Richard E. Wilcox

Tactile extinction: distinguishing between sensorimotor and motor asymmetries in rats with unilateral nigrostriatal damage.

Abstract We describe and demonstrate the usefulness of a reliable procedure for quantifying “sensorimotor” asymmetries in rats with nigrostriatal damage. Conventional tests, such as the popular Von Frey hair technique are sometimes inadequate in that positive responses are difficult to quantify, they are susceptible to subjective interpretation, and they require that the animal make head movements. Circling behavior has become established as a measure of asymmetrical nigrostriatal activity, but it does not measure stimulus-directed movement. One group of rats was given unilateral microinfusions of 6-OHDA into the nigrostriatal system which decreased levels of dopamine in the ipsilateral striatum and yieded an ipsilateral turning bias. Another group was given electrolytic lesions in the substantia nigra area, which yielded a severe contralateral circling bias. Small pieces of adhesive paper were applied to various parts of the limbs or snout and latencies to remove the stimuli were recorded. Because head and body movements were not required, assessment of stimulus-directed movement asymmetries could be quantified rapidly without the interference of stimulus-independent (“spontaneous”) postural and motor asymmetries. In the 6-OHDA-treated rats there was ipsilateral bias in stimulus-directed activity. Indeed, there was ipsilateral hyper-reactivity relative to sham controls. In rats with electrolytic nigra lesions, despite a strong contralateral “spontaneous” motor bias, there likewise was an ipsilateral bias in stimulus-directed movement, which could only be detected using the adhesive removal test. This similarity in stimulus-directed movement asymmetry between the two groups may reflect a common asymmetry in ascending dopaminergic pathways. The opposite circling bias following electrolytic nigra lesions may reflect differential damage to non-dopaminergic efferent projections. The bilateral adhesive removal (tactile extinction) test appears to permit the separate quantification of stimulus-directed and stimulus-independent movement asymmetries.

Posture-independent sensorimotor analysis of inter-hemispheric receptor asymmetries in neostriatum

Nigrostriatal dopaminergic neurons are thought to be critically important for somato-sensorimotor behavior. Following unilateral irreversible elimination of these neurons, an animal shows an ipsiversive postural bias and permanently fails to orient its head toward tactile stimuli placed on the contralateral side of the body. In response to apomorphine, a dopamine agonist, these rats display contraversive circling. This effect is thought to reflect denervation-induced proliferation of dopamine receptors in the ipsilateral striatum. We have developed a sensitive procedure that measures sensorimotor function independent of postural and circling biases. We record the latencies to remove small pieces of adhesive stimuli placed onto the snout or radial surface of the forelimbs. The stimuli are placed symmetrically and simultaneously, which is analogous to tactile-extinction procedures used clinically. In the first study we found that rats with unilateral 6-hydroxydopamine (6-OHDA)-induced lesions of the nigrostriatal pathway showed a contralateral sensorimotor bias in response to doses of apomorphine below those necessary to produce contraversive circling. In a second study, unilateral striatal microinjections of kainic acid (KA) were used to destroy the neurons on which the postsynaptic dopaminergic receptors of the nigrostriatal system are contained. Compared to 6-OHDA, KA produced unexpected results in standard orientation tests. None of the KA-treated rats showed contralateral neglect, and some even showed ipsilateral deficits. However, the standard orientation tests are confounded by postural asymmetries, which were irregular in the KA-treated group. Using again the posture-independent sensorimotor procedure, we found that all KA-treated rats, like the 6-OHDA-treated rats, uniformly displayed ipsilateral sensorimotor biases. Sensorimotor function relating to inter-striatal asymmetries may be more specifically assessed with the bilateral-adhesive tests.

Reactive capacity: A sensitive behavioral marker of movement initiation and nigrostriatal dopamine function.

Thirty-two Long Evans male rats with sham operations or unilateral 6-OHDA-induced damage to meso-telencephalic dopaminergic neurons were evaluated on a reactive capacity task that demanded high speed movement initiation. The task required lever manipulation to avoid signalled shock. The interval between the warning and the shock was incrementally reduced. A one-sleeved vest provided the opportunity to measure movement initiation of each limb independently. Extent of lesion was assessed by [3H]DA uptake, [3H]spiroperidol binding, or DA levels. Movement initiation latencies for each forelimb were found to be linearly related to interhemispheric striatal DA asymmetry induced by microinjections of 6-OHDA. Even those lesions resulting in small to moderate decreases in DA function, including deficits causing no chronic posture or sensory asymmetries, resulted in reactive capacity deficits and greatly slowed reaction time in the paw contralateral to the lesion. Following severe lesions, small yet substantial deficits were also seen in ipsilateral paw performance, which may be related to DA depletions found in the non-lesioned striatum. Thus, a reactive capacity task which requires the animal to react with maximal speed appears to be a potentially good index of nigrostriatal dopamine integrity even when the depletion is not severe.

