Pig fetal skeletal muscle development is associated with genome-wide DNA hypomethylation and corresponding alterations in transcript and microRNA expression.

Fetal myogenesis represents a critical period of porcine skeletal muscle development and requires coordinated expression of thousands of genes. Epigenetic mechanisms, including DNA methylation, drive transcriptional regulation during development; however, these processes are understudied in developing porcine tissues. We performed bisulfite sequencing to assess DNA methylation in pig longissimus dorsi muscle at 41- and 70-days gestation (dg), as well as RNA- and small RNA-sequencing to identify coordinated changes in methylation and expression between myogenic stages. We identified 45 739 differentially methylated regions (DMRs) between stages, and the majority (N = 34 232) were hypomethylated at 70 versus 41 dg. Integration of methylation and transcriptomic data revealed strong associations between differential gene methylation and expression. Differential miRNA methylation was significantly negatively correlated with abundance, and dynamic expression of assayed miRNAs persisted postnatally. Motif analysis revealed significant enrichment of myogenic regulatory factor motifs among hypomethylated regions, suggesting that DNA hypomethylation may function to increase accessibility of muscle-specific transcription factors. We show that developmental DMRs are enriched for GWAS SNPs for muscle- and meat-related traits, demonstrating the potential for epigenetic processes to influence phenotypic diversity. Our results enhance understanding of DNA methylation dynamics of porcine myogenesis and reveal putative cis-regulatory elements governed by epigenetic processes.

Hypermagnesemia does not increase brain intracellular magnesium in newborn swine.

Phosphorus-31 magnetic resonance spectroscopy was used in 2-day (n = 4) and 40-day (n = 4) miniswine to determine whether plasma hypermagnesemia alters brain intracellular magnesium concentration and if the plasma-brain intracellular magnesium relationship changes with age. At control, brain intracellular magnesium concentration was similar in the 2-day (0.24 +/- 0.04 mM) and 40-day groups (0.21 +/- 0.01 mM). Intravenous infusions of magnesium sulfate (MgSO(4), 60 minute) raised plasma magnesium concentration to 4-6 mM in both groups. During and for 3 hours after MgSO(4) infusions, there were no changes in brain intracellular magnesium concentration in either group and no correlation between plasma and brain intracellular magnesium (r = 0.11 and 0.08 for 2- and 40-day groups, respectively). Brain intracellular magnesium concentration appears to be tightly regulated.

Differences in brain temperature and cerebral blood flow during selective head versus whole-body cooling.

OBJECTIVE: To compare brain temperature and cerebral blood flow (CBF) during head and body cooling, with and without systemic hypoxemia. METHODS: Seventeen newborn swine were studied for either measurement of brain temperature alone (n = 9) or measurement of brain temperature and CBF (n = 8). All animals were ventilated and instrumented, and temperature probes were inserted into the rectum, into the brain at depths of 2 and 1 cm from the cortical surface, and on the dural surface. Blood flow was measured with microspheres. The protocol consisted of a control period, an interval of either head or body cooling, and cooling with 15 minutes of superimposed hypoxia. After a 1-hour recovery period, animals were exposed to the same sequence except that the alternate mode of cooling was evaluated. RESULTS: Head cooling with a constant rectal temperature resulted in an increase in the temperature gradient across the brain from the warmer central structures to the cooler periphery (brain 2 cm - dura temperature: 1.3 +/- 1.1 degrees C at control to 7.5 +/- 3.5 degrees C during cooling). Hypoxia superimposed on head cooling decreased the temperature gradient by at least 50%. In contrast, body cooling was associated with an unchanged temperature gradient across the brain (brain 2 cm - dura temperature: 1.5 +/- 1.2 degrees C at control to 1.1 +/- 0.9 degrees C during cooling). Hypoxia superimposed on body cooling did not change brain temperature. Both modes of brain cooling resulted in similar reductions of global CBF ( approximately 40%) and O(2) uptake. CONCLUSION: Brain hypothermia achieved through head or body cooling results in different brain temperature gradients. Alterations in systemic variables (ie, hypoxemia) alters brain temperature differently in these 2 modes of brain cooling. The mode of brain cooling may affect the efficacy of modest hypothermia as a neuroprotective therapy.

