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Biomedical subjects

T J Sick

Publications and source records attributed to T J Sick.

At least 55 records · Page 3Linked to original sources
PubMed1989

Indo-1 measurements of intracellular free calcium in the hippocampal slice: complications of labile NADH fluorescence.

The use of fluorescent calcium indicator, Indo-1, was evaluated for measuring changes in intracellular free calcium during electrical stimulation and anoxia in hippocampal slices. Fluorescence was measured from slices illuminated with brief (3 ns) light pulses (337 nm wavelength) from a nitrogen laser. This method of illumination produced more intense fluorescence than illumination with light from filtered xenon or mercury arc lamps, and prevented loss of electrical excitability encountered following continuous UV illumination. Background fluorescence in control slices (without Indo-1) was considerable, often approaching 50% of that obtainable after dye loading. A more serious concern, however, was that a large fraction (approximately 10%) of the background fluorescence was labile to both electrical stimulation and anoxia. This fluorescence results from changes in the reduction/oxidation (redox) state of nicotinamide adenine dinucleotide (NAD), which fluoresces in its reduced (NADH) but not oxidized (NAD+) form. Qualitative changes in free calcium could be measured by first determining the ratio of change in NADH fluorescence at 405 and 485 nm (the wavelengths of light used to measure calcium with Indo-1) prior to dye loading. Any arbitrary baseline could be selected as long as the ratio for the baseline at 405 and 485 nm was identical to that determined for labile NADH. By application of this compensation procedure, it was determined that intracellular calcium rose abruptly during the onset of anoxia and again during the spreading depression-like loss of ion homeostasis which inevitably occurred during anoxia in these slices.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Determination of intracellular pH in the in vitro hippocampal slice preparation by transillumination spectrophotometry of neutral red.

A method is presented for measuring intracellular pH (pHi) spectrophotometrically in hippocampal slices using the pH dye indicator, Neutral red (NR). Measurements of pHi by NR were compared directly with the creatine kinase (CK) equilibrium method. Slices were bathed in artificial cerebrospinal fluid buffered with bicarbonate/CO2. Intracellular pH in hippocampal slices was found to be more alkaline (approximately 0.3 pH units) than buffer pH and far more alkaline (0.5 pH units) than extracellular pH. Resting intracellular alkalinity was observed by using both the NR and the CK equilibrium methods. The method may be useful for studies of pH regulation in intact functioning tissues in vitro where rapid and repeated measurements are necessary or where cell size precludes measurements with pH-sensitive microelectrodes.

Adenosine Triphosphate↗
PubMed1989

Energy metabolism, ion homeostasis, and evoked potentials in anoxic turtle brain.

Unlike the mammalian brain, the turtle brain maintains ion homeostasis and avoids anoxic depolarization during prolonged anoxia. The mechanisms by which turtle brains avoid and recover from anoxic depolarization were investigated by relating changes in cellular ATP levels to extracellular K+ (K+o) and evoked potential activities after 6 h of anoxia and during anoxic depolarization induced by superfusion of the brain surface with iodoacetate during anoxia. ATP levels were maintained during 6 h of anoxia despite marked decreases in energy production, but ATP was lowered during inhibition of both oxidative phosphorylation and glycolysis. K+o activity was unchanged during 6 h of anoxia but increased when ATP levels decreased. Ion homeostasis in the turtle brain appears to be linked to ATP levels but is unaffected by rate of energy production. In contrast, amplitudes of evoked potentials decreased when ATP levels were maintained, but rate of energy production declined. ATP and K+o levels recovered after anoxic depolarization, but evoked potential activity was not restored. These data suggest that the basic strategy for turtle brain to survive anoxia is to avoid anoxic depolarization by maintaining ATP levels.

Animals↗
PubMed1989

Ion leakage is reduced during anoxia in turtle brain: a potential survival strategy.

