Hindlimb Grip Strength Research Articles

SUBCHRONIC TOXICITY OF BENZOTHIOPHENE ON RATS FOLLOWING DIETARY EXPOSURE

The systemic and neurobehavioral effects of benzo\\[b]thiophene (routinely referred to as benzothiophene) were studied in rats following 13-wk oral exposure. Male (170 +/- 16 g) and female (146 +/- 12 g) Sprague-Dawley rats (10 animals per group) were fed diet containing 0.5, 5, 50, or 500 ppm benzothiophene for 13 wk. Control animals were given rat feed plus vehicle ( corn oil) only. No clinical signs of toxicity and neurobehavioral effects were observed using screening tests that included cage-side observations, righting reflex, open field activities, and forelimb and hindlimb grip strength. Elevated serum aspartate aminotransferase activity and bilirubin level were observed in highest dose females. Except for a statistically significant decrease in hematocrit in the highest dose males, benzothiophene exerted no marked effects on hematological parameters. Benzothiophene exposure did not result in alterations in hepatic alkaline phosphatase activity, or the typical hepatic phase I (aniline hydroxylase, ethoxyresorufin O -deethylase, pentoxyresorufin O -dealkylase, aminopyrine N -demethylase) and phase II (UDPglucuronosyltransferase, glutathione S -transferase) drug-metabolizing enzyme activities. No significant elevation in urinary ascorbic acid, protein, and N -acetylglucosaminidase activity was detected in the treated animals. Peribiliary fibrosis was the most significant histological change and occurred in the liver of females in the 50 and 500 ppm groups. Mild epithelial hyperplasia in the renal pelvis was detected in the majority of 5 and 50 ppm females, with epithelial hyperplasia in the urinary bladder observed in the 50 ppm females. In males, increased incidence and severity of mild binucleation of hepatocytes and mild thickening of the basement membrane in kidney cortex were observed at 500 ppm. Benzothiophene was not detected in the urine of high-dose animals at the termination of the experiment. Based on the kidney, hepatic, and hematocrit changes, the no-observed-adverse-effect level (NOAEL) in the diet was determined to be 0.5 ppm (0.04 mg/kg/ d) for females and 50 ppm (3.51 mg/kg/ d) for males.

Inhalation toxicity of methanol/gasoline in rats: effects of 13-week exposure.

The subchronic inhalation toxicity of a methanol/gasoline blend (85% methanol, 15% gasoline, v/v) was studied in rats. Sprague Dawley rats (10 animals per group) of both sexes were exposed to vapours of methanol/gasoline at 50/3, 500/30 and 5000/300ppm for 6 hours per day, 5 days per week, for 13 weeks. Control animals inhaled filtered room air only. Control recovery and high dose recovery groups were also included which inhaled room air for an extra 4 weeks following the treatment period. No clinical signs of toxicity were observed in the treatment group and their growth curves were not significantly different from the control. Except for decreased forelimb grip strength in high dose females, no treatment-related neurobehavioural effects (4-6 hours post inhalation) were observed using screening tests which included cage-side observations, righting reflex, open field activities, and forelimb and hindlimb grip strength. At necropsy, the organ to body weight ratios for the liver, spleen, testes, thymus and lungs were not significantly different from the control group. There were no treatment-related effects in the hematological endpoints and no elevation in serum formate levels. Minimal serum biochemical changes were observed with the only treatment-related change being the decreased creatinine in the females. A dose-related increase in urinary ascorbic acid was detected in males after 2, 4 and 8 weeks of exposure, but not after the 12th week, and in females only at week-2. Increased urinary albumin was observed in treated males starting at the lowest dose and at all exposure periods, but not in females. A treatment-related increase in urinary beta 2-microglobulin was detected in males at week-2 only. Except for mild to moderate mucous cell metaplasia in nasal septum B, which occurred more often and with a slightly higher degree of severity in the low dose groups of both sexes, and presence of a minimal degree of interstitial lymphocyte infiltration in the prostate glands in the high dose males. No other significant microscopic changes were observed in the tissues of treated animals. Based on the marked increase in urinary ascorbic acid and albumin in the high dose males and the decreased forelimb grip strength in the high dose females, we concluded that the no-observed adverse effect level (NOAEL) of methanol/gasoline vapour is 500/30 ppm.

Physiological characterization of Taxol-induced large-fiber sensory neuropathy in the rat.

