Dysregulation of striatal dopamine release in a mouse model of dystonia

Abstract

Dystonia is a neurological disorder characterized by involuntary movements. We examined striatal dopamine (DA) function in hyperactive transgenic (Tg) mice generated as a model of dystonia. Evoked extracellular DA concentration was monitored with carbon-fiber microelectrodes and fast-scan cyclic voltammetry in striatal slices from non-Tg mice, Tg mice with a positive motor phenotype, and phenotype-negative Tg littermates. Peak single-pulse evoked extracellular DA concentration was significantly lower in phenotype-positive mice than in non-Tg or phenotype-negative mice, but indistinguishable between non-Tg and phenotype-negative mice. Phenotype-positive mice also had higher functional D2 DA autoreceptor sensitivity than non-Tg mice, which would be consistent with lower extracellular DA concentration in vivo. Multiple-pulse (phasic) stimulation (five pulses, 10-100 Hz) revealed an enhanced frequency dependence of evoked DA release in phenotype-positive versus non-Tg or phenotype-negative mice, which was exacerbated when extracellular Ca(2+) concentration was lowered. Enhanced sensitivity to phasic stimulation in phenotype-positive mice was reminiscent of the pattern seen with antagonism of nicotinic acetylcholine receptors. Consistent with a role for altered cholinergic regulation, the difference in phasic responsiveness among groups was lost when nicotinic receptors were blocked by mecamylamine. Together, these data implicate compromised DA release regulation, possibly from cholinergic dysfunction, in the motor symptoms of this dystonia model.

Fig.1. Single-pulse (1 p) or five-pulse (5 p) evoked DA release in the striatum is lower in Phe(+) than Phe(−) or non-Tg mice

A. Representative 1 p evoked [DA]_o recorded in striatal slices from non-Tg mice, Phe(−) and Phe(+) mice in 2.4 mM [Ca²⁺]_o. B. Mean 1 p evoked [DA]_o for each group (*p < 0.05 non-Tg vs. Phe(+) mice; ^#p < 0.05 Phe(−) vs. Phe(+) mice; n = 49-52 sites per group). C. Time-course of 1 p evoked [DA]_o in striatal slices from non-Tg, Phe(−) and Phe(+) mice, with peak evoked [DA]_o taken as 100% and error bars omitted for clarity. The falling phase of averaged 1 p release records were indistinguishable indicating similar DA clearance (p > 0.05, n = 5 each). D. Mean 5 p evoked [DA]_o at 10, 25 or 100 Hz in each group of mice (*p < 0.05 non-Tg vs. Phe(+) mice; ^#p < 0.05 Phe(−) vs. Phe(+) mice; n = 49-52 sites per group).

Fig. 2. D2 receptor regulation of striatal DA release is enhanced in Phe(+) mice

A.Representative records of 1 p evoked [DA]_o in a non-Tg mouse under control conditions and in increasing concentrations of the D2 receptor agonist, quinpirole (1 nM to 10 μM). B. Mean concentration-response curves for the inhibition of DA release by quinpirole in non-Tg, Phe(−) and Phe(+) mice. Values are expressed as % inhibition of DA release (mean ± SEM, n = 5 for each group) against log concentrations of quinpirole; dashed lines indicate the quinpirole concentration at which peak 1 p evoked [DA]_o was inhibited by 50%. Actual IC₅₀ and I_max values for each group (see Table 1) were calculated from fitting one-component sigmoidal curves to concentration-response data from individual experiments.

Fig. 3. Enhanced responsiveness of striatal DA release in Phe(+) mice to phasic versus tonic stimulation is Ca²⁺ dependent

A. Mean evoked [DA]_o elicited by 5 pulse (5 p) stimulus trains at 10, 25 and 100 Hz in striatal slices from each group, normalized to 1 p evoked [DA]_o in 2.4 mM [Ca²⁺]_o (n = 49-52 sites). B. Mean evoked [DA]_o elicited by 5 pulse (5 p) stimulus trains at 10, 25 and 100 Hz in striatal slices from each group, normalized to 1 p evoked [DA]_o in 1.5 mM [Ca²⁺]_o (n = 24-27). Significance of differences among the groups for each frequency in either A) 2.4 mM [Ca²⁺]_o or B) 1.5 mM [Ca²⁺]_o is indicated by *p < 0.05, **p < 0.01, ***p < 0.001 for non-Tg vs. Phe(+) or Phe(−) mice; ^#p < 0.05, ^##p < 0.01, ^###p < 0.001 for Phe(−) vs. Phe(+) mice (one- and two-way ANOVA for repeated measures with Bonferoni post hoc tests). C. Lowering [Ca²⁺]_o from 2.4 to 1.5 mM selectively amplifies the phasic (5 p) to tonic (1 p) ratio of evoked [DA]_o at 10, 25, and 100 Hz in Phe(+) mice (**p < 0.01 for 1.5 mM [Ca²⁺]_o vs. same site response in 2.4 mM [Ca²⁺]_o; n = 27-49 sites per point). No Ca²⁺ dependence of 5 p to 1 p ratio was seen in non-Tg or Phe(−) mice (n = 24-52 sites per group) (two-way ANOVA for all comparisons).

Fig. 4. Differences in phasic versus tonic DA release between Phe(+) and non-Tg or Phe(−) mice are lost when nAChRs are blocked

A. Representative 1 p and 5 p evoked [DA]_o recorded in striatal slices from non-Tg, Phe(−) and Phe(+) mice in the presence of mecamylamine (10 μM), an antagonist of nAChRs. B. Mean 1 p or 5 p evoked [DA]_o in mecamylamine in each group of mice (*p < 0.05 vs. non-Tg mice; ^#p < 0.05 Phe(−) vs. Phe(+); n = 17-22 per group). C. Ratio of 5 p to 1 p evoked [DA]_o under control conditions showing enhanced sensitivity to phasic simulation in Phe(+) mice (*p < 0.05, ***p < 0.001 vs. non-Tg mice; ^#p < 0.05, ^###p < 0.001 vs. Phe(−) mice (n = 17-22). D. Differences among the three groups were lost in mecamylamine (p > 0.05 for all comparisons; two-way ANOVA).