Effects of instilled combustion-derived particles in spontaneously hypertensive rats. Part I: Cardiovascular responses.

Epidemiological studies have reported statistically significant associations between the levels of ambient particulate matter (PM) and the incidence of morbidity and mortality, particularly among persons with cardiopulmonary disease. While similar effects have been demonstrated in animals, the mechanism(s) by which these effects are mediated are unresolved. To further investigate this phenomenon, the cardiovascular and thermoregulatory effects of an oil combustion-derived PM (HP-12) were examined in spontaneously hypertensive (SH) rats. The particle used in this study had considerably fewer water-soluble metals than the residual oil fly ash (ROFA) particles widely used in previous animal toxicity studies, with Zn and Ni constituting the primary water-leachable elements in HP-12. Rats were surgically implanted with radiotelemeters capable of continuously monitoring electrocardiogram (ECG), heart rate (HR), systemic arterial blood pressure (BP), and core temperature (T(co)). Animals were divided into four dose groups and were administered one of four doses of HP-12 suspended in saline vehicle (0.00, 0.83, 3.33, 8.33 mg/kg; control, low, mid, and high dose, respectively) via intratracheal instillation (IT). Telemetered rats were monitored continuously for up to 7 days post-IT, and were sacrificed 4 or 7 days post-IT. Exposures to mid- and high-dose HP-12 induced large decreases in HR (decreasing 30-120 bpm), BP (decreasing 20-30 mmHg), and T(co) (decreasing 1.2-2.6 degrees C). The decreases in HR and BP were most pronounced at night and did not return to pre-IT values until 72 and 48 h after dosing, respectively. ECG abnormalities (rhythm disturbances, bundle branch block) were observed primarily in the high-dose group. This study demonstrates substantial dose-related deficits in cardiac function in SH rats after IT exposure to a low-metal content, combustion-derived particle.

Effects of instilled combustion-derived particles in spontaneously hypertensive rats. Part II: Pulmonary responses.

A consistent association between exposure to high concentrations of ambient particulate matter (PM) and excess cardiopulmonary-related morbidity and mortality has been observed in numerous epidemiological studies, across many different geographical locations. To elicit a similar response in a controlled laboratory setting, spontaneously hypertensive rats were exposed to an oil combustion-derived PM (HP-12) and monitored for changes in pulmonary function and indices of pulmonary injury. Rats were implanted with radiotelemeters to monitor electrocardiogram, heart rate, systemic arterial blood pressure, core temperature, and activity. Animals were divided into four groups and exposed via intratracheal instillation (IT) to suspensions of HP-12 (0.0, 0.83, 3.33, and 8.33 mg/kg; control, low, mid, and high dose, respectively) in saline vehicle. Telemetered rats were monitored continuously for 4-7 days post-IT and pulmonary function was examined using a whole-body plethysmograph system for 6 h/day on post-IT days 1-7. At 24, 96, and 192 h post-IT, bronchoalveolar lavage fluid (BALF) was obtained from subsets of nontelemetered animals in order to assess the impact of HP-12 on biochemical indices of pulmonary inflammation and injury. Immediate dose-related changes in pulmonary function were observed after HP-12 exposure, consisting of decreases in tidal volume (decreasing 12-41%) and increases in breathing frequency (increasing 52-103%), minute ventilation (increasing 12-25%), and enhanced pause (increasing 113-187%). These functional effects were resolved by 7 days post-IT, although some average BALF constituents remained elevated through day 7 for mid- and high-dose groups when compared to those of the saline-treated control group. This study demonstrates significant deficits in pulmonary function, along with significant increases in BALF indices of pulmonary inflammation and injury in SH rats after IT exposure to HP-12.

Cardiovascular and thermoregulatory effects of inhaled PM-associated transition metals: a potential interaction between nickel and vanadium sulfate.

