Planarians are traditional model invertebrates in regeneration and developmental biology research that also display a variety of quantifiable behaviors useful to screen for pharmacologically active compounds. One such behavior is the expression of seizure-like movements (pSLMs) induced by a variety of substances. Previous work from our laboratory showed that cocaine, but not nicotine, induced pSLMs in intact but not decapitated planarians. Interestingly, as decapitated planarians regenerated their heads, they gradually recovered their sensitivity to cocaine. These results suggested a method to assess planarian brain regeneration and a possible way of identifying compounds that could enhance or hold back brain regeneration. In the present work, we demonstrate that the cholinergic agent cytisine is a suitable reference compound to apply our method. Cytisine induces pSLMs in a concentration-dependent manner in intact (but not decapitated) planarians of the species Girardia tigrina. Based on our data, we developed a behavioral protocol to assess planarian brain regeneration over time. We tested this method to measure the effect of ethanol on G. tigrina’s brain regeneration. We found that ethanol slows down the rate of planarian brain regeneration in a concentration-dependent manner, consistently with data from other research groups that tested ethanol effects on planarian brain regeneration using different behavioral protocols. Thus, here we establish a general method using cytisine-induced pSLMs as an indicator of brain regeneration in planarians, a method that shows potential for assessing the effect of pharmacologically active compounds in this process.

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One of the best-characterized planarian behaviors induced by various compounds is the change in locomotor velocity. Previous work from our laboratory showed that the sesquiterpene lactone parthenolide and the local anesthetic cocaine, reduce planarian motility. Parthenolide reverses the cocaine-induced motility decrease and vice versa. However, the exact mechanism of the cocaine/parthenolide antagonism for this specific planarian behavior is still unknown. Here we report the results of a Schild analysis to determine whether the parthenolide/cocaine relationship is orthosteric or allosteric. Our results suggest an orthosteric relationship between these two compounds in the planarian Girardia tigrina. The simplest interpretation of our data is a shared binding site for cocaine and parthenolide. Still, we cannot rule out the possibility of distinct yet overlapping binding sites with the data available.