Anoxic Response Research Articles

Neuronal selectivity of ATP-sensitive potassium channels in guinea-pig substantia nigra revealed by responses to anoxia.

1. Two sub-populations of pars compacta substantia nigra neurones were identified with very different electrophysiological properties and rostral-caudal distribution. Both cell types were identified by biocytin intracellular dye injection and found to be located within pars compacta containing tyrosine hydroxylase-positive cells. These sub-populations displayed distinctly different responses to transient anoxia. 2. The first group ('Phasic' neurones) exhibited a low threshold calcium conductance LTS gCa associated with bursts of action potentials, were located at the level of the mammillary bodies and were highly sensitive to anoxia. The second group ('rhythmic' neurones) fired in a rhythmic pattern, were located at the level of the accessory optic tract and were relatively insensitive to anoxia. 3. The anoxic response of phasic cells was characterized by membrane hyperpolarization (mean 12 mV), a decrease in input resistance (mean 36%) and cessation of action potential firing. The axonic response of these neurones was not blocked by TEA (5-10 mM), haloperidol (100 microM), the removal of extracellular calcium or depletion of endogenous dopamine. However, this effect was blocked by both the sulphonylurea tolbutamide (50-500 microM), and also by quinine (100 microM) and could be mimicked by application of diazoxide (1 mM). 4. Rhythmic cells displayed a variable response to anoxia consisting of either modest depolarization, hyperpolarization or no change in membrane potential, in all cases accompanied by little or no change in input resistance. The polarity of the membrane potential shift during anoxia was reversed by TEA (5-10 mM) or the removal of calcium. These cells were also relatively insensitive to diazoxide (1 mM). 5. It is concluded from the neuronal responses to anoxia and the pharmacological modification of these responses, that the ATP-sensitive potassium channel (KATP channel) is functionally operative in the substantia nigra and is primarily distributed on the phasically discharging cells of the rostral pars compacta. The relevance of this recently discovered ionic channel is discussed with regard to the normal and abnormal functioning of the substantia nigra.

Hypoxia-induced functional alterations in adult rat neocortex.

1. Brief periods of hypoxia (2-7 min) were induced in rat neocortical slices maintained in an interface-type recording chamber at 34-35 degrees C by changing the aerating gas from 95% O2-5% CO2 to 95% N2-5% CO2. Field potential (FP) and intracellular recordings were obtained in layers II/III of primary somatosensory cortex. Intracellular injection of biocytin revealed the characteristic morphology of supragranular spiny pyramidal neurons. 2. Excitatory synaptic transmission reversibly decreased by 45% as estimated from FP responses to orthodromic stimulation of the underlying white matter/layer VI. Excitatory postsynaptic potentials (EPSPs) were suppressed by 36% in amplitude and recovered within 2-3 min after reoxygenation. During the recovery period, EPSPs showed a reversible increase in duration by 72%. 3. Inhibitory synaptic transmission was completely blocked as determined in FP responses with a paired-pulse inhibition protocol. The fast inhibitory postsynaptic potential (IPSP) declined by 58% during hypoxia. The long-lasting IPSP was suppressed by 75% and showed incomplete recovery. During hypoxia, the amplitude of both IPSPs was significantly more strongly suppressed than the EPSP. 4. In 40% of the cells, hypoxia induced an early anoxic hyperpolarization with a reversal potential of E = -80.8 mV, followed by a postanoxic hyperpolarization (E = -89.4 mV). In a second group of cells (37%), a gradual anoxic depolarization with E = -57.5 mV was observed instead of an early hyperpolarization. In both groups of cells, the anoxic response was associated with a marked decrease in input resistance, by 42 and 31%, respectively. 5. The spike discharge frequency was reversibly suppressed by 71% during hypoxia. A transient hyperexcitability accompanied with a rise in input resistance and discharge rate was observed in 38% of the cells on reoxygenation. 6. The reversal potential of the anoxic hyperpolarization was unaffected by tetrodotoxin (TTX) but was significantly altered by application of the ATP-sensitive K+ channel (KATP) blocker gliquidone. Application of gliquidone additionally resulted in a significantly smaller hypoxia-induced decline in paired-pulse inhibition. 7. Increases in tissue high-energy phosphates induced by preincubating the slices in 25 mM creatine for greater than 2 h had a pronounced protective effect on excitatory and inhibitory synaptic transmission. 8. These data suggest a selective vulnerability of the neocortical inhibitory system during hypoxia. Our results further indicate that hypoxia activates a pre- and postsynaptic KATP conductance because of the decline in intracellular ATP.

Effect of glibenclamide, a selective blocker of an ATP-K+ channel, on the anoxic response of hippocampal neurones

Effect of glibenclamide, a selective blocker of an ATP-K+ channel, on the anoxic response of hippocampal neurones

Effects of nitroglycerin, dipyridamole, nifedipine, verapamil and diltiazem on canine coronary arterial rings contracted with 5-hydroxytryptamine and anoxia.

Anoxia has been shown to potentiate the constrictor effects of 5-hydroxytryptamine (5HT) in isolated vascular tissue. In the present study, canine coronary arterial rings were incubated with various treatments and exposed to 5HT (4 X 10(-7) M) and anoxia (95% N2 and 5% CO2). Developed tension was increased by 250 +/- 40 mg by 5HT alone and 2,000 +/- 90 mg by 5HT and anoxia. Calcium (5 mM) potentiated, while inorganic (lanthanum, 10(-2) M) and organic calcium antagonists (nifedipine, verapamil and diltiazem; IC50 = 7 X 10(-9), 7.3 X 10(-8) and 2.4 X 10(-7) M, respectively) blocked the anoxic potentiation. Anoxia alone decreased resting tension (RT). Methysergide 3 X 10(-5) M inhibited both the 5HT- and anoxia-potentiated responses. Nitroglycerin decreased RT and inhibited the anoxic response (IC50 = 7.6 X 10(-6) M), while dipyridamole decreased RT and did not affect the anoxic response. These data suggest that the potentiation of 5HT contraction by anoxia is dependent upon extracellular calcium influx and is linked to a 5HT receptor. In addition, inhibition of the anoxic response can be achieved at other sites and is not a property common to all coronary vasodilators.

