Synaptic dysfunction implicated in leucine-rich repeat kinase-2 (LRRK2) R1441G knockin mice

Abstract

BACKGROUND: Physiological function of LRRK2 is unknown, but may be related to synaptic vesicle trafficking. We hypothesize that striatal DA uptake may gradually deteriorate with age, and LRRK2 mutations perturb DA turnover leading to increased oxidative stress and neurodegeneration. METHODS: We generated C57BL/6 mice with complete knockin (KI) of LRRK2 R1441G mutation. Locomotor activity, relative population of DA neurons and DA uptake in isolated striatal synaptosomes in young (3-month) and aged (18-month) KI were compared with wild-type (WT) littermates. Viability and morphology of DA neurons were examined by immunohistochemistry. mRNA expression in FACS-sorted DA neurons was determined by real-time PCR array. RESULTS: KI mice genotype was verified by sequencing. No phenotypic abnormality, motor deficit or dopaminergic neurodegeneration were observed up to 18 months of age. Expression of mut-LRRK2 in KI was similar to wt-LRRK2 in WT mice. However, synaptosomal DA uptake was significantly reduced in both aged WT and KI mice. Synaptosomes isolated from young KI mice exhibited lower DA uptake when treated with VMAT2 inhibitor, reserpine, compared with WT. Striatal protein expression of TH, DAT and VMAT2 were similar between KI and WT mice. CONCLUSIONS: Similar to R1441C (1), our R1441G KI mice show no obvious parkinsonian phenotype up to 18 months. However, mutant synaptosomes were more vulnerable to vesicular uptake inhibition by reserpine as shown by lower total DA uptake. As LRRK2 may regulate synaptic vesicle trafficking, mutant mice may be deficient in sequestrating cytosolic free DA into synaptic vesicles, leading to increased DA autooxidation and oxidative stress. Our data suggest that LRRK2 R1441G mutation may perturb DA turnover, resulting in synaptic dysfunction and neurodegeneration.