Species: Rat
Genes: Lrrk2
Modification: Lrrk2: Knock-Out
Disease Relevance: Parkinson's Disease
Strain Name: LEH-Lrrk2^tm1sage, HsdSage: LE-Lrrk^tm1sage, formerly known as TGRL4620
Genetic Background: Long Evans Hooded
Availability: Available through Inotiv, Live and cryopreserved.

Summary

Developed in collaboration with the Michael J. Fox Foundation, homozygous Lrrk2 knockout (KO) rats develop mild histological abnormalities in peripheral organs, most notably in the kidney, but also extending to the liver, lung, and spleen. These differences start early and are progressive, although they do not appear to shorten lifespan or adversely affect organ function. KO rats are partially protected from neurodegeneration in paradigms using intracranial injection of lipopolysaccharide (LPS) or α-synuclein (Daher et al., 2014). The vendor reports normal motor function at 12 months of age, as assessed by performance on the Rotarod (Inotiv, Sep 2023).

Homozygous Lrrk KO rats appear normal early on, with the exception of being larger and heavier than wild-type Long Evans rats by 1 month of age. This difference does not appear to be due to differences in food consumption (Ness et al., 2013; Baptista et al., 2013). Of note, weight differences between KOs and wild-type rats are more pronounced in males than in females (Gu et al., 2023). In homozygous rats, there is complete knockdown of the protein  (Daher et al., 2014).

Abnormalities in peripheral organs develop early. By 2 to 4 months of age, the kidneys appear larger and darker due to the presence of a brown pigment and eosinophilic hyaline droplets, as well as due to an accumulation of oxidatively modified hemoglobin and lipofuscin components (Boddu et al., 2015). The overall architecture of the kidney remains normal up to 12 months of age (Boddu et al., 2015). However, as the KO rats age, cells in the proximal tubular epithelium of the kidney develop cytoplasmic vacuolization and lysosomes proliferate in the tubules (Ness et al., 2013; Baptista et al., 2013). Indeed, ultrastructural differences have been observed between KO and wild-type rats in kidney glomeruli and tubular epithelial cells, but these differences depend on age, sex, and diet (Gu et al., 2023). In addition, basement membranes are thickened at 4, 8. and 12 months of age along with cylindrical structures in the urine called hyaline casts (Ness et al., 2013; Baptista et al., 2013). Researchers also find that KO kidneys, but not other organs measured, have macrophage accumulation (twofold higher than in wild-type rats) at 3 months of age, suggesting inflammation (Boddu et al., 2015).

Subtle changes in kidney function are detectable as early as 1 month of age. Urine analysis reveals abnormalities in urine specific gravity, total volume, urine potassium, creatinine, sodium, and chloride. Levels of di-docosahexaenoyl (22:6) bis(monoacylglycerol) phosphate (di-22:6-BMP), a biomarker of lysosomal dysregulation, are known to be decreased in KO rats (Andrew West, personal communication) and mice (Fuji et al., 2015). In one study, however, baseline serum creatinine levels did not differ compared to wild-type ratss at 3 months of age (Boddu et al., 2015). Another study did not find differences between wild-type and KO rats in serum creatinine, blood urea nitrogen, or renal safety biomarkers, but did find that KO rats had albuminuria at 3 to 4 months of age (Gu et al., 2023). Lrrk2 KO rats are protected against rhabdomyolysis, with KO kidneys showing less tubular damage and better renal function compared to wild-type rats after glycerol-inducing acute kidney injury (Boddu et al., 2015). Consistent with these findings, lipocalin-2 (NGAL) levels, a marker upregulated in kidney injury, are significantly lower in the both the urine and blood in Lrrk2 KO rats (Ness et al., 2013). Lrrk2 KO mice are likewise protected from both acute kidney injury and chronic kidney disease challenges (Zhang et al., 2023).

The liver, lung, and spleen are also affected. In the liver, hepatocytes show mild vacuolation along with accumulation of lipid droplets. Some liver-function tests were abnormal, including reduced liver transaminases and elevated sorbitol dehydrogenase. In the lung, Type II alveolar cells have greater numbers of lamellar bodies, which are secretory organelles that release pulmonary surfactant (Baptista et al., 2013; Miklavc et al., 2014). In the spleen, minor differences in cellular composition were noted at a young age (Ness et al., 2013).

