Effect of hypoxia on expression of selected proteins involved in regulation of apoptotic activity in striatum of newborn piglets.

The levels of selected neuroregulatory proteins that inhibit or promote apoptotic cell death were measured in the striatum of piglets subjected to precisely controlled 1 h hypoxic insult followed by 0, 2 and 4 h recovery and compared to sham operated animals. The anti-apoptotic proteins: there were increases in Survivin at 0 (157%, P = 0.031) and 4 h (171%, P = 0.033), in Bcl-XL at 0 (138%, P = 0.028) and 4 h (143%, P = 0.007), in VEGF at 4 h (185%, P = 0.019) and Hsp27 at 2 h (144%, P = 0.05) and 4 h (143%, P = 0.05). The pro-apoptotic proteins: caspases-1 and 7 increased at 4 h (135%, P = 0.05) and (129%, P = 0.038), respectively. Bim increased after 4 h (115%, P = 0.028), Apoptosis Inducing Factor after 2 h (127%, P = 0.048) and Calpain after 4 h (143% of control, P = 0.04). Hypoxia causes increase in levels of both anti- and pro-apoptotic proteins. Their relative activity determines the outcome in terms of cell damage and neuronal deficit.

Early regional response of apoptotic activity in newborn piglet brain following hypoxia and ischemia.

Responses of selected neuroregulatory proteins that promote (Caspase 3 and Bax) or inhibit (Bcl-2, high Bcl-2/Bax ratio) apoptotic cell death were measured in the brain of piglets subjected to precisely controlled hypoxic and ischemic insults: 1 h hypoxia (decreasing FiO2 from 21 to 6%) or ischemia (ligation of carotid arteries and hemorrhage), followed by 0, 2 and 4 h recovery with 21% FiO2. Protein expression was measured in cortex, hippocampus and striatum by Western blot. There were no significant differences in expression of Caspase-3 between sham operated, hypoxic and ischemic groups. There were significant regional differences in expression of Bcl-2 and Bax in response to hypoxia and ischemia. The changes in Bcl-2/Bax ratio were similar for hypoxia and ischemia except for striatum at zero time recovery, with ischemia giving lower ratios than hypoxia. The Bcl-2/Bax ratio was also lower for the striatum than for the other regions of the brain, suggesting this region is the more susceptible to apoptotic injury.

Regional expression of heat shock protein 72 mRNA following mild and severe hypoxia in neonatal piglet brain.

The present study examined the effect of hypoxia on expression of 72-kDa heat shock protein (hsp72) mRNA in the newborn brain. The studies were carried out in anesthetized and mechanically ventilated newborn piglets, age 3-5 days. Hypoxic insult was induced by decreasing the fraction of inspired oxygen (FiO2) from 21% to 6% or 10% for 1 h. Oxygen pressure in the microvasculature of the cortex (cortical pO2) was measured by oxygen dependent quenching of the phosphorescence of phosphor dissolved in blood. Following the two hours of normoxic recovery, regional expression of the 72-kDa heat shock protein (hsp72) mRNA was determined using in situ hybridization and autoradiography. Two grades of hypoxia were studied. Mild hypoxia (cortical pO2 = 10-30 mm Hg) induced the expression of hsp72 mRNA predominantly in the subcortical white matter. In individual animals of this group, the extent of expression varied from isolated regions to widespread involvement of the white matter. Severe hypoxia (cortical pO2 = 3-10 mm Hg) induced the expression of hsp72 mRNA in both white and gray matter regions, with strong expression occurring in the cerebral cortex of individual animals. The present results indicate that immature white matter is more sensitive than gray matter to the hypoxia induced expression of hsp72 mRNA. Further, increased expression of hsp72 mRNA may be an indicator of a pathologic degree of hypoxic stress, and the observed increase may indicate that in the newborn brain the immature white matter is particularly sensitive to injury by hypoxia-ischemia and reperfusion.

Excitatory amino acid receptor antagonists decrease hypoxia induced increase in extracellular dopamine in striatum of newborn piglets.