The cancer chemotherapeutic agent Taxol (paclitaxel) causes a dose-related peripheral neuropathy in humans. We produced a dose-dependent large-fiber sensory neuropathy, without detrimental effects on general health, in mature rats by using two intravenous injections 3 days apart. Tests of other dosing schedules demonstrated the dependence of the severity of the neuropathy and of animal health on both the dose and the frequency of dosing. Pathologically, severe axonal degeneration and hypomyelination were observed in sections of dorsal roots, whereas ventral roots remained intact. Electrophysiologically, H-wave amplitudes in the hindlimb and amplitudes of predominantly sensory compound nerve action potentials in the tail were reduced. These effects persisted for at least 4 months after treatment. Motor amplitudes were not affected, but both motor and sensory conduction velocities decreased. The ability of rats to remain balanced on a narrow beam was impaired, indicating proprioceptive deficits. Muscle strength, measured by hindlimb and forelimb grip strength, and heat nociception, measured by tail-flick and hindlimb withdrawal tests, were not affected by Taxol. This model of Taxol-induced neuropathy in mature rats, with minimal effects on general health, parallels closely the clinical syndrome observed after Taxol treatment in humans.

Neurotoxicological evaluation of methyl tertiary-butyl ether in rats.

Methyl tertiary-butyl ether (MTBE) is an oxygenate that is added to gasoline to boost octane and enhance combustion, thereby reducing carbon monoxide and hydrocarbon tailpipe emissions. The acute and subchronic neurotoxicity of MTBE were evaluated in rats using a functional observation battery (FOB), measures of motor activity (MA) and a neuropathological evaluation. In the acute study, rats were exposed once to 0, 800, 4000 or 8000 ppm MTBE by inhalation for 6 h and then evaluated three times over a 24-h period. In the FOB evaluations, treatment-related effects were seen at the 1-h session immediately following exposure and were indicative of transient central nervous system (CNS) depression. Effects were most apparent in the high-dose group (8000 ppm) but were also evident to a lesser extent in the mid-dose (4000 ppm) group. Labored respiration, ataxia, duck-walk gait and decreases in muscle tone, hind-limb grip strength and treadmill performance were the most frequently noted findings. No significant effects were observed in the FOB when testing was conducted at 6 h and 24 h post-exposure. The pattern of motor activity measured in the different dose groups following exposure was also in keeping with a reversible CNS-depressant effect of MTBE. In the subchronic study, rats were exposed to 0, 800, 4000 or 8000 ppm MTBE for 6 h a day, 5 days per week, for 13 weeks. No persistent or cumulative effects on neurobehavioral function were found. Body weights and absolute brain weights were reduced in the 8000 ppm group, however there were no differences among groups when brain weight was expressed relative to body weight. No histopathological changes were noted in the brains or peripheral nervous tissues of MTBE-exposed animals. In summary, MTBE produced signs of acute reversible CNS depression following exposure to 8000 ppm and, to a lesser extent, to 4000 ppm vapor. The no-observed-adverse-effect level for these effects was 800 ppm in the present study. No persistent or cumulative neurotoxic effects were observed following exposure to MTBE at concentrations up to 8000 ppm for 13 weeks.

No changes in behaviour, nigro-striatal system neurochemistry or neuronal cell death following toxic multiple oral paraquat administration to rats.

