Neurobehavioral effects of acute exposure to isoparaffinic and cycloparaffinic hydrocarbons.

This article reports neurobehavioral tests in rats with C5-C11 isoparaffinic and cycloparaffinic hydrocarbons. Testing, conducted shortly after exposure, evaluated the effects in several domains including clinical effects, motor activity, functional observations, and visual discrimination performance. Isopentane and cyclopentane did not produce any evidence of acute central nervous system (CNS) effects at levels up to 20 000 mg/m(3). A C(6)/C(7) mixed cycloparaffinic solvent produced minor, reversible changes in latency to response in visual discrimination testing at 14 000 mg/m(3); the no-effect level was 4200 mg/m(3). A C(8) isoparaffin produced no effects at 14 000 mg/m(3), the highest level tested. A C(9)/C(11) isoparaffinic solvent produced minor acute CNS effects at 5000 mg/m(3), with 1500 mg/m(3) as the no-effect level. A C(10) cycloparaffinic solvent did not produce any statistically significant CNS effects at 5000 mg/m(3). These studies were designed to provide data that may be useful in setting occupational exposure limits for C5-C11 isoparaffinic and cycloparaffinic hydrocarbons.

Neurobehavioral effects of acute exposure to normal (n-) paraffins.

This article reports the results of neurobehavioral tests on C(5)-C(10) normal paraffinic constituents (n-paraffins). Shortly after exposure, effects were evaluated in several domains including clinical effects, motor activity, functional observations, and visual discrimination performance. The representative C(5) n-paraffin, n-pentane, did not produce any evidence of acute central nervous system (CNS) effects at levels up to 20 000 mg/m(3). Similarly, there was no compelling evidence that n-octane (C(8)) produced CNS effects at 14 000 mg/m(3), the highest concentration tested. n-decane (C(10)) produced minor, reversible acute CNS effects at 5000 mg/m(3), with 1500 mg/m(3) as the no-effect level. Consistent with literature data, there seemed to be a relationship between increasing molecular weight up to C(10) and acute CNS effects. However, the CNS effects were reversible. Repeated exposures did not provide evidence of metabolic induction.

Neurobehavioral effects of acute exposure to aromatic hydrocarbons.

This article reports the results of neurobehavioral tests on representative aromatic constituents, specifically C(9) to C(11) species. The testing evaluated effects in several domains including clinical effects, motor activity, functional observations, and visual discrimination performance. Exposures ranging from 600 to 5000 mg/m(3), depending on the molecular weights of the specific aromatic constituents, produced minor, reversible effects on the central nervous system (CNS), particularly in the domains of gait and visual discrimination. There was little evidence of effects at lower exposure levels. There was some evidence of respiratory effects at 5000 mg/m(3) in 1 study, and there were also minor changes in body weight and temperature. The CNS effects became less pronounced with repeated exposures, corresponding to lower concentrations in the brain of 1 representative substance, 1,2,4-trimethyl benzene (TMB). At high exposure levels, the alkyl benzenes apparently induced their own metabolism, increasing elimination rates.

Neurobehavioral effects of cyclohexane in rat and human.

The neurobehavioral effects of inhaled cyclohexane in rats and humans are investigated to define relationships between internal doses and acute central nervous system effects. Rats are exposed for 3 consecutive days at target concentrations of 0, 1.4, 8, and 28 g/m(3), 8 h/d. Measurements include standardized observational measures, spontaneous motor activity assessments, and learned visual discrimination performance. Cyclohexane concentrations in blood and brain are measured to assess internal exposure. Human volunteers are exposed for 4 hours to 86 or 860 mg/m(3) in 2 test sessions. Neurobehavioral effects are measured using a computerized neurobehavioral test battery. In rats, there are slight reductions in psychomotor speed in the high-exposure group but minimal central nervous system effects. In humans, there are no significant treatment-related effects at the levels tested.

