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The mesoprefrontal dopamine neurons are sensitive to physical, pharmacological and psychological stressors. In this report, the role of these neurons in the response to classical fear conditioning was investigated. 6-Hydroxydopamine lesions to the medial prefrontal cortex reduced dopamine levels to about 13% of controls but did not alter behavior during the acquisition of fear conditioning. As expected, conditioned fear increased dopamine metabolism (3,4-dihydroxyphenylacetic acid/dopamine ratio) in the nucleus accumbens in sham-lesion rats. The medial prefrontal 6-hydroxydopamine lesions did not alter this effect. During the expression, however, lesioned rats demonstrated a delayed extinction of the conditioned response without an overall increase in the initial conditioned response. This effect was consistent in rats receiving 6-hydroxydopamine lesions before or after the acquisition period. The calculated rates of extinction showed that the 6-hydroxydopamine lesioned rats had a reduced rate of extinction, but not acquisition, of fear conditioning. The results presented in this manuscript indicate that the mesoprefrontal dopamine neurons are involved in co-ordinating the normal extinction of a fear response but do not alter the acquisition of fearful behaviors. These data are consistent with the conclusion that the mesoprefrontal dopamine neurons are involved in maintaining the animal's response adaptability with regards to stress-related changes in the external environment.

Levodopa is the most efficacious and widely used symptomatic drug for Parkinson's disease (PD). There is currently, however, a great deal of interest focused on the possibility that levodopa-induced increases in dopamine (DA) turnover may increase oxidative damage derived from the breakdown of DA. Increased oxidative damage following levodopa may contribute to the progressive degeneration of remaining host nigral neurons as well as interfere with development and function of embryonic nigral neurons in neural grafting trials. There is abundant evidence that levodopa is toxic to embryonic nigral DA neurons in both cell culture and neural grafting models. These findings have prompted a number of studies on mechanisms of levodopa toxicity to identify effective means of ameliorating potential oxidative stress related to levodopa in PD. In the current study we have utilized cultures of embryonic nigral DA neurons to address the fundamental question of whether levodopa-induced toxicity is related to DA production or whether dopa itself contributes to cell death. We compared the degree of nigral DA cell death following chronic administration of: 1) levodopa (e.g.: l-dopa); 2) its less active stereoisomer d-dopa; and 3) DA. We examined the rank order of toxicity of these compounds in two species of rats, Fisher 344 (F344) and Sprague-Dawley (SD). Results indicate a toxicity profile of: DA > l-dopa > > d-dopa. In addition, although there was no difference in response of F344 and SD cultures to l-dopa, the SD cultures were significantly more susceptible to the neurotoxic effects of DA. Taken collectively, these results suggest that levodopa-induced toxicity is related primarily to DA production rather than oxidation of dopa to toxic metabolites, and that some strain related differences do exist.

Stress consistently has been found to activate peripheral and central catecholamine systems. Dopamine (DA) turnover in the prefrontal cortex is especially sensitive to stress produced by relatively mild footshock, conditioned fear, or exposure to a novel cage. Because lesions of the central nucleus of the amygdala block the effects of both stress and fear in many experimental paradigms, the present study evaluated whether such lesions would block stress-induced increases in prefrontal dopamine turnover using either mild footshock or novelty as stressors. In Experiment 1 electrolytic lesions of the central nucleus of the amygdala attenuated the increase in the dopamine metabolite homovanillic acid (HVA) in the prefrontal cortex evaluated in post-mortem tissue normally produced by footshock. In Experiment 2 similar lesions attenuated the increase in dopamine turnover in the prefrontal cortex using a different stressor, novelty, and a different measure of dopamine turnover, DOPAC/DA ratios. These data provide further evidence for the critical role of the amygdala in stress.