Noninvasive measurement of brain temperature after stroke.

BACKGROUND AND PURPOSE: Brain temperature may be an important factor governing the extent of neuronal injury associated with stroke. The goal of this study was to develop a noninvasive method for measuring brain temperature, both to characterize the extent to which temperature changes after stroke and to test protocols designed to reduce brain temperature. We used an animal model to test the ability of 1H MR spectroscopy to measure temperature from infarcted brain tissue at 24 hours after insult. METHODS: Unilateral permanent focal ischemia in the middle cerebral artery territory was induced in adult dogs by intravascular delivery of microfibrillar collagen. MR imaging performed at 24 hours after insult was used to guide the implantation of temperature probes into the basal ganglia infarct and into the same anatomic location on the contralateral side. Serial non-water-suppressed 1H MR spectra were obtained from 1.3-cm3 voxels using an echo time of 136 and 272 ms, alternately, from the infarcted and contralateral non-infarcted tissue during a period when brain temperature was raised and lowered by whole-body heating and cooling. RESULTS: The chemical shift difference between the 1H MR spectroscopy signal of water and N-acetylaspartate or water and trimethylamines was plotted against brain temperature for two voxel locations. The slope and intercept of the plots obtained for infarcted and non-infarcted brain were not significantly different (P < .05, t test), and there was no difference between the slope and intercept of plots made from data collected with an echo time of 136 or 272 ms. CONCLUSION: The results of this study indicate that brain temperature can be measured from regions of brain containing infarcted tissue, at least up to 24 hours after ischemia. It should be possible to apply the 1H MR spectroscopy method used in the present study to measure brain temperature after stroke.

A limited interval of delayed modest hypothermia for ischemic brain resuscitation is not beneficial in neonatal swine.

This investigation determined if a short interval of modest hypothermia (1 h) initiated 30 min after brain ischemia provided neuroprotection. The rationale for the time and duration of brain cooling reflects the likelihood that the implementation of neuroprotective strategies will occur at an interval shortly after ischemia, and that long-term maintenance of normothermia is a cornerstone of neonatal stabilization. Studies were performed in 22 ventilated neonatal mini-swine in a superconducting magnet to obtain 31P magnetic resonance spectra. After a control period all animals underwent 15 min of global brain ischemia and were maintained normothermic for the first 30 min post-ischemia. In one group of 11 swine normothermia was continued. In the other group of 11 swine, modest hypothermia was initiated at 30 min post-ischemia, continued for 1 h and followed by resumption of normothermia. Animals were subsequently weaned from ventiltor support, removed from the magnet, and underwent neurobehavioral and histologic assessment at 72 h post-ischemia. Both groups had similar severity of ischemia, as indicated by identical changes in arterial blood pressure and pH, alterations in brain beta-nucleotide triphosphate (% of control where control = 100%, 32 +/- 28 vs 27 +/- 26% for normothermic and hypothermic groups, respectively), and the extent of intraischemic brain acidosis (6.13 +/- 0.19 vs 6.14 +/- 0.14 for normothermic and hypothermic groups, respectively). In both groups the distribution of stages of encephalopathy were the same: 1 normal and 10 abnormal (4 mild, 2 moderate, and 4 severe) normothermic, and, 3 normal and 8 abnormal (4 mild, 2 moderate, and 2 severe) hypothermic animals. There was no difference in the extent of neuronal injury between groups. We conclude that a 1-h interval of modest hypothermia initiated at 30 min post-ischemia does not confer neuroprotection.

Age-dependent differences in the relationship between plasma and brain extracellular fluid concentrations of magnesium after MgSO4 infusions in miniswine.