Physiologically based pharmacokinetic modeling of cyclohexane as a tool for integrating animal and human test data.

This report describes a physiologically based pharmacokinetic model for cyclohexane and its use in comparing internal doses in rats and volunteers following inhalation exposures. Parameters describing saturable metabolism of cyclohexane are measured in rats and used along with experimentally determined partition coefficients. The model is evaluated by comparing predicted blood and brain concentrations to data from studies in rats and then allometrically scaling the results to humans. Levels of cyclohexane in blood and exhaled air are measured in human volunteers and compared with model values. The model predicts that exposure of volunteers to cyclohexane at levels of 4100 mg/m(3) ( approximately 1200 ppm) will result in brain levels similar to those in rats exposed to 8000 mg/m(3) (the no-effect level for acute central nervous system effects). There are no acute central nervous system effects in humans exposed to 860 mg/m(3), consistent with model predictions that current occupational exposure levels for cyclohexane protect against acute central nervous system effects.

Model studies for evaluating the neurobehavioral effects of complex hydrocarbon solvents III. PBPK modeling of white spirit constituents as a tool for integrating animal and human test data.

As part of a project designed to develop a framework for extrapolating acute central nervous system (CNS) effects of hydrocarbon solvents in animals to humans, experimental studies were conducted in rats and human volunteers in which acute CNS effects were measured and toxicokinetic data were collected. A complex hydrocarbon solvent, white spirit (WS) was used as a model solvent and two marker compounds for WS, 1,2,4-trimethyl benzene (TMB) and n-decane (NDEC), were analyzed to characterize internal exposure after WS inhalation. Toxicokinetic data on blood and brain concentrations of the two marker compounds in the rat, together with in vitro partition coefficients were used to develop physiologically based pharmacokinetic (PBPK) models for TMB and NDEC. The rat models were then allometrically scaled to obtain models for inhalatory exposure for man. The human models were validated with blood and alveolar air kinetics of TMB and NDEC, measured in human volunteers. Using these models, it was predicted that external exposures to WS in the range of 344-771mg/m(3) would produce brain concentrations similar to those in rats exposed to 600mg/m(3) WS, the no effect level (NOEL) for acute CNS effects. Assuming similar brain concentration-effect relations for humans and rats, the NOEL for acute CNS effects in humans should be in this range. The prediction was consistent with data from a human volunteer study in which the only statistically significant finding was a small change in the simple reaction time test following 4h exposure to approximately 570mg/m(3) WS. Thus, the data indicated that the results of animal studies could be used to predict a no effect level for acute CNS depression in humans, consistent with the framework described above.

Model studies for evaluating the neurobehavioral effects of complex hydrocarbon solvents II. Neurobehavioral effects of white spirit in rat and human.

To evaluate the neurobehavioral effects of hydrocarbon solvents and to establish a working model for extrapolating animal test data to humans, studies were conducted which involved inhalation exposure of rats and humans to white spirit (WS). The specific objectives of these studies were to evaluate the behavioral effects of exposure to WS in rats and humans and to determine relationships between internal levels of exposure and behavioral effects. In both animals and volunteers, methods for assessment of similar functional effects were used to enable interspecies comparisons. A battery of tests including standardized observational measures, spontaneous motor activity assessments and learned visual discrimination performance was utilized in rat studies to evaluate acute central nervous system (CNS) depression. Groups of rats were exposed to WS at target concentrations of 0, 600, 2400 or 4800mg/m(3), 8h/day for 3 consecutive days. Blood and brain concentrations of two WS constituents; 1,2,4-trimethylbenzene (TMB) and n-decane (NDEC), were used as biomarkers of internal exposure. In a volunteer study, 12 healthy male subjects were exposed for 4h to either 57 or 570mg/m(3) WS in two test sessions spaced 7 days apart, and neurobehavioral effects were measured using a computerized neurobehavioral test battery. Blood samples were taken at the end of the exposure period to measure internal concentrations of TMB and NDEC. Results of the behavioral tests in rats indicated WS-induced changes particularly in performance and learned behavior. In humans, some subtle performance deficits were observed, particularly in attention. The behavioral effects were related to concentrations of the WS components in the central nervous system. These studies demonstrated a qualitative similarity in response between rats and humans, adding support to the view that the rodent tests can be used to predict levels of response in humans and to assist in setting occupational exposure levels for hydrocarbon solvents.

