Arthur S. Hume

A critical evaluation of thiocyanate as a biochemical index of smoking exposure

The present paper reviews the literature on thiocyanate as a biochemical index of smoking exposure. A description of thiocyanate is provided, including its characteristic effects on health, relationship to cigarette smoking, and uses in smoking research. The paper critically evaluates the various methods of measuring thiocyanate. The evaluation of different sampling methods concludes that saliva thiocyanate is the most promising method of measuring smoking exposure. Saliva thiocyanate sampling, factors which may increase measurement error, and directions for further research are discussed.

Synergistic lethality induced by the combination of carbon monoxide and cyanide

Carbon monoxide (CO) and hydrogen cyanide are recognized pyrolytic products of plastic materials in fires. Lethal blood concentrations of CO and cyanide are found in fire victims and considered to be the cause of death. However, there are reports of fire victims with sublethal concentrations of both CO and cyanide and no other attributed reason for their deaths. Thus, the purpose of this study was to investigate lethality induced by low concentrations of CO and cyanide. Male ICR mice were used in these studies. Doses of potassium cyanide (4-9 mg/kg, i.p.) were administered to animals pretreated for 3 min with either air or CO (0.63-0.66%). From these data the LD50 value of potassium cyanide (KCN) was determined in these animals pretreated with either air or CO. A significantly lower LD50 value for KCN was found in CO-pretreated animals as compared to air-pretreated animals. In another series of experiments, animals were pretreated with either saline or KCN (1.00-6.35 mg/kg, i.p.) and then placed in the chamber containing a CO atmosphere (0.325-0.375%). Ten to 20% of saline-pretreated animals were dead at the end of the monitoring period. Sublethal doses of KCN (3.5-6.35 mg/kg, i.p.) produced a synergistic lethality as compared to the saline-pretreated animals. Blood was analyzed for CO and cyanide content to determine if there were any changes that could explain this augmented lethality. There was no difference in CO or cyanide blood concentrations between these treatment groups. From these data there was demonstrated a synergistic lethality induced by the combination of CO and cyanide that could not be explained by altered CO or cyanide blood concentrations.

Severe hypoxia produced by concomitant intoxication with sublethal doses of carbon monoxide and cyanide.

It has long been known that a number of tissue hypoxicants are generated in the fire scenario. However, until recently few investigators have undertaken to correlate smoke inhalation deaths with the simultaneous exposure to histotoxic hypoxicants. Carbon monoxide and hydrogen cyanide are two histotoxic hypoxicants that are generated in nearly every fire. Prior studies performed in our laboratory have demonstrated that death can result from concomitant exposure to otherwise sublethal concentrations of carbon monoxide and cyanide. Since most smoke inhalation victims exhibit acid/base anomalies, we sought to investigate whether the death associated with simultaneous exposure to these two hypoxicants, at concentrations widely held to be nonlethal, could be explained by acid/base imbalances. Male ICR mice were exposed to 0.35% carbon monoxide immediately after having been injected ip with potassium cyanide solution, or were challenged with either agent alone. Animals challenged with cyanide or carbon monoxide alone demonstrated significant hypoxia. However, animals challenged with both agents demonstrated much greater hypoxia than could be explained by an additive effect alone. Controls demonstrated no alteration in acid/base homeostasis. Blood pH perturbations were found to be due to severe lactic acidosis coupled with inadequate respiratory compensation. Thus, it appears that the synergistic lethal effect of simultaneous administration of carbon monoxide and cyanide are related to a precipitous decrease in blood pH, the tissue hypoxia and its resulting complications.