Recent epidemiological studies have shown an association between daily morbidity and mortality and ambient particulate matter (PM) air pollution. It has been proposed that bioavailable metal constituents of PM are responsible for many of the reported adverse health effects. Studies of instilled residual oil fly ash (ROFA) demonstrated immediate and delayed responses, consisting of bradycardia, hypothermia, and arrhythmogenesis in conscious, unrestrained rats. Further investigation of instilled ROFA-associated transition metals showed that vanadium (V) induced the immediate responses, while nickel (Ni) was responsible for the delayed effects. Furthermore, Ni potentiated the immediate effects caused by V when administered concomitantly. The present study examined the responses to these metals in a whole-body inhalation exposure. To ensure valid dosimetric comparisons with instillation studies, 4 target exposure concentrations ranging from 0.3-2.4 mg/m(3) were used to incorporate estimates of total inhalation dose derived using different ventilatory parameters. Rats were implanted with radiotelemetry transmitters to continuously acquire heart rate (HR), core temperature (T(CO)), and electrocardiographic data throughout the exposure. Animals were exposed to aerosolized Ni, V, or Ni + V for 6 h per day x 4 days, after which serum and bronchoalveolar lavage samples were taken. Even at the highest concentration, V failed to induce any significant change in HR or T(CO). Ni caused delayed bradycardia, hypothermia, and arrhythmogenesis at concentrations > 1.2 mg/m(3). When combined, Ni and V produced observable delayed effects at 0.5 mg/m(3) and potentiated responses at 1.3 mg/m(3), greater than were produced by the highest concentration of Ni (2.1 mg/m(3)) alone. These results indicate a possible synergistic relationship between inhaled Ni and V, and provide insight into potential interactions regarding the toxicity of PM-associated metals.

Cardiovascular and systemic responses to inhaled pollutants in rodents: effects of ozone and particulate matter.

Striking similarities have been observed in a number of extrapulmonary responses of rodents to seemingly disparate ambient pollutants. These responses are often characterized by primary decreases in important indices of cardiac and thermoregulatory function, along with secondary decreases in associated parameters. For example, when rats are exposed to typical experimental concentrations of ozone (O(3), they demonstrate robust and consistent decreases in heart rate (HR) ranging from 50 to 100 beats per minute, whereas core temperature (T(co) often falls 1.5-2.5 degrees C. Other related indices, such as metabolism, minute ventilation, blood pressure, and cardiac output, appear to exhibit similar deficits. The magnitudes of the observed decreases may be modulated by changes in experimental conditions and appear to vary inversely with both ambient temperature and body mass. More recent studies in which both healthy and compromised rats were exposed to either particulate matter or its specific components yielded similar results. The agents studied included representative examples of ambient, combustion, and natural source particles, along with individual or combined exposures to their primary metallic constituents. In addition to the substantial decreases in HR and T(co), similar to those seen with the O(3)-exposed rats, these animals also displayed numerous adverse changes in electrocardiographic waveforms and cardiac rhythm, frequently resulting in fatal outcomes. Although there is only limited experimental evidence that addresses the underlying mechanisms of these responses, there is some indication that they may be related to stimulation of pulmonary irritant receptors and that they may be at least partially mediated via the parasympathetic nervous system.

The spontaneously hypertensive rat as a model of human cardiovascular disease: evidence of exacerbated cardiopulmonary injury and oxidative stress from inhaled emission particulate matter.