Magnesium is a potential neuroprotective agent in the treatment of head injury and ischemia whose efficacy is likely determined by increases in brain extracellular fluid (ECF) magnesium, which in turn depends on its concentration in plasma. The objectives of this study were to: 1) examine the effects of increasing plasma magnesium concentration ([Mg]plasma) to 4-6 mM on brain ECF magnesium concentration ([Mg]ECF) and 2) determine whether maturational changes occur in the transfer of magnesium into brain ECF for newborn and more mature (approximately 1 month old) miniswine. Increases in [Mg]plasma by systemic administration of MgSO4 resulted in similar maximal elevations in brain [Mg]ECF for both age groups (193+/-76% versus 253+/-106% of control for newborn and 1-month-old miniswine, respectively). Calculations of half-lives (t1/2) for the increase and decrease in magnesium concentration (t1/2 uptake and t1/2 clearance) were used to characterize magnesium kinetics in plasma and brain ECF. Plasma magnesium uptake was shorter in 1-month-old (t1/2 = 11.1+/-0.9 min) compared with newborns (12.9+/-1.7 min, p < 0.05). The faster increase in [Mg]plasma probably contributed to a faster uptake of brain [Mg]ECF in 1-month-old compared with newborn swine (t1/2 uptake = 27.9+/-12.8 versus 46.0+/-20.9 min, respectively, p < 0.05). Although plasma magnesium clearance was shorter in 1-month-old swine compared with newborn (t1/2 = 34.3+/-7.0 versus 74.7+/-33.7 min, respectively, p < 0.05), the clearance of magnesium from the brain ECF was similar for each age group. Reductions in blood pressure and heart rate occurred during hypermagnesemia and were similar in each age group. This study shows that acute elevations in [Mg]plasma to 4-6 mM result in similar relative increases in brain [Mg]ECF for both newborn and 1-month-old miniswine. However, there are maturational differences, as demonstrated by the faster rate of magnesium uptake into the ECF observed in the older miniswine.

Failure of localized head cooling to reduce brain temperature in adult humans.

Non-invasive brain temperature measurements using proton magnetic resonance spectroscopy were used to test the hypothesis that localized head cooling would reduce brain temperature in 10 normal adult humans. Temperature reductions of the head surface to 15.8+/-3.5 degrees C did not reduce brain temperature measured in the superficial cortex (36.8+/-0.5 degrees C) or thalamus (36.6+/-0.7 degrees C), as compared to measurements obtained with a head surface temperature of 34.7+/-1.6 degrees C (37.0+/-0.6 degrees C and 36.6+/-0.4 degrees C, respectively). There was no change in the temperature gradient from the superficial to deep brain locations in the presence or absence of head cooling, and brain temperature did not decrease as a function of the duration of head cooling for periods up to 50 min. There was no correlation between the scalp surface (range: 10-38 degrees C) and brain temperature at either the deep or superficial locations.

Modest hypothermia provides partial neuroprotection when used for immediate resuscitation after brain ischemia.

