Effect of mGluR2 and mGluR2/3 activators on parkinsonism in the MPTP-lesioned non-human primate.

We have previously discovered that the selective activation of metabotropic glutamate type 2 receptors (mGluR2) and concurrent stimulation of metabotropic glutamate types 2 and 3 receptors (mGluR2/3) enhance the anti-parkinsonian action of L-3,4-dihydroxyphenylalanine (L-DOPA). Here, we sought to determine the effects of the mGluR2/3 orthosteric agonists LY-354,740 and LY-404,039, as well as the effects of the mGluR2 positive allosteric modulators LY-487,379 and CBiPES on the range of movement, bradykinesia, posture and alertness as adjuncts to L-DOPA. Ten 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned marmosets entered 4 experimental streams: L-DOPA + LY-354,740 (vehicle, 0.1, 0.3 and 1 mg/kg), L-DOPA + LY-404,039 (vehicle, 0.1, 1 and 10 mg/kg), L-DOPA + LY-487,379 (vehicle, 0.1, 1 and 10 mg/kg), L-DOPA + CBiPES (vehicle, 0.1, 1 and 10 mg/kg). For each molecule, treatments were randomised, and the range of movement, bradykinesia, posture and alertness were assessed by a blinded rater. None of the tested compounds significantly altered the global range of movement. LY-404,039 and CBiPES both reduced global bradykinesia, by up to 46% (both P < 0.05). LY-354,740, LY-404,039 and CBiPES each improved global posture by 35%, 44% and 39% (each P < 0.05), respectively. LY-404,039 and CBiPES both enhanced alertness by 54% (P < 0.05) and 79% (P < 0.01), respectively. LY-487,379 did not improve any of the parameters. Our results suggest that selective mGluR2 positive allosteric modulation and combined mGluR2/3 orthosteric stimulation might benefit bradykinesia, posture and alertness in PD when added to L-DOPA, which potentially represent novel therapeutic indications for molecules acting via these mechanisms.

[3H]-NFPS binding to the glycine transporter 1 in the hemi-parkinsonian rat brain.

L-3,4-dihydroxyphenylalanine (L-DOPA) is the main treatment for Parkinson's disease (PD) but with long term administration, motor complications such as dyskinesia are induced. Glycine transporter 1 (GlyT1) inhibition was shown to produce an anti-dyskinetic effect in parkinsonian rats and primates, coupled with an improvement in the anti-parkinsonian action of L-DOPA. The expression of GlyT1 in the brain in the dyskinetic state remains to be investigated. Here, we quantified the levels of GlyT1 across different brain regions using [3H]-NFPS in the presence of Org-25,935. Brain sections were chosen from sham-lesioned rats, L-DOPA-naïve 6-hydroxydopamine (6-OHDA)-lesioned rats and 6-OHDA-lesioned rats exhibiting mild or severe abnormal involuntary movements (AIMs). [3H]-NFPS binding decreased in the ipsilateral and contralateral thalamus, by 28% and 41%, in 6-OHDA-lesioned rats with severe AIMs compared to sham-lesioned animals (P < 0.01 and 0.001). [3H]-NFPS binding increased by 21% in the ipsilateral substantia nigra of 6-OHDA-lesioned rats with severe AIMs compared to 6-OHDA-lesioned rats with mild AIMs (P < 0.05). [3H]-NFPS binding was lower by 19% in the contralateral primary motor cortex and by 20% in the contralateral subthalamic nucleus of 6-OHDA-lesioned rats with mild AIMs animals compared to rats with severe AIMs (both P < 0.05). The severity of AIMs scores positively correlated with [3H]-NFPS binding in the ipsilateral substantia nigra (P < 0.05), ipsilateral entopeduncular nucleus (P < 0.05) and contralateral primary motor cortex (P < 0.05). These data provide an anatomical basis to explain the efficacy of GlyT1 inhibitors in dyskinesia in PD.