Model studies for evaluating the acute neurobehavioral effects of complex hydrocarbon solvents I. Validation of methods with ethanol.

As a preliminary step to evaluating the acute neurobehavioral effects of hydrocarbon solvents and to establish a working model for extrapolating animal test data to humans, joint neurobehavioral/toxicokinetic studies were conducted which involved administering ethanol to rats and volunteers. The specific objectives of the present studies were to evaluate the acute central nervous system (CNS) effects of ethanol in rats and humans and to assess relationships between internal levels of exposure and behavioral effects. A more general objective was to validate a battery of neurobehavioral tests that could be used to carry out comparative studies in both species. Accordingly, a range of tests including standardized observational measures, spontaneous motor activity assessments and learned visual discrimination performance was utilized in rat studies to evaluate acute CNS effects. Groups of rats were given ethanol at levels of approximately 0.5, 1.0 or 2.0g/kg, with blood level measurements to verify internal doses. In a volunteer study, 12 healthy male subjects were given 0.65g/kg ethanol, a level approximating the limit for motor vehicle operation in The Netherlands, and neurobehavioral effects were measured prior to and 1 and 3h after ethanol administration, with a computerized neurobehavioral test battery. Blood and air measurements were made to quantify internal doses. Results of the behavioral tests in rats provided evidence of ethanol-induced changes in neuromuscular, sensori-motor, and activity domains. There were also significant changes in visual discrimination, particularly in the areas of general measures of responding and psychomotor speed. In humans there were small but statistically significant effects on learning and memory, psychomotor skills and attention. However, the effects were subtle and not all parameters within given domains were affected. These studies demonstrated a qualitative similarity in response between rats and humans.

An analysis of human response to the irritancy of acetone vapors.

Studies on the irritative effects of acetone vapor in humans and experimental animals have revealed large differences in the lowest acetone concentration found to be irritative to the respiratory tract and eyes. This has brought on much confusion in the process of setting occupational exposure limits for acetone. A literature survey was carried out focusing on the differences in results between studies using subjective (neuro)behavioral methods (questionnaires) and studies using objective measurements to detect odor and irritation thresholds. A critical review of published studies revealed that the odor detection threshold of acetone ranges from about 20 to about 400 ppm. Loss of sensitivity due to adaptation and/or habituation to acetone odor may occur, as was shown in studies comparing workers previously exposed to acetone with previously unexposed subjects. It further appeared that the sensory irritation threshold of acetone lies between 10,000 and 40,000 ppm. Thus, the threshold for sensory irritation is much higher than the odor detection limit, a conclusion that is supported by observations in anosmics, showing a ten times higher irritation threshold level than the odor threshold found in normosmics. The two-times higher sensory irritation threshold observed in acetone-exposed workers compared with previously nonexposed controls can apart from adaptation be ascribed to habituation. An evaluation of studies on subjectively reported irritation at acetone concentrations < 1000 ppm shows that perception of odor intensity, information bias, and exposure history (i.e., habituation) are confounding factors in the reporting of irritation thresholds and health symptoms. In conclusion, subjective measures alone are inappropriate for establishing sensory irritation effects and sensory irritation threshold levels of odorants such as acetone. Clearly, the sensory irritation threshold of acetone should be based on objective measurements.

Effects of exposure to trichloroethylene and noise on hearing in rats.

