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Title: Regulation of extracellular signal-regulated protein kinases (ERK) 1 and 2 in synaptic nerve terminals
Author: Lee, Vivian Wing Yan
Awarding Body: UCL (University College London)
Current Institution: University College London (University of London)
Date of Award: 2006
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Activation of extracellular signal-regulated kinases 1 and 2 (ERK 1 and 2) is a key signalling event in the modulation of presynaptic function. This study looks at the upstream regulation of ERK 1 and 2 signalling in rat cerebrocortical synaptosomes. Kinase activation assays based on phospho-state specific antibodies revealed two major inputs for the activation of ERK 1 and 2: i) a neurotrophin-mediated signalling to ERK 1 and 2 which is dependent on the activation of small GTP-binding protein Ras and the presence of Ca ii) a Ca -dependent activation of ERK 1 and 2, stimulated by depolarisation or by the Ca ionophore ionomycin. Treatment of synaptosomes with Ca2+ chelators showed that basal ERK 1 and 2 activation was partly supported by Ca2+ from intracellular sources, whilst depolarisation-dependent activation of ERK 1 and 2 was entirely attributable to extracellular Ca influx. Like the BDNF-mediated activation, this Ca -dependent signalling to ERK was contingent on Ras, as evinced by the use of Ras inhibitor lovastatin. Using inhibitors of Ca /CaM-dependent protein kinase II (CaMKII) and phosphatidylinositol-3-kinase (PI3K), we next showed that both kinases are involved in mediating the Ca -dependent ERK 1&2 pathway. Furthermore, the Src family kinase (SFK) inhibitor, PP2, strongly reduced Ca -dependent ERK 1 and 2 activation, suggesting a role for non-receptor tyrosine kinases (nRTKs) in upstream signalling. Finally, using okadaic acid, roscovitine and baclofen respectively, we showed that the duration and efficacy of ERK 1 and 2 activation are determined by the function of protein phosphatase 2A (PP2A), cyclin- dependent kinase 5 (cdk5) and the presynaptic Gj/0-coupled GABAb receptors. Interestingly, our data demonstrate that baclofen-mediated inhibition of ERK 1&2 can be overridden by BDNF stimulation, revealing a potential feedback and cross-talk mechanism between the excitatory ERK and inhibitory GABA cascades. Together, these studies elucidate the role of ERK 1 and 2 as a hub for signalling in the nerve terminal.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available