Abstracts

RATIONALE: Neuroserpin (NS) is a selective inhibitor of tPA that is primarily expressed in neurons in the central nervous system (CNS) in areas involved with synaptic plasticity. The aim of the present study is to examine the role of tissue-type plasminogen activator (tPA) and its natural inhibitor in the brain, neuroserpin, in the modulation of seizure spreading and cell death in an animal model of limbic seizures. At the end of this presentation the participants should be able to discuss the role of neuroserpin as a potential new treatment modality for seizures.
METHODS: Adult Sprague-Dawley rats or mice, C57BL/6J, C57BL/6J-tPA-/-, C57BL/6J-PAI-1-/- C57BL/6J-Plg-/- were injected unilaterally into the amygdala with KA, and with PBS or 16 [mu]M recombinant neuroserpin into the ipsilateral hippocampus. Some animals also underwent corpus callosotomy. TPA activity and neuroserpin antigen within the limbic system were studied at different time-points by in situ zymography and immunofluorescence. TPA activity was also quantified in rats 1 hour after KA by SDS-PAGE zymography. For quantification of seizure-induced cell loss serial sections through the dorsal hippocampus were prepared and stained with hematoxylin-eosin. For clinical evaluation, animals were observed for 120 min and seizure behavior was classified as follows: 1.- myoclonic jerks involving the head and neck; 2.- unilateral tonic-clonic activity in the limbs; 3.- generalized bilateral tonic-clonic activity. Hippocampal local field potentials (EEG) were recorded for 15 min before injection of KA and PBS or neuroserpin and for 2 hours thereafter using electrodes placed in the CA1 region of the hippocampus.
RESULTS: Following the injection of KA, tPA activity and NS antigen increased within 10 minutes in the amygdala, by 30 minutes in the ipsilateral hippocampus and by 60 minutes in the contralateral hippocampus. In the hippocampus, NS expression and tPA activity were found in the CA-2 and CA-3 layers. Injection of NS into the hippocampus in close proximity to the CA-2 and CA-3 layers immediately after KA injection into the amygdala prevented the clinical and electrographic generalization of seizures. Mice lacking tPA showed a significant delay in the propagation of the seizure activity and administration of NS did not further prolong this delay, suggesting that NS was acting through its inhibition of tPA activity. PAI-1 deficient mice did not exhibit any difference compared to wild-type animals, suggesting a specific role for NS in the CNS. Plasminogen deficient mice showed a pattern of seizure spreading and a response to NS similar to wild-type animals.
CONCLUSIONS: 1.- TPA mediates the spreading of kainic acid-induced seizures throughout the limbic system. 2.- Treatment with NS, the natural inhibitor of TPA in the brain, promotes cell survival and slows the progression of KA-induced seizures.
[Supported by: NIH Grants HL55374 and HL55747 to DAL and NS36477 to JYW.]