Previous studies of rats in our laboratory indicate that a molecule or molecules released by Schwann cells exert survival and growth-promoting effects on mesencephalic dopamine neurons. In the present study, we have begun to investigate the potential for Schwann cell augmentation of host dopamine fiber systems and embryonic dopamine neuron grafts in non-human primates. Ten adult male St Kitts African Green monkeys treated with the dopaminergic neurotoxin 1-methyl 4-phenyl 1,2,3,6-tetrahydropyridine one year previously, but behaviorally asymptomatic, served as hosts for implant studies. A segment of young adult monkey saphenous nerve was collected to serve as an implanted tissue source of Schwann cell-derived growth factors. Nerve was enclosed in a hollow semi-permeable polymer fiber for implantation into the lateral ventricle, with embryonic ventral mesencephalic tissue co-grafts containing developing dopamine neurons aimed at nearby locations in the caudate nucleus. Control implants consisted of an empty polymer fiber co-grafted with embryonic ventral mesencephalon. Our morphological observations indicate that while no clear augmentation of the morphology of grafted dopamine neurons attributable to co-grafted nerve was observed, this lack of influence may be related to the spatial separation of the co-grafted tissues. In contrast, some monkeys with nerve segments in the lateral ventricle exhibited increased tyrosine hydroxylase-positive fiber staining in the immediately adjacent lateral septal area and the ventricular wall of the caudate nucleus. This enhancement was not associated with empty polymer implants. Levels of dopamine and its metabolite homovanillic acid derived from tissue punches in the caudate nucleus and septal area support the view that monkeys exhibiting morphological enhancement of host dopamine systems also show biochemical increases in dopamine levels and changes in the direction of normalization of the homovanillic acid/dopamine ratio. Biochemical values from a single septal area tissue punch in one animal were an exception to this rule. This study suggests that while the utility of peripheral nerve as a source of dopamine graft augmentation in non-human primates remains to be demonstrated, grafted nerve has a stimulatory effect on host brain dopamine systems in adult, dopamine-depleted monkeys, and that this morphological effect can be dissociated from previously hypothesized injury-induced regeneration.

The effects of prefrontal cortical dopamine depletion on subcortical dopamine function in the rat were examined. 6-Hydroxydopamine lesions of the dopaminergic innervation of the prefrontal cortex did not alter concentrations of dopamine or its metabolite 3,4-dihydroxyphenylacetic acid in either the striatum or nucleus accumbens. Similarly, the activity of the catecholamine biosynthetic enzyme tyrosine hydroxylase in the striatal complex was not changed in animals with prefrontal cortical lesions. Animals sustaining neurotoxic lesions of the prefrontal cortex were challenged with haloperidol in order to activate submaximally tyrosine hydroxylase activity. The magnitude of the haloperidol-induced increase in enzyme activity in the nucleus accumbens was significantly greater in lesioned subjects than in control animals. These data suggest that lesions of the prefrontal cortical dopamine innervation do not result in significant alterations in basal dopaminergic function in the striatal complex. However, lesions of the dopaminergic innervation of the prefrontal cortex significantly increase the responsiveness of mesolimbic dopamine afferents to pharmacological challenge.

Using the technique of perfusion microdialysis combined with a small-bore liquid chromatography system we have measured the basal and drug-induced fluxes of extracellular dopamine in the medial prefrontal cortex of chloral hydrate-anesthetized rats and have compared our findings in the cortex to that observed in the striatum. The results were as follows. (1) At a flow rate of 2 μ1/min, the basal level of dopamine in the medial prefrontal cortex was 0.28 ±0.1 ( n = 32) fmol/μl perfusate, which was nearly an order of magnitude less than that obtained from the striatum. (2) alpha-Methyl-para-tyrosine (150 mg/kg, i.v.) significantly decreased the extracellular levels of striatal and cortical dopamine. The magnitude and duration of the responses were similar in both regions. (3) Local perfusion with 30 mM K + had a more profound effect on dopamine release in the striatum than in the medial prefrontal cortex. The K + -induced release in both regions was significantly attenuated in the absence of Ca 2+ . (4) The anxiogenic beta carboline FG 7142 (15 mg/kg, i.p.) enhanced the release of cortical dopamine by about 50% while it was without an effect in the striatum. (5) Amphetamine (1 mg/kg, i.v.) significantly elevated, while reserpine (5 mg/kg, i.p.) rapidly attenuated, the dopamine level in the medial prefrontal cortex. These studies demonstrate that perfusion microdialysis, in conjunction with small-bore liquid chromatography with electrochemical detection, can be used to measure the basal release of dopamine in the rat medial prefrontal cortex and that the dopamine release process in this region, as has been shown in the striatum, is sensitive to stimulation conditions and pharmacological manipulations. The significance of variations in the pharmacological responsiveness of the dopamine released in the medial prefrontal cortex and striatum is discussed.