The efficacy of α-ketoglutaric acid in the antagonism of cyanide intoxication

Abstract It has been reported that compounds containing carbonyl groups can readily react with cyanide. Pyruvic acid, an α-ketocarboxylic acid, has been shown to antagonize the lethal effects of cyanide. It is suggested that its mechanism of action rests in its ability to react with or “bind” cyanide. In this study, α-ketoglutaric acid, also an α-ketocarboxylic acid, was evaluated for its ability to counteract the lethal effects of cyanide. α-Ketoglutaric acidvincreased the LD50 value of cyanide (6.7 mg/kg) by a factor of five, a value statistically equivalent to that ascertained in mice pretreated with sodium thiosulfate and sodium nitrite. The combination of α-ketoglutaric acid and sodium thiosulfate increased the LD50 value of cyanide to 101 mg/kg. Addition of sodium nitrite to the α-ketoglutaric acid/sodium thiosulfate regimen increased the LD50 value of cyanide to 119 mg/kg. Unlike sodium nitrite, no induction of methemoglobin formation was observed with α-ketoglutaric acid pretreatment. It is apparent from these studies that the administration of α-ketoglutaric acid in conjunction with sodium thiosulfate resulted in fewer animal deaths than sodium nitrite and sodium thiosulfate without the dangerous formation of methemoglobin.

Behavioral and neurochemical effects of continuous infusion of cocaine in rats

The ability of continuous intravenous infusion of cocaine (60 mg/kg per day for 11 or 12 days; by osmotic minipump) to alter responses to acute injection of cocaine (20 mg/kg, i.p.; given 24 hr after termination of the infusion by minipump) was tested in conscious, tethered Sprague-Dawley rats. Extracellular levels of cocaine, dopamine and metabolites of dopamine in the striatum were determined by in vivo microdialysis. Locomotor activity and stereotyped behavior were evaluated simultaneously during dialysis sampling. Prior infusion of cocaine blunted the ability of acute challenge with cocaine to increase the efflux of dopamine in the striatum, locomotor activity and stereotypy. Increases in extracellular levels of homovanillic acid in the striatum were significantly greater in cocaine-infused rats than vehicle-infused controls, both prior to and after acute injections of cocaine. However, no differences between these two groups were observed in levels of cocaine in the striatum after acute challenge. Extracellular levels of dopamine in the striatum correlated significantly (P less than 0.05) with stereotypy in both groups but with locomotor activity only in cocaine-infused rats. The results indicate that behavioral tolerance occurred after continuous intravenous infusions of cocaine, that this was correlated with neurochemical tolerance to acute cocaine challenge and that alterations in the metabolism of cocaine did not account for the observed behavioral responses.

Mechanism of antagonizing cyanide-induced lethality by α-ketoglutaric acid

alpha-Ketoglutaric acid (alpha-KG) has been shown to be an effective antagonist for cyanide-induced lethality. The mechanism of this antagonism is hypothesized to result from alpha-KG binding with cyanide. Several investigative approaches were taken to determine the existence of this binding. First, mixtures of various molar ratios of alpha-KG:cyanide were injected into a high pressure liquid chromatograph. The addition of cyanide reduced the peak area of alpha-KG at a molar ratio of greater than 1:5. Second, blood from naive male ICR mice was spiked with alpha-KG and cyanide. Headspace above these blood samples was injected into a gas chromatograph and analyzed for released hydrogen cyanide. alpha-KG reduced the peak area of hydrogen cyanide released into the headspace at molar ratios of greater than 1:2.5. Third, the effect of cyanide on the ultraviolet spectrum of alpha-KG was determined as an indication of binding. In the presence of cyanide the absorption peak at 316 nm for alpha-KG was eliminated. Inhibition of cytochrome oxidase is an accepted target enzyme for cyanide-induced lethality. Fourth, further evidence of alpha-KGs mechanism was determined by the effect of alpha-KG on brain cytochrome oxidase (BRCYTOX) and its ability to antagonize cyanide-induced inhibition of BRCYTOX. BRCYTOX activity was determined in the presence of alpha-KG and was found to be unaffected between 0.01 and 0.06 M of alpha-KG. Greater concentrations of alpha-KG inhibited BRCYTOX activity. The complete inhibition of BRCYTOX activity by 10(-5) M cyanide was prevented with 0.05 and 0.06 M alpha-KG. Fifth, BRCYTOX activity of animals pretreated with saline and then an LD80 dose (8.5 mg/kg) of cyanide was 80% inhibited, while BRCYTOX activity of animals pretreated with 2 g alpha-KG/kg, i.p., and then an LD80 dose (7.75 mg/kg) of cyanide was not different from control values. Thus, these data suggest that alpha-KG does bind with cyanide, and this binding can account for the antagonism of cyanide-induced lethality.

