Behavioural and physiological assessments of dimethyl trisulfide treatment for acute oral sodium cyanide poisoning.

Sodium cyanide (NaCN) is a commonly and widely used industrial and laboratory chemical that is highly toxic. Its availability and rapid harmful/lethal effects combine to make cyanide a potential foodborne/waterborne intentional-poisoning hazard. Effective antidotes to cyanide poisoning are currently approved only for intravenous administration. Therefore, an effective cyanide antidote that can be administered intramuscularly in pre-hospital and/or mass-casualty settings is needed. Dimethyl trisulfide (DMTS) is a naturally occurring substance used as a flavour enhancer in foods. DMTS has shown antidotal efficacy in cyanide poisoning and is thought to act as both a sulphur donor and partial methaemoglobin inducer. In this study, an intramuscular injection of DMTS (6.25-200 mg/kg) was given to rats 1 minute after an oral dose of NaCN (98.2 mg/kg; twice the median lethal dose) to test the antidotal efficacy and safety of DMTS treatment. Toxic signs and survival were examined along with behavioural function (up to 30 hour after ingestion) using a previously established operant behavioural model. A large range of DMTS doses (6.25-100 mg/kg) increased survival after oral cyanide poisoning, and the lower DMTS doses (6.25-25 mg/kg) also proved to be behaviourally and physiologically safe. Larger DMTS doses (50-200 mg/kg) produced side effects (ie, inflammation and limping) that were more severe and protracted than those observed at lower DMTS doses. The 25 mg/kg DMTS proved to be the most efficacious (increasing survival from 20% to 75%) and also produced minimal side effects (eg, inflammation) that resolved within 24-72 hour. Thus, DMTS shows promise as an intramuscularly administered cyanide antidote useful for prompt pre-hospital or mass-casualty emergency medical treatment.

Alterations of sirtuins in mitochondrial cytochrome c-oxidase deficiency.

BACKGROUND: Sirtuins are NAD+ dependent deacetylases, which regulate mitochondrial energy metabolism as well as cellular response to stress. The NAD/NADH-system plays a crucial role in oxidative phosphorylation linking sirtuins and the mitochondrial respiratory chain. Furthermore, sirtuins are able to directly deacetylate and activate different complexes of the respiratory chain. This prompted us to analyse sirtuin levels in skin fibroblasts from patients with cytochrome c-oxidase (COX) deficiency and to test the impact of different pharmaceutical activators of sirtuins (SRT1720, paeonol) to modulate sirtuins and possibly respiratory chain enzymes in patient cells in vitro. METHODS: We assayed intracellular levels of sirtuin 1 and the mitochondrial sirtuins SIRT3 and SIRT4 in human fibroblasts from patients with COX- deficiency. Furthermore, sirtuins were measured after inhibiting complex IV in healthy control fibroblasts by cyanide and after incubation with activators SRT1720 and paeonol. To determine the effect of sirtuin inhibition at the cellular level we measured total cellular acetylation (control and patient cells, with and without treatment) by Western blot. RESULTS: We observed a significant decrease in cellular levels of all three sirtuins at the activity, protein and transcriptional level (by 15% to 50%) in COX-deficient cells. Additionally, the intracellular concentration of NAD+ was reduced in patient cells. We mimicked the biochemical phenotype of COX- deficiency by incubating healthy fibroblasts with cyanide and observed reduced sirtuin levels. A pharmacological activation of sirtuins resulted in normalized sirtuin levels in patient cells. Hyper acetylation was also reversible after treatment with sirtuin activators. Pharmacological modulation of sirtuins resulted in altered respiratory chain complex activities. CONCLUSIONS: We found inhibition of situins 1, 3 and 4 at activity, protein and transcriptional levels in fibroblasts from patient with COX-deficiency. Pharmacological activators were able to restore reduced sirtuin levels and thereby modulate respiratory chain activities.

Memory deficits associated with sublethal cyanide poisoning relative to cyanate toxicity in rodents.

