The intent of this proposal is two-fold: 1) to develop positron-emitting tracers that will quantify enzyme activity within the sympathetic nerves of the heart by means of """"""""metabolic expulsion""""""""; 2) to study by positron emission tomography(PET) the effect of cardioactive and abusive drugs on the efflux kinetics of these tracers and other neuronal markers previously developed in our laboratory. The synthetic false adrenergic neurotransmitter [11C]meta-hydroxyephedrine (MHED) and the natural neurohormone [11C]epinephrine have been successfully developed in our laboratory to map the sympathetic nerve density of the human heart. The retention of MHED in the heart is predominantly uptake(1) dependent whereas the retention of [11C]epinephrine is dependent on vesicular storage. The present proposal will shift our previous emphasis from developing metabolically-resistant tracers that map the intact, """"""""healthy"""""""" neuron to that of focusing on tracers that can specifically assess the enzymatic """"""""health"""""""" of the neuron. In particular, the cytoplasmic enzyme, monoamine oxidase (MAO-B), and the vesicular enzyme, dopamine-beta- hydroxylase (DBH) will be targeted. [11C]Phenylephrine, readily synthesized by 11C-methylation of meta-octopamine, is uniquely structured to map heart MAO-B activity; its efflux rate from the heart should represent elimination of metabolites derived solely from the action of MAO-B. This hypothesis will be validated by determining the deuterium isotope effect of alpha-di-deuteron-[11C]phenylephrine on heart efflux rate, drug blocking studies with selective MAO-B and MAO-A inhibitors, and HPLC analyses of metabolites in blood and heart tissue. A novel approach to mapping heart DBH activity will be tested which uses the neuron-specific DBH substrate beta-S-[18F]fluoro-MHED. This tracer is designed to be hydroxylated to the respective fluorohydrin which will rapidly eliminate [18F]fluoride ion. The efflux rate of [18F]fluoride from the heart will provide an index of heart neuronal DBH activity. This hypothesis will be validated by determining DBH action on the tracer in vitro, drug blocking studies in vivo with desipramine and reserpine, and HPLC analyses of blood and heart tissue. The cyclic sulfamate method of introducing 18F into the ethanolamine side chain of biogenic amines, developed in this laboratory, will be employed to synthesize beta-R-[18F]fluoro analogs of MHED, epinephrine, and norepinephrine. These tracers will not be metabolic probes of MAO-B or DBH, but will provide longer-lived radioactive mapping agents for the sympathetic neuron based on their cytoplasmic or vesicular retention. After the tracers proposed in this application are validated, the sympathetic nerve effects of possible cardioactive drugs such as amphetamine, sudafed, cocaine, bretylium MPTP, and deprenyl will be systematically evaluated by PET tracer kinetic studies in dogs.
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