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Forced use of the forelimb contralateral to a unilateral injection of the dopaminergic neurotoxin 6-hydroxydopamine can promote recovery of motor function in that limb and can significantly decrease damage to dopamine terminals. The present study was conducted to determine (1) whether a form of voluntary exercise, wheel running, would improve motor performance in rats with such lesions, and (2) whether any beneficial effects of wheel running are attributable to ameliorating the dopaminergic damage. In experiment 1, rats were allowed to run in exercise wheels or kept in home cages for 2 1/2 weeks, then given stereotaxic infusions of 6-hydroxydopamine into the left striatum. The rats were replaced into their original environments (wheels or home cages) for four additional weeks, and asymmetries in forelimb use were quantified at 3, 10, 17, and 24 days postoperatively. After killing, dopaminergic damage was assessed by both quantifying 3 beta-(4-iodophenyl)tropan-2 beta-carboxylic acid methyl ester ([(125)I]RTI-55) binding to striatal dopamine transporters and counting tyrosine hydroxylase-positive cells in the substantia nigra. Exercised 6-hydroxydopamine-infused rats showed improved motor outcomes relative to sedentary lesioned controls, effects that were most apparent at postoperative days 17 and 24. Despite this behavioral improvement, 6-hydroxydopamine-induced loss of striatal dopamine transporters and tyrosine hydroxylase-positive nigral cells in exercised and sedentary groups did not differ. Since prior studies suggested that forced limb use improves motor performance by sparing nigrostriatal dopaminergic neurons from 6-hydroxydopamine damage, experiment 2 used a combined regimen of forced plus voluntary wheel running. Again, we found that the motor performance of exercised rats improved more rapidly than that of sedentary controls, but that there were no differences between these groups in the damage produced by 6-hydroxydopamine. It appears that voluntary exercise can facilitate recovery from partial nigrostriatal injury, but it does so without evident sparing of dopamine nerve terminals.

Repeated systemic administration of moderate doses of methamphetamine (mAMPH) can result in neuronal damage. In addition to the prominent damage of forebrain dopamine and serotonin terminals, mAMPH also injures certain non-monoaminergic neuronal somata in the cerebral cortex. In previous studies, we have localized the damaged neurons to the "whisker barrels" in primary somatosensory cortex, reported the time course of their appearance, and found that sensory inputs from the mystacial vibrissae appear to play a crucial role in the mechanism of their injury by mAMPH. One common feature of these studies is that they used a single marker for neuronal injury, the fluorochrome dye Fluoro-Jade, which stains neurons injured by disparate mechanisms. Here we compare mAMPH-induced damage to somatosensory cortical neurons as assessed by Fluoro-Jade and immunohistochemical staining for phospho-c-Jun. A neurotoxic regimen of mAMPH induced phospho-c-Jun-positive neurons in both cortical whisker barrels and the substantia nigra. Neurons in the barrel cortex can be sufficiently damaged by mAMPH that they become Fluoro-Jade-positive within 2 hr after the final mAMPH injection. By contrast, phospho-c-Jun immunoreactivity does not appear until 12-24 hr after mAMPH. As reported in an earlier study, unilateral removal of vibrissae prior to mAMPH treatment affords partial protection from injury in the hemisphere contralateral to the vibrissotomy. The vibrissotomized animals show similar decreases in Fluoro-Jade staining and phospho-c-Jun immunoreactivity in the protected hemisphere. Since phospho-c-Jun indicates activation of Jun N-terminal kinase pathways, which have been implicated in apoptosis, we conclude that phospho-c-Jun provides a useful new marker for mAMPH-induced damage to cortical neurons.

