P2X7 receptors modulate outer retinal and retinal ganglion cell synaptic responses in the ex vivo mouse retina

Abstract

Purpose: P2X₇R activation is known to modulate mouse electroretinogram (ERG) responses in vivo, as well as driving photoreceptor and retinal ganglion cell (RGC) damage, a consequence of various neurodegenerative conditions. However, the complexity that P2X₇Rs are distributed throughout all layers of the retina, and their localisation on various cell types puts into question their specific site(s) of action. Using a dark-adapted, ex vivo mouse retinal wholemount preparation, the present study aimed to characterise the effect of P2X₇R activation on light-evoked outer retinal responses, and on RGC synaptic responses in the inner retina, under comparable conditions.

Methods: ERG responses were recorded from the surface of isolated retinal wholemounts (adult, female C57BL/6 mice) in response to a light stimulus (1s-duration, peak wavelength 562nm, repeated every 3s). In separate experiments, ON field excitatory post-synaptic potentials (fEPSPs) were recorded from the ganglion cell layer wholemounts. The NMDAR-mediated component of the responses was isolated with a Mg­­²⁺-free Krebs medium containing NBQX, picrotoxin, strychnine and tetrodotoxin. Additional compounds were applied to the bathing medium.

Results: In physiological Krebs medium, BzATP (300µM), a P2XR agonist, induced a marked potentiation of the ERG a-wave slope to 149.0±14% of control with a concomitant reduction in the b-wave amplitude to 80.6±4% of control (both P<0.05 vs. control, n=7). BzATP also reduced the power of the ERG oscillatory potentials. In the pharmacologically modified, Mg­­²⁺-free perfusate, BzATP reduced both the a-wave slope (70.1±8% of control) and b-wave amplitude (66.6±10% of control; both P<0.05 vs. control, n=7). The effect of BzATP on the ERG components were markedly reduced by the selective P2X₇R antagonist, A438079 (10µM; a-wave: 4±16% of control; b-wave: 58.3±16% of control; both P<0.05, n=7). The ON RGC fEPSP was reduced in the presence of BzATP (78±2% of control; P<0.0001, n=21), an effect which was significantly attenuated by A438079 (68.7 ±2% of control; P<0.05, n=6).

Conclusions: In summary, P2X₇R modulates the photoreceptor response, ON-bipolar cell output and ON-retinal ganglion cell responses through partially independent mechanisms. Understanding the physiological role of P2X₇Rs in synaptic signalling may provide clues as to how these ‘danger-sensors’ contribute to neuronal dysfunction.