A REVIEW OF NEUROBIOLOGICAL STUDIES ON

PLANARIANS

BY ALANA MOSKOWITZ

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INTRODUCTION

One of the defining features of humanity is the awareness of mortality; the knowledge that all

things inevitably come to an end. To many, it is a frightening notion. It is for that reason that mankind has

always longed for the unattainable ­ immorality.

Though humans are more aware of their mortality, there are other organisms much closer to

mastering it. These organisms are called biologically immortal. According to expert Leonard Hayflick,

“The most stringent definition of immortality is a life form capable of indefinite survival in conditions

where no changes have occurred in molecular composition from some arbitrary beginning.” Hayflick

goes on to state that “A more liberal definition of biological immortality would be the indefinite survival

of a life form whose vital life processes function indefinitely” (Hayflick, 2013).

Modern scientists are studying the different organisms that fall under this definition of

immortality, hoping to understand the mechanisms that enable these organisms to live the way they do.

For example, scientists have long been observing the regenerative abilities of hydra. However, it was not

until fairly recently that the presence of stem cells was identified as the cause (Bosch, 2008). Scientists

have also been studying Turritopsis nutricula, otherwise known a the “Immortal Jellyfish”, in an effort

to understand its ability to avoid aging. The jellyfish begins life as a polyp, but when it reaches maturity,

it reverts back to the polyp stage, therefore escaping the ailments of aging (Carla, et al. 2003). As

fascinating as these organisms are, one of the most well known biologically immortal organisms is the

planarian, which is commonly used to study stem cell biology, embryonic development, and as a model

for neurobiology (Gentile, Cebria, and Bartscherer, 2011).

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Planarians are free living flatworms, belonging to the class Turbellaria of the phylum

Platyhelminthes. They are capable of reproducing either sexually or asexually, depending on the specie.

They are very simple organisms, but still have many distinct tissues and organs (Newmark and

Sanchez­Alvarado, 2002). This makes them ideal models for experimentation.

PLANARIANS AS BIOLOGICALLY IMMORTAL ORGANISMS

Planarians are classified as biologically immortal organisms because of their ability to create

endless telomerase, an enzyme that regulates the length of telomeres. Telomeres trigger cell death and

aging at a certain point, which is referred to as the Hayflick limit (Harley, et al., 1992). As a telomere

becomes shorter, a cell becomes less able to regenerate. The cell will begin to show signs of aging, until

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it eventually dies. This does not occur in planarians, enabling them to regenerate indefinitely and

preventing any signs of aging (Tan, et al., 2012).

HISTORY OF REGENERATION

Regeneration has been documented for thousands of years. Among the earliest known examples

of regeneration are included in the writings of the ancient Greek philosopher, Empedocles. His works

were passed down from generation to generation, describing incredible animals with “necks that would

grow new heads” (Sanchez­Alvarado, 2010). These writings eventually reached Aristotle, some

hundred years later, who was the first to document his observations of lizards growing new tails. During

the Middle Ages, the topic of regeneration became popular among alchemists hoping to create

regeneration in humans. Since alchemy was entirely discredited following the Renaissance period, the

study of regeneration all but stopped until the 1600’s.

During the late 1600’s, scientist Melchisédech Thévenot presented a live lizard before the

French Academy of Sciences. To the members horror, he cut off its tale, and asked the members of the

academy to write down any observations. In the coming days, the tail regenerated (Sanchez­Alvarado,

2010). This restored interest in the study of regeneration. Soon after, Abraham Trembley began his

study of polyps. He found that the polyps he found could regenerate any missing tissue. He dubbed

these polyps “Hydra”, named for the ancient Greek monster capable of regenerating its head. Sylvia and

Howard Lenhoff' translated Trembley’s findings in their book "Hydra and the Birth of Experimental

Biology” (Sanchez­Alvarado, 2010). These early regeneration studies inspired and paved the way for

scientists in later years.

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PLANARIANS AS MODELS FOR REGENERATIVE STUDY

In 1766, Peter Simon Pallas noted that planarians were able to generate an entirely new body

from no more than a fragment of the head (Newmark and Sanchez­Alvarado, 2001). This marked the

first study of regeneration carried out using planarian flatworms. Pallas’ findings were confirmed by

French naturalist Jacques Philippe Draparnaud who found that planarians undergo asexual reproduction

through fission. Planarians will reproduce by simply ejecting part of their body in order to create an

identical organism (Newmark and Sanchez­Alvarado, 2001). In 1814, J.G. Dalyell conducted one of

the first experiments regarding planarian regeneration, declaring them to be “immortal under the

knife”(Dalyell, 1814).