Oxygen Sensitization of CHO Cells at Ultrahigh Dose Rates: Prelude to Oxygen Diffusion Studies

The response of cultured CHO cells to ultrahigh dose rate (approx.10/sup 11/ rad/sec) radiation, as measured by colony-forming ability, was studied under various oxygen concentrations with single electron pulses of 3 nsec duration. A thin-layer technique was designed for these experiments to permit rapid equilibration of the cells with the ambient oxygen-containing gas and to ensure that no segments of the cell population were shielded from the relatively low energy electrons (average energy, approx.450 keV), thus eliminating the possibility of artifacts influencing surviving fraction. Dosimetry was performed using a combination of calorimetric, charge-measuring, and thermoluminescent techniques. At a concentration of 0.44% oxygen, breaking behavior of the survival curve was observed in accord with the radiolytic depletion of intracellular oxygen by the radiation pulse. The sensitivity for doses greater than the breakpoint dose followed an anoxic response, clearly different from that observed when the cells were irradiated at conventional dose rates in the same gas under the identical geometry. The breakpoint dose was found to be approximately linear over the oxygen concentration range 0 to 0.7%. These results are consistent with recent observations of mammalian cell response under high-intensity irradiation conditions and provide a basis for the design of a double-pulsemore » experiment.« less

Two components in the radiation sensitization of bacterial spores by p-nitroacetophenone: the -OH component.

p-Nitroacetophenone (PNAP) sensitizes Bacillus megaterium spores under anoxic conditions to the lethal effects of 50 kVp X-rays. Concentrations between approximately 5 X 10(-4) M and 3-8 X 10(-3) M produce the maximum effect, an increase of about 30 per cent over the anoxic response when the spores are irradiated in water. Compounds that scavenge -OH decrease, but cannot completely eliminate, this maximum amount of sensitization. These results indicate that PNAP acts to increase spores' radiation sensitivity through two separable types of chemical reactions: one which involves -OH and one which does not. Possible mechanisms responsible for these two components of damage are discussed. In these experiments 1/15 M phosphate buffer acts in several unexpected ways. This concentration itself increased the anoxic spore response by about 9 per cent (relative to the anoxic response in water). In addition, although the maximum amounts of sensitization were the same, the amounts of sensitization from lower PNAP concentrations differed when the suspending fluid was buffer instead of water. An interaction was also seen during the PNAP-t-butanol experiments; again, the responses at low PNAP concentrations were different in buffer and in water. No mechanisms for these actions of this buffer were suggested, although somewhat similar effects may occur with other organisms. Clearly, such effects must be recognized and evaluated before quantitative analyses of the actions of sensitivity-modifying agents can be completed.

Additive Effects in the Radiosensitization of B. megaterium Spores by p-Nitroacetophenone and Norpseudopelletierine-N-Oxyl

Norpseudopelletierine-N-oxyl (NPPN) has been shown to increase the sensitivity of B. megaterium spores when irradiated in anoxic buffered suspensions. The maximum sensitization achieved by NPPN alone corresponded to a 58% increase over the simple anoxic response. This compares with a 40% increase produced by paranitroacetophenone (PNAP) and 115% increase shown by oxygen. Like PNAP, NPPN is capable of producing a small measure of sensitization by interfering with water radiolysis intermediates (indirect affects) when used at low concentrations. At high concentrations, at which NPPN exerts its maximum degree of sensitization, it is concluded that the compound operates by increasing the lethality of the direct effects of radiation on the cellular target. The use of mixtures of NPPN and PNAP produces a greater degree of sensitization than can be achieved by either alone, at least a 91% increase over the anoxic sensitivity being observed. It is concluded that these two sensitizers, chosen as representatives of...

Increased Response of AnoxicBacillus MegateriumSpores to Radiation at High Dose-rates

SummaryBacillus megaterium spores have been irradiated in aqueous suspension with 5 μs pulses of electrons. The sensitivity of anoxic spores (N2) increases with increasing dose-rate in the range 10–10 000 rads/μs. The response of oxic spores (O2) is unaffected by these changes in dose-rate. OH radical scavengers prevent part of the dose-rate effect. N2O sensitizes spores at low dose-rates but not at high dose-rates. Reduction in the pulse length over the range 5 to 0·5 μs causes a decrease in the response characteristic of high dose-rates. It is estimated that the average lifetime of the radicals effective in increasing the anoxic response is about 2 to 5 μs.

"Anoxic" radiation response in cultured mammalian cells.

There have been recent expressions of caution concerning the adequacy of methods of oxygen removal from biological systems under test for anoxic radiation response. Baker and Town (1966) and Michael, Scott and Revesz (1966) have reported that the time for liquid-gas equilibration requires hours. These communications did not contain information on cell survival to radiation during the time-course of gassing. The purpose of this letter is to present results of experiments with cultured Chinese hamster cells (V79–103A) which bear on some of the questions raised by these authors. A known number of single cells were plated on to 9 cm glass Petri dishes and allowed to grow overnight to an average colony multiplicity (N) of approximately 1·7. At the time of irradiation, medium was removed from the dish and the latter, without its cover, was placed in a holder similar to that reported by Elkind and Sutton (1960). The amount of residual medium was less than 0·5 ml. The holder allowed for temperature control below ...