The present study tested the hypothesis that the increase in extracellular striatal dopamine during hypoxia is least partly associated with activation of N-methyl-D-aspartate (NMDA) and/or non-NMDA excitatory amino acid receptors. Studies were performed in anesthetized and mechanically ventilated 2-3 days old piglets. Hypoxic insult was induced by decreasing the oxygen fraction in inspired gas (FiO2) from 22 to 7% for 1 h, followed by 1 h reoxygenation at 22%. Cortical oxygen pressure was measured optically by oxygen dependent quenching of phosphorescence, and extracellular striatal dopamine was measured using in vivo microdialysis. The microdialysis probes were perfused with Ringer solution +/- 50 microM (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801) or 50 microM 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX). One hour of hypoxia decreased the cortical oxygen pressure from 46 +/- 3 Torr to 10 +/- 1.8 Torr. In striatum perfused with Ringer, statistically significant increase in extracellular dopamine, to 1050 +/- 310% of control, was observed after 20 min of hypoxia. By 40 min of hypoxia the extracellular level of dopamine increased to 4730 +/- 900% of control; by the end of the hypoxic period the values increased to 18,451 +/- 1670% of control. The presence of MK-801 in the perfusate significantly decreased the levels of extracellular dopamine during hypoxia. At 20, 40 and 60 min of hypoxia extracellular level of dopamine increased to 278 +/- 94% of control, 1530 +/- 339% of control and 14,709 +/- 1095 of control, respectively. The presence of NBQX caused a statistically significant decrease, by about 30%, in the extracellular dopamine compared to control, only at the end of the hypoxic period. It can be concluded that in striatum of newborn piglets, the excitatory NMDA receptors but not the non-NMDA receptors may be modulating the changes in extracellular levels of dopamine. The NMDA receptor antagonist, MK-801, may exert part of its reported neuroprotective effect to hypoxic stress in striatum by decreasing the levels of extracellular dopamine.

Effect of hemorrhagic hypotension on hydroxyl radicals in cat brain.

This study investigated the relationships between blood pressure, cortical oxygen pressure and hydroxyl radicals in the brain of adult cats during hemorrhagic hypotension and retransfusion. Oxygen pressure in the blood of the cortex was measured optically by the oxygen dependent quenching of phosphorescence and hydroxyl radicals by in vivo microdialysis. Following a 2 h stabilization period after implantation of the microdialysis probe in the striatum, the mean arterial blood pressure (MAP) was decreased in a stepwise manner from 132 +/- 2 Torr (control) to 90 +/- 1 Torr, 70 +/- 3 Torr and 50 +/- 3 Torr, holding the pressure at each level for 15 min. The whole blood was then retransfused and measurements were continued for 90 min. Cortical oxygen pressure progressively decreased with decrease in MAP, decreasing from 50 +/- 2 Torr (control) to 42 +/- 1 Torr, 31 +/- 2 Torr and 22 +/- 2 Torr, respectively. The level of hydroxyl radical increased by 20-25% following first 15 min of bleeding and stay on this level during the remaining period of hypotension. Maximal increase (by 78%) in level of hydroxyl radicals was observed after 15 min of retransfusion. The present study demonstrated that during hypotension and retransfusion there was an increase in the level of hydroxyl radicals in striatum. These can be important mediators of postischemic injury to the striatum.

Effect of hemorrhagic hypotension on cortical oxygen pressure and striatal extracellular dopamine in cat brain.

This study investigated the relationships between blood pressure, cortical oxygen pressure, and extracellular striatal dopamine in the brain of adult cats during hemorrhagic hypotension and retransfusion. Oxygen pressure in the blood of the cortex was measured by the oxygen dependent quenching of phosphorescence and extracellular dopamine, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) by in vivo microdialysis. Following a 2 h stabilization period after implantation of the microdialysis probe in the striatum, the mean arterial blood pressure (MAP) was decreased in a stepwise manner from 132 +/- 2 Torr (control) to 90 Torr, 70 Torr and 50 Torr, holding the pressure at each level for 15 min. The whole blood was then retransfused and measurements were continued for 90 min. As the MAP was lowered there was a decrease in arterial pH, from a control value of 7.37 +/- 0.05 to 7.26 +/- 0.06. The PaCO2 decreased during bleeding from 32.3 +/- 4.8 Torr to 19.6 +/- 3.6 Torr and returned to 30.9 +/- 3.9 Torr after retransfusion. The PaO2 was 125.9 +/- 15 Torr during control conditions and did not significantly change during bleeding. Cortical oxygen pressure decreased with decrease in MAP, from 50 +/- 2 Torr (control) to 42 +/- 1 Torr, 31 +/- 2 Torr and 22 +/- 2 Torr, respectively. A statistically significant increase in striatal extracellular dopamine, to 2,580 +/- 714% of control was observed when MAP decreased to below 70 Torr and cortical oxygen pressure decreased to below 31 Torr. When the MAP reached 50 Torr, the concentration of extracellular dopamine increased to 18,359 +/- 2,764% of the control value. A statistically significant decrease in DOPAC and HVA were observed during the last step of bleeding. The data show that decreases in systemic blood pressure result in decrease in oxygen pressure in the microvasculature of the cortex, suggesting vascular dilation is not sufficient to result in a full compensation for the decreased MAP. The decrease in cortical oxygen pressure to below 32 Torr is accompanied by a marked increase in extracellular dopamine in the striatum, indicating that even such mild hypoxia can induce significant disturbance in brain metabolism.

