The Interaction Between Ocular and Extraocular Photosensory Systems In Planarian Flatworms
Abstract
When contemplating the central factors of the evolution of eyes, few individuals consider the impact of light perception and response opposed to the resolution of visual images. Consequently, studies considering how distinct light sensing mechanisms cause different behavioral outputs are notably minimal. However, by conducting research into differential light-sensing structures and how they produce their responses, significant questions regarding the evolution of eyes could possibly be answered. While there has been extensive research into the phylogeny of vision throughout vertebrate eyes, the evolution of light sensing in invertebrates is immensely less popular. As a result, this project sought to explore how exactly different light stimuli transform into distinct responses. In order to do this, the planarian flatworms were placed in a setting where they had to decide to go near a certain light frequency, or no light. Then, the number of planarians on each side was measured, and a ratio was calculated in order to compare the distribution of the planarians. After collecting the data for healthy, complete planarians, the data was then collected for decapitated, as well as regenerating planarians. The results were analyzed using a One-Way ANOVA, where the wavelengths acted as the groups, and the ratio of worms on each side was compared to the days after the decapitation.
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The Interaction Between Ocular and Extraocular Photosensory Systems In Planarian Flatworms
Introduction
The ability to perceive and react to light has substantially influenced the phylogeny of organisms throughout the natural world. Even so, while vision in vertebrate eyes has attracted attention over the years, the eye evolves in numerous ways. More specifically, quantitative studies on the behavioral consequences of light sensing, especially in invertebrates, have been severely understudied. However, by achieving a richer understanding of the diversity of light-induced behavior, substantial developments in eye evolution have the ability to take place [Nilsson, 2013]. In fact, the effects of light-mediated behavior have even been reasoned to be responsible for evolutionary selection in addition to sensory structures [Nilsson, 2013]. Therefore, quantitative research into how organisms process and respond to specific light inputs are valuable to grasping the evolution of the eye.
Rhabdomeric Photoreception
Light sensing systems are classified based on the photoreceptors, neural networks, and sensing structures present in the organism [Nilsson, 2013]. Numerous planarian flatworms, such as the Schmidtea mediterranea and Dugesia species, possess rhabdomeric eyes. Rhabdomeric eyes are rather simple, in that they are essentially eye pits with a single opsin or photosensor [Paskin, Jellies, Bacher, & Beane, 2014]. Nonetheless, this class of eyes is exceedingly significant. The eyes perceive the presence and direction of light as well as low-resolution vision [Nilsson, 2013]. Additionally, this eye class is present in a cornucopia of animal species- suggesting an evolutionary advance [Nilsson, 2013]. However, there is a severe lack of research involving these types of eyes, aside from elements of low-resolution image. The mechanism used when these eyes are converting light stimuli into behavioral outputs remains a mystery.
Regeneration
Planarian flatworms possess incredible regenerative abilities. For instance, planarians are able to regenerate their eyes and dorsal ganglion, which acts as their brain, within mere days [Inoue, et. al., 2004]. Although the process of regeneration has been a subject of longstanding fascination [Inoue, et. al, 2004], there seems to be an absence of knowledge regarding the relationship between regeneration and the recovery of sensory function. Hence, planarians offer the opportunity to observe how light-sensing structures process distinct types of light stimuli.
Interactions Between Ocular and Extraocular Systems
Despite the various limitations in the conceptual understanding of eyes, even less is known about the interplay between different types of responses to light, with their own evolutionary histories [Cronin, Johnesen, 2016]. In addition to eye-based perception of light, metazoans possess the ability to sense light without designated eye structures [Cronin, Johnesen, 2016]. These eye-independent perceptions of light are known as extraocular, while eye-mediated perceptions of light have been classified as ocular [Xiang, et. al., 2010]. Presentations of extraocular as well as ocular sensing systems have been exhibited in countless organisms, including Drosophila Larvae [Xiang, et. al., 2010] and Caenorhabditis Elegans [Ward, et. al., 2008]. Although ocular and extraocular systems have been researched separately, there is minimal information available concerning the interactions between such systems in the same organism.
Overall, there seems to be a few questions regarding numerous areas of research as discussed. These questions include the photosensory role of rhabdomeric eyes, the relationship between regeneration and the recovery of sensory responses, as well as the interplay between ocular and extraocular systems. Consequently, this study aims to determine how the stage of regeneration affects interactions between ocular and extraocular photosensory systems when manipulating the wavelength of the light in which the planarians are exposed to from.
Methodology
1.1 Light Source and Measurements
Light-emitting diodes (LED) with specialized wavelengths were obtained from Roithner Lasertechnik. The light-emitting diodes utilized in this study include a 350 nanometer diode, a 400 nanometer diode, a 525 nanometer diode, a 545 nanometer diode, a 590 nanometer diode, and a 625 nanometer diode. In order to keep the light-emitting diode frequencies consistent, a circuit along with a power source was constructed. Additionally, the light from the diodes were released directly above the planarians from an LED mount stand with an iPhone Six Plus camera attached to it in order to calculate the number of planarians in each section of the glass slide.
1.2 Planarian Maintenance
The species of planarian flatworms used in this study were the Schmidtea Mediterranea. There were 45 planarians that were maintained at standard laboratory conditions. Once every three days, the planarians were fed beef liver extract. However, before the start of the experiment, all of the flatworms were deprived of food for a minimum of two days. Lastly, the experiments took place in a dark environment maintained at approximately 20°C.
1.3 The Experiment
The study began with the decapitation of each of the planarian flatworms. The flatworms were decapitated at just below the eye pigments (Figure 1). The flatworms were each starved for at least two days before the commencement of the experiment. Approximately 5 milliliters of medium was added to a glass slide and a single planarian was placed in the center region (Figure 2) using a Pasteur pipette. Next, the LED light was turned on, so that Region 1 (R1) was relatively illuminated, while Region 3 (R3) was significantly darker. Once two minutes would pass, the region in which the planarian was present in was determined. This process would repeat so that all 30 worms were tested, and so that all LED wavelength settings were experimented, as well. Following the testing of all 30 worms, for each LED wavelength setting, a Discrimination Index (DI) was calculated. The DI was calculated through the formula, DI = (NR3 − NR1)/N. The term, “NR†indicates the number of worms in a region, and the term “N†simply indicates the number of total worms. A DI of 1 represents a complete aversion to the wavelength, a DI of -1 indicates a complete aversion to the dark setting, while a DI of 0 indicates no preference.