Parkinson disease (PD) is manifested as generalized abnormalities in locomotor movements (hypokinesias) (paralysis agitans) such as a stooped posture, difficulty in initiating and stopping walking, a short and shuffling gait, and a tendency to fall over [1-6]. The mean age of PD onset is about 60 years [7]; its prevalence is approximately 1% of those older than 65 years [8]. The disorder occurs throughout the world [9].

Post-mortem brains of PD patients have a decreased level of dopamine [10] that correlates with the loss of dopaminergic neurons within the substantia nigra pars compacta (SNpc) and degeneration of nerve terminals in the corpus striatum (basal ganglia) (Figure 1) [1]. Because the basal ganglia participate in regulation of neuronal transmission from the cerebral cortex to motor neurons of the spinal cord, decreased levels of dopamine disrupt the balance between dopamine and other neurotransmitters that are involved in a coordinated locomotor activity, including acetylcholine, GABA (g-aminobutyric acid), glutamate, 5-HT (5-hydroxytryptamine or serotonin), norepinephrine, and neuropeptides in striatal interneurons [11]. The SNpc target neurons express both major 7-TM-GPCR (seven-transmembrane G proteincoupled receptor) dopamine receptor (DAR) types: excitatory DAR D₁ DRD1 (transduced by stimulated formation of cAMP and phosphatidyl inositol hydrolysis) and inhibitory DAR D₂ DRD2 (transduced by decreased cAMP and modulation of K⁺ and Ca²⁺ currents). In experimentally-lesioned animals, damage to the nigrostriatal pathway results in a hyperactivity of the remaining dopaminergic neurons, increased transmitter turnover rate, and state of denervation super sensitivity [12].

Current pharmacologic management of PD involves a restoration of the balance between dopamine and other neurotransmitters, most commonly by replenishment of dopamine [13]. Since direct administration of dopamine

is ineffective or inefficient, because it does not readily cross the blood-brain barrier, indirect approaches are used. The most common is administration of the amino acid levodopa (L-DOPA) [13,14]. L-DOPA enters the brain by active amino acid transport systems [15] and is then converted to dopamine by the enzyme L-aromatic amino acid decarboxylase (DOPA decarboxylase) [16].

There are several animal models of PD [1]. Most of the models use mammals and are generally both timeand resource-intensive. We here report a convenient and facile non-mammalian model for in vivo demonstration and quantitative assessment of DOPA decarboxylase or an equivalent enzyme activity in a species with primitive nervous system and mammalian-analogous neurotransmitter and 2nd messenger systems [17-26]. The model is useful for the study of drug action and for physiological aspects of drug abuse [27-43].

Phase 1. Exposure of planarians to L-DOPA resulted in a doseand time-dependent toxicity that was manifested by a decrease in number over the 2-h observation period. At each of the doses of L-DOPA, the effect of L-DOPA was attenuated by the DOPA decarboxylase inhibitor benserazide (see Figure 2).

Phase 2. Consistent with previous results [31], untreated planarians displayed a relatively constant and a near linear spontaneous locomotor activity (pLMV) of about 10 - 15 gridlines per minute over the entire obser-

vation period of 10 minutes. L-DOPA decreased the pLMV. Importantly, the pLMV in the presence of LDOPA was linear (see Figure 3).

The present work investigated the use of a simple in vivo planarian model in order to study the in vivo effect of L-DOPA, specifically, the metabolic conversion of L-DOPA by DOPA decarboxylase or the planarian equivalent. Planarians have a simple nervous system with mammalian-like neurotransmitter systems (e.g., [17,24, 26]), including dopaminergic. Therefore, they are the lowest form of extant animal that would display relevant dopaminergic-related neurotoxicity. They have been shown to respond with quantifiable dose-related behavioral changes to drug exposure or to withdrawal (e.g., [17,19,21,22,25-27,30,33-34,37]). We and others have used planarian models for investigation of drug action and related physiological processes (for a review, see the monograph [31]).

In the present study, the dose-related L-DOPA-induced (neuro)toxicity was attenuated by benserazide, which is an inhibitor of DOPA decarboxylase. It is used clinically together with L-DOPA in order to minimize the conversion of L-DOPA to dopamine in the periphery–thereby increasing the percent of the administered dose that is converted to dopamine in the CNS (Figure 4).

These results provide in vivo evidence for the presence and functional activity of DOPA decarboxylase or for an equivalent enzyme in planarians. A D. japonica aromatic amino acid decarboxylase-like activity (DjAADCA) gene,

and mRNA and protein expression have been identified in D. japonica by Nishimura and colleagues [44]. It thus seems that D. dorotocephala shares the same or similar enzyme.

In summary, L-DOPA produced doseand time-related lethality in planarians that was attenuated by a reversible inhibitor of DOPA decarboxylase (benserazide). These results are consistent with a previous demonstration of such an enzyme in planarians [44] and they support the use of a facile planarian model to study certain biochemical aspects of anti-Parkinson drugs.

The authors thank Timothy Shickley, Ph.D., for the suggestion of Planaria as a model. This work was supported by NIH grants DA15378 (RBR) and DA022694 (SMR).