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Marc Meadows, Veeramuthu Balakrishnan, Xiaohan Wang, Henrique Von Gersdorff; Exocytosis of glycine during crossover inhibition at the AII amacrine cell synapse.. Invest. Ophthalmol. Vis. Sci. 2016;57(12):2752.
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© ARVO (1962-2015); The Authors (2016-present)
The AII amacrine cell (AII-AC) is essential for visual processing in the mammalian retina by effectively coupling the Rod- and ON-Cone bipolar cell pathways while inhibiting the OFF-Cone bipolar cell (OFF-CBCs) pathway (crossover inhibition). This interneuron sustains a continuous release of glycine from the interneuron’s proximal dendritic lobules despite the lack of specialized ribbon synapse. Our research goals aim to uncover factors modulating AII-AC exocytosis during long excitatory stimuli that are physiologically relevant for scotopic and mesopic conditions.
Retinal slices generated from C57 BL6J mice were perfused in Ames medium. AII-ACs were identified under DIC microscopy and whole cell recording were performed at 33-34 oC with cesium based internal solutions. Cells were held at -80 mV and capacitance (Cm) measurements were performed by the “sine DC” method, using a 2 kHz sinusoidal voltage command (30 mV peak to peak) and analyzed using an EPC-10 Lock-in amplifier. OFF-CBCs IPSCs were recorded from slices pre-incubated with Forskolin in a NaCl based bath solution at 32 oC. Mean and SEM values were reported from ANOVA and paired t-tests.
AII-ACs are able to sustain large levels of exocytosis in response to long step depolarizations of up to 1200 ms (184.1 ± 13.4 fF; n=4). We observed a trend of asynchronous release, although the presence and degree of asynchronous release was heterogeneous among different recordings. We also observed that dialysis of cAMP into the AII-AC increases exocytosis 2-fold for a 200 ms depolarizing pulse (control: 33.9 ± 6.9 fF; n=5; cAMP: 88.8 ± 12.9 fF; n=5; p<0.01). Similar effects were observed by stimulating intercellular cAMP with Forskolin. Furthermore, Forskolin effectively potentiated IPSC frequency in OFF-CBCs from control levels (control: 8.05 ± 4.1 Hz; Forskolin: 10.28 ± 3.9 Hz; n=8; p=0.025).
Measuring the changes in membrane capacitance (Cm) across a broad range of depolarizations has uncovered the AII-ACs capacity for continuous exocytosis in response to tonic and graded depolarizations. In some recordings we obtained clear evidence of a copious amount of asynchronous release of vesicles capable of sustaining glycine release after the end of a long depolarization. Furthermore, the exocytosis in AII-ACs is also modulated by cAMP levels, which can change in response to shifts in ambient light or during the night/day cycle.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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