Experiments measuring behavior and immediate-early gene expression in the basal ganglia can reveal interactions between dopamine (DA) and glutamate neurotransmission. Nigrostriatal DA projections influence two striatal efferent pathways that, in turn, directly and indirectly influence the activity of the substantia nigra pars reticulata (SNr). This report tests the interactions between striatal DA receptors and nigral glutamate receptors on basal ganglia function by examining both contralateral turning and Fos immunoreactivity in striatum and pallidum following unilateral intranigral microinfusions of glutamate antagonists given to intact and 6-OHDA-lesioned rats. The NMDA antagonist AP5 (1 microg), or the AMPA/kainate antagonist DNQX (0.015-1.5 microg), injected into the SNr (0.5 microl) elicited contralateral turning as well as both striatal and pallidal Fos expression. Moreover, intranigral DNQX elicited more turning and greater numbers of Fos-positive striatal neurons in 6-OHDA-lesioned animals than in unlesioned controls, suggesting that the 6-OHDA injection induces functional changes in nigral glutamate transmission. In 6-OHDA-lesioned rats, systemic injections of the DA D1 receptor agonist SKF38393 (0.5 mg/kg, i.p.) increased striatal Fos expression due to intranigral DNQX. In contrast, the D2 agonist quinpirole (0.1 mg/kg, i.p.) decreased striatal Fos expression but increased the pallidal Fos arising from intranigral AP5. In additional experiments, both intact and 6-OHDA-lesioned rats were given simultaneous intranigral and intrastriatal infusions and turning and pallidal Fos expression were measured. 6-OHDA-lesioned rats given 5 microg of intrastriatal quinpirole exhibited both turning and pallidal Fos that was significantly increased by intranigral AP5. These results indicate that the opposing influences of D2 agonists and endogenous nigral glutamate transmission are mediated by striatal D2 receptors. Finally, the behavioral effects of intranigral glutamate antagonism can be dissociated from the effects on striatal or pallidal immediate-early gene expression., (Copyright 2000 Wiley-Liss, Inc.)

Fos expression in the globus pallidus (GP) of rats was elicited by the D2 agonist quinpirole both ipsilateral and contralateral to a unilateral nigrostriatal 6-hydroxydopamine (6-OHDA) injection; however, the 6-OHDA-treated hemisphere was more sensitive to this effect. The quinpirole-induced GP Fos expression was antagonized in both hemispheres by the D2 antagonist eticlopride, but not by the D1 antagonist SCH 23390. In neurologically intact rats, the D1 agonist SKF 38393, which alone did not elicit pallidal Fos expression, augmented the quinpirole-induced Fos response. Thus, D1 agonists can synergize with D2 agonists in inducing GP c-fos; but the D2-stimulated induction does not depend on concurrent D1 agonism.

The mediation of behavior by nigral and striatal dopamine (DA) D1 and D2 receptors was investigated in rats that had sustained extensive unilateral 6-hydroxydopamine-induced injury to ascending DA neurons. Selective D1 and D2 agonists and antagonists were injected directly into the DA-denervated substantia nigra pars reticula or the caudate-putamen via a chronically indwelling cannula. Contralateral rotation resulting from unilateral stimulation of supersensitive DA receptors was quantified over 46 min. Intrastriatal apomorphine (5 micrograms) or the selective D2 agonist quinpirole (5 micrograms), but not the selective D1 agonist (+/-)-SKF 38393 (15 micrograms), induced vigorous rotation. The rotation induced by intrastriatal quinpirole was greatly diminished by systemic administration of the selective D2 antagonist eticlopride (0.5 mg/kg, i.p.) and could not be enhanced by additional injection of intrastriatal (+/-)-SKF 38393. Intranigral administration of apomorphine or (+/-)-SKF 38393, but not quinpirole (same doses as above), elicited vigorous rotation. However, the rotation induced by intranigral (+/-)-SKF 38393 could not be blocked by systemic administration of the selective D1 antagonist SCH 23390 (0.5 mg/kg, s.c.), and was mimicked by intranigral (-)-SKF 38393 (15 micrograms), which exhibits 100-fold less activity than the dextrorotatory enantiomer at the D1 receptor. In order to circumvent the problem of this drug's apparent non-D1-mediated action when injected intranigrally, rotation was induced by systemic (+/-)-SKF 38393 (2.0 mg/kg, i.p.) 10 min after intranigral administration of selective antagonists. Intranigral SCH 23390 (10 micrograms), but not eticlopride (10 micrograms), powerfully antagonized the rotation induced by systemic (+/-)-SKF 38393. Conversely, rotation induced by systemic quinpirole (0.5 mg/kg, i.p.) was potently blocked by intrastriatal eticlopride but not SCH 23390. Rotation induced by systemic apomorphine (0.25 mg/kg, i.p.) was not attenuated by either antagonist alone, regardless of intracerebral injection site. The results indicate that both nigral D1 and striatal D2 receptors mediate the behavioral effects of DA agonists. These data may be useful in elucidating the mechanism(s) underlying the D1/D2 synergism observed in neurologically intact animals, as well as in understanding the action of drugs used in the treatment of Parkinson's disease.

