Figure 3

To assess the responsiveness of the sensors to acetylcholine, (a) initial rates over 15 seconds produced by 5.6 nM solutions of the sensors in phosphate buffer saline were determined (K_m = 1.05 mM, n = 3). (b) The florescent signal of the nanosensors (380 nM) was monitored in response to repeated cycles of exposure to 50 mM acetylcholine in PBS (Black arrow) followed by PBS washes (Grey arrow) demonstrating their reversible response to acutely high concentrations of acetylcholine. The (c) selectivity of the nanosensors was examined by exposing them to neurotransmitter solutions (30 mM) (White bars) of GABA, L-glutamate, glycine and dopamine (300 μM) (ANOVA, *p < 0.005 for n = 3). The activity of the sensors towards acetylcholine in the presence of the neurotransmitters was then examined with a successive addition of 15 mM acetylcholine (Grey bars) (ANOVA, *p > 0.05 for n = 3). The acetylcholine response range for butyrylcholinesterase based sensors includes the expected millimolar synaptic acetylcholine concentrations⁹ and the sensors responses may be further enhanced by the presence of endogenous synaptic cholinesterases⁵¹. All data are shown as mean ± s.d.