Response of cortical oxygen and striatal extracellular dopamine to metabolic acidosis in newborn piglets.

This study determined the relationships of metabolic acidosis, cortical oxygen pressure, and striatal extracellular dopamine in the brain of newborn piglets. After a baseline period of 120 minutes, a 0.6 N HCl solution was infused intravenously to decrease the blood pH to about 7.0-7.05. The metabolic acidosis was then corrected by injecting sodium bicarbonate and measurements were continued for one hour. The results show that decreased blood pH to about 7.2-7.15 does not cause a statistically significant change in mean blood pressure, cortical oxygen pressure or striatal extracellular dopamine. Further decrease in pH caused significant decrease in both blood pressure and cortical oxygen pressure. By the end of the period of acidosis the cortical oxygen pressure decreased from the control value of 43 +/- 4 Torr to 22 +/- 8 Torr. Changes in the extracellular level of striatal dopamine were parallel to changes in cortical oxygen pressure. The extracellular dopamine increased to 1270% of the control on the end of HCl injection. Infusion of bicarbonate to correct the acidosis resulted in an increase of cortical oxygen and progressive decline of dopamine in the extracellular medium. It is suggested that the level of extracellular dopamine in the striatum of newborn piglets was not directly affected by decrease in pH but was dependent on changes in tissue oxygen pressure during metabolic acidosis.

The effects of induced apneic episodes on cerebral cortical oxygenation in newborn piglets.

The effect of different inspiratory oxygen levels (FiO2) on cortical oxygenation (pO2) during and after recovery from apnea, was investigated in 18 anesthetized, paralyzed, and mechanically ventilated newborn piglets. Heart rate (HR) and mean arterial blood pressure (MABP) were continuously monitored as the piglets were subjected to repeated episodes of apnea initiated by disconnecting the ventilator and terminated when HR decreased to less than 80 beats/min by reconnecting the ventilator. A closed cranial window was placed over the parietal cortex of the animals and cortical pO2 was measured optically by phosphorescence quenching. Apneic episodes induced in animals ventilated with 15%, 22% and 40% oxygen had mean duration's of apnea (time before HR decreased to less than 80 beats/min) of 80, 128 and 134 s, respectively. By the end of the apnea the MABP decreased to 82%, 64%, and 54% of control, respectively. The cortical pO2 decreased from control values of 24.1, 32.3 and 38.3 Torr at 15%, 22% and 40% oxygen, respectively, to 1.7 to 3.2 Torr at the end of the apneic episode. The duration of apnea necessary for the cortical pO2 to drop below 20.3 Torr was 18, 44 and 81 s at 15%, 22% and 40% oxygen, respectively. There was an inverse correlation between the rate of decline of cortical pO2 and baseline FiO2 levels. With reventilation, the cortical pO2 reached maximal values of 42.8, 51.9 and 57.2 Torr at 15%, 22%, and 40% oxygen, respectively, before returning to the pre-apnea values. The present results show that apnea of less than 30 s duration at an FiO2 of 22% do not result in significant cortical hypoxia in hemodynamically stable piglets. Increasing the FiO2 to above 22% may possibly increase the rate of recovery of tissue oxygenation but it also may facilitate post-hypoxic cortical hyperoxia, a factor that may predispose the immature brain to free radical injury.

The effect of hypoxia and catecholamines on regional expression of heat-shock protein-72 mRNA in neonatal piglet brain.