Intraischemic reduction in temperature of 2-3 degrees C (modest hypothermia) has been demonstrated to provide partial neuroprotection in neonatal animals. This investigation determined if modest hypothermia initiated immediately after brain ischemia provides neuroprotection. Piglets were studied with rectal temperature maintained during the 1st h after 15 min of brain ischemia at either 38.3 +/- 0.3 degrees C (normothermia, n = 11) or at 35.8 +/- 0.5 degrees C (modest hypothermia, n = 11). The severity of brain ischemia was similar between groups as indicated by equivalent reduction in mean blood pressure (90 +/- 15 to 24 +/- 3 versus 92 +/- 13 to 26 +/- 3 mm Hg), and changes in cerebral metabolites and intracellular pH (pH(i)) measured by magnetic resonance spectroscopy (beta-nucleoside triphosphate = 44 +/- 9 versus 42 +/- 18% of control, control = 100%, pH(i): 6.25+/- .15 versus 6.24 +/- 0.22 for normothermic and modestly hypothermic groups, respectively). In the first 90 min after ischemia, there were no differences between groups in the duration and extent of brain acidosis, and relative concentrations of phosphorylated metabolites. Categorical assessment of neurobehavior was evaluated at 72 h postischemia (n = 16), or earlier if an animal's condition deteriorated (n = 6). Postischemic hypothermia was associated with less severe stages of encephalopathy compared with normothermia (p = 0.05). Histologic neuronal injury was assessed categorically in 16 brain regions, and postischemic hypothermia resulted in less neuronal injury in temporal (p = 0.024) and occipital (p = 0.044) cortex at 10 mm beneath the cortical surface, and in the basal ganglia (p = 0.038) compared with that in normothermia. Modest hypothermia for 1 h immediately after brain ischemia provides partial neuroprotection and may represent an adjunct to resuscitative strategies.

Calculation of intracellular cerebral [Mg2+] during hypoxic ischemia by in vivo 31P NMR.

Several algorithms for the calculation of ionized intracellular magnesium concentration from the chemical shifts of MgATP were compared, using in vivo 31P NMR data obtained from swine brain during and following hypoxic ischemia plus i.v. MgSO4 infusion. This analysis reveals that both the absolute ionized intracellular magnesium and relative changes in magnesium may vary widely between algorithms used. The calculated intracellular pH, used in algorithms to determine ionized magnesium concentration was found to be a critical parameter that governs the extent of these differences.

Effect of hypoxia on glucose-modulated cerebral lactic acidosis, agonal glycolytic rates, and energy utilization.

Newborn and 1-mo-old swine were exposed to identical durations (18 min) and degrees of hypoxia (O2 content = 4 mL/dL), to examine the effects of hypoxia on cerebral energy metabolism and intracellular pH (pHi) in vivo, using 31P and 1H nuclear magnetic resonance spectroscopy. Hypoxia produced the same extent of reductions in phosphocreatine (PCr) (63 +/- 28% and 65 +/- 10%, newborns and 1-mo-olds, respectively) and pHi (6.93 +/- 0.06 and 6.89 +/- 0.06, respectively) for either age group. The magnitude of changes in PCr, lactate, and pHi was larger for subgroups of data collected when cardiovascular instability was present, suggesting that hypotension and possibly reduced cerebral perfusion contributed to cerebral energy failure and lactic-acidosis for either age group. There were no correlations between the blood plasma glucose concentration at 18 min of hypoxia and the extent of change in PCr, lactate, or pHi for either age group. During a subsequent period of complete ischemia induced via cardiac arrest after 20 min hypoxia, the decline in PCr and nucleoside triphosphate (NTP), and increase in lactate followed similar rates compared with previously studied age-matched animals that were normoxic before ischemia. The rate constants for the change in PCr, NTP, and lactate followed similar rates compared with previously studied age-matched animals that were normoxic before ischemia. The rate constants for the change in PCr, NTP, and lactate during ischemia showed no correlation with the blood plasma glucose concentration measured immediately before cardiac arrest. These results suggest that cerebral glycolytic rates and energy utilization during ischemia are unaffected by a preceding interval of hypoxia and that hyperglycemia does not delay cerebral energy failure during hypoxia or combined hypoxic-ischemia.

An inorganic biocide using a novel presentation of silver.

Synopsis A silver-based preservative system has been developed which overcomes difficulties often associated with metals, namely solubility, dispersion and toxicity. It comprises a silver chloride titanium dioxide composite which allows the controlled release of silver ions. Microbiological data has shown the composite to be active at concentrations less than 25 ppm against a wide range of micro-organisms, including Gram-negative non-fermentative bacteria, yeasts and moulds. The composite has satisfactory preservative activity against bacteria and fungi at application concentrations of between 50 ppm and 500 ppm, depending on the formulation. At high concentrations (>300 ppm) there may be some grey discoloration of the product, but the composite remains effective. To overcome this problem, a system of enhancing antimicrobial activity was devised, based on a sulphosuccinate salt dispersing agent. This allows lower concentrations of composite to be used whilst maintaining equivalent antimicrobial activity.