We have examined whether the widely used herbicide, paraquat (1,1'-dimethyl-4,4'dipyridylium) may accumulate in rat brain following multiple oral dosing (5 mg paraquat ion/kg/day) for 14 days and whether this dosing regime may produce signs of neurotoxicity. This dosing regime may determine whether low dose exposure to mammals may be neurotoxic. Using [14C]paraquat to measure tissue and plasma paraquat concentrations, we observed significantly higher plasma and tissue paraquat concentrations in brain, liver, lungs and kidneys of rats which received multiple doses for 14 days, as compared to paraquat concentrations in tissues of rats which received only a single paraquat dose. Brain paraquat concentrations measured 24 h after dosing were tenfold higher in rats receiving 14 daily oral doses of paraquat, as compared to concentrations following a single oral dose. A neuropathological study of the rat brain yielded no evidence that multiple paraquat dosing resulted in neuronal cell damage. Particular attention was paid to the nigrostriatal system. The paraquat treated rats gained approximately 10% less body weight over the 15 day experimental period as compared with controls demonstrating that the dose of paraquat was toxic to the animals. Measurements of locomotor activity using open field tests or activity monitors did not reveal any statistically significant differences between control animals and those receiving paraquat. Fore- and hind-limb grip strength were not significantly different between the paraquat treated and control rats at any time point during the dosing regime, nor was there any evidence for locomotor coordination deficits in any of the animals receiving paraquat. Densities of dopamine D1 and D2, NMDA, muscarinic and benzodiazepine receptors in the cerebral cortex and striatum were not significantly different between controls and rats which had received multiple paraquat doses. Concentrations of catecholamine neurotransmitters in the striatum, hypothalamus and frontal cerebral cortex were also measured to examine whether there was evidence for catecholamine depletion in these brain regions. We did not observe any significant reductions in dopamine, noradrenaline or DOPAC concentrations in any brain region of paraquat treated rats as compared with controls. On the contrary, dopamine concentrations in the striatim were significantly elevated in paraquat treated animals following a 15 day paraquat dosing regime. We attribute these changes in catecholamine concentrations to the general toxicity of paraquat which produces a stress response. In conclusion, we could not find any evidence that multiple paraquat dosing can lead to changes in locomotor activity or grip strength. In addition, the absence of neuropathology or changes in neurochemistry in the nigrostriatal tract demonstrates that paraquat does not behave like MPP+(N-methyl-4-phenylpyridinium), the neurotoxic metabolite of MPTP.

NEUROTOXICITY OF MISONIDAZOLE IN RATS FOLLOWING INTRAVENOUS ADMINISTRATION

Misonidazole is a hypoxic cell radiosensitizer that induces a peripheral neuropathy in humans after exceeding a schedule-dependent cumulative threshold dose. Clinical studies of misonidazole have been conducted using oral administration, whereas most other radiosensitizers have been administered intravenously. Since route of exposure can potentially influence the toxicity of xenobiotics, the objective of this study was to assess the neurotoxicity of misonidazole in rats following intravenous dosing using a battery of routine clinical, neurofunctional, biochemical, and histopathologic screening methods. Male Sprague–Dawley rats were administered intravenous doses of misonidazole at 0 (vehicle control), 100, 200, 300, or 400mgkg −1once per day, 5 days per week, for 2 weeks. Animals were evaluated for neurofunctional and pathological changes following termination of treatment (Days 15–17) and at the end of a 4 week observation period (Days 43–45). During the dosing phase, hypoactivity, salivation, rhinorrhea, chromodacryorrhea, rough pelage and ataxia were observed at 400mgkg −1, and body weight gain of the 300 and 400mgkg −1groups was significantly decreased relative to the vehicle controls by 24% and 49%, respectively. Corresponding reductions in food consumption were 8% and 23%, respectively. Although most 400mgkg −1animals appeared normal on Day 15 prior to the neurofunctional evaluations, rotorod testing precipitated a number of clinical signs including: ataxia, impaired righting reflex, excessive rearing, tremors, vocalization, circling, head jerking, excessive sniffing and hyperactivity. All of these animals recovered and appeared normal from Day 17 through study termination. There were no treatment-related effects on motor activity, acoustic startle response, rotorod performance, forelimb group strength, toe and tail pinch reflexes, tibial nerve β-glucuronidase activity or tail nerve conduction velocity. Although hindlimb grip strength of the 400mgkg −1group was significantly decreased by 17% relative to the vehicle controls on Day 15, this finding appeared related to the reduced food consumption and body weight gain in these animals. No microscopic changes were detected in peripheral nerves. Necrosis and proliferation of fibrillary astrocytes (gliosis) were seen in the cerebellum and medulla of the 400mgkg −1animals on Day 16. Gliosis in these same brain regions was observed in the 300 and 400mgkg −1groups on Day 44. The results show that intravenous administration of misonidazole to rats causes dose-limiting central nervous system toxicity without effects on peripheral nervous tissue. The lack of peripheral neurotoxicity was most likely due to a combination of several interrelated factors including route of administration, duration and intensity of the dosing regimen, and total cumulative dose.

