TOLBUTAMIDE ATTENUATES DIAZOXIDE-INDUCED AGGRAVATION OF HYPOXIC CELL INJURY

ATP-dependent potassium (K-ATP) channels of neurons are closed in the presence of physiological levels of intracellular ATP and open when AT P is depleted during hypoxia or metabolic damage. The present study in vestigates hypoxic alterations of purine and pyrimidine nucleotide lev els supposed to intracellularly modulate K-ATP channels. In addition, the effects of the K-ATP channel activator diazoxide and its antagonis t tolbutamide were investigated on ATP, GTP, CTP and UTP levels in sli ces of the parietal cortex. Hypoxia was evoked by saturation of the me dium with 95% N-2-5% CO2 instead of 95% O-2-5% CO2 for 5 min. Nucleoti de contents were measured by anion-exchange HPLC in neutralized perchl oric acid extracts obtained from slices frozen immediately at the end of incubation. Hypoxia per se decreased purine and pyrimidine nucleosi de triphosphate contents. Thus, ATP and GTP contents were reduced to 6 9.9 and 77.6% of the respective normoxic levels. UTP and CTP contents were even more decreased (to 60.9 and 41.6%), probably because the sal vage pathway of these pyrimidine nucleotides is less effective than th at of the purine nucleotides ATP and GTP. While tolbutamide (300 mu M) had no effect on the hypoxia-induced decrease of nucleotides, diazoxi de at 300, but not 30 mu M aggravated the decline of ATP, UTP and CTP to 51.8, 37.5 and 28.5% of the contents observed at normoxia; GTP leve ls also showed a tendency to decrease after diazoxide application. Tol butamide (300 mu M) antagonized the effects of diazoxide (300 mu M). N ucleoside diphosphate (ADP, GDP and UDP) levels were uniformly increas ed by hypoxia. There was no hypoxia-induced increase of ADP contents i n the presence of tolbutamide (300 mu M). The ATP/ADP, GTP/GDP and UTP /UDP ratios uniformly declined at a low pO(2). However, only the ATP/A DP ratio was decreased further by diazoxide (300 mu M) The observed al terations in nucleotide contents may be of importance for long- and sh ort-term processes related to acute cerebral hypoxia. Thus, hypoxia-in duced alterations of purine and pyrimidine nucleotide levels may influ ence the open state of K-ATP-channels during the period of reversible hypoxic cerebral injury. Furthermore, alterations during the irreversi ble period of cerebral injury may also arise, as a consequence of decr eased pyrimidine nucleotide contents affecting cell survival via prote in and DNA synthesis. (C) 1998 Elsevier Science B.V. All rights reserv ed.