FGF14 regulates the intrinsic excitability of cerebellar Purkinje neurons

https://doi.org/10.1016/j.nbd.2008.09.019Get rights and content

Abstract

A missense mutation in the fibroblast growth factor 14 (FGF14) gene underlies SCA27, an autosomal dominant spinocerebellar ataxia in humans. Mice with a targeted disruption of the Fgf14 locus (Fgf14−/−) develop ataxia resembling human SCA27. We tested the hypothesis that loss of FGF14 affects the firing properties of Purkinje neurons, which play an important role in motor control and coordination. Current clamp recordings from Purkinje neurons in cerebellar slices revealed attenuated spontaneous firing in Fgf14−/− neurons. Unlike in the wild type animals, more than 80% of Fgf14−/− Purkinje neurons were quiescent and failed to fire repetitively in response to depolarizing current injections. Immunohistochemical examination revealed reduced expression of Nav1.6 protein in Fgf14−/− Purkinje neurons. Together, these observations suggest that FGF14 is required for normal Nav1.6 expression in Purkinje neurons, and that the loss of FGF14 impairs spontaneous and repetitive firing in Purkinje neurons by altering the expression of Nav1.6 channels.

Introduction

The autosomal dominant spinocerebellar ataxias (SCAs) are heterogeneous neurological disorders characterized by progressive cerebellar ataxia that also often present with paroxysmal dyskinesia, tremor, abnormal eye movements, and/or cognitive impairment (Schols et al., 2004, Taroni and DiDonato, 2004, Zoghbi, 2000, Zoghbi and Orr, 2000). The unexpected findings of ataxia, dystonia and tremor in mice with a targeted disruption in the Fgf14 locus (Wang et al., 2002) suggested a role for FGF14 in regulation of motor function, and led directly to the identification of a mutation in FGF14 in a family with a new type of progressive spinocerebellar ataxia now referred to as SCA27 (Van Swieten et al., 2003). Affected individuals in this family possess a missense mutation (F145S) in the FGF14 gene on chromosome 13q34 (Brusse et al., 2005, Van Swieten et al., 2003). Interestingly, a single base pair deletion, leading to a frameshift mutation (D163fsX12) in FGF14 has also been identified in an individual patient presenting with ataxia and mild mental retardation (Dalski et al., 2005, Soong and Paulson, 2007).

FGF14 belongs to the intracellular fibroblast growth factor subfamily (iFGF) that also includes FGFs 11–14 (Itoh and Ornitz, 2008), proteins that are widely expressed in the nervous system (Smallwood et al., 1996, Wang et al., 2000, Yamamoto et al., 1998). Unlike the other members of the FGF family, iFGFs are not secreted and do not interact with classical tyrosine kinase FGF receptors (Itoh and Ornitz, 2008, Olsen et al., 2003, Ornitz and Itoh, 2001, Smallwood et al., 1996). Members of the iFGF subfamily, however, have been shown to colocalize with voltage-gated sodium (Nav) channels, to interact with the C-termini of Nav channel pore-forming (α) subunits (Goldfarb et al., 2007, Laezza et al., 2007, Liu et al., 2001, Liu et al., 2003, Lou et al., 2005, Wittmack et al., 2004), and to modulate hippocampal and granule cell excitability (Goldfarb et al., 2007, Laezza et al., 2007, Wozniak et al., 2007, Xiao et al., 2007).

Purkinje neurons are the sole output of the cerebellum, critical for motor regulation and coordination (Burgess et al., 1995, Grusser-Cornehls and Baurle, 2001, Koeppen, 2005, Kohrman et al., 1996, Levin et al., 2006, Sausbier et al., 2004, Trudeau et al., 2006). A distinctive feature of mature Purkinje neurons is the robust expression of Nav1.6-encoded sodium channels (Afshari et al., 2004, Krzemien et al., 2000, Raman et al., 1997), the channel that underlies the “resurgent” sodium current that is critical for sustaining the characteristic high frequency firing of these cells (Raman et al., 1997). Mutations in human SCN8A, which encodes Nav1.6, are associated with cerebellar atrophy and ataxia (Trudeau et al., 2006). In addition, mice in which Scn8a has been disrupted or that lack Nav1.6 specifically in Purkinje neurons display cerebellar ataxia and impaired Purkinje neuron firing (Kohrman et al., 1996, Levin et al., 2006, Raman et al., 1997). These observations suggested that the ataxia in SCA27 individuals and in Fgf14−/− mice reflects impaired Purkinje neuron firing due to alterations in Nav channel expression and/or functioning. The experiments here were designed to explore this hypothesis directly.