The Reproductive and Neural Toxicities of Acrylamide and Three Analogues in Swiss Mice, Evaluated Using the Continuous Breeding Protocol

The Reproductive and Neural Toxicities of Acrylamide and Three Analogues in Swiss Mice, Evaluated Using the Continuous Breeding Protocol. Chapin, R. E., Fail, P. A., George, J. D., Grizzle, T. B., Heindel, J. J., Harry, G. J., Collins, B. J., and Teague, J. (1995). Fundam. Appl. Toxicol. 27, 9-24. Acrylamide is a known genetic, reproductive, and neural toxicant, although it is not known if one effect is predominant. The toxicities of several structural analogues of acrylamide have been incompletely characterized, and the relative sensitivity of the second generation is not known. The present studies were designed to explore the relationship between neurotoxicity and reproductive toxicity, to further characterize the toxicities of three acrylamide analogues, and to evaluate the relative sensitivity of a second generation to these compounds. For the F 0 generation, male and female Swiss CD-1 mice were provided drinking water containing acrylamide (ACR; 3, 10, 30 ppm), N,N′-methylenebisacrylamide (MBA; 10, 30, 60 ppm), N -(hydroxymethyl)acrylamide (HMA; 60, 180, 360 ppm), or methacrylamide (MACR; 24, 80, 240 ppm) during and after a 14-week cohabitation. The last litter was reared and dosed after weaning until mating at 74 ± 10 days of age with the same level of compound given to the parents. Neurotoxicity was assessed at several times in both generations by measuring forelimb and hindlimb grip strength. In the F 0 generation, ACR caused an 11% decrease in pup number without measurable neurotoxicity; female fertility was not affected. Although both generations consumed the same amount of ACR, there were larger changes in the fertility-related endpoints in the F 1 mice than in the F 0's, with no concomitant change in organ weights or sperm parameters. In F 0 mice, MBA reduced the number of live pups and their adjusted weight, with no neurotoxicity and no change in F 0 female reproduction. MBA caused greater adverse effects in the second generation, concomitant with increased consumption. In the F 0 generation, HMA caused the largest decrease in pup number during cohabitation (26%) together with a small effect on grip strength. Female reproduction was not affected. The second generation consumed more HMA and showed slightly greater toxic effects. In both generations, MACR was negative for both neurotoxicity and reproductive toxicity. Dominant-lethal studies showed that the fertility effects for ACR, MBA, and HMA could be explained by a male-mediated increase in postimplantation loss. These studies found that dominant lethality occurred without structural effects on the reproductive system in the presence of only minor effects on grip strength and without detectable neural histopathology. Female reproduction was not significantly affected by these compounds at the doses used. Thus, these data confirm the male as the affected gender and that the reproductive toxicity was greater than motoneuron toxicity when measured as grip strength.

The Reproductive and Neural Toxicities of Acrylamide and Three Analogues in Swiss Mice, Evaluated Using the Continuous Breeding Protocol

Acrylamide is a known genetic, reproductive, and neural toxicant, although it is not known if one effect is predominant. The toxicities of several structural analogues of acrylamide have been incompletely characterized, and the relative sensitivity of the second generation is not known. The present studies were designed to explore the relationship between neurotoxicity and reproductive toxicity, to further characterize the toxicities of three acrylamide analogues, and to evaluate the relative sensitivity of a second generation to these compounds. For the F0 generation, male and female Swiss CD-1 mice were provided drinking water containing acrylamide (ACR; 3, 10, 30 ppm), N,N′-methylenebisacrylamide (MBA; 10, 30, 60 ppm), N-(hydroxymethyl)acrylamide (HMA; 60, 180, 360 ppm), or methacrylamide (MACR; 24, 80, 240 ppm) during and after a 14-week cohabitation. The last litter was reared and dosed after weaning until mating at 74±10 days of age with the same level of compound given to the parents. Neurotoxicity was assessed at several times in both generations by measuring forelimb and hindlimb grip strength. In the F0 generation, ACR caused an 11% decrease in pup number without measurable neurotoxicity; female fertility was not affected. Although both genera tions consumed the same amount of ACR1 there were larger changes in the fertility-related endpoints in the F1 mice than in the F0's, with no concomitant change in organ weights or sperm parameters. In F mice, MBA reduced the number of live pups and their adjusted weight, with no neurotoxicity and no change in F0 female reproduction. MBA caused greater adverse effects in the second generation, concomitant with increased consumption. In the F0 generation, HMA caused the largest decrease in pup number during cohabitation (26%) together with a small effect on grip strength. Female reproduction was not affected. The second generation consumed more HMA and showed slightly greater toxic effects. In both generations, MACR was negative for both neuro toxicity and reproductive toxicity. Dominant-lethal studies showed that the fertility effects for ACR, MBA, and HMA could be explained by a male-mediated increase in postimplantation loss. These studies found that dominant lethality occurred without structural effects on the reproductive system in the presence of only minor effects on grip strength and without detectable neural histopathology. Female reproduction was not significantly affected by these compounds at the doses used. Thus, these data confirm the male as the affected gender and that the reproductive toxicity was greater than motoneuron toxicity when measured as grip strength.

