Kinetics of the metabolism of four PET radioligands in living minipigs.

Most radioligands are substantially metabolised in peripheral organs during the course of positron emission tomography (PET) recordings. Accurate determination of plasma concentrations of unmetabolised radioligands is often important for quantification of data from PET studies. The fractions of untransformed radioligand and radioactive metabolites in plasma extracts must then be measured. Temporal changes in these fractions are influenced by the rate constant of appearance of total radioactive metabolites in plasma (apparent rate constant of metabolism in plasma, k(0)) and the net rate constant of elimination of all radioactive metabolites from plasma (k(-1)). In order to clarify the relationship between radioligand fractions and rate constants, plasma samples collected from Göttingen minipigs during PET recordings using four different binding site ligands were analysed by radio high performance liquid chromatography. The calculated plasma concentrations of parent compounds and their radioactive metabolites were used to calculate k(0) and k(-1) for 11C-labelled NNC 112, NS 2214, PK 11195 and raclopride in minipigs using a novel application of the tissue-slope intercept plot. In general, the apparent rate constant of metabolism in plasma was found to be greater in the minipig than in man. The reported kinetic analysis enables the apparent metabolism of PET radioligands in plasma to be quantified.

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.

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.

The synthesis of (R)- and (S)-[N-methyl-11C]beta, beta-difluoromethamphetamine for the investigation of the binding mechanism of biogenic amines in vivo.

In an attempt to elucidate the contribution of the extent of nitrogen protonation on the in vivo binding of methamphetamine in the brain, the enantiomers of [N-methyl-11C]beta, beta-difluoroamphetamine (4) were prepared for use in positron emission tomography (PET) studies. Thus, the enantiomers of beta, beta-difluoroamphetamine were prepared from trans-beta-methylstyrene, via bromination, conversion into the azirine, fluorination and resolution as the tartrate salts. (R)- and (S)-beta, beta-difluoroamphetamine (3) were then each labelled with carbon-11 (tt/2 = 20.4 min) by N-methylation of the corresponding homochiral beta, beta-difluoroamphetamine with [11C]methyl iodide. The labelled products were each synthesised, purified and formulated in 35 min, starting from [11C]carbon dioxide in 15-16% decay-corrected radiochemical yield, with a radiochemical purity of > 99% and specific radioactivity of 50-150 GBq mumol-1 at end of synthesis.