Changes in Striatal Medium Spiny Neuron Morphology Resulting from Dopamine Depletion Are Reversible

Abstract

The classical motor symptoms of Parkinson's disease (PD) are caused by degeneration of dopaminergic neurons in the substantia nigra, which is followed by secondary dendritic pruning and spine loss at striatal medium spiny neurons (MSN). We hypothesize that these morphological changes at MSN underlie at least in part long-term motor complications in PD patients. In order to define the potential benefits and limitations of dopamine substitution, we tested in a mouse model whether dendritic pruning and spine loss can be reversible when dopaminergic axon terminals regenerate. In order to induce degeneration of nigrostriatal dopaminergic neurons we used the toxicity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in C57BL/6J mice; 30 mg/kg MPTP was applied i.p. on five consecutive days. In order to assess the consequences of dopamine depletion, mice were analyzed 21 days after the last injection. In order to test reversibility of MSN changes we exploited the property of this model that striatal axon terminals regenerate by sprouting within 90 days and analyzed a second cohort 90 days after MPTP. Degeneration of dopaminergic neurons was confirmed by counting TH-positive neurons in the substantia nigra and by analyzing striatal catecholamines. Striatal catecholamine recovered 90 days after MPTP. MSN morphology was visualized by Golgi staining and quantified as total dendritic length, number of dendritic branch points, and density of dendritic spines. All morphological parameters of striatal MSN were reduced 21 days after MPTP. Statistical analysis indicated that dendritic pruning and the reduction of spine density represent two distinct responses to dopamine depletion. Ninety days after MPTP, all morphological changes recovered. Our findings demonstrate that morphological changes in striatal MSN resulting from dopamine depletion are reversible. They suggest that under optimal conditions, symptomatic dopaminergic therapy might be able to prevent maladaptive plasticity and long-term motor complications in PD patients.

Keywords: dendrite morphology; spine density; spiny projection neurons; striatum.

Figure 1

Dopaminergic phenotype in the MPTP model. (A) Schematic representation of the MPTP model. Mice received either MPTP or saline only (NaCl) on five consecutive days. They were analyzed either 21 days or 90 days after the last MPTP administration. Scale bars 200 µm. (B) Representative images of TH stained coronal midbrain sections of animals 21 days or 90 days after the last MPTP injection. The framed area of the Substantia nigra pars compacta (SNc) represents the lateral area analyzed by the stereological counting of dopaminergic neurons. (C) Number of TH-positive cells after 21 days. p-value is from unpaired t-test. (D) Number of TH-positive cells after 90 days. p-value is from unpaired t-test. When panels (C) and (D) were analyzed by two-way ANOVA (Df = 1 for both factors), we observed a significant difference for treatment (F = 30.05) but no significant difference for time point (F = 2.81). (E) Striatal dopamine concentration, measured by HPLC, is relative to the mean of the NaCl condition. Two-way ANOVA (Df = 1 for both factors) revealed a significant effect of treatment (F = 25.33) but not time point (F = 0.34). (F) Striatal concentration of dopamine metabolites, i.e., (DOPAC+HVA)/dopamine, relative to the mean of the NaCl condition. Two-way ANOVA (Df = 1 for both factors) revealed a significant effect of treatment (F = 31.74) but not time point (F = 2.58). Each marker represents one animal. Numbers of animals are in Table 1.

Figure 2

Dendritic arborization of striatal MSN 21 days after MPTP. (A) Representative images of Golgi stained MSN in the striatum analyzed 21 days after the last MPTP injection. Scale bars 50 μm. (B) Total dendritic length (TDL), (C) number of branch points (BP) per MSN, (D) segment length (SL = TDL/BP). Each marker represents one MSN. Numbers of MSN are in Table 1. p-values are from Mann–Whitney tests.

Figure 3

Spine density in MSN 21 days after MPTP. (A) Representative images of Golgi-stained MSN dendrites (arrowheads) 21 days after the last MPTP injection. Scale bars 10 μm. (B) Number of spines in a 10 µm stretch of dendrite. Each marker represents one MSN. Numbers of MSN are in Table 1. p-value is from Mann–Whitney test. (C) Illustration of the principal component analysis with size and color of the blue circles representing the importance (cos²) of the factor in that row for the dimension in that column.

Figure 4

Recovery of dendritic arborization 90 days after MPTP. (A) Representative images of Golgi stained MSN in the striatum analyzed 90 days after the last MPTP injection. Scale bars 50 μm. (B) Total dendritic length (TDL), (C) number of branch points (BP) per MSN, (D) segment length (SL = TDL/BP). Each marker represents one MSN. Numbers of MSN are in Table 1. p values are from Mann–Whitney tests.

Figure 5

Recovery of spine density 90 days after MPTP. (A) Representative images of Golgi stained MSN dendrites (arrowheads) 90 days after the last MPTP injection. Scale bars 10 μm. (B) Number of spines in a 10 µm stretch of dendrite. Each marker represents one MSN. Numbers of MSN are in Table 1. p value is from Mann–Whitney test.