Developmental Biology-Planaria Lab PDF

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Regeneration of a New Planarian Body

Introduction Planaria is known as a species that can produce sexually and asexually. Are hermaphroditic as they possess male and female parts. They can produce asexually by fragmentation and tail dropping that allows them to regenerate the missing pieces. Planarian regeneration was one of the first studies scientists looked at where the gradient concept was

developed (Adell et al. 2010). The regeneration of planarian fragments was studied from different parts of the body (Adell et al. 2010). After the planaria was cut into different fragmentations it is expected that a blastema regenerates a head while the other will form a tail (Nogi & Levin, 2005). Cell communication is done through gap-junctional communication that allows signaling of the Innexin gene to be cloned and regenerate the planaria tail (Nogi & Levin, 2005). In planarian innexins there is an expression detected that plays a role in “long-range signaling” that allows the blastema to identify itself and regenerate tail (Nogi & Levin, 2005). It is though that gap-junctional communication initiates instructive signaling throughout regeneration tail (Nogi & Levin, 2005). Day Zero 1) I think environmental variables will influence the regeneration process. The rate at which the planaria regenerate has to do with many factors such as the population density. This includes how many planaria there are in this area. There is also the physiological factors of the planaria. The physiological factors can include the last time the planaria has been cut or regenerated. Physiological factors look at how long the regeneration took the previous cut or how the fission went throughout the asexual reproduction. Other environmental issues may also cause issues such as how much light is involved. As seen in the lab planaria tend to crawl away from the light and seemed to prefer the shaded areas. Another environmental factor would be the amount of injury the planaria has seen (Pellettieri et al., 2010). While dealing with the planaria you need to be very gentle as they can die easily, this was an issue in lab. There is also how much nutrients there is available to the planaria during the regeneration period (Pellettieri et al., 2010). The death of the organism was least persistent when the animals are fed and can regenerate faster (Pellettieri et al., 2010). Factors such as

pollution, predation, unusually high temperatures and red tides would affect planarian regeneration in their wild habitats (Bely and Nyberg, 2010). With the lab experiment high temperatures and pollution would be the main concerns. 2) After the cut the mobility of each fragment appears to be different. The cut of the planaria was the 4th cut assigned and was cut into 3 parts: the head, middle and the tail. Starting with the mobility of the head it was still very mobile and was crawling away from the light. The head still has the photoreceptors therefore seemed to have the most mobility of the three. The middle did not move that much and seen under the microscope was the least mobile of the three segments. The middle had no distinct features like the head, and this could be why there was such little movement. The middle portion also had light grey fragments that looked like a cilium hanging off each end. The tail did have a bit of movement, and more than the middle segment. It had a pointy end which was the tail but did not seem to be in a hurry to get out of the light like the head. Since the head has eyespots that are sensitive the light would explain why the head had the most movement under the light of the microscope while the middle and the tail did not. Therefore, they would move differently at the first cut. 3) The head fragment prefers shade over light. The eyespots are sensitive to light and this directs the planaria to the shade. This can be tested by shining light directly onto the planarian sample and to see if it moves to the shade or stays in the light. While in the lab it is shown that the head fragment preferred the shade. When analyzing the fragment under the microscope, the planaria head was rapidly moving away from the light. Within planaria it is also known that this sensitivity to light is caused from the photoreceptors in the head fragment (Brown and Ogden, 1968).

Figure 1.0 Shows the planaria before the cut. It is about 8mm in length. The photoreceptors are visible.

Figure 1.1 The initial cut. This shows the head of the planaria. Photoreceptors were still visible. The length is about 4mm.

Figure 1.2 The initial cut. This shows the middle of the planaria. It had no visible receptors or neoblasts.

Figure 1.3 The initial cut. This shows the tail of the planaria. It has no visible photoreceptors or neoblasts. Final Day 1) Not all the fragments below the head regenerated. The fragments below the head that did not regenerate was the tail. The tail survived up until the final week, where it then died and disintegrated. The guts of the tail excreted outward and caused it to die. This could have been due to the force applied when removing the specimen to the plate. The middle