Food (cassava) linamarin is metabolized into neurotoxicants cyanide and cyanate, metabolites of which we sought to elucidate the differential toxicity effects on memory. Young 6-8 weeks old male rats were treated intraperitoneally with either 2.5 mg/kg body weight (bw) cyanide (NaCN), or 50 mg/kg bw cyanate (NaOCN), or 1 µl/g bw saline, daily for 6 weeks. Short-term and long-term memories were assessed using a radial arm maze (RAM) testing paradigm. Toxic exposures had an influence on short-term working memory with fewer correct arm entries (F(2, 19) = 4.57 p < 0.05), higher working memory errors (WME) (F(2, 19) = 5.09, p < 0.05) and longer RAM navigation time (F(2, 19) = 3.91, p < 0.05) for NaOCN relative to NaCN and saline treatments. The long-term working memory was significantly impaired by cyanide with fewer correct arm entries (F(2, 19) = 7.45, p < 0.01) and increased working memory errors (F(2, 19) = 9.35 p < 0.05) in NaCN relative to NaOCN or vehicle treated animals. Reference memory was not affected by either cyanide or cyanate. Our study findings provide an experimental evidence for the biological plausibility that cassava cyanogens may induce cognition deficits. Differential patterns of memory deficits may reflect the differences in toxicity mechanisms of NaOCN relative to NaCN. Cognition deficits associated with cassava cyanogenesis may reflect a dual toxicity effect of cyanide and cyanate.

Effect of exposure to sublethal concentrations of sodium cyanide on the carbohydrate metabolism of the Indian Major Carp Labeo rohita (Hamilton, 1822)

Experiments were designed to study in-vivo effects of sodium cyanide on biochemical endpoints in the freshwater fish Labeo rohita. Fish were exposed to two sublethal concentrations (0.106 and 0.064mg/L) for a period of 15 days. Levels of glycogen, pyruvate, lactate and the enzymatic activities of lactate dehydrogenase (LDH), succinate dehydrogenase (SDH), glucose-6-phosphate dehydrogenase (G6PDH), phosphorylase, alkaline phosphatase (ALP), acid phosphatase (AcP) were assessed in different tissues (liver, muscle and gills). Result indicated a steady decrease in glycogen, pyruvate, SDH, ALP and AcP activity with a concomitant increase in the lactate, phosphorylase, LDH and G6PD activity in all selected tissues. The alterations in all the above biochemical parameters were significantly (p<0.05) time and dose dependent. In all the above parameters, liver pointing out the intensity of cyanide intoxication compare to muscle and gills. Study revealed change in the metabolic energy by means of altered metabolic profile of the fish. Further, these observations indicated that even sublethal concentrations of sodium cyanide might not be fully devoid of deleterious influence on metabolism in L. rohita.

Modulation in the protein metabolism by subacute sodium cyanide intoxication in the freshwater fish, Labeo rohita (Hamilton).

The effects of exposure to one-third and one-fifth sublethal concentrations (0.106 and 0.064 mg/L) of sodium cyanide on protein metabolism on freshwater carp, Labeo rohita, was studied. Three functionally different tissues, namely, the liver, muscle, and gills, were studied after 5, 10, and 15 days. Exposures produced marked changes in protein metabolic profile in all tissues studied. These changes were more pronounced in the one-third sublethal concentration, suggesting a cumulative action of toxicant. This investigation revealed that the total, structural, and soluble proteins and urea content in all the three tissues were decreased, whereas free amino acids, ammonia, and enzyme activity (i.e., protease, alanine aminotransferase, and aspartate aminotransferase) exhibited elevated levels at both sublethal concentrations. Variation in protein metabolism in the fish, induced by sodium cyanide, demonstrated its toxic effects on cellular metabolism, thereby leading to impaired protein synthetic machinery. The results of the present study indicate that a mechanism of impaired energy transformation has direct action on the fish, L. rohita, and its impact is clearly evident from the change in the nutritional content of the fish.

Homicide due to intravenous metallic mercury injection followed by sodium cyanide injection.

We report a case of homicide due to intravenous mercury injection followed by meperidine and sodium cyanide injection. A 35-year-old woman was found dead in bed at home by her husband. Reportedly, she had been sick for more than 5 months. Initial death investigation revealed no evidence of foul play. Her death was believed to be natural. Therefore, her body was buried without an autopsy. Two months after death, her family requested an autopsy because they suspected her physician husband killed her. Her body was exhumed, and an autopsy was performed. Postmortem examination revealed numerous metallic mercury globules in the pulmonary arteries. Toxicological analysis revealed a high concentration of mercury in the tissue samples of the lungs, liver, heart, and kidney. In addition, cyanide and meperidine were also found in the heart and liver. The detailed case history and postmortem examination findings are described.