Cardiovascular disease is considered a probable risk factor of particulate matter (PM)-related mortality and morbidity. It was hypothesized that rats with hereditary systemic hypertension and underlying cardiac disease would be more susceptible than healthy normotensive rats to pulmonary injury from inhaled residual oil fly ash (ROFA) PM. Eight spontaneously hypertensive (SH) and eight normotensive Wistar-Kyoto (WKY) rats (12-13 weeks old) were implanted with radiotelemetry transmitters on Day -10 for measurement of electrocardiographic (ECG) waveforms. These and other nonimplanted rats were exposed to filtered air or ROFA (containing leachable toxic levels of metals) on Day 0 by nose-only inhalation (ROFA, 15 mg/m(3) x 6 h/day x 3 days). ECGs were monitored during both exposure and nonexposure periods. At 0 or 18 h post-ROFA exposure, rats were assessed for airway hyperreactivity, pulmonary and cardiac histological lesions, bronchoalveolar lavage fluid (BALF) markers of lung injury, oxidative stress, and cytokine gene expression. Comparisons were made in two areas: (1) underlying cardiopulmonary complications of control SH rats in comparison to control WKY rats; and (2) ROFA-induced cardiopulmonary injury/inflammation and oxidative burden. With respect to the first area, control air-exposed SH rats had higher lung and left ventricular weights when compared to age-matched WKY rats. SH rats had hyporeactive airways to acetylcholine challenge. Lung histology revealed the presence of activated macrophages, neutrophils, and hemorrhage in control SHrats. Consistently, levels of BALF protein, macrophages, neutrophils, and red blood cells were also higher in SH rats. Thiobarbituric acid-reactive material in the BALF of air-exposed SH rats was significantly higher than that of WKY rats. Lung inflammation and lesions were mirrored in the higher basal levels of pulmonary cytokine mRNA expression. Cardiomyopathy and monocytic cell infiltration were apparent in the left ventricle of SH rats, along with increased cytokine expression. ECG demonstrated a depressed ST segment area in SH rats. With regard to the second area of comparison (ROFA-exposed rats), pulmonary histology indicated a slightly exacerbated pulmonary lesions including inflammatory response to ROFA in SH rats compared to WKY rats and ROFA-induced increases in BALF protein and albumin were significantly higher in SH rats than in WKY rats. In addition, ROFA caused an increase in BALF red blood cells in SH rats, indicating increased hemorrhage in the alveolar parenchyma. The number of alveolar macrophages increased more dramatically in SH rats following ROFA exposure, whereas neutrophils increased similarly in both strains. Despite greater pulmonary injury in SH rats, ROFA-induced increases in BALF GSH, ascorbate, and uric acid were attenuated when compared to WKY rats. ROFA inhalation exposure was associated with similar increases in pulmonary mRNA expression of IL-6, cellular fibronectin, and glucose-6-phosphate dehydrogenase (relative to that of beta-actin) in both rat strains. The expression of MIP-2 was increased in WKY but attenuated in SH rats. Thus, SH rats have underlying cardiac and pulmonary complications. When exposed to ROFA, SH rats exhibited exacerbated pulmonary injury, an attenuated antioxidant response, and acute depression in ST segment area of ECG, which is consistent with a greater susceptibility to adverse health effects of fugitive combustion PM. This study shows that the SH rat is a potentially useful model of genetically determined susceptibility with pulmonary and cardiovascular complications.

Cardiac and Thermoregulatory Toxicity of Residual Oil Fly Ash in Cardiopulmonary-Compromised Rats.

Recent epidemiological studies have reported a positive association between levels of ambient particulate matter (PM) and daily morbidity and mortality due to respiratory or cardiovascular causes; however, toxicological evidence supporting these findings is limited. The present study compared cardiac and thermoregulatory responses to intratracheal instillations of residual oil fly ash (ROFA) in normal and cardiopulmonary-compromised male Sprague-Dawley rats. Animals (n = 64) were implanted with radiotelemetry transmitters capable of continuously monitoring heart rate, core body temperature, and electrocardiographic waveforms. Comparisons of ROFA toxicity were conducted between (1) healthy rats and rats with cardiopulmonary stress or disease, including (2) rats exposed to an ambient temperature of 10° C, (3) rats preexposed to ozone to induce pulmonary inflammation, and (4) rats pretreated with monocrotaline (MCT) to induce pulmonary hypertension and vasculitis. Animals from each regimen were instilled with 1 of 4 doses of ROFA (0, 0.25, 1.0, 2.5 mg), and telemetry data were acquired for 96 h following ROFA instillation. Dose-related hypothermia and bradycardia were observed in healthy animals following exposure to ROFA; the magnitude and duration of these responses were potentiated in all compromised models. Delayed hypothermic and bradycardic responses occurred in healthy animals receiving 2.5 mg ROFA up to 48 h following instillation. These delayed responses were exacerbated in the MCT-and 10°C-exposure models, but attenuated in the 03-preexposed group. Additional observed effects of ROFA included induction of cardiac arrhythmias and increased mortality. These results demonstrate a distinct cardiac component to ROFA toxicity that agrees with epidemiological findings of PM-related excess cardiovascular mortality. Furthermore, the dose-related hypothermia and bradycardia observed in rodents from this study may confound the interpretation of results from similar air pollution toxicology studies.

Lung injury from intratracheal and inhalation exposures to residual oil fly ash in a rat model of monocrotaline-induced pulmonary hypertension.