Section snippets

Cerebellar slice recordings

Whole-cell recordings were obtained from Purkinje neurons in 300 μm parasagittal cerebellar slices prepared from 25 to 30 day old wild type (WT) and Fgf14−/− C57BL6 mice. Vibratome sections were cut in ice-cold solution containing (in mM): 87 NaCl, 2.5 KCl, 25 NaHCO3, 1 NaH2PO4, 0.5 CaCl2, 7 MgCl2, 75 sucrose and 10 glucose, bubbled with 5% CO2/95% O2. Slices were incubated at room temperature in artificial CSF (ACSF), containing in mM: 125 NaCl, 2.5 KCl, 26 NaHCO3, 1.25 NaH2PO4, 2 CaCl2, 1 MgCl

Expression of Fgf14 in Purkinje neurons

Although it has previously been reported that Fgf14 expression is evident in the granule cell layer in the (mouse) cerebellum, the resolution was insufficient to identify expression in specific cell types (Wang et al., 2002). To determine if Fgf14 is expressed in Purkinje neurons, high resolution in situ hybridization was performed on thick sections of wild type (WT) mouse cerebellum. These experiments revealed robust expression of the Fgf14 transcript in Purkinje neurons and moderate

Spontaneous and evoked repetitive firing are attenuated in Fgf14−/−Purkinje neurons

Purkinje neurons are the sole output of the cerebellar cortex, and Purkinje neuron dysfunction or degeneration has been linked to cerebellar ataxias in human and in animal models (Burgess et al., 1995, Grusser-Cornehls and Baurle, 2001, Koeppen, 2005, Kohrman et al., 1996, Levin et al., 2006, Sausbier et al., 2004, Trudeau et al., 2006). Mature Purkinje neurons express predominantly Nav 1.6-encoded Nav channels, which generate resurgent Nav currents during action potential repolarization,

Acknowledgments

This work was supported by the Hope Center for Neurological Disorders, the National Ataxia Foundation and by an NIH Neuroscience Blueprint Center Core P30 NS057105 grant to Washington University and 1R03NS62431-1. Some of this work was performed in a facility supported by NCRR grant C06 RR015502. The monoclonal antibody FGF14N56/21 was obtained from the UC Davis/NINDS/NIMH NeuroMab Facility, supported by NIH grant U24NS050606 and maintained by the University of California at Davis.

References (37)

  • WangQ. et al.

    Subcellular and developmental expression of alternatively spliced forms of fibroblast growth factor 14

    Mech. Dev

    (2000)
  • WozniakD.F. et al.

    Impaired spatial learning and defective theta burst induced LTP in mice lacking fibroblast growth factor 14

    Neurobiol. Dis

    (2007)
  • XiaoM. et al.

    Impaired hippocampal synaptic transmission and plasticity in mice lacking fibroblast growth factor 14

    Mol Cell Neurosci

    (2007)
  • YamamotoS. et al.

    Structure and expression of a novel isoform of mouse FGF homologous factor (FHF)-4

    Biochim. Biophys. Acta

    (1998)
  • ZoghbiH.Y.

    Spinocerebellar ataxias

    Neurobiol. Dis

    (2000)
  • AfshariF.S. et al.

    Resurgent Na currents in four classes of neurons of the cerebellum

    J Neurophysiol

    (2004)
  • BrusseE. et al.

    Spinocerebellar ataxia associated with a mutation in the fibroblast growth factor 14 gene (SCA27): a new phenotype

    Mov Disord

    (2005)
  • BurgessD.L. et al.

    Mutation of a new sodium channel gene, Scn8a, in the mouse mutant ‘motor endplate disease’

    Nat Genet

    (1995)
  • Cited by (100)

    • Physiology of Dystonia: Animal Studies

      2023, International Review of Neurobiology
    • 5.09 - Sodium Channels

      2020, The Senses: A Comprehensive Reference: Volume 1-7, Second Edition
    • Ion channel dysfunction in cerebellar ataxia

      2019, Neuroscience Letters
      Citation Excerpt :

      In FGF14−/− mice, Purkinje neuron spontaneous firing is greatly disrupted, with 80% of neurons appearing silent. Additionally, Nav1.6 expression is reduced in Purkinje neurons of FGF14−/− mice, suggesting that this interaction is necessary for normal membrane expression [92]. The FGF14F145S mutation reduces Nav1.6 expression at the axon initial segment, and reduces sodium currents in hippocampal neurons, leading to early depolarization block upon current injections [58].

    View all citing articles on Scopus
    1

    These authors contributed equally to this work.

    View full text