Quantitative relationship between brain temperature and energy utilization rate measured in vivo using 31P and 1H magnetic resonance spectroscopy.

In neonatal and adult animals, modest reduction in brain temperature (2-3 degrees C) during ischemia and hypoxia-ischemia provides partial or complete neuroprotection. One potential mechanism for this effect is a decrease in brain energy utilization rate with consequent preservation of brain ATP, as occurs with profound hypothermia. To determine the extent to which modest hypothermia is associated with a decrease in brain energy utilization rate, in vivo 31P and 1H magnetic resonance spectroscopy (MRS) was used to measure the rate of change in brain concentration of phosphocreatine, nucleoside triphosphate, and lactate after complete ischemia induced by cardiac arrest in 11 piglets (8-16 d). Pre-ischemia metabolite concentrations and MRS-determined rate constants were used to calculate the initial flux of high energy phosphate equivalents (d[approximately P]/dt, brain energy utilization rate). Baseline physiologic and MRS measurements were obtained at 38.2 degrees C and repeated after brain temperature was adjusted between 28 and 41 degrees C. This was followed by measurement of d[approximately P]/dt during complete ischemia at 1-2 degrees C increments within this temperature range. Adjusting brain temperature did not alter any systemic variable except for heart rate which directly correlated with brain temperature (r = 0.95, p < 0.001). Before ischemia brain temperature inversely correlated with phosphocreatine (r = -0.89, p < 0.001), and reflected changes in the phosphocreatine-ATP equilibrium, because brain temperature inversely correlated with intracellular pH (r = -0.77, p = 0.005). Brain temperature and d[approximately P]/dt were directly correlated and described by a linear relationship (slope = 0.61, intercept = -12, r = 0.92, p < 0.001). A reduction in brain temperature from normothermic values of 38.2 degrees C was associated with a decline in d[approximately P]/dt of 5.3% per 1 degree C, and therefore decreases in d[approximately P]/dt during modest hypothermia represent a potential mechanism contributing to neuroprotection.

Neonatal ischemic neuroprotection by modest hypothermia is associated with attenuated brain acidosis.

BACKGROUND AND PURPOSE: A 2.9 degrees C reduction in the intraischemic rectal temperature of neonatal piglets is associated with less brain damage compared with animals with normothermic rectal temperatures. This investigation studied one potential mechanism for this observation: better maintenance of energy stores and less brain acidosis secondary to reduced metabolic activity associated with modest hypothermia. METHODS: 31P MR spectroscopy was used to study piglets before, during, and after 15 minutes of partial brain ischemia with intraischemic rectal temperatures of either 38.3 +/- 0.4 degrees C (n = 10, normothermic) or 35.4 +/- 0.5 degrees C (n = 10, hypothermic). Animals were followed up for up to 72 hours after ischemia and were evaluated clinically and by brain histology. RESULTS: Values for pHi remained 0.15 to 0.20 pH units greater in modestly hypothermic than in normothermic piglets during ischemia and the initial 30 minutes after ischemia (P = .049, group effect). Phosphocreatine, beta-ATP, and inorganic phosphorus were similar between groups. The relationship between the intraischemic energy state and subsequent clinical evidence of brain damage (irrespective of group assignment) revealed lower pHi over the last 7 minutes of ischemia for abnormal compared with normal piglets (5.98 +/- 0.22 versus 6.39 +/- 0.24, respectively; P = .002). In contrast, intraischemic beta-ATP (41 +/- 19% versus 57 +/- 21% of control) and inorganic phosphorus (273 +/- 31% versus 224 +/- 92% of control) for abnormal and normal piglets, respectively, did not differ between groups. CONCLUSIONS: Intraischemic modest hypothermia attenuates the severity of brain acidosis during and 30 minutes after ischemia compared with normothermic animals and supports the concept that attenuated brain acidosis is a potential mechanism by which hypothermia may reduce ischemic brain damage.