Neurotoxicity of acrylamide and 2,5-hexanedione in rats evaluated using a functional observational battery and pathological examination

The clinical effects of two neurotoxicants, acrylamide and 2,5-hexanedione, were compared in rats using a functional observational battery (FOB), which includes a series of home cage and open-field observations, sensorimotor measurements, and physiological parameters. Neurotoxicity was assessed weekly in adult male Long-Evans rats after initiation of IP administration of 9 doses of acrylamide (12, 15, or 50 mg/kg given 3 times a week) and 28 doses of 2,5-hexanedione (150, 225, and 350 mg/kg given daily). Using the FOB, it was possible to detect differences in neurotoxic effects of these two chemicals. Acrylamide significantly affected home cage posture, foot splay and time on the rotarod, whereas 2,5-hexanedione altered hindlimb grip strength and the approach response. Both compounds caused changes in ability to walk, right, and maintain agility on a rotarod within 21 days from initiation of toxicant administration. In addition, both compounds caused dose-dependent decreases in weight gain. Neuropathic changes were detectable at the highest dosages at 21 days in acrylamide-treated rats and at 28 days in rats treated with 2,5-hexanedione. Administration of acrylamide also decreased activities of neural esterases. This study indicated that the FOB could be used to detect evidence of neurotoxicity in rats treated with acrylamide and 2,5-hexanedione, with alterations evident even before pathological changes were induced by 2,5-hexanedione.

Auditory deficits and motor dysfunction following iminodipropionitrile administration in the rat

The behavioral effects of 3,3′-iminodipropionitrile (IDPN) were studied using reflex modification of the acoustic startle response and figure-eight maze activity. A number of experiments were conducted with separate groups of adult male Long-Evans hooded rats exposed to saline or 50–500 mg/kg IDPN for 3 consecutive days. Auditory thresholds (reflex modification), motor activity, and grip strength were measured 1 day, and 1, 3, and 9 weeks postdosing. Reflex inhibition was monitored daily, prior to, during, and for 7 days following exposure. Auditory thresholds for 5-and 40-kHz tones were elevated approximately 25 dB and 50 dB, respectively. The onset of this auditory dysfunction in the 200-mg/kg/day group, as demonstrated by a loss of reflex inhibition, was 2 days for the 40-kHz tone and 4 days for the 5-kHz tone. Motor activity was increased up to 400% in the 200-mg/kg group, whereas there was no alteration in hindlimb grip strength. These data demonstrate dosage- and time-dependent alterations in auditory and motor function following IDPN exposure.

The effects of 2,4-dithiobiuret on sensory and motor function

2,4-Dithiobiuret (DTB) exposure causes a delayed onset muscle weakness in rats that has been attributed to depressed neuromuscular transmission. The present study compares the effects of DTB on both sensory and motor function in rats. Adult male Long-Evans hooded rats were exposed to saline, 0.25, 0.5, or 1.0 mg/kg/day DTB, ip, for 5 consecutive days (Days 1–5). Body weights were monitored throughout the experiment. Motor activity was measured for 1 hr in figure-eight mazes on Days 0, 6, 13, and 27. Forelimb and hindlimb grip strength were assessed on Days 6, 13, and 27. Auditory thresholds were determined for 5-and 40-kHz tones using reflex modification of the startle response on Days 0, 7, 14, and 28. Visual function was examined on Day 6 in animals exposed at 0.5 mg/kg/day using flash-and pattern-elicited visual evoked potentials (FEPs and PEPs, respectively). Thermal sensitivity was measured using the hot plate procedure. All motor endpoints were decreased in a dosage-and time-dependent manner; the higher the dosage the longer the effects lasted. There were no effects on any measure of sensory function with the exception of peak N2 of the FEP. Both the amplitude and latency of FEP N2 were altered by DTB exposure. Decreases in body weight were maximal on Day 9 at 1.0 mg/kg/day (20% from control), but recovered by Day 22. Motor activity was suppressed on Day 6 only, whereas grip strength measures were decreased on both Days 6 and 13. Auditory thresholds were not significantly altered; however, baseline startle amplitude was decreased at the highest dosage on Days 7 and 14, but recovered by Day 28. Hot plate latencies were not altered by DTB treatment. These data demonstrate that DTB produces a reversible impairment of motor function, without altering auditory, thermal, or pattern visual function. FEP N2, which is thought to arise from activity generated in the superficial layers of visual cortex, was diminished by DTB treatment, indicating that DTB can alter the function of the CNS, although effects on the motor system are more pronounced.