To investigate the development of denervation supersensitivity to serotonin (5-hydroxytryptamine, 5-HT) in the amygdala (AMYG) and the ventral lateral geniculate nucleus (vLGN), single cell recordings, microiontophoretic, histochemical and biochemical techniques were used in the present study. 5-HT projections to the vLGN and the AMYG were destroyed by 5,7-dihydroxytryptamine (5,7-DHT, a relatively selective toxin for 5-HT neurons) injected directly into the lateral ventricle or the ascending 5-HT pathway in the ventromedial tegmentum area. Enhanced responsiveness of cells to the inhibitory effect of microiontophoretically applied 5-HT (ionto-5-HT) began to develop within 24 h and approached a maximum 7 days after 5,7-DHT pretreatment. In general, the time courses for the reduction in both the density of 5-HT fluorescent varicosities and synaptosomal 5-HT uptake activity parallelled the time course for the development of denervation supersensitivity to 5-HT. During the first 2 days after 5,7-DHT, the enhanced sensitivity was selective for 5-HT; responses to d -lysergic acid diethylamide (LSD), norepinephrine (NE) and γ-aminobutyric acid (GABA) were unchanged. Seven or more days after 5,7-DHT there was a marked increase of the responsiveness of neurons in the vLGN and the AMYG to both 5-HT and LSD (a 5-HT agonist which is not a substrate for the high affinity 5-HT uptake system). At these later times, the responsiveness of cells in the AMYG to NE and to a lesser extent GABA was also increased. In contrast to the marked supersensitivity seen after 5,7-DHT induced denervation, chronic administration of parachlorophenylanine, a 5-HT synthesis inhibitor, failed to induce 5-HT supersensitivity.

Administration of the neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, to primates produces an excellent behavioral model of idiopathic Parkinson's disease. In the vervet monkey, regional biochemical differences in the striatum of two 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated groups were examined one to two months after treatment and compared with controls; one group displayed no observable gross motor abnormalities after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine treatment (asymptomatic), whereas the other group became markedly parkinsonian (symptomatic). In both 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated groups massive depletions of dopamine and homovanillic acid concentrations were observed in the striatum; generally, dopamine losses in the symptomatic group (> 95%) were greater than in the asymptomatic group (> 75%). However, in striatum, a marked heterogeneity in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine susceptibility was found; certain striatal regions having 99% depletion of dopamine even in asymptomatic monkeys. Overall, in ventromedial regions of striatum the losses of dopamine and homovanillic acid concentrations were less than in dorsolateral regions at the same coronal level. There was a significant negative correlation between control homovanillic acid/dopamine ratios and susceptibility of examined regions to 1-methyl-4-phenyl 1,2,3,6-tetrahydropyridine toxicity. Unlike idiopathic, but similar to postencephalitic, Parkinson's disease, dopamine and homovanillic acid levels in caudate nucleus were not spared relative to putamen; in fact, in the asymptomatic group caudate nucleus dopamine and homovanillic acid concentrations were depleted to a greater extent than in putamen. The relatively small, but distinct, differences in dopamine and homovanillic acid depletions between asymptomatic and symptomatic monkeys in some regions indicates the potential of reversing parkinsonian motor abnormalities with intrastriatal grafts or implants that produce relatively low amounts of dopamine.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) administration is able to produce nigrostriatal damage and motor disabilities in primates similar to those seen in Parkinson's disease. Two months after MPTP treatment in African Green monkeys, significant depletions of dopamine (DA) and/or homovanillic acid (HVA) were found in the dorsal ventral tegmental area, and septum, but not in the ventral part of the ventral tegmental area or nucleus accumbens. However, DA losses were greater at all examined sites in the striatum. In putamen and caudate nucleus the decreases in DA and HVA appeared more marked dorsolaterally than ventromedially. After MPTP treatment the ratio HVA/DA was elevated in the septum and all striatal regions; in the striatum the increases in ratio were greater in the dorsolateral than in the ventromedial samples. NE concentration was not significantly altered by MPTP in the mesolimbic system. In control animals the HVA concentration and the ratio HVA/DA were higher in the putamen than in the caudate nucleus. A longitudinal study showed that CSF HVA and 3-methoxy-4-hydroxyphenylglycol were reduced by MPTP and remained below baseline level for 12 months after MPTP treatment. This biochemical study indicates that in the monkey MPTP is able to induce selective damage within both the nigrostriatal and mesolimbic DA systems.

