The Ice Bucket Challenge – How Did It Contribute to Scientific Research?

The Ice That Got the Gene

Remember the ice bucket challenge? Of summer 2014? Where what seemed like a lot of people all of a sudden had the urge to chuck ice cold water over themselves and film it? Well it turned out this wasn’t just another new crazy internet fad. The ice bucket challenge was started by Nancy Frates from the UK after her son was diagnosed with the condition ALS (amyotrophic lateral sclerosis); it was to gain awareness of the progressive neurodegenerative disease and raise money to fund new research. There were over 17million people who uploaded videos of their challenge onto social media sites including many celebrities such as Bill Gates and Mark Zuckerberg. A staggering £87.7M was raised for ALS, funding 6 new research projects! This has led to the scientific discovery of new genes associated with both the hereditary and sporadic causes of the disease that will lead to new therapeutic targets for potential drugs.

Project MinE

Project MinE largely funded by the ice bucket challenge is the largest ever study of inherited ALS. More than 80 researchers from over 11 different countries conducted searches for risk genes in ALS affected families. Bernard Muller and Robert Jan Suit, entrepreneurs from the Netherlands, were both diagnosed with ALS in 2010 and 2011. They made a decision to turn their business skills to finding a solution and so founded Project MinE. The project started with thousands of untested blood samples from ALS patients that were sat gathering dust in a Netherlands lab. Project MinE was chosen to be the recipient of the funds raised by the ice bucket challenge which enabled them to fund their project and commence the analysis of the blood samples. The researchers used arrays of common single nucleotide polymorphisms (SNPs) to genotype 15,156 ALS patients and 26,224 healthy controls from many different countries totalling more than 18 million SNPs tested. Some 1,861 had whole genome sequencing, which involves reading every single one of the six billion letters in the human genome (A, G, C, T).

What is a Single Nucleotide Polymorphism?

A nucleotide is a single building block of DNA. There are 4 building blocks of DNA: adenine (A), guanine (G), cytosine (C), thymine (T). A single nucleotide polymorphism is the most common form of variation in the genetic code and involves the change of one nucleotide. For example, in a certain segment of DNA a SNP may replace adenine with cytosine. These changes occur once in every 300 nucleotides throughout the genome averaging around 10 million SNPs. These nucleotide changes can affect a genes function, but most have no effect on a person’s health or development. SNPs can be used by scientists to discover information within the genome such as being a biological marker allowing scientists to locate genes associated with disease. They can be used to track disease inheritance, predict a person’s response to certain drugs, and susceptibility to toxins.

 

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New Risk Gene

A new risk gene now associated with and believed to be in amongst the most common genes to contribute to ALS is the NEK1 gene. This gene encodes for the… wait for it, it’s a bit of a mouthful: serine/threonine kinase NIMA (never in mitosis gene A)- related kinase. The NEK1 gene has many diverse functions within a cell such as mitosis (production of identical daughter cells from cell division), microtubule stability, internal transportation of proteins, formation of primary cilia that sense mechanical and chemical stimuli, regulation of the permeability of the mitochondrial membrane, and assists with DNA repair. Disruption of these cellular functions is linked to the onset of ALS. The role of NEK1 in ALS is not fully understood yet; according to Dr Lucie Bruijn from the ALS Association, they are unsure whether it is the NEK1 gene itself that is connected to ALS development or mutations within the gene, with another possibility being a link between this gene combined with mutations in another gene. Including the NEK1 gene there is a total of 7 genes recently found in association with ALS which are TBK1, CCNF, GLE1, MATR3, TUBA4A, CHCHD10. This discovery has provided researchers with a new target for therapy development and a new avenue of research to look down to gain a better understanding of what causes the disease. So to all of you who participated – bravo!

Author: Laura Ellis

Editor: Molly Campbell

References

Bettencourt, C. and Houlden, H. (2015). Exome sequencing uncovers hidden pathways in familial and sporadic ALS. Nature Neuroscience, 18(5), pp.611-613.

Cirulli, E., Lasseigne, B., Petrovski, S., Sapp, P., Dion, P., Leblond, C., Couthouis, J., Lu, Y., Wang, Q., Krueger, B., Ren, Z., Keebler, J., Han, Y., Levy, S., Boone, B., Wimbish, J., Waite, L., Jones, A., Carulli, J., Day-Williams, A., Staropoli, J., Xin, W., Chesi, A., Raphael, A., McKenna-Yasek, D., Cady, J., Vianney de Jong, J., Kenna, K., Smith, B., Topp, S., Miller, J., Gkazi, A., Al-Chalabi, A., van den Berg, L., Veldink, J., Silani, V., Ticozzi, N., Shaw, C., Baloh, R., Appel, S., Simpson, E., Lagier-Tourenne, C., Pulst, S., Gibson, S., Trojanowski, J., Elman, L., McCluskey, L., Grossman, M., Shneider, N., Chung, W., Ravits, J., Glass, J., Sims, K., Van Deerlin, V., Maniatis, T., Hayes, S., Ordureau, A., Swarup, S., Landers, J., Baas, F., Allen, A., Bedlack, R., Harper, J., Gitler, A., Rouleau, G., Brown, R., Harms, M., Cooper, G., Harris, T., Myers, R. and Goldstein, D. (2015). Exome sequencing in amyotrophic lateral sclerosis identifies risk genes and pathways. Science, 347(6229), pp.1436-1441.