Rats given unilateral injections of 6-OHDA along the course of the mesotelencephalic dopaminergic projection show impairments in contralateral sensorimotor functions from which they often recover. Such rats also display an enhanced sensitivity to DA receptor stimulants, e.g. apomorphine, as revealed by contralateral turning, and an increased binding of neuroleptic compounds (e.g. [3H]spiroperidol) to the denervated striatum. This research examines the relationship of these receptor changes to both behavioral supersensitivity and recovery of sensorimotor functions by quantifying the time course of each phenomenon after injury. The supersensitivity to apomorphine and the behavioral recovery developed with a similar time course after injury, being evident within 1.5-3 days and reaching nearly maximal levels by 2 weeks postoperatively. A significant increase in in vivo [3H]spiroperidol binding to the denervated striatum occurred by 4 days postoperatively, and the magnitude of this change increased linearly during the first postoperative month. In contrast, the in vitro binding of this ligand to membranes of the denervated striatum was not increased until 3 weeks after the lesion. The results suggest that a proliferation of DA receptors may contribute to the pharmacological supersensitivity and the recovery of function, and that these early receptor changes may be revealed with greater sensitivity using in vivo binding techniques.

Unilateral damage to the mesotelencephalic dopaminergic projection of the rat produces impairments in sensorimotor functions, including an inability to localize contralateral somatosensory stimuli. Many rats with 6 hydroxydopamine injections along this pathway show a gradual improvement in somatosensory localization during the first post-operative month. One mechanism that may contribute to this behavioral recovery is the proliferation of dopamine receptor sites in the affected neostriatum. The role of these binding site changes in the recovery was tested by chronic administration of lithium to rats. Rats given lithium in their drinking water for 4 weeks after the 6-hydroxydopamine injection showed a greatly attenuated recovery of sensorimotor functions, compared to brain-damaged rats drinking unadulterated water. When rats that were given lithium for the first four postoperative weeks were subsequently given unadulterated water to drink, they recovered normally. Lithium treatment did not prevent the augmentation of [3H]spiroperidol binding in the neostriatum ipsilateral to the lesion, relative to the contralateral control neostriatum, at 4 or 8 weeks post-operatively. Thus, lithium treatment dissociates behavioral recovery after nigrostriatal injury from the lesion-induced relative proliferation of neostriatal [3H]spiroperidol binding sites. However, the lithium treatment decreased the absolute amount of specific [3H]spiroperidol binding to both the ipsilateral and contralateral neostriata. The significance of these findings for the neural mechanism underlying this recovery sequence is discussed.