Nondestructive measurement of retinal glucose transport and consumption in vivo using NMR spectroscopy.

The cellular events underlying various retinopathies are poorly understood but likely involve perturbation of retinal glucose metabolism. Current methods for assessing this metabolism are destructive, thus limiting longitudinal studies. We hypothesize that following an intravitreous injection, the clearance rate of a glucose analogue will be a nondestructive index of retinal glucose transport and metabolism in vivo. First, radiolabeled glucose analogues were injected into the vitreous. After 40 min, the dominant clearance path was posterior via the retina and was consistent with a facilitated transport mechanism. Next, either [6,6-2H2]glucose or 3-deoxy-3-fluoro-D-glucose was injected into the vitreous of rabbit eyes, and the clearance rate of each analogue was determined over 40 min using, respectively, 2H or 19F NMR. These rates were interpreted as a function of the retinal glucose transport and consumption. From the NMR data, the rate of retinal glucose consumption was approximately 16 times slower than the transport of glucose. These data demonstrate that NMR measurements of glucose analogue clearance rate from the vitreous can provide a nondestructive index of retinal glucose transport and consumption in vivo.

Validation of a noninvasive method to measure brain temperature in vivo using 1H NMR spectroscopy.

The goal of this study was to evaluate the potential of using the difference between the 1H NMR frequencies of water and N-acetylaspartic acid (NAA) to measure brain temperature noninvasively. All water-suppressed and non-water-suppressed 1H NMR spectra were obtained at a field strength of 4.7 T using a surface coil. Experiments performed on model solutions revealed a decrease in the difference between NMR frequencies for NAA and water as a linear function of increasing temperature from 14 to 45 degrees C. Changing pH in the range 5.5-7.6 produced no discernible trends for concurrent changes in the slope and intercept of the linear relationship. There were minor changes in slope and intercept for solutions containing 80 or 100 mg of protein/ml versus no protein, but these changes were not considered to be of sufficient magnitude to deter the use of this approach to measure brain temperature. The protein content of swine cerebral cortex was found to remain constant from newborn to 1 month old (78 +/- 12 mg/g; n = 41). Therefore, data collected for the model solution containing 80 mg of protein/ml were used as a calibration curve to calculate brain temperature in eight swine during control, hypothermia, ischemia, postischemia, or death, over a temperature range of 23-40 degrees C. A plot of 61 temperatures determined from 1H NMR versus temperatures measured from an optical fiber probe sensor implanted 1 cm into the cerebral cortex showed excellent linear agreement (slope = 1.00 +/- 0.03, r2 = 0.96). We conclude that 1H NMR spectroscopy presents a practical means of making noninvasive measurements of brain temperature with an accuracy of better than +/- 1 degree C.

Evaluation of potential effectors of agonal glycolytic rate in developing brain measured in vivo by 31P and 1H nuclear magnetic resonance spectroscopy.