The turtle brain maintains ATP levels, ion homeostasis, and some neural function during prolonged anoxia despite significant decreases in the rate of ATP production. To test the hypothesis that reduced ATP production during anoxia was compensated partly by conserving energy through reduced ion leakage, the rate of K+ leakage was measured in normoxic and anoxic turtle brains in which Na(+)-K(+)-adenosinetriphosphatase was inhibited with ouabain. Rates of K+ leakage were significantly lower in brains subjected to 2 h of anoxia than in normoxic brains. These data support the suggestion that reduced ion leakage is an important mechanism for energy conservation during anoxia.

Acclimatization↗
PubMed1988

MPP+-induced increases in extracellular potassium ion activity in rat striatal slices suggest that consequences of MPP+ neurotoxicity are spread beyond dopaminergic terminals.

MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) produces symptoms similar to idiopathic Parkinson's disease in primates. A metabolite of MPTP, MPP+ (1-methyl-4-phenylpyridinium), is actively accumulated by dopaminergic (DA) terminals and selectively destroys nigrostriatal DA neurons. The mechanism of this effect remains unknown but reports that MPP+ inhibits electron transport in isolated mitochondria and increases oxidation of cytochrome b in striatal slices suggest that depression of ATP production is involved. To relate metabolic effects of MPP+ with tissue electrophysiology, extracellular potassium ion activity [K+]o was measured by microelectrodes simultaneous to optical monitoring of reduction/oxidation (redox) activity of cytochrome b during superfusion of MPP+ onto rat striatal and hippocampal slices. MPP+ increased oxidation of cytochrome b and increased [K+]o in slices of striatum. These increases were greater than expected from a selective effect of MPP+ on DA terminals which likely comprise no more than 3% of the total striatal mass. These effects of MPP+ were slowed by a dopamine uptake inhibitor (mazindol) and did not occur in hippocampal slices. These findings indicate that MPP+ influences ion transport as well as metabolic activity and that these actions require the presence of functioning DA terminals. However, the large amplitudes of the MPP+-induced changes suggest that consequences of MPP+-neurotoxicity are not ultimately confined to DA terminals. Two hypothesis are proposed: that energy failure in DA terminals results in leakage of neurotoxic substances or metabolites altering membrane conductance properties of adjacent cells and thereby placing additional demand upon ion transport pumps and mitochondrial oxidative phosphorylation; or that there is secondary uptake of MPP+ leading to mitochondrial inhibition in cells neighboring DA terminals.

1-Methyl-4-phenylpyridinium↗
PubMed1988

Calcium-sensitive recovery of extracellular potassium and synaptic transmission in rat hippocampal slices exposed to brief anoxia.

We examined the possibility that Ca2+-sensitive inhibition of synaptic transmission following anoxia involves compromise of ion transport activity. Rat hippocampal slices were superfused with artificial cerebrospinal fluids containing different concentrations of CaCl2, and subjected to short anoxia. Durations of anoxia were sufficient to provoke anoxic depolarization, indicated by a sudden rise in extracellular K+ (K+o). Following anoxia, apparent K+ transport was assessed by measuring the magnitude of subnormal K+o (the K+o undershoot) in hippocampal region CA1. Recovery of synaptic transmission 1 h after anoxia was determined by evaluation of the magnitudes of the orthodromically stimulated population spike recorded from CA1 pyramidal cells. K+o undershoots and recovery of synaptic transmission decreased as CaCl2 or the duration of anoxic depolarization increased. These data suggest: (1) that increased artificial cerebrospinal fluid CaCl2 compromised K+ reaccumulation after anoxia; and (2) that ion transport dysfunction may inhibit recovery of synaptic transmission.

Action Potentials↗
PubMed1988

Depth profile of local oxygen tension and blood flow in rat cerebral cortex, white matter and hippocampus.