ALSA.org. (2016). Homepage – ALS Association. [online] Available at: http://www.alsa.org/ [Accessed 19 Sep. 2016].

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Neuroanatomical Mechanisms Involved in Cue Induced Relapse to Alcohol

Drug addiction can be defined as a chronically relapsing disorder due to the difficulty for addicts to maintain abstinent from a particular drug (Koob and Volkow 2010). This is due to an unconscious mechanism within the brain that drives people to seek for a particular drug even when they intend to remain abstinent. In this article I am going to be looking at cue-induced relapse to alcohol.

Alcohol addiction is a major problem in the UK, costing the NHS £3.5billion per year. 9% of adult men and 4% of adult woman in the UK are dependent on alcohol, but only 1% of alcoholics actually seek treatment (statistics on alcohol). Alcohol also causes 4.6% of global disease and injury (Ryan et al, 2013). The current treatments for alcoholism prescribed by the NHS are aimed at preventing relapse or limiting the effects of alcohol. Some examples are naltrexone and acamprosate (NHS). These treatments, however, are unsatisfactory as there are still high relapse rates, in addition to compliance issues (Ryan et al, 2013).

To understand why the treatment of alcoholism is so difficult, the behaviour of an alcohol addicted individual compared to that of a non-addicted drinker needs to be understood. This difference highlights how the seeking of alcohol becomes an unconscious and uncontrollable drive. Addiction occurs once there is a shift from drinking being a goal directed behaviour (going out for a drink with a friend on a night out), to an automatic habitual behaviour (drinking every day). When people start drinking they are seeking positive reinforcement (this is a goal directed behaviour) and have control over their decision to drink alcohol. On the other hand habit-seeking behaviours are not due to the expectation of a positive reward, but are instead driven by the memory of previous, reinforcing, alcohol associated history; thus the action of seeking alcohol becomes automatic and involuntary. This is where environmental cues start to come into play. Cues such as the sight of a pub trigger memories of drinking, causing craving symptoms due to the establishment of habitual pathways in the brain. Therefore, addicts will take a drink when these cues are presented. The other sign that drinking has become habitual is that immediate negative effects associated with drinking have no effect on preventing alcohol seeking behaviours. The brain region involved in mediating these two behaviours is the striatum. Goal-seeking behaviour is mediated by the dorsomedial striatal circuitry whereas habitual control takes place in the dorsolateral striatum, and evidence shows that there is a shift of control from the dorsomedial to the dorsolateral striatum as habitual learning takes place and addiction occurs (Corbit and Janak, 2016).

Relapse is an enduring problem, even once withdrawal symptoms have disappeared and alcohol addicted patients have undergone long periods of abstinence. Although there are many factors involved in relapse (such as stress) a major problem for alcoholics is the constant exposure to alcohol and alcohol related cues, due to the legal nature of alcohol and its wide availability. Some of the main areas in the brain that have been shown to be involved with cue-induced relapse to alcohol-seeking are the medial prefrontal cortex (mPFC), the nucleus accumbens (NAc) core and shell, the ventral tegmental area (VTA), the basolateral and central amygdala and the cornu ammon regions of the hippocampus. Two major neurotransmitters have been implicated in signalling to these brain regions and triggering relapse due to the presentation of an alcohol related cue. These are Orexin-A and relaxin-3.

There are two forms of orexin: orexin A and orexin B, which bind to the receptors orexin-1 (OX1) and orexin-2 (OX2). Orexin A and its cognate receptor OX1 have been identified as being involved in the act of alcohol seeking. Orexins are hypothalamic neuropeptides and their neurones originate in the lateral, dorsomedial and perifornical areas of the hypothalamus. Orexins play a role in many autonomic functions such as feeding and arousal, as well as being implicated in the reward system. Their axons project from the hypothalamus to the mesocortical limbic system.

