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. 2008 Apr 8;105(14):5614-9.
doi: 10.1073/pnas.0801689105. Epub 2008 Apr 2.

Dicer loss in striatal neurons produces behavioral and neuroanatomical phenotypes in the absence of neurodegeneration

Trinna L Cuellar  1 , Tigwa H Davis, Peter T Nelson, Gabriel B Loeb, Brian D Harfe, Erik Ullian, Michael T McManus
Affiliations

Dicer loss in striatal neurons produces behavioral and neuroanatomical phenotypes in the absence of neurodegeneration

Trinna L Cuellar et al. Proc Natl Acad Sci U S A. .

Abstract

MicroRNAs (miRNAs) are small noncoding RNAs that can act to repress target mRNAs by suppressing translation and/or reducing mRNA stability. Although it is clear that miRNAs and Dicer, an RNase III enzyme that is central to the production of mature miRNAs, have a role in the early development of neurons, their roles in the postmitotic neuron in vivo are largely unknown. To determine the roles of Dicer in neurons, we ablated Dicer in dopaminoceptive neurons. Mice that have lost Dicer in these cells display a range of phenotypes including ataxia, front and hind limb clasping, reduced brain size, and smaller neurons. Surprisingly, dopaminoceptive neurons without Dicer survive over the life of the animal. The lack of profound cell death contrasts with other mouse models in which Dicer has been ablated. These studies highlight the complicated nature of Dicer ablation in the brain and provide a useful mouse model for studying dopaminoceptive neuron function.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Behavioral deficits and decreased lifespan in Dcrflox/flox;DR-1 cre mice. (A) Schematic of Dcr conditional targeting construct. (B) Clasping phenotype observed in Dcrflox/flox;DR-1 Cre animals. Dcrflox/wt;DR-1 Cre animals were used as controls for all experiments. (C) Kaplan-Meier survival curves of Dcrflox/flox;DR-1 Cre animals as compared with controls. Females have a median lifespan of 69 days (n = 22) and males have a median lifespan of 78 days (n = 21). (D) Body weights at death (between 10 and 12 weeks of age); Dcrflox/flox;DR-1 Cre animals exhibit wasting and loss of body mass as compared with controls. ***, P < 0.0001 for females (n = 13) and males (n = 14), Student's t test. SEM is shown. (E) Footprint analysis of Dcrflox/flox;DR-1 Cre animals reveals abnormal gait. (F) Reduced stride length in Dcrflox/flox;DR-1 Cre animals as compared with controls (n = 8; **, P = 0.0004, Student's t test). SEM is shown.
Fig. 2.
Fig. 2.
Reduced brain size and weight in Dcr flox/flox;DR-1 Cre mice. (A) A representative brain from a control Dcr flox/wt; DR-1 Cre mouse (Left) as compared with a brain from a Dcr flox/flox; DR-1 Cre animal (Right) (littermates, 10 weeks old). (B) Brain weights of dying Dcr flox/flox; DR-1 Cre animals (empty bars) and controls (filled bars) from males (n = 7) and females (n = 6). Student's t test, ***, P = 0.0004 and 0.0007, respectively, SEM is shown. Brains were obtained from animals between 10 and 12 weeks of age.
Fig. 3.
Fig. 3.
Reduced levels of miRNAs in the striatum of Dcr flox/flox; DR-1 Cre mice. (A) Northern blotting for miR-124a, miR-132, and miR-134 miRNAs in the striatum of Dcr flox/flox; DR-1 Cre mouse and a control mouse (lane 1, Dcr flox/wt; DR-1 Cre; lane 2, Dcr flox/flox; DR-1 Cre). RNA was obtained from 10- to 12-week-old mice. (B) Taqman qRT-PCR analysis for miR-124a, miR-132, and miR-134 in Dcr flox/flox; DR-1 Cre (empty bars) mice as compared with Dcrflox/wt; DR-1 Cre controls (filled bars). RNA was obtained from 10- to 12-week-old mice. (C) miR-124a LNAish in the striatum of a representative dying (10 weeks old) Dcr flox/flox; DR-1 Cre mouse and a control. (Scale bar: 100 μM.)
Fig. 4.
Fig. 4.
Astrogliosis in the striatum of Dcr flox/flox; DR-1 Cre mice. GFAP immunostaining from a representative Dcr flox/flox; DR-1 Cre mouse and control at 10 weeks of age (n = 4 for all genotypes). (Scale bars: 100 μM.)
Fig. 5.
Fig. 5.
NeuN staining of neurons in the striatum demonstrates smaller neuronal size but no loss of neurons in Dcr flox/flox; DR-1 Cre mice. (A) (Upper) Schematic of striatal regions stained for NeuN/DAPI. Regions 1–5 correspond to approximate Bregma coordinates: +074, +0.38, +0, −0.82, and −1.0 mm. For each coordinate, five images were taken representing most of the striatal region of each image, and all NeuN/DAPI cells were counted. (Lower) Average NeuN/DAPI-positive neurons counted per image in Dcr flox/wt; DR-1 Cre animals (filled bars) and Dcr flox/flox; DR-1 Cre animals (empty bars) for each coordinate shown above. (B) Representative images of NeuN/DAPI-stained sections from region 3 of a Dcr flox/flox; DR-1 Cre mouse and a control. n = 3 for all genotypes. (C) LacZ staining demonstrates no loss of DR-1 neurons in the striatum of Dcr flox/flox; DR-1 Cre animals, as compared with controls. (D) Average number of LacZ-positive neurons counted per image in Dcr flox/wt; DR-1 Cre animals (filled bar) and Dcr flox/flox; DR-1 Cre animals (empty bar). (Scale bars: B and C, 100 μM.)
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