‘Tis the season to be jolly

Sunday 18 December 2022

Last night I saw a fabulous performance of Handel’s Messiah in the beautiful 16^th century Scuola Grande dei Carmini, and this morning I woke up to blue sky and radiant sunshine in Venice. What’s not to like?

After a couple of weeks of cold, rain, fog and empty backstreets, Venice has suddenly come alive in the run up to Christmas. A funfair has appeared near our apartment, the Christmas tree is sparkling in Piazza San Marco, magical decorations adorn the shopping streets, and the bars and restaurants are now buzzing with banter and laughter. Every morning I go for a short, gentle jog along the waterfront with sights such as the Basilica di Santa Maria della Salute, Palazzo Ducale (Doge’s Palace) and Campanile (bell tower) laid out majestically in front of me. I pinch myself that I am in this amazing place.

As a bonus, my daughter Rosa came out to visit for a couple of days last week. She is taking a year out before going to university to study theology and philosophy, and has a job in the local Paperchase shop to get some experience – and money – under her belt before doing some travelling in the spring. Needless to say, we had a fantastic time exploring Venice together, and enjoying some great Italian cuisine.

Clare comes out again next week to join me for Christmas and New Year before I return to normal life at the end of the first week of January. I had mixed feelings about trying to live in a foreign city on my own for a few weeks, but it’s turned out to be a great experience. Although I’ve all but given up trying to learn Italian, with my brain seemingly unable to retain the simplest vocabulary, I’ve managed to survive in a foreign country without any major challenges. Shopping can be a bit tricky as can doing simple things in the kitchen like opening bottles and jars. Food packaging in general is a bitch. But I take my time and find creative ways to do things. And I pump myself full of medication every day.

So for now, la vita è bella. Life is good. It won’t always be like this of course. But the positive news is that listening to great music and enjoying the warmth of the sun on my face are two things I will still be able to do even when I am in the advanced stages of Parkinson’s.

Return to fabulous Venice

Sunday 20 November 2022

There are few places in the world more wonderful than Venice on a sunny autumn afternoon. 

As I wander through the ancient streets and piazzas, across countless small bridges, I marvel at the timeless grandeur set against crisp azure skies. As I turn each corner, something unexpected greets me: an idyllic canal, a magnificent church, a crumbling palatial facade, a burst of sunlight.

Although Venice is an exquisite museum piece, taking one back to the golden era of the great city states of The Renaissance, it is also a living, vibrant hub of tourism and commerce. I gawp at the luxuries gleaming in the windows of Gucci, Prada, Ferragamo, Versace, Cartier and Louis Vuitton; I marvel at the dazzling displays of Murano glass; my mouth waters at all the tempting culinary offerings; I study the modern art sculptures that are dotted around as part of the Biennale festival; and I imagine wearing one of the Venetian masks I see hanging outside the souvenir shops during the Carnival. Everywhere I go there is the mellifluous chatter of Italians, all wearing designer sunglasses, all stylishly attired and all, like me, enjoying life on a fabulous day in Venice.

By the time I return to our rented apartment by the Arsenale, the old shipbuilding area of the city, I am hobbling with cramp and dystonia in my feet, despite periodically popping the grey and blue pills that I carry everywhere. This reminds me that the reason I am here is to do precisely that: take in as much enjoyment as possible from life until my body stops me.

I will be here for the next seven weeks, with Clare periodically joining me. We have rented a (surprisingly cheap) apartment as an experiment to see if living in a different European country for a couple of months could be an option for the next two or three years. I can do my work anywhere that has reliable WiFi so why not do it from Venice, or Rome, or Seville, or the French Riviera? Of course it may not work. I may struggle too much on my own or get lonely or bored, especially when the weather is cold and damp.

But today I don’t concern myself with the future. I live in the present and squeeze as much enjoyment out of the day as I can.

Time for an espresso. And some chocolate.