The interactive effects of hypoxemia, hyperoxia, and temperature on the gill morphology of goldfish (Carassius auratus).

Acclimation of crucian carp and goldfish to temperatures below 15°C causes covering of the gill lamellae by a mass of cells termed the interlamellar cell mass (ILCM). Here we explore the cues underlying gill remodeling (removal or growth of an ILCM) and specifically test the hypotheses that 1) depletion of internal O(2) stores in the absence of any change in external O(2) status can trigger the removal of the ILCM in goldfish acclimated to 7°C, 2) exposing fish acclimated to 25°C to an abundance of O(2) (hyperoxia) can reverse the gill remodeling (i.e., cause the covering of lamellae by an expansion of the ILCM), and 3) neuroepithelial cells (NECs) are involved in signaling the shedding of the ILCM. Hypoxemia induced by phenylhydrazine (anemia) or 5% CO caused a decrease in the ILCM from 80% to 23% and 35%, respectively. Hyperoxia exposure at 25°C caused an increase to 67% of total ILCM and a smaller decrease in the size of the ILCM when fish were transferred from 7 to 25°C. Daily sodium cyanide injections were used to stimulate NECs; this treatment led to a significant decrease in the ILCM. Thus, the three major conclusions of this study are 1) that gill remodeling can occur during periods of internal hypoxemia, 2) that O(2) supply and demand may be a significant driving force shaping gill remodeling in goldfish, and 3) the NECs may play a role in triggering the shedding of the ILCM during hypoxia.

The control of breathing in goldfish (Carassius auratus) experiencing thermally induced gill remodelling.

At temperatures below 15°C the gill lamellae of goldfish (Carassius auratus) are largely covered by an interlamellar cell mass (ILCM) which decreases the functional surface area of the gill. The presence of the ILCM in goldfish acclimated to cold water conceivably could lead to a covering of the neuroepithelial cells (NECs), which are believed to be important for sensing ambient O2 and CO2 levels. In this study we tested the hypothesis that goldfish with covered lamellae (and presumably fewer NECs exposed to the water) exhibit a decreased capacity to hyperventilate in response to hypoxic stimuli. Measurements of ventilation amplitude and frequency were performed during exposure to acute hypoxia (P(w(O2))=30 mmHg) or following injections of the O2 chemoreceptor stimulant NaCN into the buccal cavity or caudal vein of fish acclimated to 25°C (uncovered lamellae) or 7°C (covered lamellae) to stimulate predominantly the externally or internally oriented NECs, respectively. The results demonstrated no significant differences in the response to hypoxia, with each group exhibiting similar percentage increases in ventilation amplitude (90-91%) and frequency (34-43%). Similarly, with the exception of a rightward shift of the ventilation frequency dose-response in the fish acclimated to 7°C, there were no significant differences between the two groups of fish in the ED(50) values. These findings suggest that goldfish with covered lamellae retain the capacity to sense external hypoxic stimuli. Using immunohistochemistry to identify serotonin-enriched NECs, it was demonstrated that the presence of the ILCM results in the NECs being redistributed towards the distal regions of the lamellae. In 25°C-acclimated fish, the NECs were distributed evenly along the length of the lamellae with 53±3% of them in the distal half, whereas in fish acclimated to 7°C, 83±5% of the NECs were confined to the distal half. Using the neuronal marker antibody ZN-12, it was demonstrated that the NECs at the distal edges of the lamellae are innervated by nerve fibres. Thus, it is hypothesised that the capacity to sense external hypoxic stimuli in goldfish acclimated to cold water is maintained despite the increasing coverage of the gill epithelial surfaces because of a redistribution of innervated NECs to the exposed distal regions of the lamellae.

Two-pore K+ channels, NO and metabolic inhibition.

Ischemic preconditioning is a potent endogenous mechanism protecting many organs from the devastating effects of prolonged ischemia. In the heart, NO is one mediator of this myoprotective response thought to involve activation of the K(ATP) channel. Ischemic preconditioning is known to be induced by metabolic inhibition using sodium cyanide (NaCN) in single cardiomyocytes. In the present study, we show for the first time that the end effector channel activated by NaCN has been incorrectly identified. The channel activated is not K(ATP) but instead belongs to the relatively new family of two-pore domain potassium channels (K2P). Further when activated by metabolic ischemia, the amplitude of K2P current is directly modulated by activators and inhibitors of the NO pathway.