A rat model of monocrotaline (MCT)-induced pulmonary injury/hypertension has been recently used in particulate matter (PM) health effects studies, however, results have been equivocal. Neither the mechanism by which mortality occurs in this model nor the variation in response due to differences in PM exposure protocols (i.e., a bolus dose delivered intratracheally versus a similar cumulative dose inhaled over three days) have been fully investigated. Sprague Dawley rats (SD, 60 d old; 250-300 g) were injected with either saline (healthy) or MCT, 60 mg/kg, i.p. (to induce pulmonary injury/hypertension). Ten days later they were exposed to residual oil fly ash (ROFA), either intratracheally (IT; saline, 0.83 or 3.33 mg/kg) or by nose-only inhalation (15 mg/m3 x 6 h/d x 3 d). Lung histology, pulmonary cytokine gene expression (0 and 18 h postinhalation), and bronchoalveolar lavage fluid (BALF) markers of injury were analyzed (24 and 96 h post-IT; or 18 h post-inhalation). Data comparisons examined three primary aspects, 1) ROFA IT versus inhalation effects in healthy rats; 2) pulmonary injury caused by MCT; and 3) exacerbation of ROFA effects in MCT rats. In the first aspect, pulmonary histological lesions following ROFA inhalation in healthy rats were characterized by edema, inflammatory cell infiltration, and thickening of alveolar walls. Increases in BALF markers of lung injury and inflammation were apparent in ROFA-IT or nose-only exposed healthy rats. Increased IL-6, and MIP-2 expression were also apparent in healthy rats following ROFA inhalation. In regards to the second aspect, MCT rats exposed to saline or air showed perivascular inflammatory cell infiltrates, increased presence of large macrophages, and alveolar thickening. Consistently, BALF protein, and inflammatory markers (macrophage and neutrophil counts) were elevated indicating pulmonary injury. In regards to the third aspect, 58% of MCT rats exposed to ROFA IT died within 96 h regardless of the dose. No mortality was observed using the inhalation protocol. ROFA inhalation in MCT rats caused exacerbation of lung lesions such as increased edema, alveolar wall thickening, and inflammatory cell infiltration. This exacerbation was also evident in terms of additive or more than additive increases in BALF neutrophils, macrophages and eosinophils. IL-6 but not MIP-2 expression was more than additive in MCT rats, and persisted over 18 h following ROFA. IL-10 and cellular fibronectin expression was only increased in MCT rats exposed to ROFA. In summary, only the bolus IT ROFA caused mortality in the rat model of lung injury/hypertension. Exacerbation of histological lesions and cytokine mRNA expression were most reflective of increased ROFA susceptibility in this model.

Cardiac arrhythmia induction after exposure to residual oil fly ash particles in a rodent model of pulmonary hypertension.

Recent epidemiological studies have reported a positive association between exposure to ambient concentrations of particulate matter (PM) and the incidence of cardiopulmonary-related morbidity and mortality. The present study examined the effects of fugitive residual oil fly ash (ROFA) PM on cardiac arrhythmia induction in healthy and cardiopulmonary-compromised rodents. Male Sprague-Dawley rats were implanted with radiotelemetry transmitters capable of monitoring the electrocardiogram and were subjected to one of two treatment regimens. Rats in the first treatment regimen (n = 16) served as normal control animals whereas rats in the second treatment regimen (n = 16) were injected with monocrotaline (MCT, 60 mg/kg, ip) to induce pulmonary vascular inflammation and hypertension and served as a model of cardiopulmonary disease. Rats within each treatment regimen were equally divided into four dose groups (0.0, 0.25, 1.0, 2.5 mg ROFA), instilled intratracheally, and monitored for 96 h. In the animals in the first treatment regimen, ROFA instillation caused dose-related increases in the incidence and duration of serious arrhythmic events that appeared to be associated with impaired atrioventricular conduction and myocardial hypoxia. There were no lethalities in the normal animals following ROFA instillation. The frequency and severity of arrhythmias were greatly exacerbated in the MCT-treated animals in the second treatment regimen and were accompanied by one, three, and two deaths in the low-, medium-, and high-dose groups, respectively. The results of the present study demonstrate substantial cardiac effects in normal and compromised rats after exposure to ROFA PM and implicate both conductive and hypoxemic arrhythmogenic mechanisms in the observed cardiac-related lethalities. These results support previous epidemiological studies that suggest a link between preexisting cardiopulmonary disease and potentiation of adverse health effects following exposure to anthropogenic particulates.

Impact of the hypothermic response in inhalation toxicology studies.

