INTRODUCTION (996 words)
Parkinson’s disease (PD) is a progressive, age-associated neurodegenerative disorder characterized by a depletion of dopamine in the striatum due to the loss of dopaminergic neurons of the substantia nigra compacta (SNc; Fahn et al., 2004; Olanow et al., 2004). L-DOPA (3,4-dihydroxyphenyl-L-alanine) is considered the most effective and well-tolerated noninvasive therapy for the treatment of PD motor symptoms. However, over time this effect decreases and more than 75% of L-DOPA-treated PD patients develop incapacitating abnormal involuntary movements (AIMs) referred to as L-DOPA-induced dyskinesia (LID) (Aquino and Fox, 2015; Cenci et al., 2020; Cenci and Olanow, 2017; Espay et al., 2018; Fahn et al., 2004; Olanow et al., 2004). As such, there has been a number of studies conducted to try and find alternative therapies that do not result in the pro-dyskinetic side effects or can reduce the dyskinetic signs.
The mechanisms involved in the pathogenesis of LID are complex and are not entirely understood. In the nigrostriatal circuitry, there are several alterations associated with LID manifestation, for example, the expression of specific genes (ERK1/2 extracellular signal-regulated kinase 1/2; DARP32 dopamine- and cAMP-regulated neuronal phosphoprotein; Fos, Finkel–Biskis–Jinkins osteosarcoma) (Cenci et al., 2007; Cenci, 2014; Cenci and Crossman 2018; You et al., 2018), and altered synaptic plasticity (Cenci et al., 2007, 2020; Cenci, 2014) with modifications in the long-term potentiation of the cortico-striatal pathway (Borgkvist et al., 2018; Calabresi et al., 2015; Picconi et al., 2011; 2003). Pre-clinical and clinical evidence has emerged to support the use of a number of compounds to treat LID. Amantadine (the non-selective N-methyl-D-aspartate receptor antagonist - Blanchet et al., 1998; Lundblad et al., 2002) is the only compound with robust evidence of anti-dyskinetic effects in patients. However, its therapeutic success is limited (Bortolanza et al., 2015; Cenci et al., 2020; 2014).
Various neural pathways such as the serotonin, acetylcholine, γ-aminobutyric acid, nitric oxide, or cannabinoid interact with dopaminergic pathways have been proposed as possible pharmacological targets for LID suppression (Dos-Santos-Pereira et al., 2016; Espadas et al., 2020; Huot et al., 2013; Johnston et al., 2018; Vijayakumar and Jankovic, 2016). In rodent PD models, neuroinflammation has been reported to be associated with the development and onset of LID (Barnun et al., 2008; Boi et al., 2019 Bortolanza et al., 2015 a, b; Munoz et al., 2014; Mulas et al., 2016; for review see Del Bel et al., 2016; Carta et al., 2017). In pre-clinical studies, pharmacological based therapy with anti-inflammatory compounds as corticosterone (Barnum et al., 2008), cannabidiol (dos-Santos-Pereira et al., 2016, Espadas et al., 2020), nitric oxide synthase inhibitor (Bortolanza et al., 2016; Padovan-Neto et al., 2009, 2015; Solís et al., 2015), thalidomide (Boi et al., 2019; Mulas et al., 2016), or IRC-82451 a multitargeting molecule (Aron-Badin et al., 2013), reduced the severity of dyskinesia and was associated with a reduction in inflammation. Amantadine may also be somewhat beneficial by inhibiting the inflammatory activation of microglia (Kim et al., 2012).
Recently, it was hypothesized that the striatum in LID, evolves from the combination of dopaminergic terminals degeneration, inadequate dopamine presence following the administration of L-DOPA and excessive glutamate concentration (Barcia et al., 2003; McGeer et al., 2003; Picconi et al., 2011; 2003; Tansey et al., 2007). In vivo brain imaging of patients with PD reveals an association between widespread microglial activation and the pathological process (Bartels et al., 2007; Gerhard et al., 2006; Ouchi et al., 2005). Inflammatory reaction in the basal ganglia of both parkinsonian rats and patients with PD that have a history of dyskinesia also revealed angiogenesis, vascular endothelial growth factor up-regulation and altered brain-blood barrier properties (Janelidze et al., 2015; Lerner et al., 2017; Ohlin et al., 2011, 2012). Furthermore, long after the neurotoxin, 6-hydroxydopamine (6-OHDA) has been cleared from the brain the striatum of lesioned rats receiving L-DOPA treatment displayed sustained neuroinflammation and neurodegeneration (Bortolanza et al., 2015a and b; Muñoz et al., 2014; Spinnewyn et al., 2011; Teema et al., 2016). Therefore, targeting neuroinflammation might be a pharmacological strategy to limit LID (Del Bel et al., 2016; Huot et al., 2013;).
Tetracycline antibiotics are increasingly being used, largely due to their anti-inflammatory features (Bortolanza et al., 2018; Reglodi et al., 2017). The second-generation tetracycline antibiotic, doxycycline (6-deoxy-5-hydroxytetracycline, a Food and Drug Administration approved compound), protects dopaminergic neurons from degeneration induced by 6-OHDA (Lazzarini et al., 2013), 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP, Du et al. 2001; Wu et al. 2003) and lipopolysaccharide (LPS, Zhang et al., 2015). This neuroprotective effect seems to be associated with its anti-inflammatory properties (Lazzarini et al., 2013; Stoilova et al., 2013). At a clinical level, the therapeutic rationale for targeting inflammation in PD is supported by the observation of the reduced incidence of PD in patients using tetracycline for rosacea treatment (Egeberg et al., 2016). Moreover, these rosacea patients did not present any prominent side effects related to the antimicrobial activity (Gompels et al. 2006; Langevitz et al. 1992; NINDS NET-PD Investigators 2006; Payne et al. 2011; Smith et al. 2011). Although it is unclear what drives increased inflammation in patients with PD, tetracycline and analog compounds have emerged as a potential treatment for PD and other neurodegenerative disorders (Johnston et al., 2019; Socias et al., 2018).
Due to the well-known anti-inflammatory effects of doxycycline, we hypothesized that this drug might reduce LID displayed inflammatory processes and, as a result, alleviate the expression of LID. In addition, to avoid side effects, such as bacterial resistance, COL-3 (CMT-3,6-demethyl-6-deoxy-4-de[dimethylamino]-tetracycline formerly known as incyclinide) a chemically modified tetracycline (Edan et al., 2013; Liu et al., 2001) might be efficacious in the treatment of LID.
In the present study, we examined the potential effect of doxycycline and COL-3 to modify LID induced by chronic L-DOPA treatment in 6-OHDA lesioned rats. We analyzed whether doxycycline and/or COL-3 given once dyskinesia was already present decreased dyskinetic signals and also whether the co-administration of doxycycline with L-DOPA from day one inhibited the appearance of LID. Using an immunohistochemistry analysis, we examined dopamine depleted striatum changes due to L-DOPA and doxycycline treatment.