Neurotoxicologic evaluation of rats after 13 weeks of inhalation exposure to dichloromethane or carbon monoxide

Male and female Fischer 344 rats were exposed to dichloromethane (methylene chloride, DCM) or carbon monoxide (CO) for 6 hr/day, 5 days/week, for 13 weeks. Since oxidative metabolism of DCM to CO and CO 2 is a saturable process, DCM exposure concentrations were selected clearly below saturation (50 ppm), just below saturation (200 ppm), and well above saturation (2000 ppm). At saturation of metabolism, metabolic CO causes about 10% carboxyhemoglobinemia (COHb). Therefore, as a control for CO effects, a separate group of rats was exposed to 135 ppm CO to induce approximately 10% COHb. Postexposure functional tests included an observational battery, hindlimb grip strength, and a battery of evoked potentials (flash, auditory brainstem, somatosensory, caudal nerve). After functional tests were completed, rats from all groups were perfused with fixative and a comprehensive set of nervous tissues from the high DCM exposure group and from controls were examined by light microscopy. Although some miscellaneous functional and morphologic variations were recorded, none were related to treatment. Thus, subchronic exposures as high as 2000 ppm DCM or 135 ppm CO had no deleterious effects on any of the measures of this study.

ZPT-related distal axonopathy: Behavioral and electrophysiologic correlates in rats

These experiments examined the relationship between behavioral and electrophysiologic signs of neuromuscular dysfunction in rats with zinc pyridinethione (ZPT)-induced neurotoxicity. ZPT added to the diet of adult rats at 50 ppm produced a moderate (approximately 200 g) reduction in forelimb and hindlimb grip strength which occurred during the second week of dosing. Other behavioral tests of peripheral nervous system toxicity were affected inconsistently. Electrophysiologic changes included a marked (maximum 95%) reduction of indirect muscle-evoked potential (M-wave) and a decrement (maximum 30%) during repetitive M-wave elicitation. Electrophysiologic changes were greater in hindlimb than in forelimb. Needle electromyography revealed denervation potentials in ZPT-treated rats which appeared after M-wave changes and recovered faster than did M-wave amplitude. Caudal nerve conduction velocity was unaffected, indicating that peripheral neurotoxic effects were confined to the neuromuscular junction. When ZPT exposure was discontinued, grip strength recovered in about 1 week. In contrast, electrophysiologic measures required 42 days to recover completely. These results indicate that deficits in neuromuscular junction physiology are a sensitive index of ZPT-related distal axonopathy.

Behavioral Indices of Neurotoxicity

There are many classes of chemicals widely used in a number of commercial and industrial processes having a potential to affect adversely the nervous system. Because of their relative sensitivity to some agents and general noninvasive characteristics, functional measurements of neurotoxicity are being used with greater frequency, especially at the level of hazard identification. The neurobehavioral test battery used by the National Toxicology Program (NTP) includes motor activity, fore- and hindlimb grip strength, acoustic startle response, responsiveness to an adverse thermal stimulus and general health and clinical measures (body weight, autonomic signs, tremor, convulsions). These tests have been used in nearly 40 studies involving various dosing regimens with rats and mice. The NTP battery shows several salient features of an effective screen, including the ability to differentiate known neurotoxicants from nonneurotoxicants, identify certain types of neurological sequelae or profiles of neurotoxic effects and construct dose- and time-response data in a screening context. The extent to which the NTP battery has predictive validity is still being evaluated.