Microregional oxygenation and blood flow were measured in rat cerebral cortical lamina, subcortical white matter and hippocampus. This was done to examine relationships between oxygenation and blood flow at a local level, to determine effects of craniotomy, and to consider whether flow/oxygenation relationships might be predictive of selective vulnerability known to accompany anoxia or ischemia. Blood flow and oxygen tension were measured with closely apposed polarographic microelectrodes. Oxygen tension was highest in white matter, lower in the region of cortical laminae IV-V and lowest in the hippocampus. Blood flow in the hippocampus was higher than that in white matter or laminae IV-V of cerebral cortex. Ratios of blood flow to oxygenation were similar throughout the cortex, higher in white matter but oxygenation in hippocampus was significantly less than expected from measurement of hippocampal blood flow reflecting increased oxygen consumption or relative hypoxia due to increased diffusion distances for oxygen in hippocampus. Comparison of data from closed vs opened skull animals indicated that diffusion of oxygen and hydrogen influenced data to approximately 1000 micron below the cerebral surface.

Animals↗
PubMed1988

1-Methyl-4-phenylpyridinium (MPP+) increases oxidation of cytochrome-b in rat striatal slices.

Effects of 1-methyl-4-phenylpyridinium, (the active metabolite of the neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine), on reduction/oxidation activity of mitochondrial cytochromes were studied in rat striatal slices using scanning spectrophotometry. The objective was to test the hypothesis that the neurotoxin alters electron transport in the mitochondrial respiratory chain. Incubation of rat striatal slices with MPP+ (1 microM) produced a time-dependent oxidation of Cytochrome-b in a manner consistent with the concept of a block in electron transport in the intramitochondrial respiratory chain between nicotinamide adenine dinucleotide (NAD) and Cytochrome-b. This effect of MPP+ was decreased by co-incubation with a potent dopamine uptake inhibitor (mazindol), or when studied in a tissue with low dopaminergic innervation (hippocampus). The amplitude of Cytochrome-b oxidation was greater than that expected from a selective effect of MPP+ on dopaminergic neurons suggesting that neighboring cells are influenced secondary to the MPP+ effect on dopaminergic terminals.

1-Methyl-4-phenylpyridinium↗
PubMed1988

Inhibition of glycolysis alters potassium ion transport and mitochondrial redox activity in rat brain.

To examine the relationships between brain glycolysis, ion transport, and mitochondrial reduction/oxidation (redox) activity, extracellular potassium ion activity (K+0) and redox shifts of cytochrome oxidase (cytochrome a,a3) were recorded previous to and during superfusion of rat cerebral cortex with the glycolytic inhibitor iodoacetic acid (IAA). IAA produced oxidation of cytochrome a,a3, increased local oxygenation, increased K+0, and, in response to neuronal activation, slowed rates of K+0 reaccumulation. Rates of rereduction of cytochrome a,a3, after the oxidation of this cytochrome by stimulation, were also slowed by IAA. These effects of IAA demonstrate the dependence of K+0 reaccumulation on the integrity of glycolysis, support the concept that active processes are involved in brain ion transport, and suggest a link between ATP supplied by glycolysis and ion transport activity. These data are also compatible with the suggestion that residual dysfunctions after brain ischemia result from derangements in glycolytic functioning rather than from limitations in oxygen availability or oxidative metabolic activity.

Animals↗
PubMed1988

Potassium ion homeostasis and mitochondrial redox activity in brain: relative changes as indicators of hypoxia.