Is urea pulsing in toadfish related to environmental O2 or CO2 levels?

The neurochemical, serotonin (5-hydroxytryptamine; 5-HT) is involved in the regulation of toadfish pulsatile urea excretion as well as the teleost hypoxia response. Thus, the goal of this study was to determine whether environmental conditions that activate branchial chemoreceptors also trigger pulsatile urea excretion in toadfish, since environmental dissolved oxygen levels in a typical toadfish habitat show significant diel fluctuations, often reaching hypoxic conditions at dawn. Toadfish were fitted with arterial, venous and/or buccal catheters and were exposed to various environmental conditions, and/or injected with the O(2) chemoreceptor agonist NaCN or the 5-HT(2) receptor agonist alpha-methyl-5HT. Arterial PO(2), as well as ammonia and urea excretion were monitored. Natural fluctuations in arterial PO(2) levels in toadfish did not correlate with the occurrence of a urea pulse. Chronic exposure (24 h) of toadfish to hyperoxia was without effect on nitrogen excretion, however, exposure to hypoxia caused a significant reduction in the frequency of urea pulses, and exposure to hypercapnia resulted in a reduction in the percentage of nitrogen waste excreted as urea. Of toadfish exposed acutely to hypoxia, 20% pulsed within 1 h, whereas none pulsed after normoxic or hypercapnic treatments. Furthermore, 20% of fish injected intravenously with NaCN pulsed within 1 h of injection, but no fish pulsed after injection of NaCN into the buccal cavity. To test whether environmental conditions affected 5-HT(2) receptors, toadfish were injected with alpha-methyl-5HT, which elicits urea pulses in toadfish. No significant differences in pulse size occurred among the various environmental treatments. Our findings suggest that neither the environmental conditions of hypoxia, hyperoxia or hypercapnia, nor direct branchial chemoreceptor activation by NaCN play a major role in the regulation of pulsatile urea excretion in toadfish.

Carotid chemoreceptor reflex modulation by arginine-vasopressin microinjected into the nucleus tractus solitarius in rats.

BACKGROUND: In addition to their role of sensing O2, pH, CO2, osmolarity and temperature, carotid body receptors (CBR) were proposed by us and others to have a glucose-sensing role in the blood entering the brain, integrating information about blood glucose and O2 levels essential for central nervous system (CNS) metabolism. The nucleus tractus solitarius (NTS) is an important relay station in central metabolic control and receives signals from peripheral glucose-sensitive hepatoportal afferences, from central glucose-responsive neurons in the brainstem and from CBR and arginine-vasopressin (AVP)-containing axons from hypothalamic nuclei. METHODS: In normal Wistar rats anesthetized with pentobarbital, permanent cannulas were placed stereotaxically in the NTS. Glucose changes were induced in vivo after CBR stimulation with sodium cyanide (NaCN-5 microg/100 g), preceded by an infusion of AVP [(10 or 40 pmol/100 nL of artificial cerebrospinal fluid) aCSF] or an antagonist for V1a receptors (anti-glycogenolytic vasopressin analogue-VP1-A) (100 pmol/100 nL of aCSF) into the NTS. RESULTS: CBR stimulation after an AVP infusion (larger dose) into the NTS resulted in a significantly higher arterial glucose and lower brain arterial-venous glucose difference. In the same way, VP1-A administration in the NTS significantly decreased the effects observed after AVP priming before CBR stimulation or preceding the CBR stimulation, alone. CONCLUSIONS: We propose that AVP in the NTS could participate in glucose homeostasis, modulating the information arising in CBR after histotoxic-anoxia stimulation.

Chemoreceptor plasticity and respiratory acclimation in the zebrafish Danio rerio.

