Modulation of Retinal Ganglion Cell Function and Implications for Neuroprotection

• Main Author: Martins, João Filipe da Costa
• Format: Dissertation
• Language: English
• Published: ProQuest Dissertations & Theses 01-01-2014
• Subjects: Neurosciences; Ophthalmology
• ISBN: 9798607324377

Neuropeptide Y (NPY) is a neuromodulator in central nervous system (CNS) that can exert neuroprotective effects. NPY is expressed in mammalian retina but the location and potential modulatory effects of NPY receptor activation remains largely unknown. Retinal ganglion cell (RGC) death is a hallmark of several retinal degenerative diseases, particularly glaucoma. In purified RGCs, we detected immunoreactivity and mRNA for NPY and NPY receptors in these cells. Using cultured purified RGCs and ex vivo retinal preparations we have evaluated the effect of NPY receptor activation on changes in RGC intracellular free calcium concentration – [Ca2+]i and RGC spiking activity. RGC spike recordings were performed by a multi-electrode array (MEA). We found that NPY application attenuated the increase in the [Ca2+]i triggered by glutamate in purified RGCs, possibly via Y1 receptor activation. Moreover, Y1/Y5 receptor activation increased the initial burst response of OFF-type RGCs, though no effect was observed in the RGC spontaneous spiking activity. The Y1 receptor activation was able to modulate directly RGC responses by attenuating the N-Methyl-D-aspartic acid (NMDA)-induced increase in RGC spiking activity. These results suggest that Y1 receptor activation at the level of inner or outer plexiform layers leads to modulation of RGC receptive field properties. Using in vitro culture of retinal explants exposed to NMDA, we found that NPY pre-treatment prevented NMDA-induced cell death through activation of Y1 and Y5 receptors. In an animal model of retinal ischemia-reperfusion (I-R) injury, pre-treatment with NPY was not able to prevent cell death or rescue RGCs. In summary, we found clear modulatory effects of NPY at the level of RGCs, and Y1 receptor appears to have a predominant role. However, further studies are needed to evaluate whether NPY neuroprotective action translates to in vivo models of retinal degenerative diseases. Sildenafil (ViagraTM), a cyclic guanosine monophosphate (cGMP)-specific phosphodiesterase type 5 inhibitor, is widely used for the treatment of erectile dysfunction and pulmonary arterial hypertension. Clinical studies reported transient visual impairments in patients after a single dose sildenafil ingestion, suggesting the implication of RGCs, since these cells convey visual information to the brain centres of visual processing. However, the effect of sildenafil on the RGC light responses is not fully understood. Using a MEA technique, in the second part of this study, we evaluated the effect of sildenafil on RGC light responses in ex vivo retinas. Under continuous perfusion, sildenafil citrate (0.3 to 30 µM) was applied to retinal preparations during 10 to 60 min followed by sildenafil washout. High concentration (30 µM) of sildenafil reversibly decreased the magnitudes of both ON- and OFF-type RGC light responses, and in 50% of RGCs, light responses were completely supressed. Sildenafil also greatly increased the latency of ON- and OFF-types of light responses. We provide the evidence that extended exposure to sildenafil and repeated light stimulation potentiates drug effects and delays recovery. In conclusion, we show that MEA recordings in ex vivo retinas might be a valuable method to understand how RGC circuitry can be affected by different drug treatments. This understanding is relevant to the development of neuroprotective strategies needed for retinal degenerative diseases, namely glaucoma, where no available treatment can effectively stop the progression of RGC death.