Squids as a Potentially Useful Model for Studying Neurodegeneration and Dementia in Mammals
This blog is based off of the paper “Squid (Loligo Pealei) Giant Fiber System: A Model for studying Neurodegeneration and Dementia?” by Philip Grant, Yali Zheng, and Harish C. Pant.
The pathology that causes many neurodegenerative disorders is fairly well known and understood. It is known that the disorders that result in memory loss and dementia are likely caused by an accumulation of proteins in the forms of extracellular plaques and intracellular filamentous tangles, which contain hyperphosphorylated cytoskeletal proteins. Scientists believe that this hyperphosphorylation happens as a result of deregulation of a normal pattern of topographic phosphorylation. It is believed that this is essentially a shift of the cytoskeletal protein phosphorylation, which normally occurs within the axonal compartment, but shifts to the cell bodies (figure 1).
Implications of the study
As neurodegenerative disorders are very prevalent in society, this topic is huge in the research industry right now. To put the wide spreadedness of this disease into perspective, according to the Alzheimer’s association, in 2010 alone, the global cost of Alzheimers was $604 billion, which is 1% of global GDP, and about $200 billion more than the annual revenue of Wal-Mart or Exxon Mobile. In addition to their larger neuronal anatomy, cephalopods have been used for years in the neurobiology field. Their brains support both learning and memory, and can be trained to identify visual cues. Although to this day there have been no documented cases of neurodegenerative diseases in squids, the normal nerve function and motor systems of these animals can still serve as a successful model for the research of the pathology behind the neurodegenerative disorders.
What kind of studies are scientists doing in this field? And how do squids play a role?
The current studies in this field are focusing on factors regulating compartment specific patterns of the cytoskeletal protein phosphorylation, mostly of the neurofilaments. The proteins inclusions include tau, neurofilatments, synuclein, and huntingtin. These accumulations could be at fault for the neuronal cell death, a shared common feature amongst many neurodegenerative disorders (Goedert 1999), although their rise and specific function in the cell death is not yet clear. This will allow researchers to establish a baseline from which to base further studies.
The ability to fully understand the phosphorylation mechanisms could potentially allow researchers to make more progress in the development of treatments for a multitude of neurological disorders. Unfortunately, the neurons of mammals are far too small for researchers to be able to separate the cell body and axon compartments, therefore this is where the giant axons and nervous systems of squids come into use. The stellate ganglion of the giant squid (figure 2) is very close neurologically to the mammalian systems, and scientists can easily separate the cell body and the axon compartments, and can do so in amounts that are conducive to study. In Grant et al., researchers use this portion of the squid’s anatomy to create a model to use for the study of neurodegeneration and dementia in mammals.
To study the factors which regulate the topographic phosphorylation of NF proteins, scientists extracted axoplasm from the axons, as well as giant fiber lobe cell bodies. Axoplasm is the cytoplasm that is inside the axon component of a neuron. The axoplasm had not been contaminated by myelin sheath or glial cells. These two tissue samples were then analyzed by comparison of their kinase assays to help to determine what differences exist in the compartment activities. The NF proteins consist of three subunits, called NF-220, NF-60, and NF-70.
Results of the Study
NF-220 was only found in the axon, where as NF-60 and NF-70 were found in both compartments, as can be seen in the Western Blot analysis in figure 3 part a. In addition, it was determined that the higher molecular weight proteins are only phosphorylated in the axoplasm, as can be seen in the right of figure 3a.
In addition, findings demonstrated that there may not be the same set of kinases in both the axon and giant fiber lobe (GFL), since the western blot shows that they higher weight molecular proteins are phosphorylated in the cytoplasm. Interestingly enough, it was also determined that the substrates histone and casein (figure 3b) were phosphorylated in the GFL, which equivocates the lack of phosphorylation of the proteins shown by the previous western blot. Therefore it can be concluded that although the kinases aren’t identical in the compartments, there is roughly equal kinase activity in both cell compartments (table 1).
The researchers in Grant et al., have proposed a model for the regulation of the sites of the neurons that are thought to be degraded during these disorders. This model was created by researchers to further study the sites of deregulation that may lead to the diseases that exhibit neurodegeneration, dementia, and neuronal cell death.
Strengths and Weaknesses
Researchers are still unable to answer whether abnormal hyperphosphorylation of the NF within the cell body is the reason for the formation of aggregate proteins, or whether the abnormally phosphorylated proteins are a result of the conformational changes in the folding of the substrates that are present in the cell bodies, as discussed by Kosik and Shimura (2005).
Another weakness of this study is the fact that transcription and translation regulation of NF gene expression are not dealt with in the model, the studies focus on post-translational processing, MF assembly, and transport from the cell body compartment to the axon. The model simplifies this and assumes all kinases, regulators, and substrates are synthesized in the cell bodies of the GFL.
In addition, in Rishal et al., a newly developed method of extracting mammalian axoplasm is described. It would be interesting for the researchers of Grant et al., to repeat this study using the method described by Rishal et al., which is based on the incubation of separated short segements of nerve fascicles in a medium to help separate and purify the axon components. Next, the axoplasm is extracted. The extracted axoplasm is free of extra cellular contents such as serum and glial cells, so that these cells can be accurately analyzed without any contaminants.
Although the authors of Grant et al. still aren’t exactly 100% sure what the origin of the abnormally phosphorylated proteins that give rise to neurodegenerative disorders, they were able to come up with some answers as to the composition and phosphorylation activity of the compartments of the neurons. Further studies should be completed in order to successfully determine the origin of these proteins, so that scientist can understand how to either prevent and or treat these accumulations, leading to more answers for the treatment or cure of these diseases.