The goals of this study were to assess the respiratory consequences of exposing adult zebrafish Danio rerio to chronic changes in water gas composition (hypoxia, hyperoxia or hypercapnia) and to determine if any ensuing effects could be related to morphological changes in branchial chemoreceptors. To accomplish these goals, we first modified and validated an established non-invasive technique for continuous monitoring of breathing frequency and relative breathing amplitude in adult fish. Under normal conditions 20% of zebrafish exhibited an episodic breathing pattern that was composed of breathing and non-breathing (pausing/apneic) periods. The pausing frequency was reduced by acute hypoxia (Pw(O)2<130 mmHg) and increased by acute hyperoxia (Pw(O)2>300 mmHg), but was unaltered by acute hypercapnia. Fish were exposed for 28 days to hyperoxia (Pw(O)2>350 mmHg), or hypoxia (Pw(O)2=30 mmHg) or hypercapnia (Pw(CO)2=9 mmHg). Their responses to acute hypoxia or hypercapnia were then compared to the response of control fish kept for 28 days in normoxic and normocapnic water. In control fish, the ventilatory response to acute hypoxia consisted of an increase in breathing frequency while the response to acute hypercapnia was an increase in relative breathing amplitude. The stimulus promoting the hyperventilation during hypercapnia was increased Pw(CO)2 rather than decreased pH. Exposure to prolonged hyperoxia decreased the capacity of fish to increase breathing frequency during hypoxia and prevented the usual increase in breathing amplitude during acute hypercapnia. In fish previously exposed to hyperoxia, episodic breathing continued during acute hypoxia until Pw(O)2 had fallen below 70 mmHg. In fish chronically exposed to hypoxia, resting breathing frequency was significantly reduced (from 191+/-12 to 165+/-16 min(-1)); however, the ventilatory responses to hypoxia and hypercapnia were unaffected. Long-term exposure of fish to hypercapnic water did not markedly modify the breathing response to acute hypoxia and modestly blunted the response to hypercapnia. To determine whether branchial chemoreceptors were being influenced by long-term acclimation, all four groups of fish were acutely exposed to increasing doses of the O(2) chemoreceptor stimulant, sodium cyanide, dissolved in inspired water. Consistent with the blunting of the ventilatory response to hypoxia, the fish pre-exposed to hyperoxia also exhibited a blunted response to NaCN. Pre-exposure to hypoxia was without effect whereas prior exposure to hypercapnia increased the ventilatory responses to cyanide. To assess the impact of acclimation to varying gas levels on branchial O(2) chemoreceptors, the numbers of neuroepithelial cells (NECs) of the gill filament were quantified using confocal immunofluorescence microscopy. Consistent with the blunting of reflex ventilatory responses, fish exposed to chronic hyperoxia exhibited a significant decrease in the density of NECs from 36.8+/-2.8 to 22.7+/-2.3 filament(-1).

Calcium ions: its role in cyanide neurotoxicity.

Cyanide is a well-established poison known for its rapid lethal action and toxicity. The central nervous system is one of the main target sites for cyanide toxicity. Cyanide also alters the brain biogenic amine levels. In the present study the role of calcium ions in cyanide toxicity was evaluated using the calcium channel blocker diltaizem (DIL) in Wistar strain albino rats. This study showed that DIL pretreatment prevented cyanide induced changes in the dopaminergic and serotonergic system in the corpus striatum.

Peripheral O2 chemoreceptors mediate humoral catecholamine secretion from fish chromaffin cells.

This study addressed the hypothesis that the secretion of catecholamines from trout (Oncorhynchus mykiss) chromaffin cells, during hypoxia, is triggered by stimulation of O(2) chemoreceptors located within the gills. Sodium cyanide was administered into the inspired water (external cyanide) or injected into the gill circulation (internal cyanide) to pharmacologically stimulate external (water sensing) or internal (blood sensing) O(2) chemoreceptors, respectively. Both of these treatments caused an elevation of circulating catecholamine levels. The response to external, but not internal, cyanide was abolished by removal of the first gill arch. Hypoxia produced an increase in circulating catecholamine levels that was unaffected by removal of the first gill arch or by denervation of the pseudobranch. Cyanide and hypoxia both caused the well-documented cardiorespiratory reflexes normally observed in this species. This study demonstrates, for the first time, that gill O(2) chemoreceptors can initiate the reflex that leads to catecholamine release from the chromaffin cells and that stimulation of internally oriented O(2) receptors on all gill arches appears to be the physiologically important mechanism for initiating release.

Effects of carotid and aortic chemoreceptor denervation in newborn piglets.