Metabotropic glutamate receptors in Parkinson's disease.

Parkinson's disease (PD) is a complex disorder that leads to alterations in multiple neurotransmitter systems, notably glutamate. As such, several drugs acting at glutamatergic receptors have been assessed to alleviate the manifestation of PD and treatment-related complications, culminating with the approval of the N-methyl-d-aspartate (NMDA) antagonist amantadine for l-3,4-dihydroxyphenylalanine (l-DOPA)-induced dyskinesia. Glutamate elicits its actions through several ionotropic and metabotropic (mGlu) receptors. There are 8 sub-types of mGlu receptors, with sub-types 4 (mGlu4) and 5 (mGlu5) modulators having been tested in the clinic for endpoints pertaining to PD, while sub-types 2 (mGlu2) and 3 (mGlu3) have been investigated in pre-clinical settings. In this book chapter, we provide an overview of mGlu receptors in PD, with a focus on mGlu5, mGlu4, mGlu2 and mGlu3 receptors. For each sub-type, we review, when applicable, their anatomical localization and possible mechanisms underlying their efficacy for specific disease manifestation or treatment-induced complications. We then summarize the findings of pre-clinical studies and clinical trials with pharmacological agents and discuss the potential strengths and limitations of each target. We conclude by offering some perspectives on the potential use of mGlu modulators in the treatment of PD.

Anti-parkinsonian effect of the mGlu2 positive allosteric modulator LY-487,379 as monotherapy and adjunct to a low L-DOPA dose in the MPTP-lesioned marmoset.

In previous experiments, we have discovered that positive allosteric modulation of metabotropic glutamate 2 (mGlu2) receptors enhances the anti-parkinsonian action of an optimal dose of L-3,4-dihydroxyphenylalanine (L-DOPA). Whether selective mGlu2 positive allosteric modulation would also alleviate parkinsonian disability as monotherapy or as adjunct to a sub-optimal dose of L-DOPA has not been determined. Here, we assessed the anti-parkinsonian effect of mGlu2 positive allosteric modulation as monotherapy and adjunct to a sub-optimal dose of L-DOPA in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned marmosets. The highly selective positive allosteric modulator (PAM) LY-487,379 was utilised to activate mGlu2 receptors. When administered as monotherapy, LY-487,379 10 mg/kg diminished global parkinsonism by 48% (P < 0.001) and increased duration of on-time by 7-fold, when compared to vehicle treatment (P < 0.05). When added to a sub-optimal dose of L-DOPA, LY-487,379 10 mg/kg decreased global parkinsonism by 44% (P < 0.001) and extended duration of on-time by 2.5-fold (P < 0.01). Our results indicate that selective mGlu2 positive allosteric modulation elicits anti-parkinsonian benefits as monotherapy and as adjunct to sub-optimal dose of L-DOPA paradigms, potentially suggesting that mGlu2 PAMs may have a therapeutic niche early in the treatment of PD as DOPA-sparing agents.

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Effect of glycine transporter 1 inhibition with bitopertin on parkinsonism and L-DOPA induced dyskinesia in the 6-OHDA-lesioned rat.