Previous studies from this laboratory showed that the decreases in Tco and associated functional parameters often observed in rodents following exposure to xenobiotic agents are capable of modulating the subsequent toxic response and that the magnitude of this induced hypothermic response may itself be modified by a number of experimental conditions. A moderate hypothermic response, characterized by a temperature drop of approximately 2 degrees C, appears to afford the optimal protection. Studies in which exposures occur through inhalation of harmful gases or particles present a special set of problems. In such studies, the dose of the toxic agent to which the animal is exposed is a function of the concentration of the agent in the atmosphere and the minute ventilation of the animal. Although ambient concentrations is generally held constant in laboratory studies, minute ventilation varies directly with metabolism, and both of these parameters may change significantly across experimental conditions. Thus, at low Tas, metabolism and minute ventilation are relatively high and uptake of inhalable toxic agents is increased. However, the development of the hypothermic response during the exposure entails a directly correlated reduction in these parameters and, presumably, in dose. For the most part, inhalation toxicological studies are conducted using resting animals or exercising humans. Animals are sometimes concurrently exposed to CO2 to simulate the increased ventilation of exercise and more closely mimic human studies. The experimental protocols employed in the above inhalation studies permitted examination of (1) the impact of species, size, handling stress, and changes in Ta on both the induced hypothermic response and the concomitant pulmonary toxicity; (2) the additive impact of exercise stress on O3 toxicity; and (3) the toxicity of ambient-derived particulate matter in normal rats and in rats with preexisting pulmonary inflammation. The results of these studies demonstrate that the magnitude of the induced hypothermic response is directly proportional to the uptake of the toxic agent by the lung and inversely proportional to the mass of the animal and the ambient temperature at which the exposure is conducted. The hypothermic response is sensitive to a number of experimental stresses including handling and changes in cage conditions. Exercise attenuates the hypothermic response, whereas CO2-stimulated increases in ventilation employed as an exercise surrogate may potentiate the response. Toxic exposures conducted in animals with lung disease or compromised pulmonary function may induce a severe hypothermic response while comparable exposures in normal animals produce only mild or moderate responses. In general, the development of the hypothermic response in the presence of ambient pollutants serves to decrease the minute ventilation of the animal and therefore limits the uptake and dose of the airborne toxicant. The results of these inhalation studies support our previous conclusions concerning the impact of the hypothermic response on toxicity and emphasize the need to monitor and incorporate these changes in functional parameters into analyses of toxicological data. Furthermore, because humans do not demonstrate a robust hypothermic response following exposure to toxic agents, extrapolation of the results obtained from animal studies and comparisons with data from human studies are considerably more complicated.

Mouse strain differences in ozone dosimetry and body temperature changes.

Strain differences in susceptibility to inhaled ozone (O3) have been observed in mice, with C57BL/6J (B6) mice reported to be more sensitive than C3H/HEJ (C3) mice when exposed to equal concentrations of O3. To determine whether differences in the delivered dose of O3 to the lung could help explain these differences, C3 and B6 mice were exposed to 18O-labeled ozone (18O3), and the resulting 18O concentrations in pulmonary tissues were monitored as an indicator of O3 delivered dose. Body core temperatures (Tco) of similarly treated mice were measured during O3 exposures (using surgically implanted temperature probes) in an effort to correlate lung O3 dose to changes in basal metabolism. Immediately after exposure to 18O3, C3 mice had 46% less 18O (per mg dry wt) in lungs and 61% less in tracheas than B6 mice. Nasal 18O tended to be lower in the C3 mice, but these differences were not significant. Although both strains responded to the O3 exposure with significant decreases in Tco, C3 mice had a 70% greater mean temperature x time product decrease during the exposure than B6 mice. These results suggest that the strain differences in O3 susceptibility may be due to differences in O3 dose to the lung, which may be related to differences in the ability of the mice to lower their Tco in response to O3 exposure.

Ambient particulate matter and respiratory and cardiovascular illness in adults: particle-borne transition metals and the heart-lung axis(,).

Epidemiological studies have consistently shown associations of exposure to ambient particulate matter (PM) with severe health effects, including mortality and hospitalization, in adults. From the standpoints of both relative risk and attributable risk, the public health burden of ambient PM exposure is potentially greatest in elderly adults with underlying cardiopulmonary illness. Recent experimental data suggest that PM-borne transition metals have toxicity that could be mechanistically relevant to PM-related epidemiological findings. These data may prove to be especially relevant in elderly adults with cardiopulmonary illness. At the same time, important uncertainties remain in the epidemiological and experimental databases, such that the true degree of correspondence between the two is not yet known. In our opinion, this combination of emerging experimental-epidemiological coherence and remaining uncertainty imparts high priority to further research into the health effects of PM-borne transition metals. This research should not be confined to the respiratory system. Rather, it should examine the entire heart-lung axis and should probably consider other body systems (e.g. the vascular system) as well. In this research, close interdisciplinary communication should be sustained and experimental and epidemiological approaches should be coordinated to the maximum feasible extent.

