52 MODIFICATION OF BRAIN CELL MEMBRANE Na + ,K + -ATPASE FOLLOWING RECOVERY FROM SEVERE FROM SEVERE ISCHEMIC AND HYPERCANIC INJURY IN NEWBORN PIGLETS

The present study tests the hypothesis that changes in the brain cell membrane Na+, K+-ATPase are reflected in the kinetic characteristics of the enzyme molecule and are specific to the active sites for ATP, Na+, K+ and strophanthidin. Studies were performed in 7 normoxemic piglets hyperventilated to PaCO2 of 8± 1 mmHg for 1 hr and allowed to recover for 6 hrs (ischemic injury) and 8 normoxemic piglets with administration of 7% CO2 for 90 min (PaCO2 84±7 mmHg) followed by 2 hrs recovery (hypercapnic injury). Brain cell membranes were prepared and the affinities of the Na+.K+-ATPase for ATP, K+, Na+(Km in mM) and strophanthidin (IC50 in mM) were determined in the hypo- and hypercapnic states, and their respective recoveries and compared to control. For ATP, Km decreases from 0.78±0.15 in controls to 0.62±0.11 during hypocapnia and 0.61 ±0.12 during hypercapnia; for K+ the Km changes from 2.36±0.56 in controls to 4.06 ± 0.37 during hypocapnia and 1.13±0.09 during hypercapnia, and for Na+ the Km increases from 5.43 ±0.57 in controls to 8.33±0.72 during hypocapnia and 6.97±0.41 during hypercapnia. The IC50 for strophanthidin decreases from 1×10−6M in controls to 6×10−7 M during hypocapnia and 7×10−7 M during hypercapnia. The recovery phases yielded affinities which were not statistically different from the experimental conditions. These results indicate that the active sites of Na+, K+-ATPase are modified during hypo- and hypercapnia. The differential alteration of the affinity for K+ suggests that during ischemia, dephosphorylalion and therefore energy release are decreased, while hypercapnia favors enzyme dephosphorylation. We suggest that the K+ site is the critical regulatory site for enzyme function in brain cell membranes of newborn piglets.