The unilateral injection of 6-hydroxydopamine (8 micrograms) into the ventral tegmental area of awake rats produced a rapidly developing and irreversible sensory neglect to contralateral tactile stimuli. This neglect developed in a caudal to rostral direction on the affected body surface and coincided with significant elevation in the concentrations of dopamine and two of its metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in the ipsilateral neostriatum. The unilateral injection of procaine or gamma-hydroxybutyric acid (GHB) into the substantia nigra of awake animals also produced a contralateral neglect that developed in a caudal to rostral direction, but the behavioral effect of these drugs diminished within 1 hr. Concentrations of dopamine, dihydroxyphenylacetic acid and homovanillic acid in the neostriatum were markedly elevated during continuous infusions of procaine or gamma-hydroxybutyric acid. The extent of sensory neglect and changes in dopamine metabolism in the neostriatum varied according to the amount of gamma-hydroxybutyric acid injected into the nigra and according to the proximity of injections of gamma-hydroxybutyric acid to the pars compacta. The rapid onset of sensory neglect following microinjections of 6-hydroxydopamine, procaine or gamma-hydroxybutyric acid is consistent with the ability of each of these drugs to block the conduction of impulses in mesostriatal neurons and suggests that concomitant increases in levels of dopamine, dihydroxyphenylacetic acid and homovanillic acid in the neostriatum resulted from decreases in the release of dopamine coupled with increased synthesis of dopamine. These findings also indicate that the catabolism of dopamine to dihydroxyphenylacetic acid or homovanillic acid may originate intraneuronally, without prior release of dopamine and its recapture by mesostriatal terminals, if the flow of impulses in this pathway has been blocked.

Rats with unilateral 6-hydroxydopamine injections along the mesotelencephalic dopaminergic projection showed a profound impairment in localizing somatosensory stimuli on the contralateral body surface at 3 days postoperatively. Approximately one-half of the affected animals recovered the ability to localize tactile stimuli during 6 weeks postoperatively, whereas the remainder did not. When it occurred, the recovery of sensorimotor function began between the third and fifth day postoperatively and plateaued between days 14 and 21. Unilateral damage to these dopaminergic neurons resulted in hemispheric asymmetries of [14C]2-deoxyglucose incorporation at 3 days postoperatively. For structures that normally receive a dopaminergic innervation (e.g., neostriatum, nucleus accumbens septi, olfactory tubercle) the autoradiographic density of the injured side was decreased relative to the intact hemisphere. For structures that receive striatal inputs (globus pallidus, entopeduncular nucleus, substantia nigra pars reticulata), the autoradiographic density was increased on the side of the injury. This pattern of altered [14C]2-deoxyglucose incorporation was still present at 6 weeks postoperatively in animals that showed no recovery of somatosensory localization during that time. In contrast, rats that did recover showed no hemispheric asymmetries within the anterior neostriatum, globus pallidus, or substantia nigra pars reticulata at 6 weeks postoperatively, and the time course of normalization of metabolic activity in these structures was similar to that for behavioral restoration. These results directly demonstrate the importance of the neostriatum and particular structures efferent to it in the recovery of sensorimotor functions after striatal dopamine depletion. The types of neuronal plasticity within this basal ganglia circuitry responsible for the normalization of [14C]2-deoxyglucose incorporation and behavior are discussed.

The binding of [3H]spiroperidol to rat coronal sections in vitro was investigated using two procedures: swabbing studies, in which the tissue sections are wiped from the microscope slides after incubation in the presence of [3H]spiroperidol, and autoradiographic studies, in which the autoradiographic negatives are analyzed using computer-assisted densitometry. In the swabbing studies, the pharmacological and kinetic properties of butaclamol-displaceable binding were investigated, and the following results suggest that [3H]spiroperidol binds specifically to only a single site within the basal forebrain of tissue sections and that the site is the dopamine D-2 receptor. The pseudo-first order and first order plots for the rate of association to and dissociation from tissue sections appeared to be linear. Dopamine antagonists, such as haloperidol and butaclamol, were much more effective than dopamine agonists or the serotonin S-2 ligand, ketanserin, in inhibiting [3H]spiroperidol binding. The ability of dopamine agonists to inhibit [3H]spiroperidol binding was markedly reduced by the guanine nucleotide, Gpp(NH)p. Saturation analysis of specific [3H]spiroperidol binding revealed a Kd and Bmax of 0.93 nM and 447 fmol/mg protein, and a Hill coefficient of 1.05. The findings are also compatible with the possibility that [3H]spiroperidol binds to several sites that have identical affinities for this ligand. Densitometric studies were used to assess the effect of lesions on [3H]spiroperidol binding in the neostriatum. Intrastriatal injection of kainic acid substantially reduced 1 microM (+)--butaclamol-displaceable binding, indicating that the receptors are in large part on intrinsic striatal neurons. Neostriatal [3H]spiroperidol binding was investigated 7 days after destruction of the mesotelencephalic dopamine system by the ventral tegmental injection of 6-hydroxydopamine. As determined by saturation analysis, the average values for Kd and Bmax were 0.66 nM and 1212 fmol/mg protein in the intact striatum, and 0.82 nM and 1504 fmol/mg in the denervated striatum. The finding of a significant 23.8% increase in receptor density by the end of the first postoperative week, a period during which behavioral supersensitivity to apomorphine increases rapidly, supports the hypothesis that a proliferation of D-2 receptors underlies the behavioral manifestations of denervation supersensitivity.