Animals in Parkinson's research

Saturday 22 October 2022 

Continuing with the theme of animal models, here is an essay I wrote as part of my MSc neuroscience about the use of animals in Parkinson’s research. Many people, myself included, feel uncomfortable about the use of animals in brain research, an issue for which there are no easy answers. There is quite a bit of technical terminology in here, but you can easily skim over the fancy words without losing the overall meaning.

 

Describe and discuss the relevance of animal models in Parkinson’s disease

Introduction

When James Parkinson published the first description of the medical condition that now bears his name, he speculated that a cure would “ere long be discovered” (Parkinson 1817). He was overly optimistic: over 200 years later there is no clinically proven way to halt or reverse the progression of Parkinson’s disease (PD) (Vijiaratnam et al. 2021). There was, however, a major breakthrough in 1957 when Swedish neuropharmacologist Arvid Carlsson performed a seminal experiment first on mice and then on rabbits. He administered reserpine to the animals which reduced their dopamine levels, effectively tranquilising them, then demonstrated that treatment with levodopa (L-dopa) could rapidly reverse the effect (Carlsson et al. 1957). Today, as a direct result of Carlsson’s animal experiments, levodopa remains the “gold standard” drug to treat symptoms of PD, and is used by many of the estimated 10 million people in the world with the condition (Schapira et al. 2009). 

This essay first surveys the animal models used for PD research then explores their role across various therapeutic strategies, including pharmaceutical development, deep brain stimulation and neuronal regeneration. But, given that over 60 years since the discovery of levodopa, there are still no disease-modifying therapies generally available for PD, this essay also addresses the question of whether too much reliance has been placed on animal models in recent decades and what their role should be in the future.

Animal models used in PD research

In the context of a neurodegenerative condition such as PD, the purpose of an animal model is to replicate some aspect of the condition in a simplified way that can be used to elucidate its pathophysiology and/or develop potential therapies. Given that the aetiopathogenesis of PD is not yet fully understood, a number of different mechanisms have been exploited for the development of animal models. These fall into four categories (pharmacological depletion of dopamine, neurotoxins, oxidative stress and genetic factors) and are summarised in Table 1. The corresponding roles these play in selected postulated mechanisms of PD are illustrated in Figure 1.

 

Table 1. Summary of animal models commonly used in PD research (adapted from Betarbet et al. 2002, and Konnova & Swanberg 2018).

Primary target mechanism	Model	Main symptoms induced	Comments
Pharmacological dopamine depletion	Reserpine	Akinesia, catalepsy
Environmental toxin	MPTP	Akinesia, rigidity	Commonly used model
	Rotenone	Akinesia, rigidity, tremor	Pesticides and herbicides; human toxicity debated
	Paraquat / Maneb	Reduced motor activity
Oxidative stress	6-OHDA	Akinesia	Commonly used model
	Methamphetamine	No clear PD symptoms
Genetic risk factors	PINK1, Parkin etc.	Various

Figure 1. Selected postulated pathogenic mechanisms in PD and associated models

(adapted from Przedborski 2017 and Dawson et al. 2010)

 

In terms of species used, 85% of PD experiments using animal models are performed on mice and rats (Figure 2). These are the laboratory animals of choice because they are easy to breed and manipulate, are relatively inexpensive, and because degeneration of nigrostriatal dopaminergic neurons leads to impaired motor function mirroring the effect PD has on motor function in humans (Konnova & Swanberg 2018).

 

Figure 2. Proportion of species used for animal models of PD in 23,000 research articles from January 1990 to June 2018 (reproduced from Konnova & Swanberg 2018).

 

One of the most commonly used PD models in recent decades has been the administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). This was originally synthesised as a type of heroin which caused severe Parkinsonism in a number of narcotics users in California in 1982. A researcher described the effect as like a textbook case of advanced PD, saying:

“Patients exhibited virtually all of the motor features of typical Parkinson’s disease, including tremor and asymmetry of findings. They even exhibited non-motor aspects of the disease such as facial seborrhea and mild deficits in higher cognitive function. Furthermore, these patients all had a dramatic and near immediate response to L-dopa.” (Langston 2017).

