A rat model of neurobehavioral sensitization to toluene.

Some individuals report that, following either a single high-level or repeated lower-level exposures to chemicals (initiation), subsequent exposure to very low concentrations of chemicals (triggering) produces a variety of adverse effects, including disruption of cognitive processes. Our objective was to model this two-step process in a laboratory animal. Two groups of 16 rats, eight male and eight female, received whole-body inhalation exposure to toluene, either at 80 ppm for 6 h/day for 4 weeks (Repeat group) or to 1600 ppm for 6 h/day on one day only (Acute group). Two other groups (Trigger group and Clean group) of 16 were sham-exposed. After 17 days without toluene exposure, the Acute, Repeat and Trigger groups began a series of daily toluene 'trigger' exposures (10 ppm for 1 h) followed immediately by testing on an operant repeated-acquisitions task requiring learning within and across sessions. The Clean group was sham-exposed prior to operant testing. Trigger or sham exposures and operant testing continued 5 days/week for 17 sessions. Analysis of variance revealed a variety of statistically significant (P < 0.05) differences between treatment groups. Furthermore, the patterns of differences between groups differed (P < 0.05) for female and male rats. For example, male rats of the Trigger group made the most responses, and female rats of the Repeat group responded most slowly. The observation of important changes in the operant behavior of female and male rats previously exposed to toluene, at relatively low concentrations (80 or 1600 ppm) and then later re-exposed at very low concentrations (10 ppm), is consistent with the experiences of humans reporting cognitive difficulties following acute or chronic exposures to chemicals.

Chronically indwelling venous cannula and automatic blood sampling system for use with nonhuman primates exposed to 60 Hz electric and magnetic fields.

An automated blood sampling system was developed for use with tethered baboons (Papio cynocephalus) during concurrent exposure to 60 Hz 30 kV/m electric fields and 0.1 mT (1.0 G) magnetic fields. The system was controlled by a FORTH-based microcomputer, which operated a pump, a fraction collector, and two pinch valves. A swivel mechanism at the end of the tether allowed the baboons to move freely in their cages. The hardware and software were designed for fail-safe operation. Heparinized saline was infused at a rate of 0.5 mL/min until a sample cycle was initiated. Then, blood was drawn from the animal into a storage tube at a rate of 12.5 mL/min, a sample of undiluted blood was taken from the end of the storage tube near the baboon, and the blood remaining in the storage tube was then flushed back into the animal. Use of the storage tube prevented the peristaltic pump rollers from pressing on tubing containing blood, and return of the blood diluted with saline limited the blood wasted per sample to less than 0.5 mL. The system functioned reliably in three experiments, collecting samples as scheduled 97% of the time. Although it was initially designed for and used successfully with primates in an electric and magnetic field environment, this type of system could be employed in many areas of biomedical research or medical treatment.

Regularly scheduled, day-time, slow-onset 60 Hz electric and magnetic field exposure does not depress serum melatonin concentration in nonhuman primates.

Experiments conducted with laboratory rodents indicate that exposure to 60 Hz electric fields or magnetic fields can suppress nocturnal melatonin concentrations in pineal gland and blood. In three experiments employing three field-exposed and three sham-exposed nonhuman primates, each implanted with an indwelling venous cannula to allow repeated blood sampling, we studied the effects of either 6 kv/m and 50 microT (0.05 G) or 30 kV/m and 100 microT (1.0 G) on serum melatonin patterns. The fields were ramped on and off slowly, so that no transients occurred. Extensive quality control for the melatonin assay, computerized control and monitoring of field intensities, and consistent exposure protocols were used. No changes in nocturnal serum melatonin concentration resulted from 6 weeks of day-time exposure with slow field onset/offset and a highly regular exposure protocol. These results indicate that, under the conditions tested, day-time exposure to 60 Hz electric and magnetic fields in combination does not result in melatonin suppression in primates.

A 60 Hz electric and magnetic field exposure facility for nonhuman primates: design and operational data during experiments.