Four groups of rats (n=8 per group) were exposed to either 3000 ppm trichloroethylene (TCE) alone or to 95 dB SPL noise alone or to the combination of TCE and noise or to control conditions. Exposure was carried out 18 hours/day, 5 days/week for 3 weeks. Exposure to TCE alone resulted in hearing loss at 4, 8, 16 and 20 kHz, but not at 24 and 32 kHz. Hearing loss due to exposure to noise alone occurred at frequencies of 8, 16 and 20 kHz. In general, combined exposure to TCE and noise resulted in larger auditory threshold changes than that produced by either TCE alone or noise alone when measured 1 and 2 weeks after the completion of exposure. For frequencies of 8, 16 and 20 kHz, hearing loss due to combined TCE-noise exposure was not larger than the algebraic sum of hearing loss due to exposure to TCE or noise alone. However, at a frequency of 4 kHz, hearing loss due to combined exposure was significantly larger than that produced by TCE exposure alone or noise alone, which itself had no effect at this frequency. These results suggest evidence of an interaction of combined exposure to TCE and noise at the lower edge of the range of frequencies affected.

Multivariate time of peak effects assessment for use in selecting time of testing in acute neurotoxicity studies.

One of the aims of conducting observational assessments shortly following administration of a test compound is to provide information regarding the profile of acute neurotoxic effects. By limiting the time of peak effects (TOPE) determination to a time range-finding study using only gait and arousal as the end-points for determining time of peak effects, as was proposed in the IPCS/WHO Collaborative Study on Neurobehavioral Screening Methods protocol, it is possible that the time of testing selected for the acute study proper may underestimate other neurotoxic effects which show a different time course. We explored the feasibility of including measures of autonomic activity as well as clonic/tonic movements in the TOPE determination in two experiments using chlorpyrifos and carbaryl as test compounds. A scoring system based on the original operational definitions provided in the IPCS/WHO protocol was devised. Results indicated that there were considerable differences in the time course for autonomic effects and convulsive behavior in comparison to effects on gait and arousal. It is concluded that the use of a multivariate approach for TOPE determination may provide a more comprehensive empirical basis for selecting a testing time for studies designed to profile acute neurotoxic effects.

Changes in regional brain GFAP levels and behavioral functioning following subchronic lead acetate exposure in adult rats.

Adult male WAG/Rij/MBL rats were dosed with lead acetate at 0, 4.0, 8.0 or 12.5 mg/kg, 5 days per week for 4 weeks. Animals were assessed prior to exposure, at the end of the 4-week exposure period and after a 2-week recovery period using a functional observational battery (FOB) and motor activity assessment. Rats were sacrificed two weeks after the last test session and glial fibrillary acidic protein (GFAP) concentrations were measured in eight selected brain regions. A dose-dependent decrease in motor activity was observed immediately following the end of the exposure period with no differences observed 2 weeks after cessation of exposure. Alterations in gait, decreased fore- and hindlimb grip strength, and decreased arousal were also found. Behavioral changes were accompanied by reduced weight gain and decreased body temperature during the course of exposure. GFAP concentrations were elevated in the frontal cortex, occipital cortex, striatum' and hippocampus but not in thalamus, cerebellum or brain stem. These results indicate that lead causes functional effects in the adult rat which can be detected by neurobehavioral methods. Furthermore, region-specific alterations in brain GFAP concentrations provided evidence of specificity of lead neurotoxicity in the adult brain.

A change in the phosphorylation pattern of the 30000-33000 Mr synaptonemal complex proteins of the rat between early and mid-pachytene.