One experiment (Brown et al) looked at the involvement of orexin A innervation of the prelimbic cortex and the VTA and its involvement in alcohol-seeking behaviour in rats due to reinstatement of alcohol linked triggers. The prelimbic cortex is a brain region located in the prefrontal cortex and has been linked to alcohol seeking behaviour. The VTA is a major brain region involved in the dopaminergic reward pathway (mesolimbic pathway) and also has a role in alcohol-seeking behaviour. For this experiment rats where trained to administer an ethanol containing solution by pressing a lever. The cues present that were to become associated with the reward of alcohol included a light placed above the lever and vanilla essence scent. The rats then underwent extinction of alcohol (they were put in the chamber without alcohol being delivered when the lever was pressed and no cues where present) and then after a period of extinction the cues where reinstated and the rats where either given a vehicle or an OX1 inhibitor (SB-334867) to see whether by preventing orexin signalling there would be a decrease in the tendency of rats to relapse due to cue-induced reinstatement of alcohol. Selective inhibition of either the VTA or the prelimbic cortex both significantly reduced alcohol reinstatement in rats.

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Figure 1a depicts a significant decrease in alcohol seeking in rats when the SB3 antagonist was applied to the VTA compared to when rats where only given the vehicle. 1b also shows a significant decreased in lever presses when SB3 was injected into the prelimbic cortex.

Orexin neurones project from the lateral hypothalamus to the prelimbic cortex. These neurones are involved in sending appetitive signals to the prelimbic cortex, triggering alcohol-seeking behaviour. The role of the orexin system in the VTA is more centred on reward value, but the results clearly show that this pathway is also implicated in cue-mediated reinstatement of alcohol seeking. Jupp et al also investigated orexin mediated alcohol seeking by examining Fos expression. They also applied the OX1 inhibitor SB-334867, but at two stages, either during immediate reinstatement to alcohol after a period of extinction or after 5 months or extinction. They found that Fos expression increased (when presented with alcohol related cues) in the infra-limbic cortex, the prelimbic cortex, orbitofrontal and piriform cortices, NAc core and shell, basolateral and central amygdala, lateral and dorsomedial hypothalamus and the BNST. SB-3 inhibited Fos expression showing that orexin signalling circuitry is involved in cue-reinstatement, but depending on the time of relapse they found a change in the amount of Fos inhibition, indicating that the orexin circuitry involved in reinstatement may change over time. In immediate reinstatement the brain areas involved are the orbitofrontal and prelimbic cortex and the accumbens core but after protracted abstinence there is a shift to the cortical locus.

The second neurotransmitter involved in cue-reinstatement of alcohol is Relaxin-3. Relaxin-3 is a highly conserved neuropeptide, throughout species, and is the true ancestor of the relaxin peptide family. Its cognate receptor is relaxin peptide 3 receptor (RXFP3). Relaxin-3 networks have been linked with arousal functions such as stress, feeding, sleep/wake states and motivation and reward. Relaxin-3 is expressed in GABAergic neurones in the hindbrain nucleus incertus. These neurones then project to the forebrain areas: the amygdala, bed nucleus of the stria terminalis (BNST), hippocampus and the lateral hypothalamus. The previous experiment was repeated by Ryan and team, however the rats were injected with either an antagonist for RXFP3 called R3(B1-22)R or a vehicle. Rats that were injected with R3(B1-22)R had significantly reduced lever presses then rats injected with the vehicle.

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Figure 2a demonstrates that application of the R3(B1-22)R antagonist significantly reduced lever presses when alcohol cues where reinstated.

These experiments demonstrate the importance of orexin and relaxin-3 in drug seeking behaviors and cue induced triggers. These experiments also highlight the importance of understanding these complicated systems so that new pharmaceuticals can be developed to help prevent alcoholics from relapsing again and to make their attempts to remain abstinent easier. Further research into the involvement of these two neurotransmitters and their involvement in relapse is crucial so that our understanding of alcoholism can develop, allowing scientist to help alcoholics, and the treatment they receive, improve.

Author: Lara Cornish 

References: 

 

Alcohol concern (2016). Statistics on alcohol. Available at https://www.alcoholconcern.org.uk/help-and-advice/statistics-on-alcohol/. Accessed on the 5th of September.

Brown, R. M., Kim, A. K., Khoo, S. Y. S., Kim, J. H., Jupp, B. & Lawrence, A. J. (2016) Orexin-1 receptor signalling in the prelimbic cortex and ventral tegmental area regulates cue-induced reinstatement of ethanol-seeking in iP rats. Addiction Biology, 21(3), 603-612.

Corbit, L. H. & Janak, P. H. (2016) Habitual Alcohol Seeking: Neural Bases and Possible Relations to Alcohol Use Disorders. Alcoholism-Clinical and Experimental Research, 40(7), 1380-1389.