Previously we have shown that hypercarbia produces a larger decrease in agonal glycolytic rate in 1-month-old swine than in newborns. In an effort to understand the mechanism responsible for this difference, we tested the hypothesis that hypercarbia produces age-related changes in the concentration of one or more effectors of phosphofructokinase activity. Specifically, in vivo 31P and 1H NMR spectroscopy was used to compare changes in lactate levels, intracellular pH, free magnesium concentration, and content of phosphorylated metabolites for these two age groups at three intervals during the first 1.5 min of complete ischemia in the presence or absence of hypercarbia (PaCO2 = 102-106 mm Hg). Hypercarbia produced the same drop in intracellular brain pH for both age groups, but the decrease in phosphocreatine level and increase in inorganic phosphate content were greater in 1-month-olds compared with newborns. During ischemia there was no difference between the magnitude of change in intracellular pH and levels of phosphocreatine and inorganic phosphate in hypercarbic 1-month-olds versus newborns. Under control conditions, i.e., normocarbia and normoxia, the free Mg2+ concentration was lower and the fraction of magnesium-free ATP was higher for newborns than 1-month-olds. However, there was no change in these variables for either age group during hypercarbia and early during ischemia. Thus, age-related differences in the relative decrease in agonal glycolytic rate during hypercarbia could not be explained by differences in intracellular pH, inorganic phosphate content, or free magnesium concentration. The [ADP]free at control was higher in newborns compared with 1-month-olds, and there was no age-related difference in [AMP]free. These variables did not change for newborns when exposed to hypercarbia, but for 1-month-olds [ADP]free and [AMP]free increased during hypercarbia relative to control values. High-energy phosphate utilization during ischemia for hypercarbic 1-month-olds was reduced by 74% compared with normocarbic 1-month-olds during ischemia, whereas the reduction in energy utilization (14%) was not significant for hypercarbic versus normocarbic newborns during ischemia.(ABSTRACT TRUNCATED AT 400 WORDS)

Age-related changes in swine brain creatine kinase-catalyzed 31P exchange measured in vivo using 31P NMR magnetization transfer.

31P exchange rates through the creatine kinase-catalyzed interconversion of phosphocreatine and gamma-ATP were measured in a total of 27 miniature swine ranging in age from 5 days preterm to 5 weeks old. A steep increase in the forward rate constant for 31P exchange from phosphocreatine (PCr) to gamma-ATP was observed between 2 days preterm and 3 days postterm, with a more gradual increase for older ages. In contrast, the [PCr]/[NTP] ratio measured by in vivo 31P nuclear magnetic resonance (NMR) remained constant throughout this age interval and close to unity. Forward and reverse rate constants and the rate of flux for 31P exchange were equal to each other for both preterm and 5-week-old animals, suggesting that the creatine kinase reaction is near-equilibrium for this span of age. Multifrequency steady-state saturation of P(i) and PCr compared to single-frequency saturation of PCr produced the same extent of saturation transfer to gamma-ATP, and the saturation of P(i) alone had no effect on the gamma-ATP 31P NMR signal. These results suggest that even for immature swine brain, creatine kinase activity should be adequate to buffer against changes in [ATP] when there is a mismatch between energy supply and energy demand, during conditions such as ischemia or hypoxia. The results from the present study indicate the unlikelihood that previously reported discrepancies between forward and reverse 32P flux rates in rat brain (Shoubridge et al., FEBS Lett 140:288-292, 1982) were due to neglect of gamma-ATP to P(i) exchange.(ABSTRACT TRUNCATED AT 250 WORDS)

Modest hypothermia provides partial neuroprotection for ischemic neonatal brain.

Hypothermia is a frequent occurrence in newborns, and thermoregulatory management is a fundamental part of medical stabilization. Although modest reduction in brain temperature (2-3 degrees C) before ischemia provides neuroprotection in adults, the effect of modest hypothermia on immature brain has not been examined. Nine-day-old swine were exposed to 15 min of incomplete global brain ischemia, with intraischemic rectal temperatures of either 38.3 +/- 0.4 degrees C (n = 10, normothermic) or 35.4 +/- 0.5 degrees C (n = 10, hypothermic). The relationship between rectal and brain temperature was delineated in preliminary experiments on four swine. Animals with intraischemic rectal temperatures maintained at either 39.5 degrees C or 35.5 degrees C were associated with a similar magnitude of difference in brain temperature. Therefore, rectal temperature was used to monitor brain temperature for 20 animals studied subsequently. Ischemia was induced by combining neck compression with hemorrhagic hypotension and resulted in similar group values for mean arterial pressure and changes in pH and blood gases at the completion of ischemia. A clinical overall performance score and brain tissue structure were evaluated after 72 h (or earlier if animals died prematurely). Hypothermic animals had less severe stages of impairment compared with the normothermic group (p = 0.023). Hypothermic piglets had less histologic damage in the neocortex at 0.5 cm beneath the brain surface (p = 0.048), the caudate nucleus (p = 0.038), and the pons/midbrain (p = 0.04) and the same direction of effect in neocortex at 1 cm beneath the surface (p = 0.07) and the cerebellum (p = 0.07) as compared with normothermic animals.(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of systemic glucose concentration on brain metabolism following repeated brain ischemia.

