Protection of Adult Pig Retinal Ganglion Cells from Glutamate-Induced Excitotoxicity
Glaucoma is a neurodegenerative disease characterized by the death of optic nerve fibers and consequent blindness (Glaucoma Research Foundation, 2005). One theory suggests that increased intraocular pressure evokes the release of excess glutamate. The overabundance of this excitatory neurotransmitter causes an excitotoxic effect, which kills the retinal ganglion cells (RGCs). Data indicates that acetylcholine (ACh) released from amacrine cells in the retina can prevent excitotoxicity by acting as an agonist for nicotinic ACh receptors (nAChRs) on RGCs (Wehrwein et al, 2004). PNU 282987 is a synthetic drug that is a nicotinic agonist selective for the ±7 nAChR (Tocris, 2006). To determine if PNU has neuroprotective effects against glutamate-induced excitotoxicity, RGCs were isolated from adult porcine eyes using a two-step panning technique, which utilized an antibody selective for RGCs. The isolated cells were plated on culture dishes at consistent densities. Some dishes received no treatment (control), other dishes received glutamate (excitotoxicity), and the remaining dishes received glutamate and agonist (PNU or nicotine) at various concentrations (neuroprotection). Cell survival was assayed after constant exposure to treatment for three days. Live cells were labeled with a fluorescent dye and microscopic images were captured and analyzed. Results showed that PNU displayed significant neuroprotective effects at concentrations of 10nM and 100nM. Similar results were obtained with nicotine, with maximal effects seen at 500nM. When ±-bungarotoxin, a nicotinic antagonist specific for ±7 nAChRs, was pre-applied with PNU, the efficacy of PNU was reduced supporting the concept that the neuroprotective effect of PNU is mediated through alpha7 nAChRs.
Faculty Mentor: David Linn
Page last modified May 13, 2011