In 1898, the “Father of Genetics”,

Thomas Hunt Morgan published his

article “Experimental studies of the

regeneration of Planaria maculata”. In

his article, he describes some of the

various experiments he performed in

testing the regenerative abilities of

planarian flatworms. He found that

planarians has a natural sense of

polarity, enabling them to identify which way was “up” during regeneration. In addition, he found that

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planarians could be divided in any direction and still be able to regenerate. Not only could they be

divided in multiple directions, but they could be divided into numerous pieces ­ 279, as Morgan

concluded (Morgan, 1898). The experiments conducted by T. H. Morgan inspired many scientists

during the turn of the century to experiment with planarians.

MECHANISMS OF REGENERATION

Scientists in the developmental biology field have been utilizing planarians flatworms as a way of

understanding the mechanisms of regeneration. This is because planarian flatworms contain pluripotent

stem cells called neoblasts throughout their body, which enable them to recreate any missing tissue

through the processes of epimorphosis and morphallaxis (Wagner, Irving, Wang, and Reddien, 2011).

During epimorphosis, undifferentiated cells, called neoblasts, form over a wound site. This mass

of cells is known as the blastema. Within the blastema, genetic information develops to meet the needs

of the wound ­ a new tail or part of the head may regenerate from the initial wound site. (Agata, Saito,

and Nakajima, 2007). Morphallaxis differs from epimorphosis in that morphallaxis deals with the

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regeneration of an entire new body. During morphallaxis, a mass of undifferentiated cells will develop

into new tissue to replace absent tissue and create a new body. Within these undifferentiated cells,

genetic information develops and is eventually dispersed and made proportionate, thus creating new

cells and tissues (Reddien and Sánchez Alvarado, 2004).

The determining factor in the type of regeneration that a planarian undergoes is dependent on

the type of wound. Based on the wound type, the planarian will respond with one of two responses. It

determines which type of regeneration through a two wave mitotic response. The neoblasts emit two

different mitotic peaks, signaling to the body what the appropriate response is. The body will then act on

these signals and regenerate cells as necessary (Wenemoser and Reddien, 2010).

Many scientists believe that in regeneration, the way that cells are able to maintain polarity and

differentiate is by receiving chemical signals through signal pathways. In this way, information can be

transported throughout the body, and the body is able to respond to injury. These pathway are called

hedgehog (Hh) pathways (Rink, 2009, Ingham, Nakano, and Seger, 2009, Gurley, et al. 2010).

It is the theory of many scientists the RNA is a factor in cell replication and regeneration. This

idea originated from the controversial studies of James V. McConnell. A scientist in McConnell’s lab

suggested that RNA is involved in memory retention, using that as an explanation for how memories

were preserved through head regeneration. To test this, the scientists treated a conditioned planarian

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flatworm with ribonuclease, an enzyme that breaks down RNA, and cut it in two. The progeny resulting

from the tail end had no memory; the progeny from the head portion however, did. It was suggested

that some memory is stored in the central nervous system, and some is stored in RNA. In an attempt at

memory transfer, McConnell made unconditioned planarians eat the ground up remains of conditioned

planarians (McConnell, 1962). This practice was seen as unethical, the experiments proved difficult if

not impossible to repeat, there were no listed citations, and the article was never peer­reviewed before

being released to the public. For these reasons, many scientists came to the conclusion that

McConnell’s findings were not to be taken seriously (Rilling, 1996).

Though McConnell’s works have been largely discredited, a few studies have shown a link

between RNA and regeneration. The article “Double­stranded RNA specifically disrupts gene

expression during planarian regeneration”, describes how dsRNA acts like a restriction enzyme,

preventing specific genes from being expressed during planarian regeneration. This shows how RNA

can be used to manipulate genes (Sanchez­Alvarado and Newmark, 1999). In addition, “Gene

Knockdown in Planarians Using RNA Interference” explains how by using RNAi, a type of RNA that

interferes with gene transcription, scientists have been able to study gene function during regeneration

(Oviedo, et al., 2007).

USE OF PLANARIANS IN BEHAVIORAL STUDIES

In the words of planarian experts, Philip A. Newmark and Alejandro Sanchez­Alvarado, “…

planarians have key anatomical features (mesoderm, central nervous system (CNS) and excretory

system) that might have been platforms for the evolution of the complex and highly organized tissues and

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organs found in higher organisms” (Newmark and Sanchez­Alvarado, 2002). In synaptic organization

and in many features of its neurons, the planarian brain is similar to the brain of a vertebrate organism.