Ozone toxicity in the rat. III. Effect of changes in ambient temperature on pulmonary parameters.

Pulmonary toxicity of ozone (O3) was examined in adult male Fischer 344 rats exposed to 0.5 parts/million O3 for either 6 or 23 h/day over 5 days while maintained at an ambient temperature (Ta) of either 10, 22, or 34 degrees C. Toxicity was evaluated by using changes in lung volumes and the concentrations of constituents of bronchoalveolar lavage fluid that signal lung injury and/or inflammation. Results indicated that toxicity increased as Ta decreased. Exposures conducted at 10 degrees C were associated with the greatest decreases in body weight and total lung capacity and the greatest increases in lavageable protein, lysozyme, alkaline phosphatase activity, and percent neutrophils. O3 effects not modified by Ta included increases in residual volume and lavageable potassium, glucose, urea, and ascorbic acid with exposure at 34 degrees C. Most effects were attenuated during the 5 exposure days and/or returned to normal levels after 7 air recovery days, regardless of prior O3 exposure or Ta. It is possible that Ta-induced changes in metabolic rate may have altered ventilation and, therefore, the O3 doses among rats exposed at the three different Ta levels.

Ozone toxicity in the mouse: comparison and modeling of responses in susceptible and resistant strains.

Previous studies from this laboratory have demonstrated a concentration-related hypothermia and increases in bronchoalveolar lavage (BAL) fluid indexes of toxicity in the rat after exposure to environmentally relevant levels of ozone (O3). In similar studies with C57BL/6J (B6) and C3H/HeJ (C3) mice, other investigators have reported differential effects on BAL toxicity indexes between the two strains after O3 exposure. The present study investigated the relationship between the reported strain differences in BAL parameters in B6 and C3 mice exposed to O3 and the induced hypothermic response. Male 80-day-old mice (n = 94, 47/strain) were used for these studies. Subsets (n = 8/strain) of these animals were surgically implanted with radiotelemetry transmitters that permitted continuous monitoring of core body temperature and activity. All telemetry animals and an equal number of nontelemetry animals (n = 8/strain) were exposed to filtered air for 24 h followed by a 2-h exposure to 2 parts/million 16O3. With use of a similar protocol, groups of nontelemetry mice (n = 12/strain) were exposed to either filtered air or 2 parts/million 16O3 for 2 h. At 0 or 22 h postexposure, mice were anesthetized with halothane and intubated, and their lungs were lavaged with 37 degrees C saline. Although both strains of mice exhibited significant abrupt decreases in core body temperature on exposure to O3 and both recovered rapidly after cessation of the O3 exposure, the response of the C3 mice was more dynamic than that of the B6 mice. Similarly, both strains showed characteristic changes in biomarkers of O3 toxicity; however, the increases in BAL fluid protein and cells at 22 h postexposure were significantly greater and the percentage of neutrophils was significantly less in B6 mice than in C3 mice. It is possible that the strain differences in BAL constituents may be related to the differences in the hypothermic response.

Ozone toxicity in the rat. II. Modeling changes due to ambient temperatures and duration.

Previous studies involving exposures to xenobiotic agents have demonstrated decreases in physiological parameters such as heart rate (HR) and core body temperature (Tco) and have shown that these toxic responses are modulated by changes in ambient temperature (Ta). We recently published the results of a study in which male Fischer 344 rats were implanted with radiotelemetry transmitters that permitted continuous monitoring of HR, Tco, and motor activity. These animals were divided into nine treatment groups (n = 4-5/group) composed of combinations of one of three O3-exposure regimens [0.0 parts/million (ppm) O3 x 24 h/day x 5 days; 0.5 ppm O3 x 6 h/day x 5 days; or 0.5 ppm O3 x 23 h/day x 5 days] at one of three Ta values (10,22, or 34 degrees C). We now report on statistical approaches for the modeling and analyses of these data. The models utilized were dependent on the treatment combinations. Circadian rhythms of Tco during air control periods were fit by cosine models. Overall effects of O3 in the 6- and 23-h exposure groups were best modeled by modified damped-sine and one-compartment models, respectively, for both HR and Tco. These results demonstrate improved methods for the evaluation of biorhythmicity.