Further investigation revealed that MPTP was converted to MPP+ (1-methyl-4-phenylpyridinium) in the brain which is toxic to mitochondria (Langston et al. 1984). Around the same time, it was demonstrated that MPTP had the same effect on monkeys (Burns et al. 1984), causing a rapid depletion of dopaminergic neurons in the substantia nigra pars compacta whilst leaving the ventral tegmental area largely unaffected. Before the end of 1984, the effect had also been demonstrated in mice (Heikkila et al 1984), and thus MPTP was established as an apparently realistic model for PD and has been widely used since.

6-Hydroxydopamine (6-OHDA) causes oxidative stress but does not cross the blood-brain barrier so has to be injected directly. When administered to the substantia nigra, dopaminergic neurons rapidly degenerate; and when administered to the striatum it produces a much slower degeneration of the nigrostriatal system (Betarbet et al. 2002). The 6-OHDA model does not mimic the clinical features of PD as closely as MPTP but is nevertheless also widely used.

A number of pesticides, herbicides and fungicides are now generally believed to be risk factors for PD (Pezzoli & Cereda 2013) and in particular the herbicides paraquat and maneb, and the pesticide rotenone, have been used in rodent models of PD. However, whilst they have been shown to induce clear PD symptoms in rodent models, the level of toxicity in humans is debated (Blandini & Armentero 2012).

Reserpine and methamphetamine both cause dopamine depletion with little impact on the cell bodies (Betarbet et al. 2002) though the mechanisms are different, with methamphetamine believed to cause oxidative stress whereas reserpine interferes with action of dopamine transporters (Yaffe et al. 2018).

Finally, there are genetic models. As of 2019, Approximately 90 mutations, many of them single nucleotide polymorphisms, had been identified as PD risk factors in humans (Nalls et al. 2019). Of these a number have been replicated in animal models (Table 2), though their ability to replicate the dopaminergic neurodegeneration or clinical features of PD is limited (Blandini & Armentero 2012).

Table 2. PD mutations that have been used in animal models (reproduced from Konnova & Swanberg 2018).

Locus	Gene	Mutation	Inheritance
PARK1	SNCA	A30P, A53T, E46K	Dominant
PARK2	Parkin	Various	Recessive
PARK4	SNCA	Duplication and triplication	Dominant
PARK5	UCH-L1	I93M, S18Y	Dominant
PARK6	PINK1	G309D, exonic deletions	Recessive
PARK7	DJ-1	L166P, homozygous exon deletion	Recessive
PARK8	LRRK2	G2109S, R1441C/G	Dominant

 

Applications of PD animal models

The models described above have been used for the pre-clinical testing of a large number of candidate drug treatments for PD. This includes dopamine agonists, of which pramipexole is a recent example having been licensed by the US Food and Drug Administration[1] and European Medicines Agency[2] in 1997. This had been tested on a variety of models, including the MPTP model on monkeys, the 6-ODHA model on rats (Dooley & Markham 1998), the MPTP model on mice (Kitamura et al. 1997) and the methamphetamine model on mice (Hall et al. 1996).

Animal models are used to test theories about PD pathogenesis. For instance, experiments transplanting gut microbiota from humans with PD to mice resulting in parkinsonian motor impairment, lend support to the recent idea that PD may, in some cases, start in the gut (Sampson et al. 2016, Liddle 2018).

