Rescue of locomotor impairment in dopamine D2 receptor-deficient mice by an adenosine A2A receptor antagonist

Abstract

In Parkinson's disease a degeneration of dopaminergic neurons of the nigrostriatal pathway is observed. Loss of dopaminergic regulation of striatal neuron activity results in altered motor functions. Adenosine A2A (A2AR) and dopamine D2 (D2R) receptors are colocalized in striatal medium spiny neurons. It has been proposed that adenosine binding to A2AR lowers the affinity of dopamine for D2R, thus modulating the function of this receptor. Absence of D2R in knockout mice (D2R-/-) results in impaired locomotion and coordinated movements. This indicates that absence of dopamine in Parkinson's disease might principally affect D2R-mediated effects with regard to locomotor functions. A2AR-selective antagonists have been demonstrated to have anti- parkinsonian activities in various models of Parkinson's disease in rodents and nonhuman primates. In this article, D2R-/- mice were used to explore the possibility that an A2AR antagonist might reestablish their motor impairment. Interestingly, blockade of A2AR rescues the behavioral parameters altered in D2R-/- mice. In addition, the level of expression of enkephalin and substance P, which were altered in D2R-/-, were also reestablished to normal levels after A2AR antagonist treatment. These results show that A2AR and D2R have antagonistic and independent activities in controlling neuronal and motor functions in the basal ganglia. They also provide evidence that selective A2AR antagonists can exhibit their anti-parkinsonian activities through a nondopaminergic mechanism.

Fig. 1.

Expression of adenosine A2AR mRNA in striatum of WT, heterozygote, and D2R−/− mice. A,B, Adenosine A2AR in situ hybridizations on coronal sections from WT and D2R−/− mice (KO).C, Northern blot analysis of A2AR expression in striata from WT, heterozygote D2R+/− (Hetero), and D2R−/− (KO) mice. Values are presented as percentage of WT control.

Fig. 2.

Binding of [³H]CGS21680 to mouse striatal membranes. Scatchard analysis of the saturation binding experiment using [³H]CGS21680 in WT (▪) and D2R−/− (▴) mice. Maximum binding capacities (B_max) in fmol/mg protein = 1175 ± 141.4 for WT; 1233 ± 37.4 for D2R−/−; dissociation constant (K_d) in nm = 6.5 ± 0.6 for WT; 7.5 ± 0.6 for D2R−/−. Values are mean ± SEM of four independent experiments performed in duplicate.

Fig. 3.

Effect of KW-6002 treatment on striatal neuropeptide expressions. D2R−/− and WT mice were treated with KW-6002 (2.5 mg/kg) or vehicle. Mice were killed 3 hr after treatment. Neuropeptide mRNA expression levels in each treatment are expressed as percentage of control (WT treated with vehicle) ± SEM from 4–11 individuals in each group. **, p < 0.01 versus wild type with vehicle treatment. ^†,p < 0.05; ^†††,p < 0.001 versus D2R−/− mice with vehicle treatment by Student's t test.

Fig. 4.

Effect of KW-6002 on locomotor activities of WT and D2R−/− mice. Horizontal (A) and vertical (B) movement of WT and D2R−/− mice in 10 min. After 1 hr habituation, mice were administered vehicle or KW-6002 (2.5 mg/kg), and their locomotor activities were counted for 5 min. Values represent sum of 5 min counting at 0 and 15 min after administration. *, p < 0.05, **, p < 0.01, ***, p < 0.001 versus WT vehicle-treated,^†, p < 0.05, ^†††,p < 0.001 versus vehicle-treated D2R−/− mice; Student's t test.

Fig. 5.

Effects of KW-6002 on coordinate and spontaneous movements. Rotarod (A) and ring (B) tests were conducted 2.5 hr after administration with vehicle or KW-6002. The rod was rotated at eight turns per minute for a maximum of 3 min. The time that mice spent on the rod without falling down was measured, and the best performance among three trials was used for analysis. For the ring test, the duration ot time that mice did not show any voluntary movements on a 5.5-cm-diameter ring was measured. Values represent mean ± SEM fall latency or time immobile (in seconds). *, p < 0.05, **, p < 0.01, ***, p < 0.001 versus WT vehicle-treated mice. ^††,p < 0.01, ^†††,p < 0.001 versus vehicle treated D2R−/− mice by Student's t test.