Hematological abnormalities are also present. In general, Lrrk2 KO rats have lower red blood cell counts, hemoglobin levels, and hematocrit values, but these changes are mild, variable, and not phenotypically significant (Baptista et al., 2013). Circulating levels of some hormones are also reduced (e.g., adiponectin, adrenocorticotropic hormone, prolactin), whereas others are elevated (e.g., galanin, insulin, insulin-like growth factor, and luteinizing hormone). Blood cholesterol (primarily high-density lipoproteins) is increased. Taken together, these changes indicate Lrrk2 deficiency may lead to a variable and potentially insignificant mild metabolic hormonal dysregulation (Ness et al., 2013). 

Hypertension has also been observed in Lrrk2 KO rats. At 3 months of age, compared with wild-type rats, KO rats have higher diastolic, systolic, and mean arterial pressure as detected by tail cuff (Boddu et al., 2015). Heart rate is reduced in KO rats compared with wild-type rats (Boddu et al., 2015).

In the brain, stereological counts of tyrosine hydroxylase (TH)-positive neurons confirmed no difference in dopaminergic neuron numbers in the substantia nigra under basal conditions. Interestingly, Lrrk2 KO rats were resistant to dopaminergic neuron loss elicited by LPS or viral overexpression of α-synuclein. Specifically, when LPS was injected into the substantia nigra pars compacta of wild-type rats, it triggered a 40 percent loss of TH-positive neurons. However, Lrrk2 KO rats exhibited only about a 20 percent loss under the same conditions. Likewise, viral overexpression of α-synuclein in the substantia nigra results in a 30 percent loss of dopaminergic neurons in wild-type rats at 4, 8, and 12 weeks after viral injection. In contrast, no significant neuronal loss is observed in Lrrk2 KO rats. Prominent α-synuclein pathology is observed in both wild-type and KO rats following viral overexpression of α-synuclein (Daher et al., 2014).

The neuroprotection reported in Lrrk2 KO rats is associated with a decrease in the number of cells staining positive for CD68, a marker of activated microglia, macrophages, and monocytes. Wild-type rats, but not Lrrk2 KO rats, exposed to LPS or α-synuclein overexpression have high levels of CD68-positive cells in the substantia nigra. Many of these CD68-positive cells also expressed iNOS and Lrrk2 (Daher et al., 2014).

Neurotransmitter release has also been assessed by in vivo microdialysis in the striata of Lrrk2 KO rats (Creed et al., 2019). Basal levels of neurotransmitters (dopamine, glutamate, acetylcholine) and dopamine metabolites (3,4-dihydroxyphenylacetic and homovanillic acid) are not different from wild-type rats at 4, 8, and 12 months of age (Creed et al., 2019). However, the basal level of homovanillic acid decreases over time in Lrrk2 KO rats and is reduced in 12-month-old rats compared with 4- and 8-month-old rats (Creed et al., 2019). Evoked release of dopamine, glutamate, acetylcholine, GABA, and glycine does not differ between Lrrk2 KO and wild-type rats, nor do the release patterns change across ages in KO rats. However, evoked release of 5-hydroxyindoleacetic acid (a 5-HT metabolite) is decreased at 8 and 12 months of age compared with 4 months of age in KO but not in wild-type rats (Creed et al., 2019).

Synaptic vesicle endocytosis has been measured in primary striatal neuron cultures from Lrrk2 KO rats, and compared with wild-type rats, endocytosis is impaired, showing slower endocytic recovery (Arranz et al., 2015).

Modification Details

Zinc finger nuclease (ZFN) technology was used to generate a deletion of 10 base pairs in exon 30 of the rat Lrrk2 gene. This resulted in a frameshift and a premature stop codon in the same exon.

Phenotype Characterization

COMMENTS / QUESTIONS

No Available Comments

Make a comment or submit a question

To make a comment you must login or register.