The objective of the present study was to test the hypothesis that in neonatal piglets there would be no hypoventilation after sham denervation or aortic denervation (AOD) alone, but there would be transient hypoventilation after carotid body denervation (CBD) and the hypoventilation would be greatest after combined carotid and aortic denervation (CBD+AOD). There was a significant (P < 0.05) hypoventilation in CBD and CBD+AOD piglets denervated at 5, 15, and 25 days of age. The hypoventilation in CBD+AOD piglets denervated at 5 days of age was greater (P < 0.05) than that of all other groups. Conversely, sham-denervated and AOD piglets did not hypoventilate after denervation. Injections of sodium cyanide showed that aortic chemoreceptors were a site of recovery of peripheral chemosensitivity after CBD. This aortic sodium cyanide response was abolished by prior injection of a serotonin 5a receptor blocker. Residual peripheral chemosensitivity after CBD+AOD was localized to the left ventricle. We conclude that 1) aortic chemoreceptors contribute to eupneic breathing in piglets that were carotid denervated at 5 days of age and 2) there are multiple sites of residual peripheral chemosensitivity after CBD.

Pre-Bötzinger complex functions as a central hypoxia chemosensor for respiration in vivo.

Recently, we identified a region located in the pre-Bötzinger complex (pre-BötC; the proposed locus of respiratory rhythm generation) in which activation of ionotropic excitatory amino acid receptors using DL-homocysteic acid (DLH) elicits a variety of excitatory responses in the phrenic neurogram, ranging from tonic firing to a rapid series of high-amplitude, rapid rate of rise, short-duration inspiratory bursts that are indistinguishable from gasps produced by severe systemic hypoxia. Therefore we hypothesized that this unique region is chemosensitive to hypoxia. To test this hypothesis, we examined the response to unilateral microinjection of sodium cyanide (NaCN) into the pre-BötC in chloralose- or chloralose/urethan-anesthetized vagotomized, paralyzed, mechanically ventilated cats. In all experiments, sites in the pre-BötC were functionally identified using DLH (10 mM, 21 nl) as we have previously described. All sites were histologically confirmed to be in the pre-BötC after completion of the experiment. Unilateral microinjection of NaCN (1 mM, 21 nl) into the pre-BötC produced excitation of phrenic nerve discharge in 49 of the 81 sites examined. This augmentation of inspiratory output exhibited one of the following changes in cycle timing and/or pattern: 1) a series of high-amplitude, short-duration bursts in the phrenic neurogram (a discharge similar to a gasp), 2) a tonic excitation of phrenic neurogram output, 3) augmented bursts in the phrenic neurogram (i.e., eupneic breath ending with a gasplike burst), or 4) an increase in frequency of phrenic bursts accompanied by small increases or decreases in the amplitude of integrated phrenic nerve discharge. Our findings identify a locus in the brain stem in which focal hypoxia augments respiratory output. We propose that the respiratory rhythm generator in the pre-BötC has intrinsic hypoxic chemosensitivity that may play a role in hypoxia-induced gasping.

Cyanide intoxication induced exocytotic epinephrine release in rabbit myocardium.

Cyanide intoxication, which has been used as a model of energy depletion at cardiac sympathetic nerve terminals, causes non-exocytotic release of norepinephrine (NE). However, the effect of cyanide intoxication on cardiac epinephrine (Epi) release remains unknown. Using cardiac microdialysis in the rabbit, we measured dialysate Epi and NE concentrations as indices of myocardial interstitial Epi and NE levels, respectively. Local administration of sodium cyanide (30 mM) through the dialysis probe increased both Epi and NE levels (from 11.3+/-2.3 to 32.3+/-4.4 pg/ml and from 33.6+/-6.1 to 389.0+/-71.8 pg/ml, respectively, mean+/-S.E., P<0.01). Local desipramine (100 microM) administration suppressed the cyanide induced NE response without affecting the Epi response. In contrast, local omega-conotoxin GVIA (10 microM) administration partially suppressed the cyanide induced NE response and totally abolished the Epi response. In conclusion, cyanide intoxication causes N-type Ca(2+) channel dependent exocytotic Epi release as well as inducing N-type Ca(2+) channel independent non-exocytotic NE release.

A reproducible nonlethal animal model for studying cyanide poisoning.

