Developmental Neurotoxicity Study in the Han Wistar Rat/MnCl2/OECD 426/Inhalation/GLP
The purpose of this study was to demonstrate potential functional and morphological effects on the developing nervous system of the rat offspring that may arise from exposure in utero and during early life. This developmental neurotoxicity study was performed according the OECD Guidelines for the Testing of Chemicals, No. 426 and OPPTS Health Effects Test Guidelines No. 870.6300.
Three groups of 27 females were treated with Manganese dichloride at 3.5, 12.3 and 17.6ug/L (corresponding to group 2, 3 and 4 respectively) from Sigma Aldrich and the control group was treated with air under the same conditions as animals exposed to the test item. Manganese dichloride were used as supplied for aerosol generation to obtain suitable particle sizes and administered by inhalation (nose only) once daily to female rats from day 6 to day 19 post coitum and from day 1 to day 20 post partum. Pups were not treated directly but exposed indirectly via maternal blood and milk during pre- and postnatal neurological development.
Offspring was randomly selected from within litters for neurotoxicity evaluation. The evaluation consisted of observations to detect gross neurologic and behavioral abnormalities, including the assessment of physical development, behavioral ontogeny, motor and sensory function, learning and memory, the evaluation of brain weights and neuropathology during postnatal development and adulthood.
Mortality / Viability
At the high dose level, one dam was killed in extremis on day 15 post coitum. This dam showed breathing noises and labored breathing (highest grading) after exposure and was in a weakened condition, with body weight loss, ruffled fur and a hunched posture. Due to the incidental occurrence, observations leading to the pre-termination were considered not to be related to the treatment.
All other dams survived until the scheduled necropsy.
At high dose level, up to 70% of the dams showed breathing noises during the gestation period with steadily decreasing numbers of affected dams towards the end of the period. Except three dams, dams showed no more clinical signs during the lactation period.
No test item-related clinical signs were observed at the low and mid dose level during the study.
Mean food consumption was dose-dependently decreased immediately after treatment started from day 6 - 11 post coitum at mid and high dose level (7% and 27%, respectively, compared to the control group) and recovered afterwards from day 11 to the end of exposure period at both dose levels.
There were no effects on food consumption during gestation and lactation for dams treated at the low dose level.
During lactation period, there were no effects on mean food consumption at any dose level.
Mean body weight gain was dose-dependently decreased after treatment started at the mid and high dose level.
At the high dose level, there was a body weight loss up to 3% within the first 5 days after treatment started. Afterwards, body weight gain increased but remained statistically significantly reduced if compared to the control values until day 17 post coitum. Furthermore, mean body weight was also reduced during the gestation period.
At the mid dose level, mean body weight gain was reduced from start of treatment until day 17 post coitum, with statistical significance until day 12 post coitum. Mean body weights were not statistically significantly different form the control group.
No effects on mean body weight gain and mean body weights were observed at the low dose level during the gestation period.
During lactation period, there were no effects on mean body weight gain and mean body weights at any dose group.
Reproduction and Breeding Data
The mean duration of gestation was similar in all dose groups and not affected by treatment with the test item.
No increased postnatal loss was observed during the lactation period at any dose level. The viability and weaning indices were similar in all groups.
There were no test item-related macroscopic findings in any group.
Litter Size and Pup Survival
Mean number of pups per litter was similar in all groups at first litter check and at the end of the lactation period and not affected by treatment with the test item. Sex ratios at first litter check and on day 21 post partum were unaffected by treatment with the test item.
No findings that were considered to be test item-related were noted at the first litter check or during pre- and post-weaning.
Food consumption of pups was considered unaffected by treatment with the test item.
Mean pup weight and its development were considered unaffected by treatment with the test item.
Developmental Indices and Sexual Maturation
There were no test item-related differences when the developmental indices (pinna unfolding, incisor eruption, onset of coat development and opening of eyes) occurred between the control and the dose groups.
Mean age and mean body weight at balanopreputial separation in males and vaginal opening in females were similar in all groups.
Detailed Clinical Observations and FOB
No findings were noted for any selected pups on days 5 and 11 post partum during the detailed clinical observations.
In male and female pups, observations and measurements conducted as part of the functional observational battery on days 21, 35, 45 and 60 post partum, did not indicate any effects of the test item in any group.
Locomotor activity was not affected by treatment with the test item during the pre-weaning (day 13, 17 and 21 post partum) and post-weaning period (day 60 post partum).
Y-Maze – Learning and Memory
On day 25 and 61 post partum, in all pups there was a clear evidence of learning of the task in the initial (learning phase) since the time taken to complete the first trial was greater than for subsequent trials. There was also a clear evidence of memory since the first trial of the memory phase was less than half of time for the first trial of the learning phase.
The mean amplitude of startle response and habituation on days 25 and 61 post partum were not affected by treatment with the test item.
There were no brain weight effects in any group on day 11, on day 22 and day 63 post partum.
Macroscopic Brain Morphometry
No findings were noted on the length of the cerebellum and forebrain of all examined groups on day 11, 22 and 63 post partum.
Microscopic Brain Morphometry and Neuropathology
The daily administration of Manganese dichloride, in utero and in early life produced no morphological findings in the tissues examined.
All of the histopathological findings encountered were considered to have arisen spontaneously or post mortem.