The present study has shown that hypoxia leads to expression of heat-shock protein in the brain of newborn piglets and this process is almost completely abolished by depletion of catecholamines prior to the hypoxic episode. The piglets were anesthetized and mechanically ventilated. One hour of hypoxia was generated by decreasing the oxygen fraction in the inspired gas (FiO2) from 22% to 6%-10%. FiO2 was then returned to the control value for a period of 2 h. Following the 2 h of reoxygenation, regional expression of the 72-kDa heat-shock protein (hsp72) mRNA was determined using in situ hybridization and autoradiography. The hypoxic insult (cortical pO2 = 3-10 mmHg) induced expression of hsp72 mRNA in regions of both white and gray matter, with strong expression occurring in the cerebral cortex of individual animals. Depleting the brain of catecholamines prior to hypoxia, by treating the animals with alpha-methyl-p-tyrosine (AMT), resulted in a major change in the hsp72 mRNA expression. In the catecholamine depleted group of animals, the intensity of hsp72 mRNA expression was greatly decreased or almost completely abolished relative to the nondepleted hypoxic group. These results suggest that the catecholamines play a significant role in the expression of the hsp72 gene in response to hypoxic insult in neonatal brain.

Effect of hypoxia and reoxygenation on regional activity of nitric oxide synthase in brain of newborn piglets.

The activity of nitric oxide synthase (NOS) was measured in homogenates from cortex, striatum, hippocampus, cerebellum, pons, thalamus and midbrain of the brain of newborn piglets and the effects of hypoxia and posthypoxic period on this activity was evaluated. The control activities were 19.7, 31.5, 26.8, 16.7, 33.6, 19.3 and 39.4 pmol/mg protein per min, respectively. A 1 h period of hypoxia (an FiO2 of 7%) resulted in statistically significant decreases in the activity of NOS in every region of the brain except for the cortex, where the activity was not significantly altered compared to control. By 2 h of reoxygenation following such a hypoxic episode, the NOS activities increased to above control levels in all regions of the brain, but this increase was statistically significant compared to control only in thalamus. Since hypoxia induced the greatest decrease in NOS activity in the cerebellum, the kinetic constants of the enzyme were measured in homogenates from this region of brain. The decreased activity following the hypoxic episode was associated with an approximately four-fold increase in the apparent affinity (KM) for arginine with no significant change in the maximal rate of reaction (Vmax). The decrease in NOS activity subsequent to a hypoxic episode may contribute to the disturbances in cellular metabolism in the immature brain induced by episodes of hypoxia-reoxygenation.

Relationships of dopamine, cortical oxygen pressure, and hydroxyl radicals in brain of newborn piglets during hypoxia and posthypoxic recovery.

The present study describes the relationships of extracellular striatal dopamine, cortical oxygen pressure, and striatal hydroxyl radicals in brain of newborn piglets during hypoxia and posthypoxic reoxygenation. Hypoxia was induced by reducing the fraction of inspired oxygen (FiO2) from 22% (control) to 7% for 1 h. The FiO2 was then returned to the control value and measurements were continued for 2 h. Cerebral oxygen pressure was measured by the oxygen dependent quenching of phosphorescence and extracellular levels of dopamine, 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and hydroxy radicals in the striatum were determined by in vivo microdialysis. Hypoxia decreased the cortical oxygen pressure from 47 +/- 2 to 9 +/- 1.3 torr (p < 0.001); the levels of extracellular dopamine in the striatum increased to 16,000 +/- 3,270% of control (p < 0.01), whereas the levels of DOPAC and HVA decreased to 25.3 +/- 6% (p < 0.001) and 36 +/- 5% (p < 0.01) of control, respectively. Compared with control, the hydroxyl radical levels at each time point were not significantly increased during hypoxia, although the sum of the measured values was significantly increased (p < 0.05). During the first 5 min after FiO2 was returned to 22%, the cortical oxygen pressure increased to control values and stayed at this level for the remainder of the measurement period. The extracellular level of dopamine declined to values not statistically different from control during 40 min of reoxygenation. During the first 10 min of reoxygenation, DOPAC and HVA further decreased and then began to slowly increase. By 70 min of reoxygenation, the values were not significantly different from control.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of the efficacy of blood and polyethylene glycol-hemoglobin in recovery of newborn piglets from hemorrhagic hypotension: effect on blood pressure, cortical oxygen, and extracellular dopamine in the brain.