The lateral elements (LEs) of synaptonemal complexes (SCs) of the rat contain major components with relative electrophoretic mobilities (Mr s) of 30000-33000, which are the products of a single gene. After one-dimensional separation of SC proteins on polyacrylamide-SDS gels, these components show up as two major bands, whereas upon two-dimensional electrophoresis they are resolved in at least 24 spots, which focus at pH 6.5 to 9.5. In this paper we show that these spots represent phosphorylation variants. For the analysis of the phosphorylation of the 30000- to 33000-Mr SC components during progression through meiotic prophase, we developed a procedure for isolation of fractions of testicular cells of the rat that are enriched in separate stages of meiotic prophase. Analysis of the 30000- to 33000-Mr SC components in these fractions by two-dimensional electrophoresis and immunoblotting showed that phosphorylated variants of the 30000- to 33000-Mr SC proteins occur throughout meiotic prophase. However, the extent of phosphorylation changes between early and mid-pachytene, when one phosphate group is probably added to each of the variants.

The gene encoding a major component of the lateral elements of synaptonemal complexes of the rat is related to X-linked lymphocyte-regulated genes.

The lateral elements of synaptonemal complexes (SCs) of the rat contain major components with relative electrophoretic mobilities (M(r)S) of 30,000 and 33,000. After one-dimensional separation of SC proteins on polyacrylamide-sodium dodecyl sulfate gels, these components show up as two broad bands. These bands contain closely related proteins, as judged from their peptide maps and immunological reactivity. Using affinity-purified polyclonal anti-30,000- and anti-33,000-M(r) component antibodies, we isolated a cDNA encoding at least one of the 30,000- or 33,000-M(r) SC components. The protein predicted from the nucleotide sequence of the cDNA, called SCP3 (for synaptonemal complex protein 3), has a molecular mass of 29.7 kDa and a pI value of 9.4. It has a potential nucleotide binding site and contains stretches that are predicted to be capable of forming coiled-coil structures. In the male rat, the gene encoding SCP3 is transcribed exclusively in the testis. SCP3 has significant amino acid similarity to the pM1 protein, which is one of the predicted products of an X-linked lymphocyte-regulated gene family of the mouse: there are 63% amino acid sequence similarity and 35% amino acid identity between the SCP3 and pM1 proteins. However, SCP3 differs from pM1 in several respects, and whether the proteins fulfill related functions is still an open question.

Mid-frequency hearing loss and reduction of acoustic startle responding in rats following trichloroethylene exposure.

Modification of auditory evoked startle responding using prepulse inhibition was used to examine the effects of trichloroethylene (TCE) exposure on auditory thresholds. Rats were exposed by inhalation to 0, 1500, or 3000 ppm TCE for 18 hours per day, 5 days a week for 3 weeks. Auditory thresholds for 5 and 20 kHz tones were measured before exposure and at 1, 3, and 6 weeks postexposure. In addition, hearing thresholds for 5 and 35 kHz tones were examined at a 5-week postexposure time-point. Results indicated that hearing thresholds for 20 kHz but not for 5 or 35 kHz prepulses were significantly increased in rats exposed to 3000 ppm TCE. These findings demonstrate a selective hearing loss in the 20 kHz range by short-term, high-level TCE exposure. With respect to effects on startle responding per se, the present study also found that compared to controls, TCE-exposed rats failed to show an increase in baseline startle with repeated testing. This difference could not be attributed to differences in body weight and was persistent throughout the postexposure period.

Interactions between simultaneously activated behavioral systems in the rat.

Interactions between electrically induced attack and teeth-chattering from 1 electrode and grooming from another were examined in male albino rats. The interaction between electrically induced attack and deprivation-induced feeding, as well as the effect of food deprivation on attack, was also studied. Results indicate that attack appears to be a dominant response, for it suppressed grooming and feeding at a low level of activation. On the other hand, it was not affected by simultaneously induced grooming or feeding. However, food deprivation decreased the threshold for attack, leaving attack latency, attack form, or bite targets unaffected. Teeth-chattering, suggested to be related to attack and flight, was also a dominant response. Results suggest that interactions between behavioral systems are in favor of the systems that must act acutely on activation in order to survive. Apparently, the regulations governing these interactions are represented in the functional organization of the brain.