Endurance training effects on striatal D2 dopamine receptor binding and striatal dopamine metabolites in presenescent older rats

Endurance training is associated with higher binding of 3H-spiperone to striatal D2 dopamine receptors of rats sacrificed 48 h following the last exercise bout (Gilliam et al. 1984). In the present study we investigated the effects of endurance training in presenescent older rats on the relationship between steady-state levels of DA and its metabolites in striatum versus the affinity and density of striatal D2 DA receptors. Citrate synthase activity of the gastrocnemius-plantaris muscle was 29.06±2.27 μmole/g wet wt in 21-month-old trained rats versus 22.88±1.13 μmole/g wet wt in 21-month-old untrained animals.DOPAC levels and DOPAC/DA ratios were greater in the old controls. Endurance training was associated with lower DOPAC levels in the 21-month-old animals. Thus, endurance training may postpone selectively changes in DA metabolism over a portion of the lifespan.As expected, the number of D2 DA binding sites was reduced with age (6 months Bmax:429±21 fmoles/mg protein; 21 months:355±20) with no change in affinity. The Bmax of old runners was significantly higher (457 ± 38 fmoles/mg protein) than that of old controls. Thus, endurance training appears to exert a protective effect on D2 dopamine receptors during the lifespan. Taken together, the present results suggest that there may be a possible reciprocal relationship between changes in DA metabolites and DA binding as a function of exercise in presenescent older rats, and that endurance training may decelerate the effects of age both on nigrostriatal dopamine neurons and on striatal D2 dopamine receptors during a portion of the lifespan.

The effects of exercise training on [3H]-spiperone binding in rat striatum

Thirty male Sprague-Dawley rats 100 days of age were divided into three groups: interval trained, endurance trained, and pair-weighted controls. Both trained groups ran up to one hour per day, 6 days per week for 12 weeks. The interval trained group ran up to 20 repeat intervals at 54 meters per minute for 30 seconds, while the endurance trained group ran at 27 meters per minute for 60 minutes. The animals were sacrificed, and the effects of aerobic training were documented by measuring cytochrome oxidase activity in the mixed quadriceps muscles. The cytochrome oxidase activity of the interval and endurance trained groups increased 49%, and 31% respectively, above the control group. [3H]-spiperone was used to label dopamine receptors in the striatum. The endurance group was not significantly different from the interval group in [3H]-spiperone receptor binding, so the two exercise groups were combined to form one group of runners. The runners had significantly higher [3H]-spiperone receptor binding than the controls, F(1,26) = 4.87, p less than 0.05. The mean and standard error for receptor binding was 89 +/- 13 fmoles/mg protein for the runners and 60 +/- 5 fmoles/mg protein for the controls.

Correlation of rates of calcium entry and endogenous dopamine release in mouse striatal synaptosomes

The time course of simultaneous Ca2+ entry and endogenous dopamine release was examined in mouse striatal synaptosomes depolarized by 30 mM KCl. Ca2+ entry and endogenous dopamine release exhibited fast and slow phase processes. The fastest rates occurred between 0 and 1 s. Ca2+ uptake and dopamine release dropped off quickly with 5-15 s rates at 13 and 10%, respectively, of the 0-1 s rate. Both processes were maintained at relatively high rates at the 1-3 and 3-5 s intervals suggesting mixed fast and slow phase processes. Uptake of Ca2+ and release of dopamine occurred in parallel over the entire 30 s measurement period; however, approximately 70% of the Ca2+ uptake and dopamine release occurred within the first 5 s following depolarization. A calculated ratio of Ca2+ entry versus dopamine release showed that approximately 1-2 Ca2+ ions were required to cause the release of one dopamine molecule. This ratio remained constant from 1 to 15 s following depolarization. Our results suggest that Ca2+ entry is coupled to endogenous dopamine release for both the fast and slow phase process.

Acute and chronic effects of barbiturates on depolarization-induced calcium influx into rat synaptosomes

Depolarization-induced 45Ca2+ influx into synaptosomes isolated from nontreated control and acutely treated rats (given 60 mg/kg phenobarbital i.p.) was significantly depressed (54 and 37%, respectively) by an in vitro challenge with pentobarbital, 0.3 mM (final concentration). However, depolarization-induced 45Ca2+ influx into synaptosomes isolated from tolerant rats (received dietary phenobarbital, 2.5 mg/g of diet, for 13 days) was not significantly altered when the synaptosomes were challenged with 0.3 mM pentobarbital. This suggests that synaptosomal membranes adapt during chronic exposure to barbiturates to allow for an enhanced Ca2+ influx subsequent to depolarization. Our data suggest that sedation may, at least in part, occur as a result of depressed stimulus-secretion coupling and that behavioral tolerance to sedation may occur because of the development of membrane tolerance to allow enhanced calcium influx.

