Minocycline inhibits glial proliferation in the H-Tx rat model of congenital hydrocephalus

Abstract

Background: Reactive astrocytosis and microgliosis are important features of the pathophysiology of hydrocephalus, and persistent glial "scars" that form could exacerbate neuroinflammation, impair cerebral perfusion, impede neuronal regeneration, and alter biomechanical properties. The purpose of this study was to determine the efficacy of minocycline, an antibiotic known for its anti-inflammatory properties, to reduce gliosis in the H-Tx rat model of congenital hydrocephalus.

Methods: Minocycline (45 mg/kg/day i.p. in 5% sucrose at a concentration of 5-10 mg/ml) was administered to hydrocephalic H-Tx rats from postnatal day 15 to day 21, when ventriculomegaly had reached moderate to severe stages. Treated animals were compared to age-matched non-hydrocephalic and untreated hydrocephalic littermates. The cerebral cortex (both gray matter laminae and white matter) was processed for immunohistochemistry (glial fibrillary acidic protein, GFAP, for astrocytes and ionized calcium binding adaptor molecule, Iba-1, for microglia) and analyzed by qualitative and quantitative light microscopy.

Results: The mean number of GFAP-immunoreactive astrocytes was significantly higher in untreated hydrocephalic animals compared to both types of controls (p < 0.001). Minocycline treatment of hydrocephalic animals reduced the number of GFAP immunoreactive cells significantly (p < 0.001). Likewise, the mean number of Iba-1 immunoreactive microglia was significantly higher in untreated hydrocephalic animals compared to both types of controls (p < 0.001). Furthermore, no differences in the numbers of GFAP-positive astrocytes or Iba-1-positive microglia were noted between control animals receiving no minocycline and control animals receiving minocycline, suggesting that minocycline does not produce an effect under non-injury conditions. Additionally, in six out of nine regions sampled, hydrocephalic animals that received minocycline injections had significantly thicker cortices when compared to their untreated hydrocephalic littermates.

Conclusions: Overall, these data suggest that minocycline treatment is effective in reducing the gliosis that accompanies hydrocephalus, and thus may provide an added benefit when used as a supplement to ventricular shunting.

Figure 1

Astrocyte density (cells/μm³) in minocycline treated and untreated animals. The number of GFAP-immunoreactive cells was significantly higher (stars) in untreated hydrocephalic animals compared to both types of control (p < 0.001). Minocycline treatment significantly reduced the number of immunoreactive cells in hydrocephalic animals (p < 0.001). No significant differences were found between the control animals receiving minocycline and those that did not receive the drug, suggesting that minocycline has no effect on the normal brain. Data are mean ± SD, n = 5, cell counts were multiplied by 10⁵ to get the actual cell number per μm³.

Figure 2

Microglia density (cells/μm³) in minocycline treated and untreated animals. The number of immunoreactive microglia in untreated hydrocephalic animals was significantly higher than in the minocycline-treated hydrocephalic group (p < 0.001). Minocycline treatment had no effect on control brains. There was a statistically significant increase between both control groups compared to the untreated hydrocephalic animals (p < 0.001). Data are mean ± SD, n = 5, cell counts were multiplied by 10^-5 to get the actual cell number per per μm³.