Abstracts

Rationale: Protoplasmic astrocytes become “reactive” and exhibit morphological changes early in the course of many CNS diseases. It has been suggested that reactive astrocytes (RAs) display different immunohistochemical changes depending on whether they are proximal or distal to the damaged area. Consistent with this, RAs in the vicinity of a stab wound demonstrate higher GFAP expression and larger cellular volume than the RAs that are far from a stab wound (Anderova et al, 2004). We studied the functional changes of RAs and their potential relevance to glial scar formation and epiletogenesis. Methods: Whole-cell patch-clamp recording was performed on astrocytes 2-7 days or one month after pilocarpine-induced seizures or stab wound in rat cortex. Gap junction coupling was examined by filling cells with 0.3% Lucifer yellow and/or 0.5% biocytin. Brain slices were fixed for further immunohistochemical staining. Results: Near areas of brain injury, most RAs were hypertrophied and had a passive current pattern. Two types of RAs were classified based on immunhistochemistry and electrophysiology. "Severe" reactive RAs have a large cellular volume, and high expression of GFAP and vimentin. They have low resting membrane potential and large cell capacitance. Glutamate and potassium uptake are severely impaired in this group of RAs, and gap junction coupling is dramatically reduced. "Moderate" RAs have smaller changes of cellular volume and GFAP expression. These cells reveal little change in RMP and potassium uptake, and only a small reduction of glutamate uptake. Gap junction communication is partially reduced in the moderate RAs. Reactive astrocytosis varied temporally and spatially following brain injury. At day 2 after seizures or stab wound, there were few severe RAs. At this early stage, mostly moderate RAs were seen, glutamate uptake function was only mildly impaired, and potassium uptake was not altered. By day 7 following injury, severe RAs predominated, and both glutamate and potassium uptake functions were dramatically reduced. Our data also showed that there were only moderate RAs in the location far (distal) from the stab wound or damaged brain area. One month after the pilocarpine insult, many RAs with a high level of GFAP exist in the area of scarring in entorhinal cortex. Few remaining RAs can be seen in the distal location of cortex. Conclusions: Our results show that, following seizure or stab wound brain injury, there is a progressive change over time of RAs in the area near injury from moderate to severe. The presence of severe RAs is correlated with the severity of brain damage. Severe RAs likely contribute to glial scar formation. Moderate reactive astrocytes distal to the injury appear able to recover to normal one month after the insult. These results suggest that transition from moderate to severe RAs may be an irreversible step in the formation of glial scar following brain injury. Prevention of progressive conversion from moderate to severe RAs may provide a therapeutic opportunity to prevent glial scar formation and epileptogenesis.