Hypothalamic substrates for brain stimulation-induced patterns of locomotion and escape jumps in the rat.

The hypothalamic response area for electrically induced locomotion was determined using moveable electrodes and discriminant analysis as an appropriate statistical technique. At 241 out of 641 stimulated sites locomotion was induced. The distribution of locomotion sites is relatively diffuse. Discriminant analysis of both positive and negative electrode localizations yields areas with high, intermediate or low probability of inducing the response. The response is considered to be mediated by fibres of the subpallido-pedunculopontine system, which includes the mesencephalic locomotor region. Different categories of exploratory and flight-directed locomotion were distinguished, and response areas for both categories were determined. In addition the response area for escape jumps was delimited. Exploratory locomotion is mainly induced from the lateral hypothalamus, while flight-directed locomotion and escape jumps are evoked from the medial hypothalamus. The response area for exploratory locomotion reflects the lateral hypothalamic distribution of the subpallidal projection to the mesencephalic locomotor region. A diffuse substrate for flight behavior seems to occupy almost the entire medial hypothalamus. It is concluded that a locomotor subroutine subserving different behavioural mechanisms can be activated at many hypothalamic sites.

Hypothalamic substrates for brain stimulation-induced attack, teeth-chattering and social grooming in the rat.

In this paper the boundaries of the hypothalamic response areas for brain stimulation-induced attack, social grooming and teeth-chattering were delimited. A total of 641 hypothalamic sites in 71 male CPW/WU Wistar rats were electrically stimulated. Positive sites for any behavioural response cluster into restricted hypothalamic areas. Discriminant analysis of both positive and negative electrode localizations yields areas with high, intermediate and low probabilities of inducing the behavioural response concerned. Each response has its own response area where probabilities are high. Neuroanatomical correlates of these response areas are discussed. The response area of attack is suggested to be an integrative processing area, stimulation of which overrules some aspects of integration and directly activates the behavioural program of attack. Although some authors consider all three responses to be part of the behavioural repertoire of aggression, the response areas are not identical. Social grooming and attack are considered to be induced from different neural systems. Similarly, attack and teeth-chattering have been shown to derive from different neural mechanisms, despite substantial overlap of both response areas. It is suggested that teeth-chattering derives from the simultaneous activation of both attack and flight tendencies. No further distinctions with respect to threshold current intensities can be made within responses areas. However, the underlying neural substrates are not homogeneous, for thresholds vary along the course of individual electrodes.

Hypothalamic substrates for brain stimulation-induced grooming, digging and circling in the rat.

Despite a great number of studies concerned with the induction of specific behavioural responses from the rat hypothalamus by electrical brain stimulation, hypothalamic response areas and underlying neural substrates have never been determined accurately. In this study the boundaries of the hypothalamic response areas for grooming, digging and circling were delimited using moveable electrodes, an enriched environment containing a variety of goal objects, and an appropriate statistical technique. A total of 641 hypothalamic sites in 71 male CPB/WU Wistar rats were electrically stimulated. Results are plotted on a detailed stereotaxic brain atlas of the rat hypothalamus. Positive sites for any behavioural response cluster into restricted hypothalamic areas. Discriminant analysis of both positive and negative electrode localizations yields areas with high, intermediate or low probabilities of inducing the behavioural response concerned. Each response has its own response area where probabilities are high, although there may be overlap. Even within response areas a distinction can be made between areas in which the response can be induced at relatively high or low threshold current intensities. Lowest threshold sites within electrode tracks are often clustered. In search of neuroanatomical correlates, grooming is related to the distribution of ACTH-immunoreactive neural elements, digging is related to the distribution of efferent fibres from the bed nucleus of the stria terminalis, and circling is related to the distribution of dopaminergic fibres of the nigrostriatal pathway. The results clearly point to the stimulation site being the most important determinant of the evoked behavioural response. Evidently behavioural specificity does exist within the hypothalamus.