A unique exposure facility was designed and constructed to generate large-scale vertical electric fields (EF) of up to 65 kV/m and horizontal magnetic fields (MF) of up to 100 microT (1G), so that the behavioral and neuroendocrine effects of 60 Hz EF or combined electric and magnetic field (E/MF) exposure could be examined using nonhuman primates as subjects. Facility design and operational problems and their solutions are presented, and representative operational data from four sets of experiments are provided. A specially designed, optically isolated, 4 cm spherical-dipole EF probe and a commercially available MF probe were used to map the EF and MF within the fiberglass animal cages. In addition, amplifiers, signal conditioners, and A/D converters provided EF, MF, and transformer signals to a microcomputer at 15 min intervals. The apparatus produced homogeneous, stable E/MF at the desired intensities, and the fiberglass cages did not produce appreciable distortion or attenuation. Levels of recognized EF artifacts such as corona and ozone were negligible. The facility worked as intended, providing a well-characterized and artifact-controlled environment for experiments with baboons (Papio cynocephalus).

Rapid-onset/offset, variably scheduled 60 Hz electric and magnetic field exposure reduces nocturnal serum melatonin concentration in nonhuman primates.

Experiments with rodents indicate that power-frequency electric field (EF) or magnetic field (MF) exposure can suppress the normal nocturnal increase in melatonin concentration in pineal gland and blood. In a separate set of three experiments conducted with nonhuman primates, we did not observe melatonin suppression as a result of 6 weeks of day-time exposure to combined 60 Hz electric and magnetic fields (E/MF) with regularly scheduled "slow" E/MF onsets/offsets. The study described here used a different exposure paradigm in which two baboons were exposed to E/MF with "rapid" E/MF onsets/offsets accompanied by EF transients not found with slowly ramped E/MF onset/offset; profound reductions in nocturnal serum melatonin concentration were observed in this experiment. If replicated in a more extensive experiment, the observation of melatonin suppression only in the presence of E/MF transients would suggest that very specific exposure parameters determine the effects of 60 Hz E/MF on melatonin.

Detection thresholds for 60 Hz electric fields by nonhuman primates.

Because responses of animals to detection of the presence of an electric field (EF) are a possible mechanism for production of biological effects, it is important to know what EF intensities are detectable. Operant methods were used to train six baboons (Papio cynocephalus) to perform a psychophysical task involving detection of EF presence. During the response phase of a trial, a subject responded on one push button to report the presence of the EF and on a different push button to report the absence of the EF. Correct reports of EF presence of absence produced delivery of food rewards. The subjects became proficient at performing this psychophysical detection task; during 35 days of testing, false alarm rates averaged 9%. The average EF detection threshold was 12 kV/m; the range of means among subjects was 5-15 kV/m. Two special test procedures confirmed that the subjects were responding directly to EF presence of absence and not to artifacts that might be associated with EF generation. The EF detection threshold of nonhuman primates is similar to thresholds reported for rats and humans.

Initial exposure to 30 kV/m or 60 kV/m 60 Hz electric fields produces temporary cessation of operant behavior of nonhuman primates.

In two separate experiments, we examined the effects of a 60 Hz electric field (EF) on performance of an operant schedule consisting of two signaled components: fixed-ratio (FR30) and differential reinforcement of low-rate (DRL20). In each experiment, 12 naive baboons (Papio cynocephalus) were assigned randomly to either an EF-exposed experimental group or a sham-exposed control group. A homogeneous vertical EF of 30 kV/m was used in one experiment; 60 kV/m was used in the other. The experimental design for both experiments included 6 week preexposure, exposure, and postexposure periods. The planned analyses indicated no evidence of statistically significant (P < .05) effects of EF exposure. However, exploratory analyses comparing performance during the last week of preexposure and the first week of exposure revealed statistically significant acute effects (work stoppage): The mean response rates of the EF-exposed groups were greatly reduced on day 1 of exposure but were normal by the end of day 2 of EF exposure. We hypothesize that introduction of a highly unusual stimulus, the EF, temporarily interfered with normal operant behavior to produce a primary work stoppage. Supplementary cross-over experiments added at the end of each main experiment indicated that work stoppage occurred again when formerly EF-exposed subjects served as sham-exposed controls, while other subjects received their first EF exposure. Presumably, reoccurrence of other stimuli correlated with initial exposure to the EF became sufficient to subsequently cause secondary work stoppage in the absence of direct EF exposure. The primary and secondary work-stoppage effects were reproducible.