Dyskinesia remains an unmet need in Parkinson's disease (PD). We have previously demonstrated that glycine transporter 1 (GlyT1) inhibition with ALX-5407 reduces dyskinesia and slightly improves parkinsonism in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned marmoset. Here, we sought to determine the effect of bitopertin, a clinically-tested GlyT1 inhibitor, on parkinsonism and dyskinesia in the 6-hydroxydopamine (6-OHDA)-lesioned rat. To do so, we assessed the effect of bitopertin on parkinsonism as monotherapy and as adjunct to a low dose of L-3,4-dihydroxyphenylalanine (L-DOPA). We then assessed the efficacy of bitopertin on dyskinesia in the context of acute challenge and chronic administration studies. Lastly, we evaluated whether de novo treatment with bitopertin, started concurrently with L-DOPA, would diminish the development of dyskinesia. We discovered that bitopertin (0.3 mg/kg), when administered alone, reduced the severity of parkinsonism by 35% (P < 0.01). As adjunct to a low dose of L-DOPA, bitopertin (3 mg/kg) enhanced the anti-parkinsonian effect of L-DOPA by 36% (P < 0.05). Moreover, the acute addition of bitopertin (0.03 mg/kg) to L-DOPA reduced dyskinesia by 27% (P < 0.001), and there was no tolerance to the anti-dyskinetic benefit after 4 weeks of daily administration. Lastly, bitopertin (0.03 mg/kg) started concurrently with L-DOPA, also attenuated the development of dyskinesia, by 33% (P < 0.01), when compared to L-DOPA alone. Our results suggest that GlyT1 inhibition may simultaneously reduce parkinsonism and L-DOPA-induced dyskinesia and represents a novel approach to treat, possibly prevent, motor complications in PD.

Cognition and serotonin in Parkinson's disease.

Cognitive impairment affects up to 80% of patients with Parkinson's disease (PD) and is associated with poor quality of life. PD cognitive dysfunction includes poor working memory, impairments in executive function and difficulty in set-shifting. The pathophysiology underlying cognitive impairment in PD is still poorly understood, but there is evidence to support involvements of the cholinergic, dopaminergic, and noradrenergic systems. Only rivastigmine, an acetyl- and butyrylcholinesterase inhibitor, is efficacious for the treatment of PD dementia, which limits management of cognitive impairment in PD. Whereas the role of the serotonergic system in PD cognition is less understood, through its interactions with other neurotransmitters systems, namely, the cholinergic system, it may be implicated in cognitive processes. In this chapter, we provide an overview of the pharmacological, clinical and pathological evidence that implicates the serotonergic system in mediating cognition in PD.

Effect of the glycine transporter 1 inhibitor ALX-5407 on dyskinesia, psychosis-like behaviours and parkinsonism in the MPTP-lesioned marmoset.

Dyskinesia and psychosis are complications encountered in advanced Parkinson's disease (PD) following long-term therapy with L-3,4-dihydroxyphenylalanine (L-DOPA). Disturbances in the glutamatergic system have been associated with both dyskinesia and psychosis, making glutamatergic modulation a potential therapeutic approach for these. Treatments thus far have sought to dampen glutamatergic transmission, for example through blockade of N-methyl-D-aspartate (NMDA) receptors or modulation of metabotropic glutamate receptors 5. In contrast, activation of the glycine-binding site on NMDA receptors is required for their physiological response. Here, we investigated whether indirectly enhancing glutamatergic transmission through inhibition of glycine re-uptake would be efficacious in diminishing both dyskinesia and psychosis-like behaviours (PLBs) in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned common marmoset. Six marmosets were rendered parkinsonian by MPTP injection. Following repeated administration of L-DOPA to induce dyskinesia and PLBs, they underwent acute challenges of the glycine transporter 1 (GlyT1) inhibitor ALX-5407 (0.01, 0.1 and 1 mg/kg) or vehicle, in combination with L-DOPA, after which the severity of dyskinesia, PLBs and parkinsonian disability was evaluated. In combination with L-DOPA, ALX-5407 0.1 and 1 mg/kg significantly reduced the severity of dyskinesia, by 51% and 41% (both P < 0.001), when compared to vehicle. ALX-5407 0.01, 0.1 and 1 mg/kg also decreased the severity of global PLBs, by 25%, 51% and 38% (all P < 0.001), when compared to vehicle. The benefits on dyskinesia and PLBs were achieved without compromising the therapeutic effect of L-DOPA on parkinsonism. Our results suggest that GlyT1 inhibition may be a novel strategy to attenuate dyskinesia and PLBs in PD, without interfering with L-DOPA anti-parkinsonian action.