BACKGROUND: Successful blood substitutes, when infused in place of an equal volume of whole blood, provide similar delivery of oxygen to the tissues without introducing abnormalities in cellular metabolism. STUDY DESIGN AND METHODS: Equal volumes of whole blood (control), polyethylene glycol-hemoglobin solution at 6 g per dL, dextran solution, and physiologic saline were compared for their ability to reverse the effects of hemorrhagic hypotension on oxygenation and dopamine metabolism in the brain of newborn piglets. The decrease in mean arterial blood pressure was used as a measure of the hemorrhagic insult. Cerebral oxygen pressure was determined optically by the oxygen-dependent quenching of phosphorescence, and the extracellular level of dopamine in the corpus striatum was determined by in vivo microdialysis. RESULTS: Following a 2-hour stabilization after implantation of the microdialysis probe in the corpus striatum, the mean arterial blood pressure was decreased from 88 +/- 7 torr (control) to 42 +/- 5 torr by the removal of blood in a stepwise manner, over a period of 60 minutes. Decrease in mean arterial blood pressure caused a progressive stepwise decrease in cortical oxygen pressure from 48 +/- 5 torr to 16 +/- 4 torr at the end of bleeding. As a consequence of the decrease in oxygen pressure, extracellular dopamine increased progressively to about 2300 percent of the control value. When a volume of blood equal to that removed was returned and bicarbonate was injected to help correct arterial pH, blood pressure, cortical oxygen pressure, and extracellular dopamine all returned within the 20- to 30-minute recovery period to values not significantly different from control values. An equal volume of polyethylene glycol-hemoglobin solution, even with significantly lower hemoglobin content than whole blood, gave results comparable to those with whole blood. CONCLUSION: Polyethylene glycol-hemoglobin solution, like whole blood but in contrast to physiologic saline or dextran solution, was capable of returning the mean arterial blood pressure, cortical oxygen pressures, and extracellular dopamine nearly to control levels after acute blood loss in newborn piglets.

Comparison of postasphyxial resuscitation with 100% and 21% oxygen on cortical oxygen pressure and striatal dopamine metabolism in newborn piglets.

The present study tests the hypothesis that ventilation with 100% O2 during recovery from asphyxia leads to greater disturbance in brain function, as measured by dopamine metabolism, than does ventilation with 21% oxygen. This hypothesis was tested using mechanically ventilated, anesthetized newborn piglets as an animal model. Cortical oxygen pressure was measured by the oxygen-dependent quenching of phosphorescence, striatal blood flow by laser Doppler, and the extra-cellular levels of dopamine and its metabolites by in vivo microdialysis. After establishment of a baseline, both the fraction of inspired oxygen (FiO2) and the ventilator rate were reduced in a stepwise fashion every 20 min over a 1-h period. For the subsequent 2-h recovery, the animals were randomized to breathing 21 or 100% oxygen. It was observed that during asphyxia cortical oxygen pressure decreased from 36 to 7 torr, extracellular dopamine increased 8,300%, and dihydroxyphenylacetic acid and homovanillic acid decreased by 65 and 60%, respectively, compared with controls. During reoxygenation after asphyxia, cortical oxygen pressure was significantly higher in the piglets ventilated with 100% oxygen than in those ventilated with 21% oxygen (19 vs. 11 torr). During the first hour of reoxygenation, extracellular dopamine levels decreased to approximately 200% of control in the 21% oxygen group, whereas these levels were still much higher in the 100% oxygen group (approximately 500% of control). After approximately 2 h of reoxygenation, there was a secondary increase in extracellular dopamine to approximately 750 and approximately 3,000% of baseline for the animals ventilated with 21 and 100%, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

3,4-dihydroxyphenylalanine (dopa) metabolism and retinoic acid induced differentiation in human neuroblastoma.

In mature cells of the sympathetic nervous system and the adrenal gland, the activity of dihydroxyphenylalanine decarboxylase (DDC) is higher than that of tyrosine hydroxylase and 3,4-dihydroxyphenylalanine (dopa) does not accumulate in the cells. On the other hand, it is known that in some neuroblastoma cells there is a relative deficiency of DDC, resulting in accumulation and secretion of dopa. Such a relative deficiency of DDC is a characteristic of neural cells at an early stage of neural crest development, suggesting the neuroblastoma are cells arrested in early neural crest development. If this were the case, it is possible that agents such as retinoic acid (RA) could induce neuroblastoma to differentiate into mature cells with respect to their metabolism of catecholamines. We have measured the effect of RA on the metabolism of dopa and expression of tyrosine hydroxylase and DDC in human neuroblastoma cell lines, CHP-126, CHP-134, IMR-32, NB-69, and LA-N-5. When the cell cultures were treated with RA, they showed wide variations in response as measured by morphological change, growth inhibition, enzyme activities and enzyme expressions. The RA treatment modulated the activities of tyrosine hydroxylase and DDC, but does not increase DDC relative to tyrosine hydroxylase. It is concluded that RA does not induce biochemical differentiation of the neuroblastoma into mature cells even when there are extensive morphological changes and suppression of growth rate.

