Planarian brain regeneration as a model system for developmental neurotoxicology

Abstract

Freshwater planarians, famous for their regenerative prowess, have long been recognized as a valuable in vivo animal model to study the effects of chemical exposure. In this review, we summarize the current techniques and tools used in the literature to assess toxicity in the planarian system. We focus on the planarian's particular amenability for neurotoxicology and neuroregeneration studies, owing to the planarian's unique ability to regenerate a centralized nervous system. Zooming in from the organismal to the molecular level, we show that planarians offer a repertoire of morphological and behavioral readouts while also being amenable to mechanistic studies of compound toxicity. Finally, we discuss the open challenges and opportunities for planarian brain regeneration to become an important model system for modern toxicology.

Keywords: Behavior; neurodevelopment; planarians; regeneration; toxicology.

Figure 1

Comparison of the two scoring systems by Grebe and Schaeffer (GS system) (Grebe & Schaeffer 1991) and Wu et al (2012a). While substantial overlap exists between the two systems, the GS system provides more readouts (18 vs. 13). The Wu system has the advantage of clear categories of shape changes.

Figure 2

Examples of planarian morphological readouts and body shapes. (A) Pharynx is extended outside of the body. Animal treated with 0.4% chloretone. (B) Body is contracted. Often associated with wrinkles/ornamentation in the periphery of the animal. Animal treated with 20 μmol/L chlorpyrifos oxon. (C) Body is curled in a C‐shape. Also referred to as a banana curl or coil. Animal treated with 0.4% chloretone. (D) Body is twisted around itself in a screw‐like fashion. Also referred to as spiraling. Animal treated with 100 mmol/L serotonin. (E) Animal is extended and moves along its side in a snake‐like motion. Animal treated with 100 mmol/L serotonin. Scale bar 0.5 mm.

Figure 3

Overview of behavioral assays employed in the literature to quantify neuronal function after toxicant exposure. (A) The planarian locomotor velocity (pLMV) method measures worm speed by counting the number of gridlines crossed in a given time. (B) Center of mass (COM) tracking to determine type of locomotion, worm velocity and exploratory behavior. (C) Phototaxis is generally tested using a linear light gradient. (D) Thermotaxis can be tested using a Peltier element to generate a cooler center, which worms prefer. Scale bar 1 cm.

Figure 4

Morphological and anatomical readouts of developmental neurotoxicity in planarians. (A) Time course of regeneration of control (top) and 15 mg/mL Triton X‐100 treated (bottom) worms. On day 1, animals are amputated along the black dotted line. On day 4, the unpigmented blastema (indicated by the white dotted line) is clearly distinguishable. On later days, the reappearance of eyes (black asterisk) and auricles (white asterisk) can be scored. (B) Brain structure is visualized by immunohistochemistry with anti‐synapsin antibody (anti‐ SYNORF1, Developmental Studies Hybridoma Bank) in control and 0.1% ethanol treated regenerating animals 2 weeks post‐amputation. Scale bar 0.1 mm.