Elsevier

Brain Research

Volume 904, Issue 2, 22 June 2001, Pages 208-217
Brain Research

Research report
Reactive nitrogen species contribute to blood–labyrinth barrier disruption in suppurative labyrinthitis complicating experimental pneumococcal meningitis in the rat

https://doi.org/10.1016/S0006-8993(01)02164-3Get rights and content

Abstract

Sensorineural hearing damage is a frequent complication of bacterial meningitis, affecting as many as 30% of survivors of pneumococcal meningitis. There is a substantial body of evidence that oxidants, such as reactive nitrogen species (RNS), are central mediators of brain damage in experimental bacterial meningitis. In the present study, we investigated whether RNS also contribute to the pathophysiology of suppurative labyrinthitis in our well-established rat model of pneumococcal meningitis. In all infected rats, but not in uninfected controls, we observed suppurative labyrinthitis. Cochlear inflammation was accompanied by severe blood–labyrinth barrier (BLB) disruption as evidenced by increased Evans Blue extravasation. Furthermore, increased cochlear expression of endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS) was detected by immunohistochemistry. Colocalization of iNOS and tyrosine nitration (a marker of RNS attack) indicated that nitric oxide (NO) produced by iNOS contributes to oxidative cochlear damage through the action of RNS. To determine the pathophysiological role of RNS in BLB disruption, rats were treated with peroxynitrite scavengers (MnTBAP and uric acid, UA). Six h after adjunctive treatment with 300 mg/kg i.p. UA or 15 mg/kg i.p. MnTBAP+100 mg/kg i.p. ceftriaxone, BLB disruption was significantly reduced compared with that in infected animals treated only with ceftriaxone. Therefore, we conclude that RNS are involved in the breaching of the BLB during meningogenic pneumococcal labyrinthitis.

Introduction

The mortality of meningitis due to Streptococcus (S.) pneumoniae is 20–30% [11], [42], [43], and up to one-third of survivors are left with significant neurologic sequelae, most commonly hearing impairment, mental retardation, and focal neurologic deficits [7], [39], [44]. The frequency of hearing impairment varies with the causative pathogen: in a prospective study of 185 children with acute bacterial meningitis, the incidence of hearing loss was 31.5% with S. pneumoniae, 10.5% with Neisseria meningitidis, and 6% with Haemophilus (H.) influenzae infections [10]. The high incidence of hearing impairment after pneumococcal meningitis was confirmed by a more recent prospective study of 180 children with pneumococcal meningitis, which showed moderate to severe hearing loss in 32% of the cases [3]. Hearing loss during bacterial meningitis is sensorineural, unilateral or bilateral, often severe, and permanent [3], [10], [36]. The site of the lesion probably is the cochlea [35], [37]. Electron microscopic histopathology showed damage to the organ of Corti (involving the hair cells, supporting cells, and nerve terminals) and the stria vascularis during experimental meningogenic bacterial labyrinthitis in the rabbit [37], [40]. Pathophysiological studies have shown that NO contributes to hearing damage during cochlear inflammation induced by microperfusion with pneumolysin [1], [2]. However, the exact pathophysiological mechanisms of cochlear damage in suppurative labyrinthitis are still largely unknown. In a well-established adult rat model of pneumococcal meningitis, we recently identified peroxynitrite as a mediator of brain damage [23]. Nitrotyrosine residues on proteins (a marker of peroxynitrite) were detected immunohistochemically in the brain of rats with pneumococcal meningitis, and pretreatment with the peroxynitrite scavenger uric acid (UA) attenuated meningeal inflammation, blood–brain barrier (BBB) disruption, and increased intracranial pressure. In this study, we investigated the integrity of the blood–labyrinth barrier (BLB) during cochlear inflammation in our rat model of pneumococcal meningitis and conducted an immunohistochemical study of the presence of eNOS, iNOS, and tyrosine nitration in cochlear inflammation. In order to determine if RNS contribute to BLB disruption, we evaluated the effect of two different peroxynitrite scavengers (UA and MnTBAP) on cochlear Evans Blue extravasation.

Section snippets

Rat model of pneumococcal meningitis

Adult male Wistar rats (270–350 g) were anesthetized with halothane (Hoechst AG, Frankfurt, Germany), and meningitis was induced by transcutaneous intracisternal injection of 150 μl 106 colony forming units ml−1 of Streptococcus pneumoniae type 3 [29]. Controls were injected with 150 μl sterile phosphate-buffered saline (PBS, pH 7.4, n=4). Each rat was put into an individual cage, allowed to wake up, and fed with a standard diet and water ad libitum. Twenty h after infection, clinical signs of

Histological characterization of labyrinthitis

Cochlear inflammation was present in all infected rats, but in none of the uninfected control animals. All ears of the infected animals showed stonger inflammation in the tympanic scale than in the vestibular scale. Within the tympanic scale, inflammation was strongest in the first basal turn of the cochlea. In all ears from infected animals, in which the cochlear aqueduct was discernible, it was infiltrated by inflammatory cells. Inflammatory cells were not observed in the scala media in any

Discussion

The main findings of this study are: (a) labyrinthitis is a regular complication in our rat model of pneumococcal meningitis, (b) labyrinthitis is accompanied by a disruption of the BLB, (c) eNOS and iNOS are possible sources of NO during cochlear inflammation and possibly contribute to oxidative cochlear damage through the formation of RNS, and (d) BLB disruption is attenuated by the peroxynitrite scavengers UA and MnTBAP.

The route of infection of the inner ear during bacterial meningitis is

Acknowledgements

We thank Dr. V. Arbusow and Dr. P. Schulz (Department of Neurology) for their advice on dissection and handling of rat cochleae. We also thank Ms. S. Walter for technical assistance and Ms. J. Benson for copyediting the manuscript. The financial support of the Volkswagenstiftung (AZ I/75 119) and of the Graduiertenkolleg (Infektion und Immunität) of the Deutsche Forschungsgemeinschaft to HWP is appreciated gratefully.

References (11)

  • K.M. Boje

    Inhibition of nitric oxide synthase attenuates blood–brain barrier disruption during experimental meningitis

    Brain Res.

    (1996)
  • F.R. Amaee et al.

    N-methyl-l-arginine protects the guinea pig cochlea from the cytotoxic effects of pneumolysin

    Acta Otolaryngol. Stockh.

    (1995)
  • F.R. Amaee et al.

    Possible involvement of nitric oxide in the sensorineural hearing loss of bacterial meningitis

    Acta Otolaryngol.

    (1997)
  • M. Arditi et al.

    Three-year multicenter surveillance of pneumococcal meningitis in children: clinical characteristics, and outcome related to penicillin susceptibility and dexamethasone use

    Pediatrics

    (1998)
  • J.S. Beckman et al.

    Nitric oxide, superoxide, and peroxynitrite: the good, the bad, and ugly

    Am. J. Physiol.

    (1996)
There are more references available in the full text version of this article.

Cited by (0)

View full text