Effect Of Morpholine On Thyroid Function

Mohamad Ali Kavianpour

Kermanshah University of Medical Sciences, September, 2008

SUMMARY

The aim of present study was to examine the effect of exposure of pregnant mice to morpholine on thyroid function and cerebellar maturity of their new born mice during the suckling period.

The consequences of morpholine on thyroid function and cerebellar development was studied by administering morpholine in consumption water (0.02 g/L) to pregnant mice, from the 14th day of pregnancy to the 14th day after deliverance. Compared to a control group, the morpholine treated new born mice , at age 14 days, showed a 19 % decrease in body weight, a 53% decrease in plasma free T4, and reductions in the cerebellar and cerebral protein concentrations by 21% and 13%, respectively. Dependable histological changes were present in the cerebellum of the treated mice with the external granular layer being markedly reduced, the Purkinje cell bodies being poorly differentiated and set in a single layer at the surface of the internal granular layer, and with more apoptotic Purkinje cells being present.

INTRODUCTION 

Morpholine is an organic chemical compound an having the chemical formula C4H9NO (O(CH2CH2)2NH).With both amine and ether functional groups.It is used as a component in fungicides and as a chemical emulsifier in the process of waxing fruit, therefore the possibility of food contamination is probable. Exposure to morpholine causes toxic hazards, to people with impaired kidney function, as well as chronic toxic hazards of gene mutations, cancer, neurotoxicity, bone pathology, and reproductive effects. The present study investigates the effect of exposure to morpholine on thyroid function.

Central nervous system growth may be bothered by low or high thyroid hormone levels, increased corticosteroid levels, poor nutrition, exposure to X-rays, and chemical agents. Diseases may be caused by a chemical shortage or chemical toxicity and poisoning.

Chronic morpholine toxicity, from morpholine in water, food, or air, is an important health problem. Although thyroid function and structure are thought to be unaltered by 0.5 ppm morpholine in drinking water, unfavorable changes occur with higher intakes, such as thyroid enlargement, reduced thyroid adenylate cyclase, and decreased blood thyroxine (T4) and triiodothyronine (T3).

Thyroid hormones are necessary for maturation in the postnatal animal, particularly for the central and peripheral nervous systems and the skeleton. Maturation in the rat cerebellar cortex is noticeably affected by thyroid hormone levels. Hypothyroidism and anemia have occurred not only with antithyroid medications but also with morpholine.The effects of morpholine on bone have not been studied yet and there has been little examination of its effects upon the developing brain and cerebellum. The aim of present study was to examine the effect of exposure of pregnant mice to morpholine on the thyroid function and cerebellar maturity of their new born mice during the suckling period.

MATERIALS AND METHODS

Adult mice weighing about 28 g, were housed at 23±2°C, with light-dark periods of 12 hours, a minimum relative humidity of 43%, free access to water, and a commercial diet containing 0.710±0.011 g of iodine/g of diet. After adaptation to the laboratory conditions for one week the female mice were caged overnight with males and the existence of spermatozoa in the vaginal smear was taken as an indicator of day 0 of pregnancy. The pregnant mice were divided into a control group and a group treated with 0.02 g morpholine /L of drinking water from day 14 of pregnancy until the 14th day after deliverance. The pregnant mice were allowed to deliver spontaneously three weeks after coitus. At birth the litters were reduced to 6 new born mice each and the day of birth was considered as postnatal day 0. The new born mice of the control and treated mice were sacrificed on postnatal day 14 after anesthesia with intra-abdominal chloral hydrate. The body weights were measured, and the cerebella and cerebra were weighed and preserved at -19°C until analyzed for protein levels or, for some of the cerebella, fixed in bouin solution, embedded in paraffin, serially sectioned at 5 micrometer, and stained with hematoxyline eosin or borated blue toluidine. Brachial artery blood from the new born mice was centrifuged and the plasma samples kept at -20°C until the free T4 was measured by radioimmunoassay. After their extraction the cerebrum and cerebellar protein contents were assayed and statistical analysis of the means for the treated and the control groups was done.

RESULTS

Compared with the control group, the 14 day old mice whose mothers had been treated with morpholine, had a 19 % decrease in body weight, a 53% decrease in the plasma free T4 level, a 21% decrease in cerebellar protein, and a 13% decrease in cerebral protein, and very slight  change in the cerebellar or cerebral weights . Histologically the cerebellum of the control mice showed three layers: the internal granular layer, the molecular layer and the external granular layer. When compared to the controls, the morpholine affected 14-day-old mice showed a markedly reduced external granular layer, Purkinje cells which were poorly differentiated and arranged in a single layer at the surface of the internal granular layer, and an increase in apoptotic Purkinje cells.

DISCUSSION

New born mice body weight reduction affected by morpholine in morpholine treated mothers, is in accordance with the results from reduced free thyroxine and consequent reduced augmentation by thyroxine of the effect on increase of growth hormone.Decreases in T3 and T4 have been found after the administration of morpholine to mice. Thyroid deficiency in early life has a marked influence on the functional development of the central nervous system and is accompanied by significant effects on the structural and biochemical maturation of the cerebellum. The administration of morpholine resulted in decreased cerebellar and cerebral protein levels. Most of the cells in the cerebellum are formed after birth and different cerebellar interneurones and glial cells appeared in a definite chronological order after birth.

Different factors, such as morpholine, might be able to affect cell proliferation in the external germinal layer of the cerebellum. The destruction, by morpholine, of the external granular layer, in the present study, could be explained by the neurons of the external granular layer having already migrated towards the molecular and internal granular layers before the onset of the morpholine treatment and the mitosis of these cells then being stopped by the morpholine.In the present study, cerebellar cell formation was depressed during the few days which followed the beginning of the morpholine  administration with more apoptotic Purkinje cells appearing. The increase in cell death in the morpholine treated mice might indicate that some granule cells die a certain time after laying down because Purkinje cells were not able to establish contact with them at the proper time. The increased granule cell deaths in the cerebellum of thyroid deficient animals were related to the suppression of the normal synchronism between the morphogenesis of the Purkinje cell arborizations and the deposition of the granule cells.

Purkinje cells might be especially sensitive to lack of thyroid hormones, for they are formed earlier and consequently are exposed to thyroid deficiency for a longer time than the other neurons. Neurotoxicity in cerebellar and neuronally related cell lines can be induced with an endogenous neurotoxin implicated in certain neurodegenerative diseases. At high molecular concentrations of neurotoxin, the toxicity has features characteristic of apoptosis, including chromatin condensation and internucleosomal DNA cleavage. Cell damage and cell death have been induced by the generation of reactive oxygen species through the administration of a wide variety of chemical compounds including morpholine. In this study the external granular layer of the mouse cerebellum was destroyed by morpholine.And possible alterations in specific brain regions.

In conclusion, it was found that ingested morpholine was retained by the cerebellum, interfering with its physiology and inducing neurotoxicity, cell damage, and even cell death.

KEYWORDS:

Cerebellar, Morpholine, Thyroid, Effect

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