Hypokinesia and Reduced Dopamine Levels in Zebrafish Lacking β- and γ1-Synucleins

Abstract

Background: Synucleins are implicated in Parkinson disease pathogenesis, but their functions are incompletely understood. Results: Zebrafish lacking ␤-and ␥1-synucleins showed hypokinesia and decreased dopamine levels. These abnormalities were rescued by human ␣-synuclein. Conclusion: Synucleins are necessary for spontaneous movement and dopaminergic function. Significance: A key functional role for synucleins in dopamine neurons may be relevant to Parkinson disease pathogenesis.

Different Mechanisms Regulate Expression of Zebrafish Myelin Protein Zero (P0) in Myelinating Oligodendrocytes and Its Induction following Axonal Injury

“…1B) was employed to map the 5Ј-end of the mpz mRNA transcript in adult zebrafish brain, using the tobacco acid pyrophosphatase (TAP) method and a gene-specific reverse primer complementary to sequence in exon 3. TAP Ϫ and RT-PCR controls were used as described previously (12,14) to verify that the boundary between the RACE adapter and the cDNA sequence occurred at the mRNA cap site. 5Ј-RACE generated a single major ϳ410-bp product, which was cloned; 12 randomly selected clones were sequenced.…”

“…For Northern blot, an mpz ORF probe template was made by PCR using primers 5Ј-ATG CTG TCC GTA CTG GCA CTG ACC-3Ј and 5Ј-CAC TAC ACC AAT AAT GGA GCC CGT-3Ј and ligated into pGEM-T. Sequence-verified plasmid clones were linearized, and digoxigenin-labeled antisense cRNA probes were synthesized by in vitro transcription, as described previously (12,14). Whole mount RNA in situ hybridization (12), RNA in situ hybridization to brain cryosections (12), and double label in situ hybridization/immunohistochemistry (15) were carried out exactly as reported in our previous work.…”

shRNA targeting α-synuclein prevents neurodegeneration in a Parkinson’s disease model

“…Furthermore, we did not detect downregulation of TH (the rate-limiting enzyme for dopamine biosynthesis) in nigral neurons to account for the change in striatal dopamine level. Reduced dopamine levels in the absence of neurodegeneration have been reported in several different synuclein knockout and knockdown models (31,45,60). Snca -/-mice showed a 15%-20% reduction in striatal dopamine, but no significant change in the ratio of dopamine to its metabolites (45); this reduction was more prominent (36%) in aged animals (31).…”

Quantification of larval zebrafish motor function in multiwell plates using open-source MATLAB applications

“…Evaluation of zebrafish larval movement is a key component of the analysis of phenotypes resulting from genetic mutations 1 , gene knockdown approaches 2 , chemogenetic lesions 3 , drugs 4 and toxins 5 that target nervous system development, structure or function. Abnormal motor responses can provide evidence regarding the functional importance of biochemical or cellular abnormalities.…”

“…The detection of more than one larval object within a well generates a “ too many objects ” error for an individual frame (the cumulative error report at the end of the assay is used as a quality control measure to ensure that a single larval object has been tracked throughout the assay in each well). However, the activity of multiple larvae within each well can be measured using pixel quantification instead of tracking 2 , since the quantification approach does not rely on identification of discrete larval objects. In this case, the centroid tracking function could potentially be removed from LSRtrack to make the program run faster.…”