Inhibition of DAT function attenuates manganese accumulation in the globus pallidus

Abstract

Manganese (Mn) is an essential nutrient, though exposure to high concentrations may result in neurotoxicity characterized by alterations in dopamine neurobiology. To date, it remains elusive how and why Mn targets dopaminergic neurons although recently the role of the dopamine transporter has been suggested. Our primary goal of this study was to examine the potential roles of the monoamine transporters, dopamine transporter (DAT), serotonin transporter (SERT) and norepinephrine transporter (NET), in neuronal Mn transport. Using striatal synaptosomes, we found that only inhibition of DAT significantly decreased Mn accumulation. Furthermore, weanling rats chronically exposed to Mn, significantly accumulated Mn in several brain regions. However, rats receiving the specific DAT inhibitor GBR12909 (1 mg/kg bw, three times/week; four weeks) had significantly lower Mn levels only in the globus pallidus compared to saline-treated rats (p<0.05). Our data show that inhibition of DAT exclusively inhibits Mn accumulation in the globus pallidus during chronic exposure.

Figure 1

Experimental design of in vivo studies. Weanling male Sprague-Dawley rats were randomly placed into one of four dietary treatment groups: control (CN), iron-deficient (ID), control with Mn supplementation (via the drinking water; 1g Mn/L) (CNMn), and ID with Mn supplementation (IDMn). Two independent experiments with two separate sets of animals were conducted (n = 24 for each experiment; n = 48 total). Within each dietary group, half of the rats received intraperitoneal injections of GBR12909, 1 mg/kg (ED-50), three times per week while the other half received saline injections of equal volume. Dietary and pharmacological intervention lasted for four weeks.

Figure 2

Mean uptake of ³H-dopamine is shown for striatal synaptosomes incubated in the absence (gray) or presence (white) of biogenic amine transporter inhibitors for dopamine (DA), norepinephrine (NE) or serotonin (5HT) at the ED50 dose (1 μM GBR12909; 100 μM desipramine; 10 μM fluoxetine) for 5 minutes.

Figure 3

Manganese concentration [μg/L] is shown for striatal synaptosomes incubated in the absence or presence of 0.5 mM manganese for 5 minutes. Synaptosomes either received no treatment (gray), 1 μM GBR12909 (dotted), 100 μM desipramine (striped), or 10 μM fluoxetine (white). The mean concentration is shown with SEM. * p-value < 0.001

Figure 4

Manganese concentration [nmol/g tissue] is shown for all brain regions: globus pallidus (GP); caudate putamen (CP); hippocampus (HC); substantia nigra (SN); and cerebellum (CB). Diets were CN (gray), CNMn (dotted), ID (striped), and IDMn (white). Mn supplementation led to a significant increase in brain regional Mn concentrations as indicated by the asterisk. The mean concentration is shown with SEM. * p-value < 0.05

Figure 5

Manganese concentrations [nmol/g tissue] are shown for CN and CNMn groups with and without 1 mg/kg GBR for three representative brain regions: globus pallidus (GP), caudate putamen (CP), and hippocampus (HC). Diets were CN (gray), CNMn (dotted), ID (striped), and IDMn (white). The mean concentration is shown with SEM. * p-value < 0.001; † p-value = 0.07

Figure 6

Iron concentration [nmol/g tissue] is shown for all brain regions: globus pallidus (GP); caudate putamen (CP); hippocampus (HC); substantia nigra (SN); and cerebellum (CB). Diets were CN (gray), CNMn (dotted), ID (striped), and IDMn (white). The mean concentration is shown with SEM. † p-value = 0.13