Combined 5-HT2A and mGlu2 modulation for the treatment of dyskinesia and psychosis in Parkinson's disease.

Antagonising the serotonin 2A (5-HT2A) receptor is an efficacious way to alleviate dyskinesia and psychosis in Parkinson's disease (PD). However, previous research indicates that there might be a limit to the effects conferred by this approach. 5-HT2A receptors were shown to form hetero-dimers with metabotropic glutamate 2 (mGlu2) receptors, in which 5-HT2A blockade and mGlu2 activation elicit equivalent effects at the downstream signalling level. We have previously shown that mGlu2 activation reduces both dyskinesia and psychosis-like behaviours (PLBs) induced by L-3,4-dihydroxyphenylalanine (l-DOPA), in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned primate. Here, we hypothesised that concurrent 5-HT2A antagonism and mGlu2 activation would provide greater anti-dyskinetic and anti-psychotic benefits than either approach alone. We conducted 3 series of experiments in the MPTP-lesioned marmoset. In the first series of experiments, the mGlu2 positive allosteric modulator LY-487,379 and the 5-HT2A antagonist EMD-281,014, either alone or in combination, were added to l-DOPA. In the second series of experiments, the mGlu2/3 orthosteric agonist LY-354,740 and EMD-281,014, either alone or in combination, were added to l-DOPA. In the last series of experiments, we investigated whether mGlu2 blockade would diminish the effects of antagonising 5-HT2A receptors. To this end, the mGlu2/3 orthosteric antagonist LY-341,495 and EMD-281,014, either alone or in combination, were added to l-DOPA. We found that the anti-dyskinetic effect of the combination LY-487,379/EMD-281,014 was greater than the ones conferred by LY-487,379 (by 35%, P < 0.05) and EMD-281,014 (by 38%, P < 0.01). The anti-dyskinetic and anti-psychotic effects of the combination LY-354,740/EMD-281,014 were also greater than the ones conferred by LY-354,740 (by 57% for dyskinesia and 54% for PLBs, both P < 0.001) and EMD-281,014 (by 61% for dyskinesia and 53% for PLBs, both P < 0.001). The anti-parkinsonian action of l-DOPA was maintained with all treatments. Lastly, the addition of LY-341,495 abolished the therapeutic effects of EMD-281,014 on dyskinesia and PLBs. Our results suggest that mGlu2 activation may enhance the anti-dyskinetic and anti-psychotic effects of 5-HT2A blockade and could provide relief to PD patients with dyskinesia and psychotic symptoms beyond what can be achieved with current therapies.

Effect of the mGlu2 positive allosteric modulator CBiPES on dyskinesia, psychosis-like behaviours and parkinsonism in the MPTP-lesioned marmoset.

Advanced Parkinson's disease (PD) is often complicated by the occurrence of dyskinesia, motor fluctuations and psychosis. To this day, few treatment options are available for each of these phenomena, and they are at times not effective or elicit adverse events, leaving some patients short of therapeutic options. We have recently shown that positive allosteric modulation of metabotropic 2 (mGlu2) receptors with the prototypical positive allosteric modulator (PAM) LY-487,379 is efficacious at alleviating both dyskinesia and psychosis-like behaviours (PLBs), while simultaneously enhancing the anti-parkinsonian action of L-3,4-dihydroxyphenylalanine (L-DOPA), in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned marmoset. Here, we assessed the effects of CBiPES, a mGlu2 PAM derived from LY-487,379, but with improved pharmacokinetic properties. Six MPTP-lesioned marmosets with reproducible dyskinesia and PLBs were administered L-DOPA in combination with vehicle or CBiPES (0.1, 1 and 10 mg/kg), after which their behaviour was rated. CBiPES 10 mg/kg reduced global dyskinesia by 60% (P < 0.0001), while peak dose dyskinesia was reduced by 66% (P < 0.001), compared to L-DOPA/vehicle. CBiPES 10 mg/kg also diminished global PLBs by 56% (P < 0.0001), while peak dose PLBs were reduced by 64% (P < 0.001), compared to L-DOPA/vehicle. Lastly, CBiPES enhanced the anti-parkinsonian action of L-DOPA, by reducing global parkinsonian disability by 43% (P < 0.01), compared to L-DOPA/vehicle. Our results provide further evidence that mGlu2 positive allosteric modulation may be an approach that could be efficacious for the treatment of dyskinesia, psychosis and motor fluctuations in PD.

