March 2012
Volume 53, Issue 14
Free
ARVO Annual Meeting Abstract  |   March 2012
The Cdk5 Inhibitor, Roscovitine, Protects Retinal Neurons From Chemical Anoxia, In Vitro, And Excitotoxicity, In Vivo
Author Affiliations & Notes
  • John P. Wood
    Ophthalmic Research Laboratories, S Australian Institute of Ophthalmology, Adelaide, Australia
  • Teresa Mammone
    Ophthalmic Research Laboratories, S Australian Institute of Ophthalmology, Adelaide, Australia
  • Glyn Chidlow
    Ophthalmic Research Laboratories, S Australian Institute of Ophthalmology, Adelaide, Australia
  • Robert J. Casson
    Ophthalmic Research Laboratories, S Australian Institute of Ophthalmology, Adelaide, Australia
  • Footnotes
    Commercial Relationships  John P. Wood, None; Teresa Mammone, None; Glyn Chidlow, None; Robert J. Casson, None
  • Footnotes
    Support  NHMRC grants #565202, #626964 and #508123
Investigative Ophthalmology & Visual Science March 2012, Vol.53, 6573. doi:
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      John P. Wood, Teresa Mammone, Glyn Chidlow, Robert J. Casson; The Cdk5 Inhibitor, Roscovitine, Protects Retinal Neurons From Chemical Anoxia, In Vitro, And Excitotoxicity, In Vivo. Invest. Ophthalmol. Vis. Sci. 2012;53(14):6573.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: : The cyclic-dependent kinase family member, cdk5, has been implicated in a wide variety of neuronal functions including development, synaptic plasticity, cell survival and cell death. Since the tissue milieu in the pathological retina is thought to be conducive to the activation of cdk5, we sought to investigate the role of this enzyme in different paradigms of injury. Furthermore, we aimed to determine whether inhibition of cdk5 with its known inhibitor, roscovitine, could prevent cell and tissue damage in such models.

Methods: : Mixed retinal cell cultures comprising glia and neurons were challenged for 24 hours with 500μM sodium azide (NaN3), a mitochondrial complex IV inhibitor (chemically mimics anoxia), in the presence or absence of 10-500μM roscovitine. Neuronal death and survival was assessed by immunocytochemistry and immunoblotting (tau, PGP9.5, GABA, MAP2, βIII-tubulin). For in vivo studies, adult SD rats were subjected to intraocular injection with either vehicle, 40nmol/eye N-methyl-D-aspartate (NMDA), 2.5μM roscovitine or NMDA/roscovitine together, and left for one week. Retinas/optic nerves were analysed by RT-PCR, immunoblot and immunohistochemistry to determine neuronal loss and/or survival. Activation of cdk5 was assessed by determining levels of this protein and of its endogenous activator p35.

Results: : Treatment of retinal cultures with 500μM NaN3 elicited a reduction of neuronal numbers to approximately 40-55% of the control level, although this varied depending on the requisite neuron marker employed (eg. 55.8% with PGP9.5, 42.4% with βIII-tubulin). Roscovitine was able to significantly attenuate neuron loss in a concentration-dependent manner (eg. 500μM roscovitine elevated NaN3-treated neuron numbers to 79.3%). In vivo, NMDA caused a decrease in proteins and mRNAs for βIII-tubulin, neurofilaments, tau and PGP9.5 in the retina and optic nerve; roscovitine-treated animals showed a relatively and significantly increased level of such markers and hence an increased neuronal survival. In both in vivo and in vitro models, an increase in the levels of p35 was detected but the levels of cdk5 remained constant.

Conclusions: : Cdk5 plays a role in retinal neuron death both in vitro in response to "chemical anoxia" and in vivo in response to an excitotoxic insult. Inhibition of this enzyme with roscovitine leads to a greater survival of neurons in both cases. The present data imply a role for cdk5 in pathological neuronal death in the retina.

Keywords: retinal degenerations: cell biology • neuroprotection • pathology: experimental 
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