Species: Mouse
Genes: LRRK2
Mutations: LRRK2 G2019S
Modification: LRRK2: Transgenic
Disease Relevance: Parkinson's Disease
Strain Name: B6;C3-Tg(PDGFB-LRRK2*G2019S)340Djmo/J
Genetic Background: Transgene introduced into C57BL/6J x C3H/HeJ embryos. Founder mice were bred with C57BL/6J mice.
Availability: Available through The Jackson Laboratory, Stock# 016575, Cryopreserved.

Summary

These transgenic mice overexpress mutant human LRRK2 throughout the brain via a promoter that drives neuronal-specific expression (Ramonet et al., 2011). Nonetheless, the mutant human protein is also overexpressed in the lung and spleen (Kozina et al., 2018).

As hemizygous mice age, they develop neurodegeneration of dopaminergic neurons in the substantia nigra, as well as autophagic and mitochondrial abnormalities. The mice have a normal lifespan, but motor deficits and anxiety/depression-like behaviors begin to appear in middle age.

Hemizygous mice are viable, fertile, and produce normal numbers of progeny. There are no differences from non-transgenic littermates in body weight or survival (Ramonet et al., 2011). Mutant LRKK2 mRNA is expressed widely in the brain, with the highest expression in the olfactory bulb, cerebral cortex, hippocampus, striatum, and cerebellum. LRRK2 protein was found at levels about 2.7-fold higher than endogenous LRRK2 levels in tyrosine hydroxylase (TH)-positive dopaminergic neurons of the substantia nigra pars compacta. In whole brain samples from 2-month-old transgenic mice, LRRK2 is expressed at levels of 3- to 4-fold higher than non-transgenic animals (Nikonova et al., 2012).

Abnormalities in motor behavior surface with age. While young (2- to 4-month old) and middle-aged (10- to 12-month old) mice performed similarly to non-transgenic mice on the Rotarod, older animals (14- to 18-month old) stayed on only about half as long (Lim et al., 2018). Locomotor activity in the open-field test, however, appeared normal at both 6 and 15 months of age (Ramonet et al., 2011). In a different cohort from another study, consistent deficits (though non-significant until 16 months of age) were observed in Rotarod performance starting at 8 months of age (Palomo-Garo et al., 2016). Moreover, on the hanging wire test, which measures muscle strength, significant motor deficits appeared as early as 8 months of age.

As assessed by multiple behavioral tests—including a light-dark test, elevated plus maze, sucrose preference test, forced swimming test, and tail suspension test—LRRK2 G2019S mice develop anxiety/depression-like symptoms at 10 to 12 months of age (Lim et al., 2018). At the same time, serotonin levels declined in the hippocampus. Moreover, the expression of 5-HT1a receptors ramped up with age in the hippocampus, amygdala, and dorsal raphe nucleus.

At 1 to 2 months of age, LRRK2 G2019S mice display normal numbers of neurons in the substantia nigra, as assessed by TH staining and Nissl staining (Ramonet et al., 2011). However, as the mice age, these neurons degenerate. By 19-21 months of age, hemizygous mice exhibit about 18 percent of TH-positive dopaminergic neurons in the substantia nigra pars compacta. There is also a 14 percent reduction in dopaminergic dendrites in the substantia nigra pars reticulata. Others, however, have not observed differences in TH immunostaining in the substantia nigra between transgenic and wild-type mice either at 6, 12, or 18 months of age (Palomo-Garo et al., 2016) or at 2 years of age (Kozina et al., 2018). Neuronal numbers in the ventral tegmental area were normal (Ramonet et al., 2011) and the number of cerebellar Purkinje cells (based on immunostaining for calbindin) did not differ between transgenic and wild-type mice at 18 months of age (Palomo-Garo et al., 2016). Elevated levels of the anti-apoptotic proteins, nucleolin and heat-shock protein 70, were observed in whole brain samples (Jang et al., 2018).

Despite the loss of dopaminergic nigral neurons, the density of TH-positive dopaminergic nerve terminals in the striatum was comparable to the density in non-transgenic mice. Also, levels of striatal dopamine and its primary metabolites were normal even at relatively advanced ages (Ramonet et al., 2011). Specifically, at 14 to 15 months of age, LRRK2 G2019S mice had wildtype levels of dopamine, 3,4-dihydroxyphenylacetic acid (DOPAC), and homovanillic acid (HVA). In the olfactory bulb, HVA and DOPAC concentrations were low, but dopamine levels were equivalent to non-transgenic controls. Moreover, transgenic mice displayed normal pre-pulse inhibition of the acoustic startle reflex, a behavioral measure of sensorimotor gating that can be modulated in part by dopaminergic neurotransmission, at 6 and 15 months.

