor 1 (IP3R) and ryanodine receptor (for review, Hernandez-Espinosa and Morton, 2006). Thus, the neuroprotection in immortalized striatal neurons induced by FK506 in the presence of STS can also be related to transcription regulation.

    • Reduced calcineurin protein levels and activity in exon-1 mouse models of Huntington's disease: Role in excitotoxicity

      2009, Neurobiology of Disease
      Citation Excerpt :

      We have previously observed that, in contrast to N-terminal exon-1 mhtt, full-length mhtt up-regulates calcineurin A mRNA levels (Xifro et al., unpublished results). The opposite results on calcineurin A protein regulation observed in full-length (Xifro et al., 2008) and N-terminal exon-1 mhtt models (present results; Lievens et al., 2002; Hernandez-Espinosa and Morton, 2006) could be related to the presence of aggregates. It has been proposed that proteins interacting with exon-1 mhtt, such as transcription factors, can be sequestered in the aggregates, which impede their function (Li and Li, 2004).

    • Infusion of FK506, a specific inhibitor of calcineurin, induces potent tau hyperphosphorylation in mouse brain

      2008, Brain Research Bulletin
      Citation Excerpt :

      Because CN is important in cellular signal transduction, it is involved in many biological processes, including immune responses and cardiac hypertrophy [19]. The studies on biological roles of CN have progressed to the important discovery that it is the target of the immunosuppressant drug cyclosporin A (CsA) and FK506, which can also be found as tacrolimus [3,13]. CsA and FK506 inhibit CN activity after forming complexes with cytoplasmic immunophilins, cyclophilins and FKBP [2].

    View all citing articles on Scopus
    View full text