I. S. Kass

Protection of hippocampal slices from young rats against anoxic transmission damage is due to better maintenance of ATP.

1. Dentate granule cells in hippocampal slices from young rats (aged 30‐40 days) are more resistant to damage from 10 min of anoxia than are granule cells from adult rats. The evoked population spike from these cells recovers to 78% of its pre‐anoxic amplitude in young animals while in adult animals it shows only 4% recovery. This increased resistance is associated with higher levels of adenosine triphosphate (ATP) during the anoxic period. 2. When the duration of anoxia in slices from young animals is increased to 15 min, ATP falls to levels found in adult tissue after 10 min of anoxia. The dentate granule cells in slices from young animals show little recovery of the evoked response (19%) after such an exposure to anoxia. 3. When slices from young animals are subjected to 10 min of anoxia in low‐glucose (2 mM) artificial cerebrospinal fluid, ATP levels fall to those found in adult tissue after 10 min of anoxia and the evoked response from the dentate granule cells again shows little recovery (10%). 4. The evoked response in the CA1 pyramidal cell layer of slices from young rats is more resistant to damage from 5 or 7 min anoxia than it is in slices from adults. Thus this region, also, shows an age‐dependent increase in susceptibility to anoxic damage. ATP levels in the CA1 region of tissue from young animals at the end of 5 and 7 min anoxia are greater than ATP levels in tissue from adult animals after these same anoxic exposures. 5. Basal levels of 45Ca accumulation are greater in CA1 and dentate gyrus from young rats. However, the percentage increases during 10 min of anoxia are less than one‐half the values in slices from adult animals. 6. The results suggest that the increased resistance of slices from young animals to anoxic transmission damage may be explained by the better maintenance of ATP in synaptic regions of these slices during anoxia. This may confer the increased resistance by lowering the anoxic increase in cell Ca2+.

The importance of sodium for anoxic transmission damage in rat hippocampal slices: mechanisms of protection by lidocaine.

1. The effect of sodium influx on anoxic damage was investigated in rat hippocampal slices. Previous experiments demonstrated that a concentration of tetrodotoxin which blocks neuronal transmission protects against anoxic damage. In this study we examined low concentrations of lidocaine (lignocaine; which do not block neuronal transmission), for their effect on recovery of the evoked population spike recorded from the CA1 pyramidal cell layer. 2. Recovery of the population spike, measured 60 min after a 5 min anoxic period, was 4 +/‐ 2% of its preanoxic, predrug level. Lidocaine concentrations of 10, 50, and 100 microM significantly improved recovery to 56 +/‐ 12, 80 +/‐ 7 and 70 +/‐ 14%, respectively. 3. Lidocaine (10 microM) did not alter the size of the evoked response before anoxia and had no significant effect on potassium levels or calcium influx during anoxia. It did, however, reduce cellular sodium levels (146 +/‐ 7 vs. 202 +/‐ 12 nmol mg‐1) and preserve ATP levels (2.17 +/‐ 0.07 vs. 1.78 +/‐ 0.07 nmol mg‐1) during anoxia. All values were measured at the end of 5 min of anoxia except those for Ca2+ influx which were measured during 10 min of anoxia. 4. High concentrations of lidocaine (100 microM) did not improve recovery significantly over that observed with 10 microM. They also had no significantly greater effects on sodium levels than 10 microM lidocaine (137 +/‐ 12 vs. 146 +/‐ 7 nmol mg‐1); however, 100 microM lidocaine significantly improved potassium (202 +/‐ 18 vs. 145 +/‐ 6 nmol mg‐1) and ATP (2.57 +/‐ 0.06 vs. 2.17 +/‐ 0.07 nmol mg‐1) levels, while reducing calcium influx (7.76 +/‐ 0.12 vs. 9.24 +/‐ 0.39 nmol mg‐1 (10 min)‐1) when compared with 10 microM lidocaine. 5. We conclude that sodium influx and ATP depletion are of major importance in anoxic damage since 10 microM lidocaine reduced these changes during anoxia and improved recovery of the population spike. In addition, our results indicate that the properties of the sodium channel are altered during anoxia, since sodium influx is blocked by a concentration of lidocaine that does not affect the population spike in the preanoxic period.

The barbiturate thiopental reduces ATP levels during anoxia but improves electrophysiological recovery and ionic homeostasis in the rat hippocampal slice

The barbiturate anesthetic thiopental enhances recovery of the evoked population spike recorded from rat hippocampal slices after short periods of anoxia. Thiopental reduces changes in sodium, potassium and calcium but enhances the fall in ATP levels during anoxia. The postsynaptic population spike recorded from the CA1 pyramidal cell region of the slices treated with thiopental (600 microM) recovered to 67% of the preanoxic amplitude after 3.5 min of anoxia. There was less recovery (24%) when a lower concentration of thiopental (250 microM) was used. Untreated slices recovered to only 10% of their preanoxic amplitude after 3.5 min of anoxia. Other studies have demonstrated that maintaining ATP levels during anoxia may be an important mechanism of protection. In contrast to those studies, thiopental was protective although it enhanced the fall of ATP levels after 3.5 min of anoxia in the CA1 region and after 3.5 and 5 min in the dentate region. Thus enhanced recovery of the population spike with thiopental is not due to its preservation of ATP levels. This result allows a clear separation of improved ATP levels during anoxia from other mechanisms of protection. We therefore looked for other mechanisms of protection. Sodium and potassium levels were measured after 10 min of anoxia. In untreated tissue, sodium levels in the slice rose and potassium levels fell significantly. In thiopental-treated tissue, changes in sodium and potassium caused by anoxia and by veratridine under normoxic conditions were significantly reduced. During anoxia calcium-45 uptake increases; thiopental significantly reduces this uptake.(ABSTRACT TRUNCATED AT 250 WORDS)

