Presynaptic depolarization differentially regulates dual neurotransmitter release from starburst amacrine cells in the mouse retina

Abstract

The retina is comprised of diverse neural networks, signaling from photoreceptors to ganglion cells to encode images. The synaptic connections between these retinal neurons are crucial points for information transfer; however, the input-output relations of many synapses are understudied. Starburst amacrine cells in the retina are known to contribute to retinal motion detection circuits, providing a unique window for understanding neural computations. We examined the dual transmitter release of GABA and acetylcholine from starburst amacrine cells by optogenetic activation of these cells, and conducted patch clamp recordings from postsynaptic ganglion cells to record excitatory and inhibitory postsynaptic currents (EPSCs and IPSCs). As starburst amacrine cells exhibit distinct kinetics in response to objects moving in a preferred or null direction, we mimicked their depolarization kinetics using optogenetic stimuli by varying slopes of the rising phase. The amplitudes of EPSCs and IPSCs in postsynaptic ganglion cells were reduced as the stimulus rising speed was prolonged. However, the sensitivity of postsynaptic currents to the stimulus slope differed. EPSC amplitudes were consistently reduced as the steepness of the rising phase fell. By contrast, IPSCs were less sensitive to the slope of the stimulus rise phase and maintained their amplitudes until the slope became shallow. These results indicate that distinct synaptic release mechanisms contribute to acetylcholine and GABA release from starburst amacrine cells, which could contribute to the ganglion cells' direction selectivity.

Keywords: EPSCs; GABA; acetylcholine; iPSCs; kinetic; optogenetic; postsynaptic currents; synapse.

Figure 1

SAC voltage changes in response to distinct kinetics of ontogenetically stimuli. (A) SAC voltage responses evoked by 10 ms, 100 ms, 300 ms, 600 ms, and 990 ms triangle rise time stimuli. Traces in black were individual sweeps, and the trace in red indicate an average of five recordings. The rise time is indicated below each stimulus. (B) A summarized graph showing the time constant (tau) of SAC response rise phase as a function of the stimulus rise time (N=5 SACs). It revealed that ChR2 responded linearly as the light stimuli temporal aspects changed.

Figure 2

EPSPs and IPSPs varied when the rising phase of SAC stimuli changed. (A) Representative Sweeps of IPSCs (upper) and EPSCs (lower) obtained from a ganglion cell in response to optogenetic stimuli of 10 ms to 990 ms rise times. (B) (left) Normalized EPSCs in ganglion cells as a function of SAC stimuli rise time (black, n=15 RGCs) with average responses shown in blue. (middle) Normalized IPSPs (black, n=9 RGCs) with average IPSC shown in red. (right) The average EPSCs and IPSCs amplitudes are plotted with asterisks displaying a p< 0.05 when comparing the IPSC and EPSC response of the same triangle rise time. (C) The latency between stimulus onset to response onset time. Five RGCs that exhibited both EPSCs and IPSCs were selected and compared their latencies. (left) EPSC latencies and average in blue. (middle) IPSC latencies and average in red. (right) The average EPSC and IPSC latencies were overlaid, displaying no differences.