Strain differences in the laboratory rat: impact on the autonomic, behavioral, and biochemical response to cholinesterase inhibition.

Intraspecies variation has been found to affect the physiological, behavioral, and biochemical responses to a variety of neurotoxicants, including the organophosphate diisopropyl fluorophosphate (DFP). However, there is little information on long-term physiological responses to neurotoxicant exposure using strain as a dependent variable. In the present study, radiotelemetry methodology was used to continuously monitor core temperature, heart rate, and motor activity for 4 d following administration of 1.5 mg/kg DFP (sc) in four common strains of rat: Sprague-Dawley (SD), Long-Evans (LE), Fischer 344 (F344), and Wistar (WST). The F344 rat was least susceptible to DFP in terms of both a minimal hypothermic response and recovery of the day-night difference in core temperature. The SD strain was unusual in that its heart rate was elevated relative to the other strains after DFP, in spite of a marked decrease in core temperature and motor activity. The LE strain exhibited the largest reduction in core temperature and heart rate following DFP. Serum and brain cholinesterase activity (ChE) measured 3 h after administration of 1.0 mg/kg DFP also indicated strain effects. The F344 showed less inhibition in these variables compared to the other strains, a response that may explain its attenuated thermoregulatory response to DFP. Overall, the inbred F344 rat demonstrated better resistance to DFP compared to the outbred strains. Therefore, the impact of genetic differences on sensitivity to neurotoxicants such as DFP could be an important tool in understanding the mechanism of action of these agents.

Restrictive lung disease in rats exposed chronically to an urban profile of ozone.

The potential for irreversible lung impairment resulting from life-long ozone (O3) exposure remains uncertain. To address this question, young adult rats (male, F-344) were exposed to a simulated urban profile of O3 for 1, 3, 13, 52, or 78 wk, after which pulmonary function tests were performed. To assess reversibility of effects, cohorts from the 13-, 52-, and 78-wk groups were evaluated, respectively, after an additional 6, 27, and 17 wk of clean air. Static and dynamic lung properties were based on measurements of lung volume apportionment, respiratory system compliance (Crs), DLCO, multibreath N2 washout, and maximum expiratory flow-volume relationships. Electrocardiography was also performed in unanesthetized, restrained rats after 52 and 78 wk, as were determinations of wet and dry lung weights, lung collagen, and associated connective tissue crosslinks. Small (< 10%) but significant reductions in TLC and RV were noted after 13, 52, and 78 wk of O3 exposure. At 13 and 52 wk, N2 washout was enhanced, though at 78 wk it was similar to control. None of these changes appeared progressive with continued O3 exposure. Post exposure to clean air did not completely reverse the reduction in TLC. Additionally, Crs, though not affected during O3 exposure, decreased during the air recovery. No O3-related changes in collagen were apparent, however. Thus, near life-long exposure of F-344 rats to a worse-case, urban profile of O3 appears to have led to a functionally restrictive, i.e. "stiffened," lung without overt fibrosis. Furthermore, certain aspects of the O3-induced effect were not fully reversible.

Ozone toxicity in the rat. I. Effect of changes in ambient temperature on extrapulmonary physiological parameters.

These studies examined the effects of exposure to near environmental levels of ozone (O3) on the unanesthetized unrestrained rat as well as the influence of changes in ambient temperature (Ta) on the observed responses. Male Fischer 344 rats were implanted with radiotelemetry transmitters that permitted continuous monitoring of electrocardiogram, heart rate, core body temperature (Tco), and activity. Telemetry animals (n = 4-5/group) were combined with nontelemetry animals to produce nine treatment groups (n = 44-50/group) composed of combinations of one of three O3 exposure regimens (0.0 ppm x 24 h/day, 0.5 ppm x 6 h/day, or 0.5 ppm x 23 h/day) paired with one of three Ta levels (10, 22, or 34 degrees C). The experimental protocol consisted of a Control Period (filtered air; 1 day), Treatment Period (O3; 5 days), and Recovery Period (filtered air; 7 days). At specific intervals during the experiment, subgroups (n = 6) of nontelemetry animals were randomly selected from each treatment group, anesthetized with urethan, and intubated, and their lungs were lavaged with warm saline. In general, results from the bronchoalveolar lavage procedure indicated that toxicity increased in magnitude and duration as the length of time of O3 exposure increased and the Ta decreased. Similarly, whereas minimal extrapulmonary effects were observed at an Ta of 34 degrees C, O3 exposures at Ta levels of 22 and 10 degrees C produced significant decreases in heart rate (160 and 210 beats/min, respectively), Tco (2.0 and 3.5 degrees C, respectively), and body weight (15 and 40 g, respectively). Decreases in these functional parameters reached their maxima over the first 2 exposure days and returned to control levels after the 3rd day of exposure. These data demonstrate the profound impact of Ta on Tco and other extrapulmonary parameters in the conscious unrestrained rat exposed to O3. Furthermore, these results suggest an integral role for both Tco and Ta in determination of the uptake of inhaled pollutants and modulation of the subsequent toxic effects and may have important implications with respect to the assessment of toxic risk.