Pentobarbital tolerance and withdrawal: Correlation with effects on the GABAA receptor

A model for the development of pentobarbital tolerance and dependence was characterized and correlated with changes in radioligand binding to the GABAA-benzodiazepine receptor chloride channel complex. While one day of pentobarbital exposure decreased the duration of loss of righting reflex, tolerance to the hypothermic effects of thiopental and barbital took 7 days to develop, indicating that pharmacokinetic and pharmacodynamic tolerance are separable. Increased sensitivity to pentylenetetrazol-induced seizures was first observed after 3 days of pentobarbital exposure, suggesting brain areas involved in seizure control develop tolerance to, and dependence on pentobarbital faster than those involved in temperature regulation. Acute exposure to pentobarbital in vivo did not affect cortical binding of [3H]muscimol in vitro, while tolerance caused a decrease in binding due to an increase in the low-affinity site KD. Pentobarbital tolerance also caused a decrease in the cortical binding of the benzodiazepine, [3H]flunitrazepam. These observations suggest that the acute effects of barbiturates on the GABAA receptor complex are reversible, while tolerance causes receptor modifications which may be related to the development of physical dependence.

Effects of subacute lead exposure on [3H]MK-801 binding in hippocampus and cerebral cortex in the adult rat

We used the NMDA receptor non-competitive antagonist, [3H]MK-801, as a ligand for an autoradiographic study to determine the effects of lead on NMDA receptor in the rat brain. Adult male rats were administered lead acetate, 100 mg/kg, or sodium acetate, 36 mg/kg (control), by i.p. for 7 days. High lead levels were detected in blood (41.1 microg/dl) and in brain (16.7-29.4 microg/g). Concentrations of lead in brain regions were not significantly different. The [3H]MK-801 binding was heterogeneously distributed throughout the rat brain with the following order of binding densities: hippocampal formation > cortex > caudate-putamen > thalamus > brainstem. Lead exposure produced a significant decrease in [3H]MK-801 binding to the NMDA receptor in the hippocampal formation including CA2 stratum radiatum, CA3 stratum radiatum, hilus dentate gyrus and presubiculum, and in the cerebral cortex including agranular insular, cingulate, entorhinal, orbital, parietal and perirhinal areas. The hippocampal formation is known as a critical neural structure for learning and memory processes, whereas, cortical and subcortical regions have been demonstrated to be involved in the modulation of complex behavioral processes. The NMDA receptor has been demonstrated to play a key role in synaptic plasticity underlying learning and memory. Lead-induced alterations of NMDA receptors in the hippocampal formation and cortical areas may play a role in lead-induced neurotoxicity.

Effects of chronic lead exposure on [3H]MK-801 binding in the brain of rat

We have used quantitative autoradiographic methods to determine the effects of chronic lead exposure on N-methyl-D-aspartate (NMDA) receptors in the brain of female rat. Rats were exposed pre- and post-natally from day 4 +/- 1 post conception with 1000 ppm lead in their drinking water. This treatment continued after weaning. No effects of lead on [3H]MK-801 binding were found at PN 28. However, lead caused a significant increase in [3H]MK-801 binding in the hippocampus including CA1 and CA2, and in the occipital and temporal cortical areas at PN 56 and at PN 112. An increase in binding was also found in the entorhinal cortex and the dentate gyrus at PN 112. Because the NMDA receptor is involved in learning and memory, the lead-induced disruption of NMDA receptors in the hippocampus and cortex may be associated with the cation-induced cognition deficits.

The reliability of Thiocyanate measurement of smoking exposure

Abstract Procedures for the determination of thiocyanate (SCN) levels in saliva samples were evaluated in a series of three experiments. Laboratory analysis procedures as well as sampling and storage conditions were examined to identify sources of measurement error in the assessment of SCN. The results of the study indicated that spectrophotometric analysis of saliva samples can be a reliable method of determining SCN levels. Also, storage containers, length of storage and storage temperatures can lead to critical differences in SCN level when saliva samples were not analyzed immediately after sampling. Guidelines for the assessment of saliva SCN are provided in a discussion of the results.