The N-methyl-d-aspartate antagonists aminophosphonovaleric acid and MK-801 reduce anoxic damage to dentate granule and CA1 pyramidal cells in the rat hippocampal slice

The effect of the N-methyl-D-aspartate antagonists, aminophosphonovaleric acid and MK-801, on irreversible transmission loss subsequent to anoxia was examined using the hippocampal slice preparation. A population spike was recorded from either the dentate granule cells or the CA1 pyramidal cells and the amplitude of this spike was compared before and 60 min following anoxia. After 10 min of anoxia the dentate granule cells recovered to 16 +/- 7% (mean +/- SE) of their preanoxic level when untreated and to 54 +/- 15% when treated with aminophosphonovaleric acid (APV). In slices treated with MK-801 the population spikes recorded from dentate granule cells recovered to 85 +/- 4% of their preanoxic level after 10 min of anoxia. Untreated CA1 pyramidal cells recovered to 8 +/- 3% of their preanoxic amplitude after 5 min of anoxia; they recovered to 59 +/- 6% when treated with MK-801 and 31 +/- 13% when treated with APV. The recovery of slices treated with the drugs was significantly different from that of untreated slices. ATP levels were measured in both the dentate and the CA1 region of slices. ATP in both regions fell less during anoxia when the slices were pretreated with either APV or MK-801. These differences between drug-treated and untreated tissue were significant with APV and MK-801. These differences between drug-treated and untreated tissue were significant with APV and MK-801 in dentate tissue after 10 min of anoxia and with MK-801 in CA1 tissue after 5 min of anoxia. This reduced fall in ATP during anoxia was accompanied by better physiological recovery after anoxia. We conclude that these NMDA antagonists provide protection against anoxic damage to dentate granule and CA1 pyramidal cells in this in vitro hippocampal preparation.(ABSTRACT TRUNCATED AT 250 WORDS)

Thiopental attenuates hypoxic changes of electrophysiology, biochemistry, and morphology in rat hippocampal slice CA1 pyramidal cells.

BACKGROUND AND PURPOSE Thiopental has been shown to protect against cerebral ischemic damage; however, it has undesirable side effects. We have examined how thiopental alters histological, physiological, and biochemical changes during and after hypoxia. These experiments should enable the discovery of agents that share some of the beneficial effects of thiopental. METHODS We made intracellular recordings and measured ATP, sodium, potassium, and calcium concentrations from CA1 pyramidal cells in rat hippocampal slices subjected to 10 minutes of hypoxia with and without 600 micromol/L thiopental. RESULTS Thiopental delayed the time until complete depolarization (21+/-3 versus 11+/-2 minutes for treated versus untreated slices, respectively) and attenuated the level of depolarization at 10 minutes of hypoxia (-33+/-6 versus -12+/-5 mV). There was improved recovery of the resting potential after 10 minutes of hypoxia in slices treated with thiopental (89% versus 31% recovery). Thiopental attenuated the changes in sodium (140% versus 193% of prehypoxic concentration), potassium (62% versus 46%), and calcium (111% versus 197%) during 10 minutes of hypoxia. There was only a small effect on ATP (18% versus 8%). The percentage of cells showing clear histological damage was decreased by thiopental (45% versus 71%), and thiopental improved protein synthesis after hypoxia (75% versus 20%). CONCLUSIONS Thiopental attenuates neuronal depolarization, an increase in cellular sodium and calcium concentrations, and a decrease in cellular potassium and ATP concentrations during hypoxia. These effects may explain the reduced histological, protein synthetic, and electrophysiological damage to CA1 pyramidal cells after hypoxia with thiopental.