Recent investigations of the function of the strionigral pathway have utilized the intranigral injection of gamma-aminobutyric acid (GABA) agonist and antagonist drugs. While the unilateral application of these substances typically produces rotational behavior, the direction of this turning (ipsilateral or contralateral to the injected hemisphere) differs in several reports. The present study determines whether the direction of this drug-induced turning depends upon the locus of migral stimulation. Picrotoxin and bicuculline methiodide were injected into either the pars compacta or the pars reticulata of the substantia nigra at several anterior-posterior levels. Injection of these drugs into the pars compacta resulted in ipsilateral turning while injection into the pars reticulata produced contralateral rotation. Both of these effects were dose-dependent and were elicited by similar threshold doses of picrotoxin. Prior 6-hydroxydopamine treatment abolished the ipsilateral but not the contralateral rotation. In contrast, muscimol injections produced contralateral turning independent of whether they were made into the pars compacta or pars reticulata. However, 6-hydroxydopamine treatment only attenuated the contralateral turning produced by pars compacta injections. These findings provide a histological basis for understanding the different types of turning behavior elicited by the intranigral injection of GABA agonists and antagonists. In addition, they suggest that GABA receptors mediate at least two independent actions in substantia nigra.

Alterations in neostriatal dopamine metabolism, release, and biosynthesis were determined 3, 5, or 18 days following partial, unilateral destruction of the rat nigrostriatal dopamine projection. Concentrations of dopamine and each of its metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and 3-methoxytyramine (3-MT) were markedly decreased in the lesioned striata at 3, 5, or 18 days postoperation. The decline in striatal high-affinity [3H]dopamine uptake closely matched the depletion of dopamine at 3 and 18 days postoperation. However, neither DOPAC, HVA, nor 3-MT concentrations were decreased to as great an extent as dopamine at any time following lesions that depleted the dopamine innervation of the striatum by greater than 80%. In these more severely lesioned animals, dopamine metabolism, estimated from the ratio of DOPAC or HVA to dopamine, was increased two- to four-fold in the injured hemisphere compared with the intact hemisphere. Dopamine release, estimated by the ratio of 3-MT to dopamine, was more increased, by five- to sixfold. Importantly, the HVA/dopamine, DOPAC/dopamine, and 3-MT/dopamine ratios did not differ between 3 and 18 days postlesioning. The rate of in vivo dopamine biosynthesis, as estimated by striatal DOPA accumulation following 3,4-dihydroxyphenylalanine (DOPA) decarboxylase inhibition with NSD 1015, was increased by 2.6- to 2.7-fold in the surviving dopamine terminals but again equally at 3 and 18 days postoperation. Thus, maximal increases in dopamine metabolism, release, and biosynthesis occur rapidly within neostriatal terminals that survive a lesion. This mobilization of dopaminergic function could contribute to the recovery from the behavioral deficits of partial denervation by increasing the availability of dopamine to neostriatal dopamine receptors. However, these presynaptic compensations are not sufficient to account for the protracted (at least 3-week) time course of sensorimotor recovery that has been observed following partial nigrostriatal lesion.