Neurotoxicity and carcinogenicity ofN‐methylolacrylamide in F344 rats and B6C3F1 mice

Toxicology and carcinogenicity studies of N-methylolacrylamide were conducted by administering the chemical by gavage in water to both sexes of F344/N rats and B6C3F1 mice 5 times per week for 16 d, 13 wk, or 2 yr. In 16-d studies, rats receiving doses of 200 mg/kg or higher and mice receiving 400 mg/kg died. In 13-wk studies, all rats given 100 mg/kg or higher doses died. Rats receiving 50 mg/kg or higher doses developed hindlimb ataxia progressing to paralysis. In neurobehavioral assessments, decreased forelimb and hindlimb grip strength occurred in rats at doses as low as 12.5 mg/kg. Landing footspread was also increased in dosed rats compared to controls. Axon filament and myelin sheath degeneration in the spinal cord and/or peripheral nerves occurred in rats receiving doses of 25 mg/kg or higher. Necrosis in the granular cell layer of the cerebellum was seen in rats given 200 mg/kg. Mice receiving 200 mg/kg in 13-wk studies died. Decreased grip strength was noted in mice at doses as low as 25 mg/kg, and rotarod performance was also affected by N-methylolacrylamide administration, but no neuropathology was seen microscopically. Testicular weights were decreased at doses as low as 12.5 mg/kg, and hepatocellular necrosis, thymic lymphocyte necrosis, and hemorrhage, necrosis, and mineralization of the zona reticularis of the adrenal gland were seen in mice that died (200 mg/kg). In 2-yr studies, survival and weight gains in male and female rats receiving doses of 6 or 12 mg/kg/d were minimally affected. No biologically important clinical signs or neoplastic or nonneoplastic lesions were attributed to N-methylolacrylamide administration to rats, suggesting that higher doses could have been tolerated. In mice, survival was not different between dosed and control groups (0, 25, or 50 mg/kg/d). Body weights were higher by as much as 25% in dosed compared to control groups. No compound-related clinical signs were observed, but increases in neoplasms of the harderian gland, liver, and lung were clearly related to chemical administration in both sexes of mice. Benign granulosa-cell neoplasms of the ovary were also increased in dosed female mice.

Lack of delayed neurotoxic effect after tri- o-cresyl phosphate treatment in male Fischer 344 rats: Biochemical, neurobehavioral, and neuropathological studies

Tri- o-cresyl phosphate (TOCP), which produces a delayed neurotoxic syndrome in humans and some animal species, was given to Fischer 344 (F344) male (18 week old) rats to determine if it causes biochemical, sensorimotor, and neuropathological effects. Animals were given TOCP by gavage in doses ranging from 10 to 100 mg of TOCP/kg daily for a period of 63 days. The rats were subjected to a series of neurobehavioral tests including fore- and hindlimb grip strength, motor activity, tremor, and latency to respond to a thermal stimulus. Central and peripheral nervous tissues were examined for damage characteristic of organophosphorous compound-induced delayed neurotoxicity (OPIDN). Brain neurotoxic esterase and acetylcholinesterase activities were inhibited in a dose-dependent fashion. A group of three chickens treated with 100 mg of TOCP/kg/day for 18 days was included as the positive control for enzymatic and histopathological alterations associated with OPIDN. Rats showed no consistent neurobehavioral changes or evidence of neuropathological damage in nervous tissues associated with treatment. In contrast, chickens treated with TOCP developed delayed neurotoxicity characterized by ataxia, which progressed to paralysis. These neurological changes included swelling, fragmentation, and degeneration of the axon and myelin in both central and peripheral nervous tissues. This study concludes that the F344 rat is not sensitive to the delayed neurotoxic effects of TOCP. When studying OPIDN in rats, care must be exercised in choosing the experimental animal since some strains, e.g., F344, are not sensitive.

Response force titration for the assessment of the neuromuscular toxicity of 2,5-hexanedione in rats

A silent, non-moving glass lever combined with strain gauges and mounted in an operant chamber measured the vertical isometric force exerted by the forepaw of rats. Every weekday the rats were subjected to a force titration schedule consisting of a sequence of discrete trials signalled by a lever light. The required force, above which a water reinforcement is delivered, was regulated by a generalized bisection algorithm. A stable force level which the rat was able to attain at 50% of the trials was quickly reached in each session by this algorithm. This technique was used to measure the neuromuscular performance decrement due to repeated 2,5-hexanedione treatment (250 and 500 mg/kg/day per os). The results were compared with fore- and hindlimb grip strength measurements on the same animals. This experiment showed that the force titration technique is able to distinguish between motivational and true force decrements, a distinction which cannot be made by the grip strength technique.