Nonhuman primates will not respond to turn off strong 60 Hz electric fields.

Using a set of six baboons (Papio cynocephalus), we conducted a series of seven experiments designed to evaluate the potentially aversive character of a 60 Hz electric field (EF). Initially, the subjects were trained, using food rewards as the reinforcer, to respond only when a cue light was illuminated. Next, an EF was presented along with the cue light; responses produced delivery of a food pellet and turned off both the cue light and the EF. Then, stimulus and reward conditions were varied. We determined that 1) presence of a strong EF does not affect operant responding for food rewards, 2) subjects will not respond at normal rates when the only reinforcer is termination of a strong EF, 3) presence of a strong EF can serve as a discriminative stimulus, 4) presence of a strong EF does not affect extinction of an appetite-motivated task, and 5) presentation of an EF can become a secondary reinforcer. The pattern of results was consistent across all experiments, suggesting that an EF of as much 65 kV/m is not aversive to nonhuman primates. Separately, we demonstrated that the average EF detection threshold for baboons in 12 kV/m. Thus, EF exposure at intensities well above the detection threshold and at species-scaled EF strengths greater than those found environmentally does not appear to be aversive.

Exposure of baboons to combined 60 Hz electric and magnetic fields does not produce work stoppage or affect operant performance on a match-to-sample task.

We examined the effects of combined 60 Hz electric and magnetic field (EMF) exposure on performance of delayed match-to-sample (MTS) procedure involving the flash rate of a light as the stimulus. Six baboons (Papio cynocephalus) fully acquired the task; four others functioned accurately only when cued. All ten subjects were assigned to EMF-exposed or sham-exposed groups of five and were used to test for a work-stoppage effect that was previously observed with initial exposure to electric fields (EF) of 30 or 60 kV/m. Here, we report the results of two experiments, each consisting of 6 week preexposure, exposure, and postexposure periods. We found no evidence of work stoppage with fields of 6 kV/m and 50 microT (0.5 G) or with 30 kV/m and 100 microT (1.0 G). In neither experiment was there evidence of an adverse effect of 60 Hz EMF exposure on MTS performance.

Effects of concurrent exposure to 60 Hz electric and magnetic fields on the social behavior of baboons.

Our previous research has demonstrated that 30 or 60 kV/m electric fields (EF) reliably produce temporary increases in the performance of three categories of baboon social behavior: Passive Affinity, Tension, and Stereotypy. The experimental design included 6 week preexposure, exposure, and postexposure periods with experimental and control groups, each with eight subjects. Here, we report two experiments that evaluated the effects of combined EF and magnetic fields (MF) on baboon social behavior. One experiment demonstrated that exposure to 6 kV/m EF and 50 microT (0.5 G) MF produced Period x Group interactions for Stereotypy and Attack, but the previously observed increases in Passive Affinity, Tension, and Stereotypy did not occur. A second experiment demonstrated that exposure to 30 kV/m EF and 100 microT 1.0 G MF did not produce the same magnitude of increases in Passive Affinity, Tension, and Stereotypy observed previously with 30 kV/m EF alone. The exposed group exhibited decreased performance rates for several behavior categories during exposure with further declines during postexposure. The control group showed fewer downward trends across periods.

Initial studies on the effects of combined 60 Hz electric and magnetic field exposure on the immune system of nonhuman primates.

