These amiR-TDP43i (or CAG-RFP-amiR-TDP43i) transgenic mice have widespread expression of both red fluorescent protein (dsRed2) and an artificial miRNA (amiR-TDP43) that targets the mouse TAR DNA binding protein mRNA for knockdown. Expression is directed by the CMV-IE enhancer/chicken β-actin/rabbit β-globin hybrid promoter. amiR-TDP43 expression in the central nervous system and periphery (including brain [both neurons and glia], spinal cord, muscle, liver and other organs) results in a modest reduction (~10-20%) of TDP-43 expression in central nervous system homogenates. dsRed2 expression may be observed by direct/native fluorescence, by mRNA (in situ) hybridization, and by antibody staining/immunohistochemistry. DsRed2 excitation and emission maxima are 563nm and 582nm, respectively. In the cortex, dsRed2 immunofluorescence is observed in ~30-40% of neurons, astrocytes and oligodendrocytes, but not in microglia. As expected, no GFP expression is reported in brain or spinal cord.
amiR–TDP43i hemizygotes are viable with no gross physical abnormalities at birth. Hemizygous mice may begin to display progressive neurological defects around the time they are weaned, including head shaking/bobbing, hyperactivity (~4-5 weeks), weight loss (~5 weeks), circling, wobbly gait (~6-7 weeks), hypoactivity (~10 weeks), paralysis (~9-20 weeks) and death by 9-20 weeks. amiR–TDP43i hemizygotes display three end-stage phenotypes: hindlimb paralysis, sudden death, and whole-body paralysis. Muscle denervation is evident by electromyography shortly after weaning. Progressive motor neuron degeneration results in the end disease stage, at which there is ~25% loss of layer 5 cortical neurons and ~60% loss of lumbar spinal cord ventral horn ChAT+ motor neurons. Many transgenic mice exhibit heart enlargement. The average lifespan is slightly shorter for males than females, but the difference is not statistically significant. The donating investigator has not attempted to generate homozygous mice to date (December 2012), and reports transgenic males breed better than transgenic females. Additionally, because the phenotype may become more severe if the mice are backcrossed with FVB/NJ for two or more generations, the colony is maintained at The Jackson Laboratory Repository by breeding transgenic males with (C57BL/6J x FVB/NJ)F1 hybrid females (Stock No. 019019) every generation.
Of note, the neuromuscular-phenotype-free, cre-inducible parental strain used to generate the amiR-TDP43i transgenic mice is also available at The Jackson Laboratory as Stock No. 017919 (amiR-TDP43u line 6 or CAG-loxp-EGFP/3xpA-loxp-RFP-amiR-TDP43).