Within-session changes in peak N160 amplitude of flash evoked potentials in rats.

The negative peak occurring approximately 160 ms after stimulation (peak N160) of flash evoked potentials (FEPs) of rats changes with repeated testing. Habituation, sensitization, and arousal have all been invoked to explain these changes, but few studies have directly tested these explanations. We examined within-session changes in peak N160 amplitude with repeated testing, and the modulatory effects of stimulus intensity and auditory white noise. Peak N160 amplitude increased with daily testing (between-session changes), and was larger at greater stimulus intensities. In contrast, peak N160 amplitude underwent within-session increases on early days and within-session decreases on later days. The within-session changes were not affected by stimulus intensity. In rats previously tested in a quiet environment, exposure to acoustic white noise increased motor activity and transiently decreased peak N160 amplitude, which then increased and subsequently decreased with continued photic and acoustic stimulation. Repeated testing in the presence of noise did not alter the within-session changes in peak N160 amplitude. Heart rate showed both within- and between-session decreases, but was unaffected by noise. The data suggest that the within-session changes in peak N160 amplitude may reflect a habituation-like response to the test environment.

Caveats regarding the use of the laboratory rat as a model for acute toxicological studies: modulation of the toxic response via physiological and behavioral mechanisms.

The rodent, specifically the laboratory rat, is the primary experimental animal used in toxicology testing. Despite its popularity, recent studies from our laboratory and others raise a number of questions concerning the rat's appropriateness as an animal model for toxicological studies. While there may be additional areas in which the rat and other small rodents fail to adequately mimic the human response to xenobiotic agents, this article will focus on the area of temperature regulation. Thus, this article will review the thermoregulatory response of the laboratory rat following acute exposure to toxic agents and examine the impact of this response on the extrapolation of toxicological data from experimental animals to humans. In general, the rat responds to acute intoxication by lowering its core temperature via both physiological and behavioral mechanisms, thereby attenuating the induced toxicity. Similar responses have not been reported in humans.

Near-lifetime exposure of the rat to a simulated urban profile of nitrogen dioxide: pulmonary function evaluation.

To investigate the potential for up to a near-lifetime exposure to high-ambient levels of nitrogen dioxide (NO2) to induce functional lung damage, groups of rats were exposed to air or a simulated urban profile of NO2 (0.5 ppm background, 1.5 ppm peak) for 1, 3, 13, 52, or 78 weeks. The dynamic, static, and diffusional characteristics of the lung were evaluated postexposure in anesthetized rats. Furthermore, for the 13-, 52-, and 78-week groups, additional animals were tested after a 6-, 26-, or 17-week period in filtered air, respectively. No significant NO2 differences between exposed and control animals were found for the nitrogen washout, compliance, lung volume, or diffusion capacity of carbon monoxide measurements. At 78 weeks, however, a reduction in delta FEF25%, an estimate of convexity in the later portion of the forced expiratory flow volume curve, was observed. Breathing patterns and mechanisms were also assessed postexposure in a parallel group of similarly exposed unanesthetized rats. These rats were examined during a filtered air, 4 and 8% carbon dioxide (CO2) challenge. In the unanesthetized rat, frequency of breathing was significantly decreased and tidal volume, expiratory resistance, and inspiratory and expiratory times tended to increase. For several of these variables, the largest response also occurred at 78 weeks and seemed to be exacerbated by CO2 challenge.(ABSTRACT TRUNCATED AT 250 WORDS)