Transgenic mice are more susceptible than non-transgenic mice to toxicity by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and by lipopolysaccharide (LPS). Following MPTP treatment, transgenic mice (3 months of age) had a greater loss of TH-positive neurons in the substantia nigra pars compacta and a greater increase in GFAP immunoreactivity in the striatum compared with non-transgenic mice (Karuppagounder et al., 2016). In addition, MPTP treatment resulted in a greater reduction of striatal dopamine levels in transgenic mice than in non-transgenic animals. Following LPS treatment, transgenic mice exhibited a greater loss of TH-positive neurons in the substantia nigra pars compacta compared with wild-type controls, and this loss was selective to dopaminergic neurons as no loss of hippocampal glutamatergic pyramidal cells was observed following LPS treatment (Kozina et al., 2018).

These mice have been assessed for a variety of neuropathological markers of abnormal protein accumulation and inflammation. Even at advanced ages (23 to 24 months), they did not exhibit abnormalities in α-synuclein, ubiquitin, tau, or GFAP in the ventral midbrain, striatum, or cerebral cortex (Ramonet et al., 2011). However, one study found α-synuclein accumulation in whole brain lysates of 12- to 19-month-old transgenic mice compared to littermates (Ho et al., 2018; also see Ho et al., 2019). In addition, following hippocampal (CA1) injection of human wild-type tau (via AAV2/6), transgenic mice exhibited enhanced neuron-to-neuron transmission of tau protein compared with non-transgenic mice (Nguyen et al., 2018).

At 17-18 months of age, transgenic mice exhibited a significant increase in autophagic vacuoles (Ramonet et al., 2011). Moreover, in transgenic mice that received an injection of a proteosome inhibitor (an autophagy-inducing condition), accumulation of ubiquitinated proteins in the striatum was exacerbated compared to non-transgenic controls (Bang et al., 2016). Greater accumulation of α-synuclein and tau was also observed in transgenic versus non-transgenic under this condition. Together, this indicates that transgenic mice are more susceptible to protein aggregate accumulation. Another study demonstrated that 12- to 19-month-old transgenic mice with the G2019S mutation have increased phosphorylation of leucyl‐tRNA synthetase, which leads to increased levels of misfolded proteins and impaired autophagy (Ho et al., 2018). In addition, levels of LAMP-1, an endolysosomal protein, were increased in the striatum and substantia nigra of transgenic versus wild-type mice (Palomo-Garo et al., 2016).

Transgenic mice also show signs of accelerated cellular senescence based on increased levels of β-galactosidase and p21 in whole-brain lysates (Ho et al., 2019).

Markers of endocannabinoid signaling (CB2 receptor, and the FAAH and MAGL enzymes) largely did not differ based on quantitative reverse transcriptase PCR in transgenic versus wild-type mice at 6, 12, or 18 months of age in the brain regions assessed (caudate putamen, globus pallidus, and substantia nigra) (Palomo-Garo et al., 2016). 

At 17 to 18 months of age, transgenic mice showed abnormally condensed mitochondria in striatal neurons (Ramonet et al., 2011). Mitochondrial volume in striatal microglia was also reduced, and a shortening and reduction of microglial processes, typical of microglial activation, were also reported (Ho et al., 2018). However, one study found that total levels of Iba-1 (a microglial marker) did not differ between transgenic and wild-type mice in the striatum or substantia nigra at 6, 12, or 18 months of age (Palomo-Garo et al., 2016) and another found no differences in Iba-1 in the substantia nigra at 2 years of age (Kozina et al., 2018). Moreover, brain lysates of 4- to 6-month-old mice had increased levels of TNF-α and CD68, a marker of microglial activation. Another study confirmed that levels of both pro-TNFα and the active, soluble form of TNFα were increased in the brains of transgenic mice compared to control littermates (Ho et al., 2019). In contrast, a different study did not find differences in mRNA levels of TNFα, ALHD1, or COX-2 in the substantia nigra of 18-month-old transgenic mice versus wild-type controls (Palomo-Garo et al., 2016).

Modification Details

Transgenic mice overexpress full-length mutant human LRRK2 with the G2019S mutation. Transgene expression is driven by a hybrid CMVe-PDGFβ promoter, and the transgene is inserted in chromosome 3 (Kozina et al., 2018).

Related Strains

LRRK2 G2019S Mouse (Tg) x α-synuclein (A53T) mice -(see Daher et al., 2012).

Phenotype Characterization

Q&A with Model Creator

Q&A with Darren Moore

What would you say are the unique advantages of this model? 
The mice develop late-onset dopaminergic neuronal loss and the accumulation of autophagic vacuoles.

What do you think this model is best used for? 
Studying molecular and cellular disease mechanisms, and neuropathology.

What caveats are associated with this model? 
The transgene is driven by an ectopic promoter and therefore human LRRK2 expression may not follow an endogenous expression profile.