Reaction time and nigrostriatal dopamine function: the effects of age and practice

Normal aged and Parkinsonian individuals lose the ability to initiate movements rapidly (increased reaction time) in parallel with changes in the nigrostriatal dopamine (DA) system. However, the ability of these individuals to improve their reaction time with practice has not been adequately assessed. We have developed a rodent model of human reaction time in which reaction time performance correlates highly with neurochemical measures of nigrostriatal DA integrity. In the present report, 15 young and 10 old male Sprague-Dawley rats were conditioned in a reaction time task to release a lever quickly in response to external stimuli in order to avoid a mild footshock. In order to examine the effects of practice on this reaction time task, the young animals were tested for 5 days at 3, 6 and 9 months of age and the old animals were tested for 5 days at 18, 21, and 24 months of age. From this well-practiced task, reaction time response latencies were measured and compared to measures of nigrostriatal DA function (steady-state levels of DA and its metabolites, D2DA receptor affinity and density). The old animals were slower in response latencies than the young animals. These age differences in response latencies, however, disappeared after several days of testing at each of the 3 test sessions, so that the old animals were not significantly slower than the young animals on days 4 and 5 of each session. As expected, the old animals showed reduced striatal D2DA receptor density with no age differences in DA receptor affinity.(ABSTRACT TRUNCATED AT 250 WORDS)

Dopamine receptor sensitivity after chronic dopamine agonists - Striatal 3H-spiroperidol binding in mice after chronic administration of high doses of apomorphine, N-n-propylnorapomorphine and dextroamphetamine

Several previous reports have demonstrated that chronic administration of both directly and indirectly acting dopamine agonists produces a supersensitive behavioral response to challenge doses of dopamine agonists when compared to the responses induced by acute administration of these drugs. That is, a given dose of a dopamine agonist will produce a greater response after chronic dopamine agonist treatment than is observed upon acute administration of that dose. A similar behavioral phenomenon resulting from chronic administration of dopamine antagonists has been suggested to be due to an increase in the number of dopamine receptors present in relevant brain areas. The same hypothesis has been put forward for the hypersensitivity induced by chronic dopamine agonist administration. The present study was designed to investigate the effect of chronic administration of high doses of both direct and indirect dopamine agonists on the dopamine receptors labeled by 3H-spiroperidol. Groups of animals (CD-1 mice) were sacrificed 1, 3 and 5 days following the last chronic injection. Striatal tissue from these mice was incubated with 3H-spiroperidol and dopamine receptor binding evaluated. Affinity of the receptors for the ligand was unaltered by treatments. The receptors labeled by 3H-spiroperidol showed no significant differences in number following the chronic administration of high doses of apomorphine (30 mg/kg). The Bmax was significantly decreased at only one time period following chronic administration of dextroamphetamine (4 mg/kg); however, these was a dramatic 30% reduction in the Bmax in striatal tissue from those mice treated with N-n-propylnorapomorphine. These results suggest that the hypersensitive behavioral response in mice following chronic administration of direct and indirect acting dopamine agonists is not due to an increase in the number of dopamine receptors in the striatum which are labeled by 3H-spiroperidol.

Apomorphine-induced stereotypic cage climbing in mice as a model for studying changes in dopamine receptor sensitivity.

We have previously confirmed in mice that apomorphine (APO) induces dopamine specific stereotypic cage climbing. Apparent changes in dopamine receptor sensitivity induced by chronic drug administration appear to be measurable by this technique. In the present experiments, murine stereotypic cage climbing was evauated as a model system for assessing the dopamine receptor supersensitivity induced by chronic administration of the potent butyrophenone neuroleptic spiroperidol. Spiroperidol induced a significantly enhanced response induced by APO (about a 7-fold increase) manifest by 48 hr (but not 24 hr) following cessation of the last chronic injection. Time-response analyses demonstrated that the action of test doses of APO (1.0 or 4.5 mg/kg, IP) was significantly prolonged in the chronic-spiroperidol animals relative to controls. The supersensitivity in the spiroperidol-treated animals lasted more than three weeks for each dose of the neuroleptic and the APO dose-response curve was shifted to the left in spiroperidol-treated animals. Results are discussed in terms of the utility of the model for establishing dose-response, time-course, and duration of effect data within the same group of animals.