Following a systemic tryptophan load, striatal extracellular fluid levels of quinolinic acid in the rat were quantified using intracerebral microdialysis. After an intraperitoneal dose of L-tryptophan (250 mg/kg), quinolinic acid levels in striatal perfusates increased by 230 fold. Peak concentrations of quinolinic acid exceeded 10(-5)M, a concentration previously shown to be neurotoxic in vitro. These results indicate that quinolinic acid is markedly precursor responsive and that its concentration in striatal extracellular fluid may reach neurotoxic levels following an acute tryptophan load.

After treatment with the neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), there was a severe loss of dopamine (DA) at all examined sites in the striatum, both in those monkeys which remained asymptomatic (77–99%) and in one monkey which developed severe parkinsonian disability (94–99%). However, the asymptomatic animals had normal DA concentration in the substantia nigra (SN); yet in the symptomatic animal DA was largely depleted in the central (86%) and medial (94%), but not lateral (8%) regions of the SN. The HVA/DA ratio was raised in the striatum of all MPTP-treated animals. In the SN though, this ratio was elevated only in the symptomatic animal, in the central and medial, but not lateral regions. The contralateral half of these brains were examined for DA histofluorescence. The SN of asymptomatic animals had a slight increase in lipofuscin fluorescence within dopaminergic neurons and a small reduction in the number of dopaminergic cells, while fluorescent intensity of individual neurons was unchanged. The SN of the symptomatic animal displayed a sharp decline in the number of DA neurons along with an increase in autofluorescent pigment granules; these changes were most pronounced in the central and medial regions of the SN. These data suggest that after MPTP the terminals of the nigrostriatal pathway are affected before the cell bodies. In the one symptomatic animal emergence of parkinsonian disability corresponded with a marked loss of DA neurons and DA Concentration in the central and medial regions of the SN. In the control monkeys a gradient in the concentration of amines and metabolites was observed within the SN; the lateral region contained the highest and the medial region the lowest concentration.

Prolonged preganglionic stimulation of the isolated hypogastric nerveas deferens preparation resulted in a 3-fold increase in the amount of NA synthesized from exogenous tyrosine. This increase was not the result of an increased effector organ activity since the increase was still apparent even after blockade of the effector organ response with Hydergine®. These results indicate that the isolated vas deferens preparation is capable of synthesizing NA from exogenous tyrosine and that local NA synthesis is in some way regulated by nervous activity and is not a steady state phenomenon independent of impulse traffic.

Previous studies have shown that p-cholorophenylalanine (PCPA), an irreversible inhibitor of serotonin (5-HT) synthesis at the tryptophan hydroxylase step, fails to prevent the l-tryptophan-induced increase in the fluorescence of 5-HT-containing neuronal perikarya of the brain stem raphe nuclei, although there is a marked depletion in whole brain 5-HT concentration. In an effort to explain this apparent discrepancy, the effect of PCPA upon the histofluorescence of raphe cell terminals in the midbrain and forebrain of rats was examined. Demonstration of the fluorescence of raphe cell terminals was aided in part by pretreatment with l-tryptophan in the presence of a monoamine oxidase inhibitor. PCPA was found to produce a marked decrease in the fluorescence of raphe terminals but, in agreement with previous results, not in the perikarya. Parallel biochemical studies showed that 5-HT was depleted by PCPA (with or without l-tryptophan loading) to a much greater extent in the forebrain (a region rich in raphe terminals), than in a block of tissue containing the raphe cell perikarya. These results indicate that the synthesis of 5-HT is more susceptible to inhibition by PCPA in the terminals than in the perikarya of raphe neurons, perhaps due to a continued synthesis of new tryptophan hydroxylase enzyme in the latter site.

Fetal substantia nigra cells of two different gestational ages were successfully transplanted into the brains of three methylphenyltetra-hydropyridine-treated monkeys with severe parkinsonian motor and behavioural deficits. Functional improvement continued for 10 weeks after cell grafts into the striata of two monkeys with substantial numbers of tyrosine-hydroxylase-positive fetal neurons at necropsy. Behavioural improvement was correlated with increases in cerebrospinal fluid (CSF) homovanillic acid (HVA) concentrations after the transplants. A control monkey with inappropriately placed transplanted cells of an earlier gestational age remained severely parkinsonian and died during a similar period. CSF HVA fell slightly in this monkey from the low level seen before the transplants. Fetal dopamine neurons of two different gestational ages appear to survive transplantation in primates and have biochemical and functional effects.