Influence of pre‐ and postnatal exposure of rats to 2.45‐GHz microwave radiation on neurobehavioral function

Rats exposed to microwaves prenatally (2,450 MHz, 10 mW/cm2, 3 h/day, days 5-20 of gestation) or perinatally (same as above plus days 2-20 postnatally) were examined by a neurobehavioral test battery on postnatal days 30 and 100. Body mass, locomotor activity, startle to acoustic and air-puff stimuli, fore- and hindlimb grip strength, negative geotaxis, reaction to thermal stimulation, and swimming endurance were assessed. The prenatally and the perinatally exposed rats (male and female) weighted more than sham-exposed rats at 30, but not at 100, days of age. In addition, the perinatally exposed animals had less swimming endurance at 30, but not at 100, days of age relative to sham-exposed rats. For the other measures, only the air-puff startle response was altered and was limited to the prenatally exposed female pups; ie, at postnatal day 30, the startle response was increased in magnitude, and at postnatal day 100, the response was decreased. No other reliable effects were observed. In a second experiment, rats treated as described above were examined for alterations in body mass, locomotor activity, reaction to air-puff stimuli, reaction to thermal stimulation, and swimming endurance at postnatal days 30-36. Again, perinatally exposed rats were larger in body mass and had less swimming endurance compared with sham-exposed rats. The latency to the air-puff startle response was longer in female pups exposed prenatally. These data indicate that altered endurance and gross motor activity result from perinatal exposure to microwave irradiation.

The temporal relationship between behavioral and hematological effects of inhaled benzene

The time-effect relationship of the behavioral and hematological changes caused by inhaled benzene was investigated in C57BL mice. Mice were exposed to air, or 100, 300, 1000, or 3000 ppm benzene in standard inhalation chambers employing dynamic vapor exposure techniques. Mice were exposed for 6 hr/day for the number of days necessary to achieve a minimum concentration × time (Ct) product of 3000 ppm-days. The intermittent exposure regimen of 6 hr/day was employed to simulate occupational exposure. The most sensitive behavioral index (milk-licking) was affected by the lowest concentration tested (100 ppm), while homecage food intake, hindlimb grip strength, and body weight were reduced only at 1000 and 3000 ppm. Some of these previously undocumented behavioral changes occurred as rapidly as hematological changes that have been considered hallmarks of benzene toxicity. A significant decrease in circulating lymphocytes occurred after exposure to all concentrations. Circulating red blood cells were variably affected by benzene, in that anemia resulted after 10 days exposure to 100 ppm and after 3 days exposure to 300 ppm but not after 3 days exposure to 1000 ppm or a single exposure of 3000 ppm. The data indicate a departure from Ct relationships, and suggest that exposure duration as well as daily dose may be an important factor in benzene toxicity.

Behavioral effects of acrylamide in the mouse

The behavioral toxicity of acrylamide was characterized in the mouse by comparing standard measures of toxicity such as body weight loss and mortality with measures of hindlimb grip strength, locomotor activity, and appetitive behaviors including episodic milk-licking. In the latter test, the mouse received 15-min access daily to a highly palatable, nonessential food substance (10% sweetened milk) that was not available in the maintenance diet. Two strains of mice, CD-1 and C57BL 6J , were injected ip five times weekly with either saline, 20, 60, or 100 mg/kg of acrylamide. Subchronic, but not acute, administration of 100 mg/kg produced weight loss, a severe neuropathy within 3 days, and 100% mortality within 2 weeks. Mice receiving 60 mg/kg subchronically had only a slight loss of body weight but developed a neuropathy within 3 weeks of dosing at which time there was a 50% mortality. Preceding these signs of toxicity, there was a highly significant increase in episodic milk-licking; this increase was significant in both strains of mice by Day 2 and remained elevated throughout the dosing period. The 20 mg/kg dose of acrylamide decreased hindlimb grip strength after 5 weeks of dosing but did not affect milk-licking or body weight even after 7 weeks of dosing. The new use of this test of appetitive behavior has documented a robust effect of acrylamide which preceded other signs of toxicity. The appearance of the various toxic signs was better predicted by the magnitude of the daily dose than the cumulative dose. This study also demonstrated the feasibility of using the mouse in behavioral toxicology.