In a pilot immunology experiment, peripheral blood samples from six baboons (Papio cynocephalus) housed as a social group were collected during week 5 of preexposure, exposure, and postexposure periods that were each 6 weeks in duration. The subjects were exposed to vertical 6 kV/m and horizontal 50 microT (0.5 G) fields for 12 h per day. Lymphocytes collected during the exposure period displayed statistically significant (p < .05) reductions in CD3+ and CD4+ counts, interleukin 2 receptor expression, and proliferative response to pokeweed mitogen. A second experiment was conducted using samples from seven subjects exposed to 30 kV/m and 100 microT (1.0 G) and eight sham-exposed subjects. Statistically significant Period x Group interactions occurred for total white blood cell count and CD4+ to CD8+ ratio, but the pattern of results was not suggestive of an exposure-related effect. Although components of the nonhuman primate immune system appear to be affected by 60 Hz electric and magnetic field exposure in one of two experiments, additional experiments are required to evaluate this possibility.

Training adaptations of baboons to light and moderate treadmill exercise.

We evaluated training adaptations by 18 baboons (Papio cynocephalus anubis) to low and moderate quadrupedal walking exercise on a motorized treadmill. Moderate training produced 47% increases in lactate threshold, 63% increases in muscle citrate synthetase activity, increases in percentage of Type IIc muscle fibers, and reduced plasma insulin concentrations. Low training produced only reduced plasma insulin concentrations. Only results indicate that the baboon response to exercise training was similar to that of Homo sapiens, and dependent on exercise intensity.

Effects of a 30 kV/m, 60 Hz electric field on the social behavior of baboons: a crossover experiment.

Using a crossover experimental design, we evaluated our earlier findings that exposure to a 30 kV/m, 60 Hz electric field for 12 hours per day, 7 days per week for 6 weeks produced significant changes in the performance rates of social behaviors among young adult male baboons. In the crossover experiment, the former control group was exposed to a 30 kV/m, 60 Hz electric field for 3 weeks. Only an extremely small, incidental magnetic field was generated by the exposure apparatus. We found that electric-field exposure again produced increases in the performance rates that index Passive Affinity, Tension, and Stereotypy. These findings, combined with results from our other electric-field experiments, indicate that exposure to strong electric fields, in the absence of associated magnetic fields, consistently produces effects that are expressed as increases in rates of performance of social behaviors in young adult male baboons.

Effects of exposure to a 60-kV/m, 60-Hz electric field on the social behavior of baboons.

We found in a previously reported study that exposure to a 30-kV/m, 60-Hz electric field had significant effects on the social behavior of baboons. However, it was not established whether or not the effects were related specifically to the 30-kV/m intensity of the field. A new experiment was conducted to determine whether or not exposure to a 60-Hz electric field at 60 kV/m would produce like changes in the baboons' social behavior. We exposed one group of eight male baboons to an electric field 12 hours a day, 7 days a week, for 6 weeks. A second group of eight animals was maintained under sham-exposure (control) conditions. Rates of performing on each of six categories of social behavior and on four categories of nonsocial behavior were used as criteria for comparing exposed with unexposed subjects and for within-group comparisons during three six-week experimental periods: Pre-Exposure, Exposure, and Post-Exposure. The results indicate that (1) during the exposure period, exposed animals exhibited statistically significant differences from controls in means of performance rates based on several behavioral categories; (2) across all three periods, within-group comparisons revealed that behaviors of exposed baboons were significantly affected by exposure to the electric field; (3) changes in performance levels probably reflect a stress response to the electric field; and (4) the means of response rates of animals exposed at 60 kV/m were higher, but not double, those of animals exposed at 30 kV/m. As in the 30-kV/m experiment, animals exposed at 60 kV/m exhibited significant differences in performances of Passive Affinity, Tension, and Stereotypy. Mean rates of performing these categories were 122% (Passive Affinity), 48% (Tension), and 40% (Stereotypy) higher in the exposed group than in the control group during exposure to the 60-kV/m field.

Effects of exposure to 30 kV/m, 60-Hz electric fields on the social behavior of baboons.

