In Vivo EPR For Dosimetry.

As a result of terrorism, accident, or war, populations potentially can be exposed to doses of ionizing radiation that could cause direct clinical effects within days or weeks. There is a critical need to determine the magnitude of the exposure to individuals so that those with significant risk have appropriate procedures initiated immediately, while those without a significant probability of acute effects can be reassured and removed from the need for further consideration in the medical/emergency system. In many of the plausible scenarios there is an urgent need to make the determination very soon after the event and while the subject is still present. In vivo EPR measurements of radiation-induced changes in the enamel of teeth is a method, perhaps the only such method, which can differentiate among doses sufficiently for classifying individuals into categories for treatment with sufficient accuracy to facilitate decisions on medical treatment. In its current state, the in vivo EPR dosimeter can provide estimates of absorbed dose with an error approximately +/- 50 cGy over the range of interest for acute biological effects of radiation, assuming repeated measurements of the tooth in the mouth of the subject. The time required for acquisition, the lower limit, and the precision are expected to improve, with improvements in the resonator and the algorithm for acquiring and calculating the dose. The magnet system that is currently used, while potentially deployable, is somewhat large and heavy, requiring that it be mounted on a small truck or trailer. Several smaller magnets, including an intraoral magnet are under development, which would extend the ease of use of this technique.

Experimental Procedures for Sensitive and Reproducible In Situ EPR Tooth Dosimetry.

In vivo electron paramagnetic resonance (EPR) tooth dosimetry provides a means for non-invasive retrospective assessment of personal radiation exposure. While there is a clear need for such capabilities following radiation accidents, the most pressing need for the development of this technology is the heightened likelihood of terrorist events or nuclear conflicts. This technique will enable such measurements to be made at the site of an incident, while the subject is present, to assist emergency personnel as they perform triage for the affected population. At Dartmouth Medical School this development is currently being tested with normal volunteers with irradiated teeth placed in their mouths and with patients who have undergone radiation therapy. Here we describe progress in practical procedures to provide accurate and reproducible in vivo dose estimates.

Natural drug extracts for a nutritive-tonic drink, promotes the induction of long-term potentiation in rat hippocampal dentate gyrus in vivo.

We have shown previously that oral administration of a nutritive-tonic drink (NTD) improves scopolamine-induced memory impairment in the passive avoidance task and Morris water-maze in mice and that this action is attributable to the natural drug extracts, rather than synthetic drugs such as taurine and caffeine, in the NTD. In order to investigate the mechanism underlying the antiamnesic effects of the natural drug extracts, the effects of the extracts on the induction of long-term potentiation (LTP) was investigated in the dentate gyrus (DG) of normal and scopolamine-treated rats. Oral administration of natural drug extracts enhanced the induction of population spike amplitude induced by weak tetanic stimulation (30 pulses at 60 Hz). Scopolamine (0.3 mg/kg, i.p.) completely inhibited the induction of LTP induced by both weak and strong tetanic stimulation (100 pulses at 100 Hz). Natural drug extracts enhanced partially but significantly the induction of LTP by strong tetanus, but had a very weak effect on that induced by weak tetanus. These results suggest that LTP induced by strong tetanus is sensitive to natural drug extracts, and that the antiamnesic effect of the NTD is at least partly attributable to the LTP-improving effect of the natural drug extracts in the DG.

Delayed whole-body cooling to 33 or 35 degrees C and the development of impaired energy generation consequential to transient cerebral hypoxia-ischemia in the newborn piglet.

