Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2017 Mar;37(3):927-937.
doi: 10.1177/0271678X16649196. Epub 2016 Jul 20.

Cerebellar Neurodegeneration in a New Rat Model of Episodic Hepatic Encephalopathy

Affiliations
Free PMC article

Cerebellar Neurodegeneration in a New Rat Model of Episodic Hepatic Encephalopathy

Teresa García-Lezana et al. J Cereb Blood Flow Metab. .
Free PMC article

Abstract

Hepatic encephalopathy has traditionally been considered a reversible disorder. However, recent studies suggested that repeated episodes of hepatic encephalopathy cause persistent impairment leading to neuronal loss. The aims of our study were the development of a new animal model that reproduces the course of episodic hepatic encephalopathy and the identification of neurodegeneration evidences. Rats with portacaval anastomosis underwent simulated episodes of hepatic encephalopathy, triggered by the regular administration of ammonium acetate, and/or lipopolysaccharide. The neurological status was assessed and neuronal loss stereologically quantified in motor areas. During the simulated episodes, ammonia induced reversible motor impairment in portacaval anastomosis rats. In cerebellum, stereology showed a reduction in Purkinje cell population in portacaval anastomosis and PCA+NH3 groups and morphological changes. An increase in astrocyte size in PCA+NH3 group and activated microglia in groups treated with ammonium acetate and/or lipopolysaccharide was observed. A modulation of neurodegeneration-related genes and the presence of apoptosis in Bergmann glia were observed. This new animal model reproduces the clinical course of episodic hepatic encephalopathy when ammonia is the precipitant factor and demonstrates the existence of neuronal loss in cerebellum. The persistence of over-activated microglia and reactive astrocytes could participate in the apoptosis of Bergmann glia and therefore Purkinje cell degeneration.

Keywords: Apoptosis; astrocytes; inflammation; microglia; neurodegeneration.

Figures

Figure 1.
Figure 1.
Stereological analysis of dopaminergic neurons in SNpc and Purkinje neurons in cerebellum. (a) Relative number of dopaminergic neurons in SNpc (black bar) and the optical density of dopaminergic prolongations in striatum (grey bar). (b) Representative image of normal Purkinje neurons in PCA+NH3. (c) Representative image of Purkinje neurons with degenerative features (red arrows) in PCA+NH3. (d) Bar diagram represents the relative number of Purkinje neurons. Gray bar correspond to normal Purkinje neurons and black bar to degenerative cells (expressed in relation to the total number of Purkinje cells). *P ≤ 0.05 total number of Purkinje cells compared to sham. #P ≤ 0.05 normal Purkinje neurons compared to sham. PCA: portocaval anastomosis; LPS: lipopolysaccharide; SNpc: substantia nigra pars compacta; TH: tyrosine hydroxylase.
Figure 2.
Figure 2.
Bergmann glia apoptosis (a–g) representative images of cleaved-caspase-3 immunolabeling. Objective 20×. (a) Sham. (b) PCA. (c) PCA+NH3. (d) PCA+LPS. (e) PCA+NH3+LPS. (f) Representative image of cleaved-caspase-3 immunolabeling and Nissl counterstain, the double stain reveals cell death of Bergmann glia (BG; arrow) in Purkinje cell layer (P). Objective 60×. (g) Bar diagram represents mean intensity value of caspase-3 inmunostaining. *P ≤ 0.05 compared to sham. PCA: portacaval anastomosis; LPS: lipopolysaccharide.
Figure 3.
Figure 3.
Reactive astrocytes in cerebellum of episodic HE model (a–e) Representative images of GFAP immunohistochemistry. Objective 40×. (a) Sham. (b) PCA. (c) PCA+NH3 activated phenotype (arrow). (d) PCA+LPS. (e) PCA+NH3+LPS. (f) Quantification of GFAP stained area divided by the number of astrocytes per image, along whole cerebellum. *P ≤  0.05 compared to sham. PCA: portacaval anastomosis; LPS: lipopolysaccharide.
Figure 4.
Figure 4.
Microglial activation in cerebellum of episodic HE model (a–e) Representative images of Iba-1 immunohistochemistry. Objective 40×. (a) Sham. (b) PCA. (c) PCA+NH3 activated phenotype (arrow). (d) PCA+LPS activated phenotype (arrow). (e) PCA+NH3+LPS activated phenotype (arrow). (f) Quantification of Iba-1 stained area along whole cerebellum. *P ≤  0.05 compared to sham. PCA: portacaval anastomosis; LPS: lipopolysaccharide.
Figure 5.
Figure 5.
Western blot of total TTR in cerebellum. Total Ttr was quantified as the addition of monomers (14 kDa), tetramers (56 kDa), and intermediate forms (28 kDa; 42 kDa) and normalized by β-actin. Liver protein extract is used as positive control. Bar diagram represents total protein levels of three replicates. *P ≤ 0.05 compared to sham. PCA: portacaval anastomosis; LPS: lipopolysaccharide; TTR: transthyretin.

Similar articles

See all similar articles

Cited by 1 article

MeSH terms

Feedback