Effect of small changes in temperature on CA1 pyramidal cells from rat hippocampal slices during hypoxia: implications about the mechanism of hypothermic protection against neuronal damage

Small reductions in temperature have been shown to improve neurologic recovery after ischemia. We have examined the effect of temperature on biochemical and physiological changes during hypoxia using rat hippocampal slices as a model system. The postsynaptic population spike recorded from the CA1 pyramidal cell region of slices subjected to 7 min of hypoxia with hypothermia (34 degrees C) recovered to 73% of its prehypoxic level; slices subjected to the same period of hypoxia at 37 degrees C did not recover. After 7 min of hypoxia ATP fell to 48% of its prehypoxic concentration at 34 degrees C and 30% at 37 degrees C. Potassium fell to 86% during 7 min of hypoxia with hypothermia, this compares to a fall to 58% at 37 degrees C. The increase in sodium after 7 min of hypoxia was also attenuated by hypothermia (133% vs. 163% of its prehypoxic concentration). When the hypoxic period was shortened to 3 min (37 degrees C) the population spike recovered to 94%. If the temperature was increased to 40 degrees C there was only 7% recovery of the population spike after 3 min of hypoxia. With hyperthermia (40 degrees C), ATP fell to 33% after 3 min of hypoxia, this compares to 81% at normothermia. Potassium fell to 76% after 3 min of hypoxia with hyperthermia, this compares to 91% at 37 degrees C. Sodium concentrations increased with hyperthermia before hypoxia, at 3 min of hypoxia there was no significant difference between the hyperthermic and normothermic tissue; there was a large increase in sodium with hyperthermia after 5 min of hypoxia (209% vs. 146%). We conclude that the improved recovery after hypothermic hypoxia is at least in part due to the attenuated changes in ATP, potassium and sodium during hypoxia and that the worsened recovery with hyperthermia is due to an exacerbation of the change in ATP, potassium and sodium concentrations during hypoxia.

Effect of α-tocopherol and free radicals on anoxic damage in the rat hippocampal slice

Abstract The effect of α-tocopherol on irreversible transmission loss subsequent to anoxia was examined using the hippocampal slice preparation. A population spike was recorded from the dentate granule cell layer after stimulation of the perforant path. The amplitude of the population spike was compared before and after anoxia. Control slices recovered to 22 ± 7% of their preanoxic amplitude after 7 min of anoxia. Slices treated with α-tocopherol showed significantly greater recovery after anoxia (P

Anesthetic protection against anoxic damage in the rat hippocampal slice.

Evoked population spikes were recorded from the dentate granule cell layer of hippocampal slices obtained from adult rats. These slices were subjected to short periods of anoxia in the presence of different anesthetics. The recovery of the population spike after anoxia was compared across treatments. Little or no recovery was found after 10 min of anoxia when no anesthetic (4 +/- 4%), 1.5% isoflurane (5 +/- 5%), or 15% isoflurane (0 +/- 0%) was present during the anoxic periods. However, the population spike did recover to 81 +/- 7% of its preanoxic amplitude within 1 h after the anoxia if thiopental 160 mg/liter was present in the perfusate during the anoxia. Fifteen percent isoflurane and 160 mg/liter thiopental were equipotent in reducing the amplitude of the evoked population spike before anoxia but only thiopental protected against the damage after 10 min of anoxia. Our results suggest that the blocking of the evoked population spike by thiopental is not the sole mechanism of its protection against anoxic damage. Isoflurane (1.5%) was able to provide a small degree of protection against shorter periods (7 min) of anoxia.

Desflurane improves the recovery of the evoked postsynaptic population spike from CA1 pyramidal cells after hypoxia in rat hippocampal slices.

Desflurane is a volatile anesthetic that allows rapid induction and emergence, reduces cerebral metabolism, and enhances tissue perfusion. We studied the effect of treatment with 4%, 6%, and 12% desflurane on hypoxic neuronal damage by comparing the size of the postsynaptic evoked population spike recorded from the cornu ammonis 1 (CA1) pyramidal cell layer of rat hippocampal slices before and 2 hours after a hypoxic insult. When the tissue was treated with 6% desflurane before, during, and after 3.5 minutes of hypoxia, recovery was significantly better in slices exposed to desflurane (37% ± 9%) compared with untreated hypoxic slices (15% ± 5%). A lower (4%) or higher (12%) concentration of desflurane did not significantly improve recovery after 3.5 minutes of hypoxia. In the period before hypoxia, 12% and 6% desflurane significantly increased the latency and decreased the amplitude of the postsynaptic population spike; 4% desflurane had a similar but nonsignificant effect on latency and amplitude. We conclude that 6% desflurane, a clinically useful concentration (1 minimal alveolar concentration), improved the recovery of postsynaptic evoked responses in rat hippocampal slices after 3.5 minutes of hypoxia. In vivo studies must be conducted to assess the potential clinical significance of 6% desfluranes neuroprotective activity.

Anoxia reduces depolarization induced calcium uptake in the rat hippocampal slice

Veratridine-induced depolarization caused a large increase in Ca uptake in the rat hippocampal slice (30.2 vs. 9.0 nM/mg dry weight). This uptake was reduced to 18.4 nM/mg when veratridine was combined with anoxia. When compared with veratridine exposure alone, the combination of anoxia and veratridine increased intracellular Na (460 vs. 380 microM/g), decreased intracellular K (30 vs. 40 microM/g) and decreased ATP levels (0.1 vs. 0.8 nM/mg). The changes in Na, K, and ATP should enhance net Ca uptake, yet Ca uptake was reduced. This suggests an effect of anoxia to block Ca channels. In summary anoxia attenuates depolarization-induced Ca uptake. This may represent a mechanism by which neurons are partially protected against anoxic damage which could be more severe if depolarization-induced Ca uptake was not limited.