We tested the hypothesis that exposure to a 30-kV/m, 60-Hz electric field produces significant change (stress) in the social behavior of adult male baboons (Papio cynocephalus anubis). One group of eight baboons was exposed to an electric field (12 hours per day, 7 days per week for 6 weeks) while a second group of eight baboons was maintained in a sham-exposure (control) condition. Exposed subjects and control subjects were compared over three, six-week experimental periods (pre-exposure, exposure, and post-exposure). Performance rates of six categories of social behaviors (passive affinity, active affinity, approach, tension, threat, and attack) and four categories of nonsocial behaviors (forage, manipulate, posture, and stereotypy) were used to compare the two groups. The results of our study indicate that 1) there were no significant differences between the two groups during the pre-exposure or post-exposure periods; 2) during the exposure period, experimental and control groups exhibited statistically significant differences in the mean performance rates of three behavior categories; 3) within-group comparisons across periods indicate that the experimentally exposed group exhibited statistically significant changes in passive affinity, tension, and stereotypy; and 4) changes in behavior performance among the exposed subjects reflect a stress response to the electric field.

Cigarette smoking, dietary hyperlipidemia, and experimental atherosclerosis in the baboon.

In separate experiments, we fed 30 male and 25 female baboons a diet enriched in cholesterol and saturated fat for periods of 3.3 and 2.6 years. Using operant conditioning with water rewards, we trained the animals to puff on smoking machines in a human-like manner. Half of the animals smoked more than 40 cigarettes per day, while the remaining animals (controls) puffed air. Initially, the diet produced twofold (males) and threefold (females) elevations from baseline levels in serum cholesterol concentrations, but over the course of the experiments, the serum cholesterol decreased to 1.5 (males) and 2.0 (females) times baseline levels in both cigarette smokers and controls. Blood carbon monoxide concentration, plasma thiocyanate concentration, and urine cotinine concentration were significantly greater in smokers than in controls. Responses to smoking in males included lymphocytosis, elevated fasting blood glucose concentration, and decreased seminal vesicle weight. In females, hemoglobin and mean corpuscular hemoglobin concentrations were elevated. The extent of atherosclerosis was examined after 2.8 (males) and 1.6 (females) years of smoking. Among males, the extent of lesions in carotid arteries was significantly greater in smokers than in controls, but there were no significant differences in atherosclerosis in the aorta or the brachial, iliac-femoral, or coronary arteries. Among females, there were no significant differences in atherosclerosis between smokers and controls in any artery. These experiments show little effect of 2 to 3 years of cigarette smoke inhalation and concurrent modest elevation of blood carboxyhemoglobin on experimental atherosclerosis in the presence of moderate hyperlipidemia.

Thyroid hormone levels and cigarette smoking in baboons.

Using a primate animal model, two studies were undertaken to examine the effects of cigarette smoking on thyroid hormone levels. In study 1, mean total triiodothyronine (total T3) and mean total thyroxine (total T4) levels were measured in two groups of baboons (Papio cynocephalus) who were taught to smoke cigarettes using operant conditioning techniques. The smokers were divided into established and naive smokers according to pack-years of exposure. A control group of never-smoker baboons was included for comparison. Blood sampling was done after long-term cigarette consumption and again 1 week after cigarette deprivation. In the naive smoker group, mean total T3 concentrations were reduced below control group values (P less than 0.05). After cigarette deprivation for 1 week, mean total T3 values returned to normal. No significant differences in total T4 levels were observed in either group. In study 2, we assessed some other indices of thyroid function. The same groups of baboons were divided into good and poor smokers by plasma cotinine and blood carboxyhemoglobin (% COHb) levels during 28 weeks of cigarette smoking activity. Immediate fluctuations and reductions in total T3 levels were observed that were not accompanied by reductions in total T4. The animals were then cigarette deprived for 1 week and blood samples were obtained every other day during this period. Significant increases in total T3 concentrations were observed in poor smokers immediately after cessation. Both groups also exhibited significant reductions (P less than 0.05) in T3 uptake and free T4 index (FT4I) when compared to control group values. These data suggest that poor smokers are more susceptible to thyroid hormone level shifts than more established smokers, since the established smokers become habituated to the compounds contained in cigarette smoke through repeated exposure.