Развитие одностороннем порядке, пораженном 6-OHDA мышиной модели болезни Паркинсона
Summary
Протокол для проведения односторонних 6-OHDA поражения медиальных пучка переднего мозга у мышей описано. Этот метод имеет низкий уровень смертности (13,3%) с 89% выживших животных показывает> 95% потерь полосатой дофамина и 90,63 ± -4,02% ipsiversive вращательное смещение в сторону поражения.
Abstract
The unilaterally lesioned 6-hyroxydopamine (6-OHDA)-lesioned rat model of Parkinson’s disease (PD) has proved to be invaluable in advancing our understanding of the mechanisms underlying parkinsonian symptoms, since it recapitulates the changes in basal ganglia circuitry and pharmacology observed in parkinsonian patients1-4. However, the precise cellular and molecular changes occurring at cortico-striatal synapses of the output pathways within the striatum, which is the major input region of the basal ganglia remain elusive, and this is believed to be site where pathological abnormalities underlying parkinsonian symptoms arise3,5.
In PD, understanding the mechanisms underlying changes in basal ganglia circuitry following degeneration of the nigro-striatal pathway has been greatly advanced by the development of bacterial artificial chromosome (BAC) mice over-expressing green fluorescent proteins driven by promoters specific for the two striatal output pathways (direct pathway: eGFP-D1; indirect pathway: eGFP-D2 and eGFP-A2a)8, allowing them to be studied in isolation. For example, recent studies have suggested that there are pathological changes in synaptic plasticity in parkinsonian mice9,10. However, these studies utilised juvenile mice and acute models of parkinsonism. It is unclear whether the changes described in adult rats with stable 6-OHDA lesions also occur in these models. Other groups have attempted to generate a stable unilaterally-lesioned 6-OHDA adult mouse model of PD by lesioning the medial forebrain bundle (MFB), unfortunately, the mortality rate in this study was extremely high, with only 14% surviving the surgery for 21 days or longer11. More recent studies have generated intra-nigral lesions with both a low mortality rate >80% loss of dopaminergic neurons, however expression of L-DOPA induced dyskinesia11,12,13,14 was variable in these studies. Another well established mouse model of PD is the MPTP-lesioned mouse15. Whilst this model has proven useful in the assessment of potential neuroprotective agents16, it is less suitable for understanding mechanisms underlying symptoms of PD, as this model often fails to induce motor deficits, and shows a wide variability in the extent of lesion17, 18.
Here we have developed a stable unilateral 6-OHDA-lesioned mouse model of PD by direct administration of 6-OHDA into the MFB, which consistently causes >95% loss of striatal dopamine (as measured by HPLC), as well as producing the behavioural imbalances observed in the well characterised unilateral 6-OHDA-lesioned rat model of PD. This newly developed mouse model of PD will prove a valuable tool in understanding the mechanisms underlying generation of parkinsonian symptoms.
Protocol
Discussion
Этот протокол описывает метод для генерации стабильного одностороннем 6-OHDA-пораженном мышиной модели болезни Паркинсона, которая является очень воспроизводимые, с высоким уровнем успеха поражением, и низкий уровень смертности. Успех 6-OHDA поражение операция может быть легко оценить пу…
Divulgazioni
The authors have nothing to disclose.
Acknowledgements
Эта работа выполнена при финансовой поддержке Министерства иностранных дел и международной торговли (Правительство Канады), Университет Торонто Connaught фонд, Канадский фонд инноваций, NSERC, Фонд Krembil и доверие Лечение болезни Паркинсона.
Materials
| Name of the reagent | Company | Catalogue number | Comments (optional) |
| desipramine HCl | Sigma-Aldrich, Oakville, ON, Canada | D125 | 25mg/kg |
| pargyline HCl | Sigma-Aldrich, Oakville, ON, Canada | P8013 | 5mg/kg |
| 6-OHDA HBr | Sigma-Aldrich, Oakville, ON, Canada | H116 | 3mg / mouse |
| stereotaxic Frame | Kopf Instruments, Tujunga, CA, USA | Model 900 | |
| mouse ear cups | Kopf Instruments, Tujunga, CA, USA | Model 921 Zygoma Ear Cups | |
| mouse incisor bar | Kopf Instruments, Tujunga, CA, USA | Model 923B | |
| mouse anaesthesia mask | Kopf Instruments, Tujunga, CA, USA | Model 923B | |
| priming kit (containing 250ml syringe) | Hamilton Company, Reno, NV, USA | PRMKIT 81120 | |
| RN compression fitting kit (1 mm) | Hamilton Company, Reno, NV, USA | 55750-01 | |
| PEEK tubing from RN compression fitting kit< (1/16th inch) | Hamilton Company, Reno, NV, USA | 55751-01 | |
| dual small hub RN Coupler | Hamilton Company, Reno, NV, USA | 55752-01 | |
| luer to small hub RN adaptor | Hamilton Company, Reno, NV, USA | 55753-01 | |
| 1ml 25S syringe model 7001KH | Hamilton Company, Reno, NV, USA | 80100 | |
| *33G removable needle (RN) pack of 6. . Custom 1 inch with 45<° bevel | Hamilton Company, Reno, NV, USA | 7803-05 | |
| Scissors | Fine Science Tools, Vancouver, BC, Canada. | 14084-08 | |
| Scalpel | Fine Science Tools, Vancouver, BC, Canada | 10003-12 | |
| Scalpel blades | Fine Science Tools, Vancouver, BC, Canada | 10035-20 | |
| Forcep | Fine Science Tools, Vancouver, BC, Canada | 11608-15 | |
| Hemostats | Fine Science Tools, Vancouver, BC, Canada. | 13004-14 | |
| Isoflurane | Abbot | 02241315 | 2-3% |
| Suters (Vicryl 4.0) | Syneture | SS-683 | |
| Steriliser | Fine Science Tools, Vancouver, BC, Canada | 18000-45 | |
| Infusion Pump | Harvard Apparatus | PhD 22/2000 | |
| Needles (27G) | Becton Dickinson | 305109 | |
| Needles (25G) | Becton Dickinson | 305127 | |
| Syringes (1ml) | BD syringe | 309692 | |
| Anaesthesia trolley | LEI medical | M2000 | |
| Baytril | CDMV, St. hyacinthe, QC | 102207 | |
| Lidocaine | CDMV, St. hyacinthe, QC | 3914 | |
| Betadine solution | CDMV, St. hyacinthe, QC | 19955 |
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