Jupp, B., Krstew, E., Dezsi, G. & Lawrence, A. J. (2011) Discrete cue-conditioned alcohol-seeking after protracted abstinence: pattern of neural activation and involvement of orexin(1) receptors. British Journal of Pharmacology, 162(4), 880-889.

Koob, G. F. & Volkow, N. D. (2010) Neurocircuitry of Addiction (vol 35, pg 217, 2010). Neuropsychopharmacology, 35(4), 1051-1051.

NHS choices (2015) alcohol misuse – treatment. Available at http://www.nhs.uk/Conditions/Alcohol-misuse/Pages/Treatment.aspx. Accessed on the 5th of Spetember 2016.

Ryan, P. J., Kastman, H. E., Krstew, E. V., Rosengren, K. J., Hossain, M. A., Churilov, L., Wade, J. D., Gundlach, A. L. & Lawrence, A. J. (2013) Relaxin-3/RXFP3 system regulates alcohol-seeking. Proceedings of the National Academy of Sciences of the United States of America, 110(51), 20789-20794.

 

 

 

Brexit – How Will It Impact Scientific Research?

The UK’s current membership in the EU has benefited scientific research through numerous avenues. A key factor is the links in the scientific community and openness which comes with collaboration between international labs that leads to breakthroughs in research faster. The idea behind this, is you can use research published by other research centres, collaborate research projects between facilities, linking equipment plus the minds of those that lead the relevant fields. The European Union (EU) allows free collaboration between research projects and the scientific community, including the Horizon 2020 scheme detailed below.

Currently students and researchers have the possibility to study and work abroad within the EU. This free movement allows the development of individuals, projects, and labs across Europe. It leads to more competition for university places, PhD placements and jobs. Higher skilled applicants leads to competition; competition in recruitment helps to maintain standards required for research to be conducted and accepted. Not only does the movement of students and workers lead to better science, but it also improves opportunities for students. Those looking to study abroad and work in international labs will gain valuable experience, enhancing their CVs, and leading to higher employment levels for UK graduates. These opportunities also encourage students to follow further education, PhDs and masters, leading to a more educated society.

For many projects, funding is dependent on schemes run by the EU. A current EU funding programme is known as Horizon 2020. It aims to provide financial and political backing to innovative projects across the EU with the aim to make funding more available and “remove the red tape” that was previously in place to progress important research. This enables projects to start quicker, and also create a more united community for research. When the UK leaves the EU then we will no longer have easy access to this network of information and funding.

Current projects which are funded through the EU include the TRANSEURO consortium which is looking to advance stem cell research into Parkinson’s disease; the European Prevention of Alzheimer’s disease consortium; the Oxford Project to Investigate Memory and Aging, the Cognitive and Aging study, the Genetic FTD initiative and the Imaging of Neuroinflammation in Neurodegenerative Diseases consortium rely on EU collaborations. These crucial projects will suffer with lower levels of funding and a lack of collaboration, and breakthroughs will take longer to reach unless funding can be found elsewhere.

If Article 50 is triggered and the UK completes the move to leave the EU, steps must be taken to ensure that science remains international for the sake of progress. The Leave campaign argued that leaving the EU will not lead to a breakdown of the scientific community currently in place, but will in fact open doors to a wider international field. It must be considered that it already was possible for these opportunities outside the EU prior to Brexit, however many projects chose collaborate in the EU. Whether negotiations will allow the UK to remain in the EU Horizon 2020 group, or give channels of communication and work for scientists remains to be seen.

The leave campaign did propose a points based immigration system which will still allow highly skilled workers such as scientific researchers to work in the UK but Brexit may also lead to labs in the UK to look further afield for research and collaborate with other projects outside Europe. This may alter the number or destination of students or researchers who choose to study/work abroad, with currently 20% of academic staff at the top universities in the UK originating from the EU (Nature 2016).

The future for science after Brexit remains uncertain. Will Brexit encourage collaborations and open wider channels of communication globally which will benefit science? The leave campaign did promise that funding for scientists and students would continue but will that promise will be held to levels equivalent to the past? Only time will tell. All we hope as scientists is that this move will not give rise to too large a challenges so that science can continue to grow, and breakthroughs be made.

 

Author: Rosie Porter

Sources:

Nature (2016) http://www.nature.com/news/uk-scientists-in-limbo-after-brexit-shock-1.20178

http://www.nature.com/news/researchers-reeling-as-uk-votes-to-leave-eu-1.20153

Neuroscience News (2016) http://neurosciencenews.com/neurodegenerative-disease-research-brexit-4653/

European Commission Horizon 2020 (2016) https://ec.europa.eu/programmes/horizon2020/en/what-horizon-2020

Scientific American (2016) http://www.scientificamerican.com/article/why-the-science-community-says-no-to-brexit/