The signal transduction and gene regulatory events which occur downstream of the precipitating causes of retinal diseases is largely unknown. Yet these downstream sites would be excellent targets for interventional therapies. The long term goal of the proposed research is to define the role of calmodulin dependent protein kinase II (CaMKII) which is likely downstream of the initiating events in eye diseases like retinal ischemia. CaMKII is upstream of the cell death and survival pathways in retinal ganglion cells (RGCs). We know that when challenged with glutamate receptor agonists: 1) retinal cytoplasmic CaMKII1 becomes autophosphorylated; 2) there is an increase in the alternatively spliced nuclear transcript, CaMKII1B; 3) caspase-3 becomes activated and 4) cells in the inner nuclear and ganglion cell layers die. We have demonstrated that the specific inhibitor of CaMKII, autocamtide inhibitory peptide (AIP), blocks these changes (1, 3 and 4) and protects all cells from cell death. Pilot data with the aid of a cell line indicates that the AIP promotes the secretion of brain derived neurotrophic factor (BDNF) which may aid the survival of challenged cells. We also know that an elevated level of CaMKII1B (2) is associated with enhanced transcription of BDNF. Since glutamate is implicated in retinal ischemia and CaMKII1 is also activated in this condition, we hypothesize that CaMKII1 plays an important role in pathways involved in the death and survival responses of RGCs during retinal ischemia. This will be tested with methods, which include RNA- interference for the CaMKII1B transcript. We will use a rat retinal ischemia-reperfusion model, as well as primary cultured rat RGCs, and ex vivo retinal explants.
Aim 1 will determine the time frame in which AIP protects RGCs during/after ischemia.
Aim 2 will explore the role of cytoplasmic CaMKII in regulating caspase-3 activation, NF:B translocation and BDNF release in RGC/retinas.
Aim 3 will explore the role of nuclear CaMKII1 in cell death/survival and the underlying mechanisms. The results of these studies will provide important insights into the signal transduction pathways and gene regulatory networks that control death and survival responses of RGCs which will be of relevance to retinal ischemia.
Retinal ischemia, a common clinical entity that has been widely studied because of its proposed role in, for example, retinal and choroidal vessel occlusion, anterior ischemic optic neuropathy, glaucoma, and diabetic retinopathy, etc, remains a common cause of blindness in the developed world due to relatively ineffective treatment. We propose to explore the signal transduction pathways linked to gene regulatory networks which control cell death and survival processes in rat retinal ganglion cells, whose death is a major contributing factor for visual impairment in these diseases. Knowledge of such pathways and networks is important for the development of practical therapeutic interventions. ? ? ?
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