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Niamh M C Connolly


Medicine & Nursing

neurodegenerative diseases amp activated protein kinases cerebrovascular disorders computational modelling therapeutic targets medicine and health sciences stress physiological neurotoxins

AMP-Activated Protein Kinase (AMPK) Activation and Signalling Following Neuronal Excitotoxicity (2013)

Abstract Toxic stimulation of neurons, excitotoxicity, is a pathological process implicated in ischaemic stroke and neurodegenerative diseases. Excess glutamate in the synaptic cleft leads to severe ionic influx into the neuron, energetic stress and the activation of the AMP-activated protein Kinase (AMPK), a key regulator of energetic homeostasis. Interestingly, AMPK activation during excitotoxicity can be both cytoprotective and cytotoxic, but the molecular switches that determine whether neurons undergo necrosis or apoptosis or tolerate an excitotoxic insult are not well understood.Using an interdisciplinary approach combining biochemistry, single-cell imaging and computational modelling, we here investigated some of the mechanisms governing neuronal fate following transient glutamate excitotoxicity. We characterised neuronal bioenergetics in rat primary neurons at single-cell level using novel fluorescent sensors for intracellular glucose, ATP and AMPK activity. We identified ATP depletion and recovery to energetic homeostasis, along with AMPK activation, as acute, surprisingly rapid responses following the onset and termination of excitotoxicity. Interestingly, glutamate exposure also induced an accumulation of intracellular glucose, providing an additional source of energy during and after glutamate-induced bioenergetic stress. Surprisingly, cells that more quickly recovered their glucose levels to baseline survived longer, indicating that the ability to regulate the glutamate-mediated glucose accumulation may be beneficial for neuronal viability.Employing computational modelling of neuronal bioenergetics and AMPKmediated survival signalling in excitotoxicity, we correctly resembled the rapid single-cell kinetics of ATP levels and AMPK activity, but failed to render glucose dynamics, indicating the presence of additional glucose regulatory mechanisms during excitotoxicity. Further single-cell experiments revealed that neither inhibition of AMPK, inhibition of glucose transport, nor removal of extracellular substrate completely abolished the glucose increase, suggesting the presence of an intracellular glucose source in neurons, releasable during excitotoxic stress and independent of glucose transport and AMPK activity.Development of a second computational model, of AMPK-mediated apoptotic signalling activated during excitotoxicity, revealed a network motif (coherent feed-forward loop) capable of filtering the effects of short-term AMPK activity on the expression of the pro-apoptotic protein Bim. This motif may prevent unwanted Bim expression and apoptosis during transient or physiological bioenergetic stress, allowing AMPK to mediate its pro-survival effects. Biochemistry experiments validated model predictions that Bim expression was determined by the duration of AMPK activity following transient neuronal excitotoxicity.We conclude that our research increased insight into the molecular mechanisms that govern neuronal fate following transient excitotoxicity, which may eventually help to predict potential therapeutic targets capable of reducing neuronal damage in stroke and neurodegenerative diseases.
Collections Ireland -> Royal College of Surgeons in Ireland -> PhD theses
Ireland -> Royal College of Surgeons in Ireland -> Theses and Dissertations

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Niamh M C Connolly

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Niamh M C Connolly
Royal College of Surgeons in Ireland
Total Publications: 14