Effect of hemorrhagic hypotension on extracellular level of dopamine, cortical oxygen pressure and blood flow in brain of newborn piglets.

The present study describes the relationships between extracellular striatal dopamine, cortical oxygen pressure and blood flow in brain of newborn piglets during hemorrhagic hypotension. Cerebral oxygen pressure was measured optically by the oxygen dependent quenching of phosphorescence; extracellular dopamine by in vivo microdialysis; striatal blood flow was monitored by a laser Doppler. Following a 2 h stabilization period after implanting the microdialysis and laser Doppler probes in the striatum, the mean arterial blood pressure (MABP) was decreased in stepwise manner from 87 +/- 4 Torr (control) to 35 +/- 5 Torr, during 63 min. The whole blood was then reinfused and measurements were continued for 45 min. Statistically significant decrease in blood flow, 10%, was observed when arterial blood pressure decreased to about 53 Torr. With further decrease blood pressure to 35 Torr, blood flow decreased to about 35% of control (P < 0.01). Cortical oxygen pressure decreased almost proportional to decrease in blood pressure. The progressive decrease in MABP from 87 +/- 4 Torr to 65 +/- 6, 52 +/- 7, and 35 +/- 5 Torr resulted in cortical oxygen pressure decreasing from 45 +/- 4 Torr to 33 +/- 3 Torr (P < 0.05), 24 +/- 4 Torr (P < 0.01) and 13 +/- 3 Torr (P < 0.01). The levels of extracellular dopamine in the striatum increased with decreasing cortical oxygen pressure. As cortical oxygen decreased, the extracellular dopamine increased to 230%, 420% and 3200% of control, respectively. Our results show that in mild hypotension total blood flow is well maintained but oxygen pressure in the microvasculature decreases, possibly due to heterogeneity in the regulatory mechanism.(ABSTRACT TRUNCATED AT 250 WORDS)

Relationship of extracellular dopamine in striatum of newborn piglets to cortical oxygen pressure.

The present studies describes the relationship between extracellular dopamine in striatum of newborn piglets and cortical oxygen pressure. The extracellular level of dopamine was measured by in vivo microdialysis and the oxygen pressure in the cortex was measured by phosphorescence lifetime of oxygen probe in the blood. Controlled, graded levels of hypoxic insult to the brain of animals were generated by decreasing of the oxygen fraction in the inspired gas (FiO2) from 21% to 14%, 11%, and 9%. This resulted in decrease in the cortical oxygen pressure from 31-35 Torr to about 24 Torr, 15 Torr and 4 Torr, respectively. The changes in extracellular level of dopamine, DOPAC and HVA were dependent on changes in cortical oxygen pressure. Stepwise decrease in the cortical oxygen pressure (see above) caused increases in extracellular dopamine of about 80%, 200% and 550%, respectively. The levels of DOPAC and HVA progressively decreased and when cortical oxygen decreased to 4-6 Torr were about 50% and 70% of control, respectively. After return of FiO2 to control (21%), the cortical oxygen pressure rapidly increased to above normal, then returned to control values. The extracellular levels of dopamine, DOPAC, and HVA recovered more slowly, attaining control values in about 30 minutes. The data show that extracellular levels of dopamine increase with even very small decreases in oxygen pressure. Thus, there is no "oxygen reserve" which protects dopamine release and metabolism from decrease in oxygen pressure.

Effect of hypoxia and reoxygenation on the activity of transglutaminase in brain of newborn piglets.

The transglutaminase activity in five regions of the brain of newborn piglets was measured and the effects of hypoxia and posthypoxic period on this activity evaluated. Enzyme activity was measured in homogenates from cortex, hippocampus, striatum, thalamus and midbrain. The control activities were 7.2, 6.2, 6.0, 5.7 and 4.6 pmol/mg protein/min, respectively. The activities at the end of an 18 min period of hypoxia induced by an FiO2 of 9% were not significantly different from control activities. By 3 h after the hypoxic episode, however, the transglutaminase activities were significantly above control levels in all five regions of the brain. Measurements of the kinetic constants of tranglutaminase indicated that increases in enzyme activity were associated with an increase in Vmax with no significant change in the apparent affinity of the enzyme for the substrate, putrescine. The increased activity of transglutaminase during the posthypoxic period, with no changes immediately after hypoxia, suggest that the increases could be due to increased enzyme synthesis rather than activation of existing enzyme. The rise in transglutaminase activity subsequent to a hypoxic episode may contribute significantly to the long-term disturbances in cellular metabolism in the immature brain induced by hypoxic episodes.