Modeling cerebellar limb dysmetria and impaired spatial memory in rats using lamivudine: A preliminary study

Abstract

Background and aim: Neurodegeneration has been associated with the use of combination antiretroviral therapy (cART). This study is aimed at determining if any constituent of cART can induce cerebellar limb dysmetria and spatial memory impairments.

Materials and methods: Forty adult male Wistar rats were randomly grouped into four (n = 10): control (distilled water 0.5 mL); Tenofovir (6 mg/kg); Lamivudine (6 mg/kg) and Efavirenz (12 mg/kg). The following neurobehavioral studies were conducted: open field, beam walk, and Morris water maze. Immunohistochemistry of CD 68 and GFAP were used to test for neuroinflammation and neurodegeneration.

Results: There was marked increase in pyknotic pyramidal cells of the hippocampus and ghost Purkinje cells in the cerebellum of treatment groups. There was also a significant increase in oxidative stress in lamivudine and efavirenz groups. In addition, Lamivudine caused a significant increase of microglial and astrocytic activity (p < 0.001, 0.05 respectively) compared to control. The open field test showed a significant decrease (p < 0.0001) of the line crossing performance in the efavirenz, lamivudine and tenofovir (with means: 26.4, 4.6, 17.4 respectively) compared to control (50.6). There was also a significant decrease in the grooming (p < 0.05) and rearing (p < 0.01) in lamivudine group. Whereas, walk latency increased in efavirenz (p < 0.01), and lamivudine (p < 0.0001) compared to control. While hind limb slips significantly increased in efavirenz (p < 0.05) and lamivudine (p < 0.0001) compared with control group. Likewise, Lamivudine and Tenofovir exposed groups experienced a significant delay in the time to identify the hidden platform in compared to control (p < 0.05).

Conclusion: Lamivudine altered efferent stimuli along the cerebellospinal tracts thereby causing motor impairments. The degenerating Purkinje fibers may have induced marked neurodegeneration in the hippocampus resulting in impaired spatial memory.

Keywords: Antiretrovirals; Astrocytes; Ataxia; Cognition; Memory; Microglia; Movement disorders.

Fig 1,

Beam walk neurobehavioral test on 17mm and 28mm beams showing (a) Latency (time taken to cross from one point to the other, (b) Number of right hindlimb slips, (C) Number of left hindlimb slips, (d) open field test, (e) Morris water maze test. *p<0.05, **p<0.01, ***p<0.0001 against control. # p<0.05 between pre-treatment and post-treatment.

Fig 2,

Cerebellar Oxidative stress markers (a) Superoxide dismutase, (b) reduced glutathione, (c) Catalase, (d) Malondialdehyde. *p<0.05, **p<0.01, ***p<0.0001 against control.

Fig 3,

Oxidative stress markers of the hippocampus- (a) Superoxide dismutase, (b) Reduced glutathione, (c) Catalase, (d) Malondialdehyde. *p<0.05, **p<0.01, ***p<0.0001 against control.

Fig 4,

(a) Relative cerebellar weight, (b) Relative volume fraction of Purkinje cells. *p<0.05, **p<0.01, ***p<0.0001 against control.

Fig 5:

showing the hippocampus with H and E stain x40. The area is the CA3 region at x1000 showing marked pyknosis in treatment groups

Fig 6:

showing the hippocampus with Nissl stain x40. The CA3 region at x1000 showing less nissl substance in treatment groups compared to control

Fig 7:

showing the cerebellum with H and E stain x100 and x 1000. Showing high cell senescence among Purkinje cells in treatment groups (arrows pointing at ghost cells) compared to control

Fig 8:

showing the cerebellum with Giemsa stain x100 and x1000. Showing loss of Purkinje dendritic cells and chromatolysis in treatment groups compared to control

Fig 9:

Distribution of glial fibrillary acidic proteins (GFAP) positive astrocytes. Immunohistochemistry of GFAP in the cerebellum x1000, *p<0.05 against control.

Fig 10:

Distribution of CD68 positive microglial cells. Immunohistochemistry of CD 68 in the cerebellum x1000, **p<0.01 against control.