Since systemic glucose concentration is an important determinant of ischemic brain metabolism in neonates, we sought to determine if the systemic glucose concentration influences brain metabolic alterations following repeated partial ischemia. A group of hyperglycemic piglets (n = 12) were compared to a group of modestly hypoglycemic piglets (n = 12) using in vivo 2H and 31P magnetic resonance spectroscopy to simultaneously measure cerebral blood flow and phosphorylated metabolites before, during and 30 min after two 10-min episodes of ischemia (i.e. Recovery 1 and 2). For both groups, beta-ATP levels at Recovery 1 and 2 were lower than Control (91 +/- 11 and 83 +/- 15% of Control, respectively for both groups combined, P = 0.002 vs Control). Inorganic phosphorus was elevated in hyperglycemic piglets at Recovery 1 and 2 (117 +/- 15 and 118 +/- 10% of Control). In contrast, in modestly hypoglycemic piglets inorganic phosphorus progressively rose from Recovery 1 (131 +/- 24% of Control) to Recovery 2 (149 +/- 37% of Control), and differed from the hyperglycemic group (P = 0.02). These changes did not correlate with post-ischemic cerebral blood flow, cerebral O2 delivery or cerebral glucose delivery. In both groups phosphocreatine and intracellular pH returned to Control values during Recovery 1 and 2. The progressive increase in inorganic phosphorus post-ischemia in hypoglycemic piglets suggests that modest hypoglycemia during and following repeated partial ischemia adversely affects immediate brain metabolic recovery.

The effect of hypercarbia on age-related changes in cerebral glucose transport and glucose-modulated agonal glycolytic rates.

This study examined the effect of hypercarbia on cerebral agonal glycolytic rates and brain lactate accumulation after complete ischemia induced by cardiac arrest. Before cardiac arrest, the blood plasma glucose concentration in seven newborn (113 d postconception; normal gestation, 115 d) and seven 1-mo-old (144 d postconception) piglets was adjusted to a specific value (range, 1 to 64 mM), and then inspired ventilation gases were changed to 10:50:40 CO2:O2:N2 for 20 min. The agonal glycolytic rate was measured by monitoring the rate of cerebral lactate formation in vivo using proton nuclear magnetic resonance spectroscopy, and postmortem brain lactate concentrations were measured biochemically in tissue extracts obtained 40 to 45 min after cardiac arrest. These data were compared with 21 normocarbic piglets of similar age, nine examined as part of the present study and 12 examined previously (Corbett RJT, Laptook AR, Ruley JI, Garcia D: Pediatr Res 30:579-586, 1991). There was a nonlinear relationship between the final postmortem brain lactate concentration and preischemia blood plasma glucose concentration that was most prominent in newborn piglets and previously had gone unnoticed. When analyzed using a steady-state model for glucose transport, this relationship revealed that normocarbic newborns had a lower preischemia affinity constant for the transport mechanism for glucose (2.8 +/- 1.5 mM) and lower cerebral glucose utilization rate relative to transport rate (0.12 +/- 0.04), compared with 1-mo-olds (4.5 +/- 1.4 mM and 0.30 +/- 0.03, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)