The planarian central nervous system is typically compared to that of mice and flies, though it also shares

characteristics of the human brain. In fact, some scientists believe that the brain of the planarian flatworm

evolved into the human brain, as evidenced by the numerous homologues between human genes and

planarian genes, and the presence of enzymes in planarian brians found in human brains (Sarnat H.B.

and Netsky M.G., 1985).

The impact of drugs on the central nervous is currently being experimented with in regards to

planarians. In 2008, scientists at Temple University explored the addictiveness of cannabinoid, an

extract of cannabis, in comparison to that of cocaine and amphetamines. Ultimately, the scientists found

that abstinence­induced withdrawal occurred with cocaine and amphetamines, but not with cannabinoid.

This conclusion suggested that cannabinoid is less addictive than some other drugs (Raffa, et al., 2008).

Scientists have also been looking into reversing the impacts of harmful drugs by using planarians

as models. One scientist in particular, Oné Pagan, has reversed the unusual behavior exhibited by

planarians exposed to cocaine (Pagan, 2008). He has also stopped nicotine withdrawal in planarians

given tobacco. This novel research could help lessen the effects of withdrawal and reduce addiction in

cases of drug abuse in humans (Pagan, 2009). Additionally, in 2014, an article was published describing

scientists using Schmidtea mediterranea as a model to understand nervous system development in

infants suffering from fetal alcohol syndrome.

PLANARIAN MEMORY STUDIES

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One of the most well known planarian experts is James V. McConnell. The experiments carried

out by McConnell during the early 1960’s are still discussed and debated by the scientific community

today because of their controversial nature. McConnell’s experiments began by conditioning planarians

to a certain response, something that had previously been thought to be impossible to do in

invertebrates. He did this by training the flatworms to find their way through a maze, guided by electric

shocks. McConnell called this process “worm running”. To test just how complete the regeneration of

the central nervous system was in planarian flatworms, he cut off their heads and then put them through

the maze again. To his surprise, after a very short amount of time, the worms were able to find their way

through the maze, suggesting that memory was retained through regeneration. This led to a string of

experiments involving memory retention through regeneration. McConnell believed that if the tail of a

conditioned worm was able to generate a head complete with the old head’s memory, memory was not

stored solely in the central nervous system (McConnell, 1962).

McConnell’s work is currently being looked into by behavioral scientists. The article “An

automated training paradigm reveals long­term memory in planaria and its persistence through head

regeneration” by Michael Levin and Tal Shomrat. The Tufts University scientists trained planarians to

become familiar with a specific environment. This was done through a complicated procedure involving

planarians natural sensitivity to light. Once the planarians had been properly conditioned, the scientists

amputated their heads. The regenerated offspring was then put into the same environment, and the

scientists tested their familiarity with that environment. What they discovered is that some memory is in

fact retained through head regeneration. While this study did not support all of McConnell’s claims, it is

a huge step in the study of memory retention in planarian flatworms (Levin and Shomrat, 2013).

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USE OF PLANARIANS

TO TEST IMPACT OF CAFFEINE ON LONG TERM MEMORY IN A NEWLY

DEVELOPED CENTRAL NERVOUS SYSTEM

Studies have been conducted showing a direct correlation between caffeine and long term

memory. By inhibiting adenosine production, memories are retained for longer periods of time (Kopf, et

al., 1999). In the article “Caffeine in floral nectar enhances a pollinator’s memory of reward”, bees were

conditioned to respond to a specific odor. Bees given caffeine were able to remember this odor and

responded appropriately three times longer than those which were not (Wright, et al., 2013). In mice,

not only has it been shown to improve long term memory by inhibiting adenosine, but it has been

suggested that when caffeine is given to an adult mouse, it can prevent alzheimers (Arendash G.W. and

Cao C., 2010). While caffeine has been tested on planarian flatworms to study regeneration, its effect

on long term memory has not been explored (Collins, 2007).

SIGNIFICANCE

Stem cells show a lot of promise for the future of medical research. Scientists involved

in developmental biology are constantly exploring new ways to study stem cells. By using planarians,

scientists are able to study not only stem cells, but developing central nervous systems. This enables

scientists to observe how stem cells impact a central nervous system in embryonic stages and also in

regeneration, the latter of which is becoming increasingly important. Some scientists suggest that stem

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cells will be used to treat neurodegenerative diseases, such as alzheimers (Abdel­Salam, 2011). This

would involve the reconstruction of brain tissue, which could potentially be damaging to the patient. By

understanding the mechanisms of regeneration, long term memory retention, and the substances that

could improve memory, one would be able to potentially save many memories that could be lost in such

a procedure.

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