In addition to direct models of PD, animals have also been key to the understanding of brain structures compromised in PD, notably the basal ganglia, and this led serendipitously to the discovery and clinical development of deep brain stimulation (DBS), an increasingly common surgical procedure to alleviate motor symptoms in the more advanced stages of PD. In the 1970s and 1980s, a series of experiments on monkeys revealed the physiology of the basal ganglia-thalamocortical (BGTC) circuitry (Vitek & Johnson 2019), in particular the role of the so-called direct and indirect pathways in controlling motor function. This led to the use for a few years from the mid-1980s of pallidotomy as a treatment for PD (Laitinen et al. 1992). Although successful in some patients in relieving the motor symptoms of PD, there were frequent complications including cognitive changes, and in other cases, PD symptoms were either not improved or worsened (Vitek & Johnson 2019). However, the use of electric stimulation during these procedures as a testing mechanism led to the idea that the application of small electric currents in certain areas of the BGTC circuit via surgically implanted electrodes might directly alleviate PD motor symptoms. This resulted in DBS, a procedure that gives “significant improvement in motor function in patients with Parkinson’s disease where the condition cannot further be improved with medical therapy” (Obeso et al. 2001). Exactly how DBS works is not fully known, and monkey models continue to be used to improve understanding (e.g., Hashimoto et al. 2003).

Finally, animals can also play a role in neuronal regeneration therapies, for example in grafting dopaminergic cells into the striatum of a patient with PD (cells are typically grafted into the striatum where the dopamine is actually used rather than the substantia nigra pars compacta). In this case, animal models can be used to test ideas such as implantation of induced pluripotent stem cells (Schweitzer et al. 2020) or in vivo cell reprogramming (Heinrich et al. 2015) prior to human experimentation. Animals can also be sources of cells for xenotransplantation given the ethical and practical challenges with using human embryonic stem cells. For example, this has been tried with some success with transplantation of neural cells from pigs into humans (Deacon et al. 1997).

 

Discussion: are animal models still relevant?

Over recent years there have been numerous candidate pharmaceuticals for PD that have shown promise in animal models only to fail to demonstrate efficacy in clinical trials in humans. Examples of recent clinical trial failures following success in mouse models include nilotinib (Simuni et al. 2021, Tanebe et al. 2014) and venglustat (Veil et al. 2021). As of mid-2021, there were at least 44 drugs in Phase 1 or Phase 2 trials (Prasad & Hung 2021), many with pre-clinical testing on rodent models of PD – will all of these also fail to meet their primary outcome targets?

Taking the MPTP model as an example, in hindsight it is clear that a drug that rapidly causes the death of dopaminergic neurons in the substantia nigra is different from a disease that results in gradual degeneration over decades and over many regions of the brain (Epsay & Stecher 2020, Vijiaratnam et al. 2021). The reality is that animal models of PD, particularly in rodents, do not fully mirror the condition in humans, where neurodegeneration can be found not just in the dopaminergic system but also in the noradrenergic, serotonergic and cholinergic systems in addition to the cerebral cortex, olfactory system and autonomic nervous system (Dauer & Przedborski 2003). Moreover, PD has many non-motor symptoms (Chaudhuri et al. 2006), is heterogeneous in its clinical presentation and likely has several underlying pathogenic mechanisms (Elkouzi et al. 2019), meaning that different animal models are potentially needed for different subtypes of PD. Taking all these factors into account, there is a significant risk of wasted effort in experimentation with inadequate models, and a risk of ethical challenges regarding the unnecessary use of animals.

Nevertheless, when focussed on symptomatic relief, animal models have directly resulted in the development of life-changing therapies for many PD patients, namely treatment with levodopa, dopamine agonists and deep brain stimulation, as well as significantly furthering understanding of related neurophysiology.

Conclusion: the future of animal models in PD

The sensible approach for the future appears to be: continue to use animal models, but selectively. Existing animal models have proven invaluable in the development of therapeutic strategies to provide relief from the symptoms of PD. However, the same models are ineffective at replicating the exact biochemical mechanisms underlying PD in humans and should be used in the development of disease-modifying agents only after careful consideration. Preferably, better models need to be found, most likely using transgenic approaches, or experiments need to be performed directly on humans. Using models like MPTP to try and develop cures for PD will probably continue to lead down costly and time-consuming blind alleys.

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[1] https://www.drugs.com/monograph/pramipexole.html

[2] https://www.ema.europa.eu/en/medicines/human/EPAR/sifrol