Previous studies using bolus intravenous injections of sodium cyanide have been used to model the sudden exposure to high concentrations of cyanide that could occur on the battlefield. This study was designed to develop a model that would simulate the type of exposure to cyanide gas that could happen during actual low-level continuous types of exposure and then compare it with the bolus model. Cardiovascular and respiratory recordings taken from anesthetized dogs have been used previously to characterize the lethal effects of cyanide. The intravenous, bolus injection of 2.5 mg/kg sodium cyanide provides a model in which a greater than lethal concentration is attained. In contrast, our model uses a slow, intravenous infusion of cyanide to titrate each animal to its own inherent end point, which coincides with the amount of cyanide needed to induce death through respiratory arrest. In this model, therapeutic intervention can be used to restore respiration and allow for the complete recovery of the animals. After recovery, the same animal can be given a second infusion of cyanide, followed again by treatment and recovery, providing a reproducible end point. This end point can then be expressed as the total amount of cyanide per body weight (mg/kg) required to kill. In this study, the average dose of sodium cyanide among 12 animals was 1.21 mg/kg, which is approximately half the cyanide used in the bolus model. Thus, titration to respiratory arrest followed by resuscitation provides a repetitive-use animal model that can be used to test the efficacy of various forms of pretreatment and/or therapy without the loss of a single animal.

Effects on breathing of carotid body denervation in neonatal piglets.

The purpose of these studies was to test the hypothesis that carotid chemoreceptor activity is necessary for postnatal maturation of the ventilatory control system. By using a lateral surgical access, 17 piglets were carotid body denervated (CBD) and 14 were sham denervated at 3-25 days of age. After surgery, there was no irregular breathing in any group. There was no significant hypoventilation when CBD was performed at less than 5 days of age (n = 5) and only a mild (arterial PCO(2) 5 Torr; P < 0.05) to moderate, transient (arterial PCO(2) 8 Torr; P < 0.5) hypoventilation in piglets denervated at 10-15 (n = 6) and 20-25 (n = 6) days of age, respectively. Three weeks after surgery, both breathing of a hypoxic gas mixture and jugular venous NaCN injections elicited a hyperpnea in the CBD piglets that was attenuated compared with that in sham CBD piglets. In the CBD piglets, there was no response to injections of NaCN in the carotid arteries, but there was a response to NaCN injected into the proximal descending aorta, suggesting the residual peripheral chemosensitivity was of aortic origin. Carotid chemoreceptor-intact piglets had carotid and aortic NaCN chemosensitivity by 2 days of age. The carotid response persisted for the 40 days of the study, but the aortic reflex persisted only until approximately 8 days of age. We conclude that 1) the major effect of CBD per se in neonatal piglets is age-dependent hypoventilation and 2) there is a high degree of plasticity in peripheral chemosensitivity in neonates that may contribute to minimizing the changes in breathing after CBD.

Important role of carotid afferents in control of breathing.

The purpose of the present study was to determine the effect on breathing in the awake state of carotid body denervation (CBD) over 1-2 wk after denervation. Studies were completed on adult goats repeatedly before and 1) for 15 days after bilateral CBD (n = 8), 2) for 7 days after unilateral CBD (n = 5), and 3) for 15 days after sham CBD (n = 3). Absence of ventilatory stimulation when NaCN was injected directly into a common carotid artery confirmed CBD. There was a significant (P < 0.01) hypoventilation during the breathing of room air after unilateral and bilateral CBD. The maximum PaCO2 increase (8 Torr for unilateral and 11 Torr for bilateral) occurred approximately 4 days after CBD. This maximum was transient because by 7 (unilateral) to 15 (bilateral) days after CBD, PaCO2 was only 3-4 Torr above control. CO2 sensitivity was attenuated from control by 60% on day 4 after bilateral CBD and by 35% on day 4 after unilateral CBD. This attenuation was transient, because CO2 sensitivity returned to control temporally similar to the return of PaCO2 during the breathing of room air. During mild and moderate treadmill exercise 1-8 days after bilateral CBD, PaCO2 was unchanged from its elevated level at rest, but, 10-15 days after CBD, PaCO2 decreased slightly from rest during exercise. These data indicate that 1) carotid afferents are an important determinant of rest and exercise breathing and ventilatory CO2 sensitivity, and 2) apparent plasticity within the ventilatory control system eventually provides compensation for chronic loss of these afferents.