The DaNeX Study of Embryonic Mesencephalic, Dopaminergic Tissue Grafted to a Minipig Model of Parkinson's Disease: Preliminary Findings of Effect of MPTP Poisoning on Striatal Dopaminergic Markers

Abstract

A multicenter study is under way to investigate the efficacy of allografting of embryonic mesencephalic neurons in a pig model of Parkinson's disease. We have first established that a stable parkinsonian syndrome can be established by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) intoxication of adult male Göttingen minipigs. We are now using positron emission tomography (PET) methods for testing the physiological responses to MPTP intoxication and the time course of the response to several treatment strategies. We now report preliminary results obtained in 11 pigs employed in the initial phase of the study; the completed study shall ultimately include 30 pigs. Animals were randomly assigned to one of five groups: 1) Control, 2) MPTP intoxication, 3) MPTP intoxication followed by allograft, 4) MPTP intoxication followed by allograft with immunosuppression, and 5) MPTP intoxication followed by allograft with immunosuppression and co-grafting of immortalized HiB5 cells, which had been manipulated to secrete glia cell line-derived neurotrophic factor (GDNF) (approximately 2 ng GDNF/h/10(5) cells). MPTP was administered (1 mg/kg/day, SC) for 7-10 days until the pigs had developed mild parkinsonian symptoms of muscle rigidity, hypokinesia, and impaired coordination, especially of the hind limbs. Approximately 2 weeks after the last MPTP dose, animals received a T1-weighted magnetic resonance imaging (MRI) scan, and a series of dynamic PET recordings. After the first series of PET scans, four grafts of porcine embryonic mesencephalic tissue (E28 days) were placed in each striatum of some MPTP-intoxicated pigs, using MRI-based stereotactic techniques. Immunosuppression of some animals with cyclosporin and prednisolone began just prior to surgery. Two more series of PET scans were performed at 4-month intervals after surgery. After the last scans, pigs were killed and the brains were perfused for unbiased stereological examination of cytological and histochemical markers in striatum and substantial nigra. The behavioral impairment of the animals (the "Parkinson's score") had been evaluated throughout the 8-month period. Kinetic analysis of the first set of PET scans has indicated that the rate constant for the decarboxylation of FDOPA in catecholamine fibers was reduced by 33% in striatum of the mildly parkinsonian pigs. The rate of association of [11C]NS-2214 to catecholamine uptake sites was reduced by 62% in the same groups of pigs. No significant difference was found in the binding potential of [11C]raclopride to the dopamine D2-like receptors in striatum of the MPTP-intoxicated versus control pigs. These preliminary results are suggestive that the activity of DOPA decarboxylase may be upregulated in the partially denervated pig striatum.

A MRI-compatible stereotaxic localizer box enables high-precision stereotaxic procedures in pigs

“…The Göttingen minipig is used for basal ganglia research, as an alternative to: (1) primates because of less economical and ethical burdens; (2) rats due to the larger brain size allowing imaging with conventional MR and PET-scanners and surgical implantation of fetal dopaminergic tissue or deep brain stimulation (DBS) electrodes Cumming et al, 2001;Danielsen et al, 1998Danielsen et al, , 2000Mikkelsen et al, 1999). A large frontal sinus pneumatizes the minipig skull making the use of bony sutures unreliable for stereotaxic implantation, as the distance between these and the brain structures may vary considerably inter-individually .…”

“…A large frontal sinus pneumatizes the minipig skull making the use of bony sutures unreliable for stereotaxic implantation, as the distance between these and the brain structures may vary considerably inter-individually . Accordingly, we perform a two-step procedure where MR-visible markers are placed in the skull followed by MRI enabling relation of the brain target to the implanted markers Danielsen et al, 2000). The animal would then be placed in a stereotaxic device for the stereotaxic implant procedure, demanding that the fixated head position corresponds exactly to the head position during previously obtained stereotaxic MRI.…”

Semi‐micro liquid chromatography of aromatic amino acid metabolites using isocratic elution and Column switching

“…This new system was tested for the analysis of catecholamine-related compounds in the pig brain, as pigs are a common animal model used for transgenic manipulation of neural genes (Erlander et al, 1985; Agarwal et al, 1993;Leshin et al, 1996;Danielsen et al, 2000;Cumming et al, 2001). Samples of 350 mg of fresh pig brain obtained from Tokyo Shibaura Zoki (Tokyo, Japan) were homogenized with 2.0 mL of 1.0 M HClO 4 .…”

The substantia nigra pars compacta of the Göttingen minipig: an anatomical and stereological study

“…The Göttingen minipig has gained increased interest during the last decade as a non-primate large experimental animal, also including its use as an animal model in PD (Mikkelsen et al 1999;Danielsen et al 2000;Sørensen et al 2000;Watanabe et al 2001;Dall et al 2002;Bjarkam et al 2004Bjarkam et al , 2005Bjarkam et al , 2008aDalmose et al 2004Dalmose et al , 2005Lind et al 2007;Lindvall and Wahlberg 2008;Jensen et al 2009;Rosendal et al 2009). For further refinement and use of the animal model in PD research, detailed knowledge of the SNc is crucial.…”