Second applications of freshly prepared CCh solutions do not affect activity characteristics or synaptic growth.

<p>Two experiments (out of three in total) in which a second bolus of freshly prepared CCh solution was added to the MEA dishes 143 h (left, same experiment as <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040980#pone-0040980-g001" target="_blank">Figure 1</a>) and 137 hours (right) after the experiment was started (45 and 89 hours after first CCh application, respectively). <b><i>A</i></b>) Spontaneous activity rates. <b><i>B</i></b>) Burstiness index. <b><i>C</i></b>) Mean synaptic size (left - 5 sites; right - 8 sites).