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References

Paper Citations

1. Ramonet D, Daher JP, Lin BM, Stafa K, Kim J, Banerjee R, Westerlund M, Pletnikova O, Glauser L, Yang L, Liu Y, Swing DA, Beal MF, Troncoso JC, McCaffery JM, Jenkins NA, Copeland NG, Galter D, Thomas B, Lee MK, Dawson TM, Dawson VL, Moore DJ. Dopaminergic neuronal loss, reduced neurite complexity and autophagic abnormalities in transgenic mice expressing G2019S mutant LRRK2. PLoS One. 2011;6(4):e18568. PubMed.
2. Kozina E, Sadasivan S, Jiao Y, Dou Y, Ma Z, Tan H, Kodali K, Shaw T, Peng J, Smeyne RJ. Mutant LRRK2 mediates peripheral and central immune responses leading to neurodegeneration in vivo. Brain. 2018 Jun 1;141(6):1753-1769. PubMed.
3. Nikonova EV, Xiong Y, Tanis KQ, Dawson VL, Vogel RL, Finney EM, Stone DJ, Reynolds IJ, Kern JT, Dawson TM. Transcriptional responses to loss or gain of function of the leucine-rich repeat kinase 2 (LRRK2) gene uncover biological processes modulated by LRRK2 activity. Hum Mol Genet. 2012 Jan 1;21(1):163-74. Epub 2011 Oct 4 PubMed.
4. Lim J, Bang Y, Choi JH, Han A, Kwon MS, Liu KH, Choi HJ. LRRK2 G2019S Induces Anxiety/Depression-like Behavior before the Onset of Motor Dysfunction with 5-HT1A Receptor Upregulation in Mice. J Neurosci. 2018 Feb 14;38(7):1611-1621. Epub 2018 Jan 5 PubMed.
5. Palomo-Garo C, Gómez-Gálvez Y, García C, Fernández-Ruiz J. Targeting the cannabinoid CB2 receptor to attenuate the progression of motor deficits in LRRK2-transgenic mice. Pharmacol Res. 2016 Aug;110:181-192. Epub 2016 Apr 6 PubMed.
6. Jang J, Oh H, Nam D, Seol W, Seo MK, Park SW, Kim HG, Seo H, Son I, Ho DH. Increase in anti-apoptotic molecules, nucleolin, and heat shock protein 70, against upregulated LRRK2 kinase activity. Anim Cells Syst (Seoul). 2018;22(5):273-280. Epub 2018 Sep 12 PubMed.
7. Karuppagounder SS, Xiong Y, Lee Y, Lawless MC, Kim D, Nordquist E, Martin I, Ge P, Brahmachari S, Jhaldiyal A, Kumar M, Andrabi SA, Dawson TM, Dawson VL. LRRK2 G2019S transgenic mice display increased susceptibility to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-mediated neurotoxicity. J Chem Neuroanat. 2016 Oct;76(Pt B):90-97. Epub 2016 Jan 22 PubMed.
8. Ho DH, Kim H, Nam D, Sim H, Kim J, Kim HG, Son I, Seol W. LRRK2 impairs autophagy by mediating phosphorylation of leucyl-tRNA synthetase. Cell Biochem Funct. 2018 Dec;36(8):431-442. Epub 2018 Nov 8 PubMed.
9. Ho DH, Seol W, Son I. Upregulation of the p53-p21 pathway by G2019S LRRK2 contributes to the cellular senescence and accumulation of α-synuclein. Cell Cycle. 2019 Feb;18(4):467-475. Epub 2019 Feb 6 PubMed.
10. Nguyen AP, Daniel G, Valdés P, Islam MS, Schneider BL, Moore DJ. G2019S LRRK2 enhances the neuronal transmission of tau in the mouse brain. Hum Mol Genet. 2018 Jan 1;27(1):120-134. PubMed. Correction.
11. Bang Y, Kim KS, Seol W, Choi HJ. LRRK2 interferes with aggresome formation for autophagic clearance. Mol Cell Neurosci. 2016 Sep;75:71-80. Epub 2016 Jun 28 PubMed.
12. Ho DH, Je AR, Lee H, Son I, Kweon HS, Kim HG, Seol W. LRRK2 Kinase Activity Induces Mitochondrial Fission in Microglia via Drp1 and Modulates Neuroinflammation. Exp Neurobiol. 2018 Jun;27(3):171-180. Epub 2018 Jun 30 PubMed.
13. Daher JP, Pletnikova O, Biskup S, Musso A, Gellhaar S, Galter D, Troncoso JC, Lee MK, Dawson TM, Dawson VL, Moore DJ. Neurodegenerative phenotypes in an A53T α-synuclein transgenic mouse model are independent of LRRK2. Hum Mol Genet. 2012 Jun 1;21(11):2420-31. PubMed.

External Citations

1. The Jackson Laboratory, Stock# 016575

Further Reading