Blood–Brain Barrier Breakdown in Cold‐Injured Brain Is Linked to a Biphasic Stimulation of Ornithine Decarboxylase Activity and Polyamine Synthesis: Both Are Coordinately Inhibited by Verapamil, Dexamethasone, and Aspirin

Abstract

An early increase in ornithine decarboxylase (ODC) activity and polyamine levels in rat cerebral capillaries was previously implicated in the mediation of blood–brain barrier (BBB) breakdown in cold-injured brain. A time course study in rat cerebrum indicated that cold injury evokes a biphasic increase in ODC activity and polyamine levels in perilesional cortex. ODC activity rose sharply (fourfold) within 1 min, remained elevated for 5 min, and then returned to the basal level by 10 min. A transient rise in polyamine concentration followed in the rank order of putrescine > spermidine > spermine. A secondary rise in ODC activity commenced in perilesional tissue at 2–6 h and peaked (8.8-fold) at 48 h. Major increases in the content of putrescine (330%), spermidine (103%), and spermine (50%) developed at 48–72 h. α-Difluoromethylornithine (DFMO), a specific irreversible inhibitor of ODC, suppressed the evoked increase in ODC activity and abolished the associated increase in content of polyamines, findings indicating that the accumulation of polyamines in cryoinjured brain reflects enhanced synthesis resulting from an ODC-mediated increase in putrescine content. Cycloheximide and actinomycin D were without effect on the early increase in ODC activity but inhibited the delayed increase in ODC activity, an observation suggesting that the initial increase in activity reflects an activation of a cryptic ODC via a posttranslational process, whereas the delayed increase in activity results from ODC synthesis mainly under transcriptional control. Because membrane phospholipid degradation, release of diacylglycerol and free arachidonic acid, and prostaglandin formation are early events in cold-injured brain, we assessed the effects of verapamil (a calcium channel blocker), dexamethasone (which inhibits arachidonic acid release), and aspirin (a cyclooxygenase inhibitor). These agents resembled DFMO in that they inhibited the early (2-min) and delayed (24-h) increase in ODC activity and polyamine concentrations and concurrently attenuated BBB breakdown in the perilesional cortex, as monitored by fluorescein transport. Exogenous putrescine nullified the protective effect of verapamil, dexamethasone, and aspirin on BBB breakdown following cryogenic injury. These results implicate Ca²⁺ influx via calcium channels, phospholipid hydrolysis, and prostaglandin synthesis in cryogenically induced stimulation of ODC activity and further strengthen the evidence linking polyamines to BBB breakdown. Changes in ODC-regulated polyamine synthesis in brain cells may play an important role in other aspects of the pathophysiology of cerebral injury.