References

Paper Citations

1. Daher JP, Volpicelli-Daley LA, Blackburn JP, Moehle MS, West AB. Abrogation of α-synuclein-mediated dopaminergic neurodegeneration in LRRK2-deficient rats. Proc Natl Acad Sci U S A. 2014 Jun 24;111(25):9289-94. Epub 2014 Jun 9 PubMed.
2. Ness D, Ren Z, Gardai S, Sharpnack D, Johnson VJ, Brennan RJ, Brigham EF, Olaharski AJ. Leucine-rich repeat kinase 2 (LRRK2)-deficient rats exhibit renal tubule injury and perturbations in metabolic and immunological homeostasis. PLoS One. 2013;8(6):e66164. Print 2013 PubMed.
3. Baptista MA, Dave KD, Frasier MA, Sherer TB, Greeley M, Beck MJ, Varsho JS, Parker GA, Moore C, Churchill MJ, Meshul CK, Fiske BK. Loss of leucine-rich repeat kinase 2 (LRRK2) in rats leads to progressive abnormal phenotypes in peripheral organs. PLoS One. 2013;8(11):e80705. Epub 2013 Nov 14 PubMed.
4. Gu YZ, Vlasakova K, Miller G, Gatto NT, Ciaccio PJ, Kuruvilla S, Besteman EG, Smith R, Reynolds SJ, Amin RP, Glaab WE, Wollenberg G, Lebron J, Sistare FD. Early-Onset albuminuria and Associated Renal Pathology in Leucine-Rich Repeat Kinase 2 Knockout Rats. Toxicol Pathol. 2023 Jan;51(1-2):15-26. Epub 2023 Apr 20 PubMed.
5. Boddu R, Hull TD, Bolisetty S, Hu X, Moehle MS, Daher JP, Kamal AI, Joseph R, George JF, Agarwal A, Curtis LM, West AB. Leucine-rich repeat kinase 2 deficiency is protective in rhabdomyolysis-induced kidney injury. Hum Mol Genet. 2015 Jul 15;24(14):4078-93. Epub 2015 Apr 22 PubMed.
6. Fuji RN, Flagella M, Baca M, Baptista MA, Brodbeck J, Chan BK, Fiske BK, Honigberg L, Jubb AM, Katavolos P, Lee DW, Lewin-Koh SC, Lin T, Liu X, Liu S, Lyssikatos JP, O'Mahony J, Reichelt M, Roose-Girma M, Sheng Z, Sherer T, Smith A, Solon M, Sweeney ZK, Tarrant J, Urkowitz A, Warming S, Yaylaoglu M, Zhang S, Zhu H, Estrada AA, Watts RJ. Effect of selective LRRK2 kinase inhibition on nonhuman primate lung. Sci Transl Med. 2015 Feb 4;7(273):273ra15. PubMed.
7. Zhang S, Qian S, Liu H, Xu D, Xia W, Duan H, Wang C, Yu S, Chen Y, Ji P, Wang S, Cui X, Wang Y, Shen H. LRRK2 aggravates kidney injury through promoting MFN2 degradation and abnormal mitochondrial integrity. Redox Biol. 2023 Oct;66:102860. Epub 2023 Aug 22 PubMed.
8. Miklavc P, Ehinger K, Thompson KE, Hobi N, Shimshek DR, Frick M. Surfactant secretion in LRRK2 knock-out rats: changes in lamellar body morphology and rate of exocytosis. PLoS One. 2014;9(1):e84926. Epub 2014 Jan 21 PubMed.
9. Creed RB, Menalled L, Casey B, Dave KD, Janssens HB, Veinbergs I, van der Hart M, Rassoulpour A, Goldberg MS. Basal and Evoked Neurotransmitter Levels in Parkin, DJ-1, PINK1 and LRRK2 Knockout Rat Striatum. Neuroscience. 2019 Jun 15;409:169-179. Epub 2019 Apr 25 PubMed.
10. Arranz AM, Delbroek L, Van Kolen K, Guimarães MR, Mandemakers W, Daneels G, Matta S, Calafate S, Shaban H, Baatsen P, De Bock PJ, Gevaert K, Vanden Berghe P, Verstreken P, De Strooper B, Moechars D. LRRK2 functions in synaptic vesicle endocytosis through a kinase-dependent mechanism. J Cell Sci. 2015 Feb 1;128(3):541–52. PubMed.

External Citations

1. Inotiv
2. I
3. notiv

Further Reading