Stereological investigation of 5-HT3 receptors in the substantia nigra and dorsal raphe nucleus in the rat.

Serotonin (5-HT) is a common neurotransmitter in mammals, playing a central role in the regulation of various processes such as sleep, perception, cognitive and autonomic functions in the nervous system. Previous studies have demonstrated that 5-HT type 3 (5-HT3) receptors are expressed in either or both the substantia nigra (SN) and the dorsal raphe nucleus (DRN) in humans, marmosets, rats and Syrian hamsters. Here, we quantify the distribution of 5-HT3 receptors across these regions in the adult rat. Fluorescent immunohistochemistry was performed on sections of rat brain covering the entire rostro-caudal extent of the SN and DRN with antibodies specific to the 5-HT3A receptor subunit, as well as others targeting the monoaminergic markers tyrosine hydroxylase (TH) and the 5-HT transporter (SERT). The number of 5-HT3A receptor-positive, TH-positive (n = 28,428 ±â€¯888, Gundersen's m = 1 coefficient of error [CE] = 0.05) and SERT-positive (n = 12,852 ±â€¯462, CE = 0.06) cells were estimated in both the SN and the DRN using stereology. We found that 5-HT3A receptor-positive cells are present in the SNr (n = 1250 ±â€¯64, CE = 0.24), but they did not co-localise with TH-positive cells, nor were they present in the SNc. In contrast, no 5-HT3A receptor-positive cells were found in the DRN. These results support the presence of 5-HT3 receptors in the SN, but not in the DRN, and do not support their expression on monoaminergic cells within these two brain areas.

Effect of the selective 5-HT2A receptor antagonist EMD-281,014 on L-DOPA-induced abnormal involuntary movements in the 6-OHDA-lesioned rat.

L-3,4-Dihydroxyphenylalanine (L-DOPA) is the most effective therapy for motor symptoms of Parkinson's disease (PD); however, with repeated administration, as many as 94% of PD patients develop complications such as L-DOPA-induced dyskinesia. We previously demonstrated that EMD-281,014, a highly selective serotonin 2A (5-HT2A) receptor antagonist, reduces the severity of dyskinesia in the parkinsonian marmoset, without interfering with L-DOPA anti-parkinsonian benefit. Here, we assessed the effects of EMD-281,014 on L-DOPA-induced abnormal involuntary movements (AIMs) in the 6-hydroxydopamine (6-OHDA)-lesioned rat. We first determined the pharmacokinetic profile of EMD-281,014, to administer doses leading to clinically relevant plasma levels in the behavioural experiments. Dyskinetic 6-OHDA-lesioned rats were then administered EMD-281,014 (0.01, 0.03 and 0.1 mg/kg) or vehicle in combination with L-DOPA and AIMs severity was evaluated. We also assessed the effect of EMD-281,014 on L-DOPA anti-parkinsonian action with the cylinder test. We found that the addition of EMD-281,014 (0.01, 0.03 and 0.1 mg/kg) to L-DOPA did not reduce AIMs severity (P > 0.05), when compared to vehicle. EMD-281,014 did not compromise L-DOPA anti-parkinsonian action. Our results suggest that the highly selective 5-HT2A receptor antagonist EMD-281,014 is well-tolerated by parkinsonian rats, but does not attenuate L-DOPA-induced AIMs. Our results highlight differences between rodent and primate models of PD when it comes to determining the anti-dyskinetic action of 5-HT2A receptor antagonists.