OBJECTIVES: Fundamental questions remain about the precise temperature providing optimal neuroprotection after perinatal hypoxia-ischemia (HI). Furthermore, if hypothermia delays the onset of the neurotoxic cascade and the secondary impairment in cerebral energy generation, the "latent phase" may be prolonged, thus extending the period when additional treatments may be effective. The aims of this study were to investigate the effects of delayed systemic cooling at either 33 degrees C or 35 degrees C on the following: (1) latent-phase duration, and (2) cerebral metabolism during secondary energy failure itself, in the 48-hour period after transient HI. METHODS: Piglets were randomly assigned to the following: (1) HI-normothermic (HI-n) rectal temperature (Trectal; n = 12), (2) HI-Trectal 35 degrees C (HI-35; n = 7), and (3) HI-Trectal 33 degrees C (HI-33; n = 10). Groups were cooled to the target Trectal between 2 and 26 hours after HI. Serial magnetic resonance spectroscopy was performed over 48 hours. The effect of cooling on secondary energy failure severity (indexed by the nucleotide triphosphate/exchangeable phosphate pool [NTP/EPP] and phosphocreatine/inorganic phosphate [PCr/Pi] ratios) was assessed. RESULTS: Compared with HI-n, HI-35 and HI-33 had a longer NTP/EPP latent phase and during the entire study duration had higher mean NTP/EPP and PCr/Pi. The latent phase (both PCr/Pi and NTP/EPP) and the whole-brain cerebral energetics were similar for HI-35 and HI-33. During the hypothermic period, compared with HI-n, PCr/Pi was preserved in the cooled groups, but this advantage was not maintained after rewarming. Compared with HI-n, HI-35 and HI-33 had higher NTP/EPP after rewarming. CONCLUSIONS: Whole-body hypothermia for 24 hours at either 35 or 33 degrees C, commenced 2 hours after resuscitation, prolonged the NTP/EPP latent phase and reduced the overall secondary falls in mean PCr/Pi and NTP/EPP during 48 hours after HI. Reducing the temperature from 35 to 33 degrees C neither increased mean PCr/Pi and NTP/EPP nor further lengthened the latent phase.

Effect of a nutritive-tonic drink on scopolamine-induced memory impairment in mice.

The effects of a liquid nutritive and tonic drug (NTD) selected from a modification of the "Kai-xin-shou-yu-shen-qi-wan" prescription, on scopolamine-induced amnesia in mice were investigated using the passive avoidance and water-maze tasks. A popular NTD in Japan that contains 17 crude (natural) drug extracts together with synthetic drugs such as taurine, caffeine, various vitamins and ethanol, and the natural drug extracts is based on a prescription of "Kampo" origin in Chinese medicine. Scopolamine (0.4 mg/kg, i.p.) reduces the step-through latency of the passive avoidance test and fear reaction behavior at 24 and 48 h after treatment. A single oral administration of the NTD (10 ml/kg) increased the step-through latency and the fear reaction behavior score in scopolamine-treated mice. Administration of the natural drug extracts found in the NTD tended to extend the step-through latency in the retention test at 48 h, but not 24 h after the initial scopolamine trial. However, administration of the synthetic drugs found in the NTD did not improve either the step-through latency or the behavioral score. The NTD and the natural drug extracts also improved the scopolamine-induced spatial memory impairment as assessed using the Morris water-maze test. In contrast, the synthetic drugs did not affect the escape latencies. Both NTD and the synthetic drugs increased the locomotor activity in scopolamine-treated mice, whereas the natural drug extracts did not. These results suggest that NTD improves scopolamine-induced amnesia, and that this action is attributable to the natural drug extracts in the NTD.

Depth of delayed cooling alters neuroprotection pattern after hypoxia-ischemia.

Hypothermia after perinatal hypoxia-ischemia (HI) is neuroprotective; the precise brain temperature that provides optimal protection is unknown. To assess the pattern of brain injury with 3 different rectal temperatures, we randomized 42 newborn piglets: (Group i) sham-normothermia (38.5-39 degrees C); (Group ii) sham-33 degrees C; (Group iii) HI-normothermia; (Group iv) HI-35 degrees C; and (Group v) HI-33 degrees C. Groups iii through v were subjected to transient HI insult. Groups ii, iv, and v were cooled to their target rectal temperatures between 2 and 26 hours after resuscitation. Experiments were terminated at 48 hours. Compared with normothermia, hypothermia at 35 degrees C led to 25 and 39% increases in neuronal viability in cortical gray matter (GM) and deep GM, respectively (both p < 0.05); hypothermia at 33 degrees C resulted in a 55% increase in neuronal viability in cortical GM (p < 0.01) but no significant increase in neuronal viability in deep GM. Comparing hypothermia at 35 and 33 degrees C, 35 degrees C resulted in more viable neurons in deep GM, whereas 33 degrees C resulted in more viable neurons in cortical GM (both p < 0.05). These results suggest that optimal neuroprotection by delayed hypothermia may occur at different temperatures in the cortical and deep GM. To obtain maximum benefit, you may need to design patient-specific hypothermia protocols by combining systemic and selective cooling.