This study was directed at relating ion transport and mitochondrial redox activity during hypoxia, as a step toward definition of brain oxygen sufficiency. To accomplish this, extracellular potassium ion activity (K+o) was recorded by ion-selective microelectrodes while reduction/oxidation (redox) ratios of cytochrome oxidase (cytochrome a,a3) were monitored by reflection spectrophotometry in cerebral cortex of rats anesthetized with pentobarbital. In normoxia, neuronal activation by direct cortical stimulation produced transient oxidation of cytochrome a,a3 and elevation of K+o. Moderate hypoxia (PaO2 above 50 mm Hg) resulted in reduction of cytochrome a,a3 but only slight elevation of K+o. At this level of hypoxia, cytochrome a,a3 continued to respond to neuronal activation with transient shifts toward oxidation and rates of K+o reaccumulation were unchanged from control. When PaO2 was further decreased below a critical threshold, stimulus-provoked oxidative responses of mitochondrial reactants were replaced by shifts toward reduction, but rates of reaccumulation of K+, spilled into the extracellular space by neuronal activation, remained unchanged. Only during severe hypoxia (PaO2 less than 20 mm Hg) was it possible in some animals to record a slowing in the reaccumulation of K+o without provocation of spreading cortical depression. These data indicate that ion transport activity in cerebral cortex is more refractory to hypoxia than is mitochondrial redox functioning. They suggest an in vivo parallel to the "cushioning" effect of mitochondria in vitro, in which oxygen consumption remains constant despite fluctuations in oxygenation and redox ratios, and also that there may be a greater anaerobic capacity to provide energy for ion transport in mammalian brain than has previously been appreciated.

Animals↗
PubMed1988

Suppression of evoked potentials with continued ion transport during anoxia in turtle brain.

A key to turtle brain survival during anoxia is continued ion transport and avoidance of anoxic depolarization. Previous findings that ATP concentration remained constant during prolonged anoxia and calculations that ATP production decreased indicate that compensatory processes, other than consumption of energy stores or increased anaerobic glycolysis, must also contribute to ion homeostasis and brain survival. To determine whether preservation of ion transport is associated with changes in electrophysiology during loss of oxidative metabolism, the brains of pentobarbital sodium-anesthetized turtles were electrically stimulated 1) to provoke measurable increments in extracellular K+ activity (a degrees k) for determination of rates of K+ reaccumulation at the stimulus site and 2) to elicit polysynaptic extracellular field potentials (evoked potentials) recordable in the olfactory bulb. During anoxia, base-line a degrees k rose only a few millimolar, and rates of reaccumulation of K+, incremented by stimulation were slightly but not significantly slowed. In contrast, postsynaptic orthodromic responses of olfactory bulb granule cells were markedly depressed by anoxia. Monosynaptic responses of granule cells to antidromic stimulation of the lateral olfactory tract were less affected, and compound action potentials in the olfactory nerve were unchanged by anoxia. These data suggest that synaptic transmission in turtle brain, as in that of mammal, is highly dependent on oxidative metabolism and that the turtle brain may effectively conserve energy for ion transport during anoxia by depression of electrical activity.

Animals↗
PubMed1988

Orthodromic field potentials and recurrent inhibition during anoxia in turtle brain.

Recent evidence suggests that suppression of electrical activity, with the likely reduction of energy use, provides a necessary strategy for prolonged anoxic tolerance of turtle brain. The present study was aimed toward defining mechanisms underlying anoxia-induced suppression of electrical activity in turtle brain. Goals were to determine whether effects of anoxia were selective to excitatory or inhibitory processes and to test the hypothesis that enhanced inhibition underlies suppression of orthodromic field potentials in olfactory bulb. Amplitudes of "conditioning" and "test" responses, provoked by pairs of stimulus pulses to the olfactory nerve, were evaluated before, during, and after anoxic insults in the brains of turtles anesthetized with pentobarbital sodium. Suppression of conditioning responses was proportionally greater than that of test responses, indicating that inhibition was decreased and that enhanced inhibitory activity is not responsible for suppression of field potentials in turtle olfactory bulb during anoxia. When normoxia was restored, conditioning responses recovered but test responses remained suppressed. These data confirm that the inhibitory effect of a conditioning pulse on responses to a second pulse of stimulation results from a recurrent circuit and suggest that anoxia-induced changes in field potentials are independent of this recurrent circuit. Enhanced inhibition during the early period of recovery after anoxic insults may provide advantages for restoring energy status and cell physiology by suppressing electrical activity and energy use.

Animals↗
PubMed1987

Extracellular potassium ion activity and electrophysiology in the hippocampal slice: paradoxical recovery of synaptic transmission during anoxia.