and head still had photoreceptors. The head unfortunately also died when looking at the last check-up. 2) As only one of the fragments regenerated photoreceptors it is hard to compare the timing it would have regenerated in comparison to the other photoreceptors. The regeneration of different fragments would have no specific timing in which they would regenerate. The time it takes to regenerate depends on the wound. The “number of neoblasts wandering to the wound” is what causes the rate of regeneration (Brøndsted, 1954). It also depends on the age of the planaria how fast that it regenerates (Brøndsted, 1954). If a fragment is bigger or smaller it can also hinder the rate of the regeneration as there is a difference (Brøndsted, 1954). It takes a while for them to regenerate as seen in the lab as the neoblasts are not readily available for the blastema formation and they are migrated from different parts of the body (Brøndsted, 1954). 3) Since only one of the fragments developed photoreceptors there is no results in how the ability of different sections would regenerate throughout the worm. However, within the middle fragment the photoreceptors developed after about a week and created a head. As talked about earlier the rate of regeneration can be affected by the size of the fragments and the neoblasts not being readily available for immediate regeneration (Brøndsted, 1954). 4) There was a change in the mobility of the tail fragment over time. The tail was alive until the last week of analyzing the specimen. The tail never had a fast mobility and was weak from the beginning of the analysis. The tail then eventually slowed down died and disintegrated. The head had a very strong mobility up until the last checkup. Currently unaware of why the head ended up dying. Possibilities could have been due to exerted

force to the specimen. The head had the most mobility throughout until the middle of the fragments developed photoreceptors. 5) A main way to see if the photoreceptors are fully generated would be how the planaria species interacts with light. The photoreceptors are highly sensitive to light. The eye of the planaria has pigment cells and photoreceptors. The pigment cells need to form a pigment eye cup while the photoreceptors are known to be located outside of the eye cup (Sakai et al. 2000) Depending on the conditions of the environment the photoreceptors or eyespots can be visible after only a few days after it has been decapitated (Sakai et al. 2000). However, since there is more mechanism involved it will take a bit longer after they are visible to be fully regenerated and sensitive to the light (Sakai et al. 2000). There needs to be a regeneration of microvilli and rhabdomeres that form after the pigment eye cup (Sakai et al. 2000). This is seen in the experiment as the eyes were visible in the middle fragment before it started having a high mobility rate. 6) The color of the regenerating tissue did not change too much from the visibility of the human eye through the microscope. However, When the middle fragment was looked at individually it is seemed to have a light grey trail. The different cell types, pigment cells and tissues that are regenerating are supposed to give the planarians a brownish color (Handberg-Thorsager et al. 2008). The neoblast are also supposed to have this color. In the lab it was mostly grey or black, but this error could be caused by the glare of the camera or lighting of the microscope. Dorsally from the top the pigmented epithelial tissue is characterized by its brown color (Handberg-Thorsager et al. 2008).

Figure 2.0 This figure shows the head about 4 days after the cut. The photoreceptors are still present.

Figure 2.1 This shows the middle of the planaria about 4 days after first cut. The excess brown material is no longer shown.

Figure 2.2 This shows the tail of the planaria about 4 days after first cut. There are no new developments at this stage.

Figure 2.3 This figure shows the head about week after initial cut. It has not grown too much but has slowed down in movement.

Figure 2.4 This figure shows the middle. A tail is now developing. It has some sign of photoreceptors. Neoblast is present.

Figure 2.5 This image shows the head at the final checkup. The head here is most likely dead. There was no movement seen.

Figure 2.6 This image shows the middle at the final checkup (2-week point). This shows how the middle-regenerated photoreceptors and a tail (neoblast).

Sources Adell, T., Cebrià, F., & Saló, E. (2010). Gradients in planarian regeneration and homeostasis. Cold Spring Harbor perspectives in biology, 2(1), a000505. Bely, A. E., & Nyberg, K. G. (2010). Evolution of animal regeneration: re-emergence of a field. Trends in ecology & evolution, 25(3), 161-170. Brøndsted, H. V. (1954). Size of fragment and rate of regeneration in planarians. Development, 2(1), 49-54.

Brown, H. M., Ito, H., & Ogden, T. E. (1968). Spectral sensitivity of the planarian ocellus. The Journal of general physiology, 51(2), 255-260. Handberg-Thorsager, M., Fernandez, E., & Salo, E. (2008). Stem cells and regeneration in planarians. Front Biosci, 13, 6374-6394. Nogi, T., & Levin, M. (2005). Characterization of innexin gene expression and functional roles of gap-junctional communication in planarian regeneration. Developmental biology, 287(2), 314-335. Pellettieri, J., Fitzgerald, P., Watanabe, S., Mancuso, J., Green, D. R., & Alvarado, A. S. (2010). Cell death and tissue remodeling in planarian regeneration. Developmental biology, 338(1), 76-85. Sakai, F., Agata, K., Orii, H., & Watanabe, K. (2000). Organization and Regeneration Ability of Spontaneous Supernumerary Eyes in Planarians—Eye Regeneration Field and Pathway Selection by Optic Nerves—. Zoological science, 17(3), 375-381....