Simultaneous NIR-EPR spectroscopy of rat brain oxygenation.

Changes in cerebral oxygenation were simultaneously monitored by electric paramagnetic resonance (EPR) oximetry and near-infrared spectroscopy (NIRS). The tissue oxygen tension (t-pO2) was measured with an L-band (1.2 GHz) EPR spectrometer with an external loop resonator and the concentration of oxyhemoglobin [HbO2] and deoxyhemoglobin [Hb] were measured with a full-spectral NIRS system. Mean cerebral hemoglobin saturation (SmcO2) was calculated from the absolute [HbO2] and [Hb]. Six adult male rats were implanted with lithium phthalocyanine (LiPc) crystals into the left cerebral cortex. The change in oxygenation of the brain was induced by altering the inspired oxygen fraction (FiO2) in air from 0.30 at baseline to 0.0, 0.05, 0.10, and 0.15 for 1, 2, 5, and 5 minutes, respectively, followed by reoxygenation with an FiO2 = 0.30. Although both t-pO2 and SmcO2 values showed a decrease during reduced FiO2 followed by recovery on reoxygenation, it was found that SmcO2 recovered more rapidly than t-PO2 during the recovery phase. The recovery of t-pO2 is not only related to blood oxygenation, but also to delivery, consumption, and diffusion of oxygen into the tissue from the vascular system. Further studies will be required to determine the exact mechanisms for the delay between the recovery of SmcO2 and t-pO2.

Effect of a nutritive-tonic drink on stress-induced serum glucocorticoid in the mouse.

We evaluated stress during "restraint with gnawing (R+G+)" and "restraint without gnawing (R+G-)" in a mouse system. R+G- induced a higher serum glucocorticoid level than R+G+. Zena F-III (a nutritive-tonic drink prescribed as "Kampo", a traditional Japanese medicine with its origin in Chinese medicine) reduced the glucocorticoid elevation in R+G+, but not in R+G-. These results support the hypotheses that (i) activity, such as gnawing, which potentially leading to escape from distress, reduces the severity of emotional distress and (ii) Zena F-III reduces the severity of mental or emotional fatigue, or increases motivation, in a stressful situation that the animal can manage itself.

Effect of nutritive and tonic crude drugs on physical fatigue-induced stress models in mice.

The present study was undertaken to investigate the acute anti-fatigue effect of a liquid nutritive and tonic crude drugs (NTDs) on stress induced in mice. After forced walking for 3 or 6h, the NTDs (applied orally, 10 ml/kg) significantly increased locomotor activity, while the administration of NTDs after rapid eye movement (REM) sleep deprivation stress and after immobilization stress did not show a specific effect, having a similar effect as the vehicle with added vitamins, taurine and caffeine. The administration of NTDs after freezing due to electric shock stress showed a specific effect which was not seen in other control groups, water, vehicle (ethanol) and vehicle including vitamins, taurine and caffeine and so resemble the specific effect of NTDs in the stress of forced walking. The present results indicate that the NTDs produced an anti-fatigue effect on the decreased locomotor activity after forced walking and immobility induced by electric stimulation. However, the crude drugs were not effective in improving immobility after sleep deprivation or immobilization stress.

Delayed hypothermia prevents decreases in N-acetylaspartate and reduced glutathione in the cerebral cortex of the neonatal pig following transient hypoxia-ischaemia.

The effects of normothermia and delayed hypothermia on the levels of N-acetylaspartate (NAA), reduced glutathione (GSH) and the activities of mitochondrial complex I, II-III, IV and citrate synthase were measured in brain homogenates obtained from anaesthetized neonatal pigs following transient in vivo hypoxia-ischaemia. In the normothermic animals there was a significant decrease in complex I activity and in the levels of GSH and NAA when compared to the controls. Delayed hypothermia preserved NAA and GSH at control levels and enhanced the rate of complex II-III activity. There was correlation (R = 0.79) between GSH and NAA levels when data from all three experimental groups were analyzed. Citrate synthase activity was not significantly different in the three groups, indicating maintenance of mitochondrial integrity. These data suggest that delayed hypothermia affords protection of integrated mitochondrial function in the neonatal brain following transient hypoxia-ischaemia.