The relationship between extracellular potassium ion activity and neuronal excitability during anoxia was investigated in hippocampal slices in vitro. Extracellular field potentials and K+ activity were measured with double-barreled ion-selective microelectrodes placed either in the stratum pyramidale or stratum radiatum of field CA1. Orthodromic spike activity of CA1 pyramidal cells and field excitatory postsynaptic potentials (f-EPSPs) failed rapidly after anoxia with little change in potassium ion activity and without failure of the Schaffer collateral prevolley or antidromic responses of pyramidal cells. As [K+]o approached 8-10 mM, f-EPSPs and orthodromic spike activity recovered spontaneously. Continued anoxia resulted in massive release of K+ into the extracellular space and complete electrical silence. Presynaptic activity and antidromically elicited spike activity recovered promptly upon reoxygenation after anoxia, but synaptic transmission remained blocked for many minutes. Spontaneous recovery of f-EPSPs and spike activity suggests that a simple mechanism involving depolarization or hyperpolarization of neuronal elements cannot account for failure of synaptic transmission observed during anoxia. However, continued elevation of [K+]o and the associated loss of pre- and postsynaptic excitability with more prolonged anoxia indicated that depolarization was responsible for the eventual electrical silence as anoxia progressed.

Action Potentials↗
PubMed1987

Role of pulmonary edema in phasic changes of cerebral oxygenation during serial seizures.

To examine whether pulmonary dysfunction leads to episodes of cerebral hypoxia during recurrent seizures, measurements were made of arterial blood pressure, blood-gases, cerebral pO2, and relative changes in cytochrome a,a3 redox levels in anesthetized, paralyzed rats. Seizures were induced serially with bicuculline (BIC) or pentylenetetrazol (PTZ). During early seizures, cerebral oxygenation increased phasically. As seizures continued, a transition often occurred following which seizures were accompanied by phasic decreases in cerebral oxygenation. In addition, pulmonary edema often occurred at an unpredictable point during a series of seizures. Seizure-associated pulmonary edema was less likely to occur with pentobarbital anesthesia and PTZ seizures, than with nitrous oxide anesthesia and BIC seizures. Pulmonary edema was always accompanied by prolonged increases in blood pressure and paroxysmal electrocortical activity, and by hypoxemia, acidemia, and decreased cerebral oxygen supply. Despite the severity of these physiological changes, the transition from phasic increases to decreases in cerebral oxygenation during serial seizures occurred with virtually the same frequency in rats with and without pulmonary edema. This indicates that this transition is independent of pulmonary edema.

Animals↗
PubMed1987

Recovery of synaptic transmission predicted from extracellular K+ undershoots following brief anoxia in hippocampal slices.

In rat hippocampal slices exposed to short duration anoxia, recovery of synaptic transmission was dependent upon the magnitude of the K+o undershoot (K+o drop below resting level) seen following return to normoxic conditions. Because the K+o undershoot is likely to be related to ion pump activity, our results imply that recovery of brain from stroke depends upon how well ion transport and energy production recover.

Animals↗
PubMed1987

Detection of an oxidizable fraction of cytochrome oxidase in intact rat brain.

Rapid-scanning reflectance spectrophotometry was used to evaluate the reduction-oxidation state of cytochrome oxidase in normoxic rat brain. Reflectance spectra were recorded from intact blood-perfused cerebral cortices after increased oxidative metabolic activity induced by direct cortical stimulation. Reflectance spectra taken from blood-free rat brain and from the rat ear vascular bed were used to identify cytochrome and hemoglobin components of spectra taken from intact brain. Cortical stimulation provoked shifts toward increased oxidation of cytochrome oxidase that were detectable in reflectance spectra. Observations of an oxidizable fraction of cytochrome oxidase demonstrate that a fraction of the cytochrome oxidase pool exists in a reduced state in normoxic brain. The presence of reduced cytochrome oxidase suggests that oxygen delivery to the brain is restricted by microvascular control mechanisms either as a function of brain metabolic physiology or as protection against oxygen toxicity.

Absorption↗