Repeated Monosodium Glutamate (Chinese Salt) Administration Impaired Memory Functions in Rats: Relationship with Decreased Plasma and Brain Tryptophan.
In the current study adverse effects of monosodium glutamate (MSG) on memory functions were evaluated. Our objective was to examine the association between MSG use and memory functions in male rats. Alteration of behavioral performance and changes in plasma and brain tryptophan (TRP) levels after MSG administration was also investigated. Fourteen adult locally bred male albino Wistar rats were divided into control and test groups (n=7). The test group was intraperitoneally injected with MSG (4 mg/kg/ml)for three weeks while control animals were injected with saline (1ml/kg). Water maze test and novel object recognition task was used for the assessment of memory function following MSG administration. Brain and plasma TRP levels were determined by HPLC-UV method. The result showed that MSG administration for 3 weeks significantly (pless than 0.05) impaired recognition memory in MSG treated rats. Short term memory (pless than 0.01) and long term memory (pless than 0.01) of MSG treated rats were significantly impaired in water maze test.
Repeated MSG administration for three weeks significantly decreased brain TRP (pless than 0.05) and plasma TRP (pless than 0.01) levels. Present study clearly indicates that repeated MSG administration significantly altered the neurobehavioral perfor mance in rats. Impairment in memory function following MSG administration may be due to decreased TRP levels in plasma and brain. The present study suggests that decreased plasma and brain TRP levels may be responsible for the impaired memory functions observed in MSG treated rats.
Keywords: Monosodium Glutamate; Memory; Tryptophan and Water Maze.
Currently, thousands of agents are intentionally added to food as flavor enhancer. Among them, monosodium glutamate (MSG), sodium salt of non-essential amino acid glutamate, is one of the most widely used food additives . Its role as a flavor enhancer was proposed early in the 20th century . It produced a unique taste referred to as a fifth taste, umami . Despite a widespread belief that glutamate can elicit asthma, migraine headache and has been implicated in causing adverse reaction referred as Chinese restaurant syndrome , there are no consistent clinical data to support this claim. It has also been reported to elicit asthmatic bronchospasm, urticaria, angio-oedema, and rhinitis  and is the inducer of oxidative stress .It has been reported previously that MSG treatment leads to degeneration of the neurons in certain brain areas and also causes various changes in the motor activity, sensory performance and learning abilities .
It has been shown previously that administration of MSG induced toxic effects in various regions of brain, thymus, liver and kidney .
MSG administration has been reported to significantly alter the neurobehavioral performance in rats including anxiety and memory impairment . Accumulating evidence suggests involvement of serotonin in the regulation of learning and memory functions . Previous studies from our laboratory have suggested that administration of TRP, the dietary precursor of 5-HT, increases brain 5-HT levels and improves memory functions . The most consistent observation following manipulation of serotonin levels in human and animals is that low extracellular 5-HT levels are associated with impaired memory functions . Reports have shown that 5-HT synthesis and release is directly related to tryptophan availability .Tryptophan depletion, a challenge tool for 5-HT has been shown to induce memory deficits in humans  but on affective behavior its effect is modest .
The purpose of the study was aimed to investigate the effects of repeated monosodium glutamate administration on memory functions in male rats. The study was also aimed to determine the neurochemical basis of the behavioral change after MSG intake by determining the alteration in plasma and brain tryptophan levels in rats.
Fourteen locally bred albino wistar male rats weighing 180-200 grams were used in this study. All animals were 4 month old and they were housed individually in their home cages. They were given free access to standard rodent diet and tap water for at least 4 days before experimentation so that rats could adapt themselves to the new environment. All experiments were conducted according to a protocol approved by Institutional Review Committee, Federal Urdu University, Karachi. In the beginning of experiment animals were randomly divided into two groups (n=7) as control and MSG treated rats. MSG at a dose of 4 mg/kg/ml was used. Test group received MSG intraperitoneally while control group was injected with saline (1ml/kg) for 3 weeks. Behavioral activities of rats were monitored after three weeks of drug administration. Rats were decapitated after monitoring the behavioral activities to collect the plasma and brain samples.
Fresh brains were dipped in chilled saline and they were stored at low temperature (-70C) until the analysis of brain TRP by HPLC-UV.
Analysis of brain and plasma TRP levels was carried out by HPLC-UV. A 5-II Shim-Pack ODS separation column of 4.0 mm internal diameter and 150 mm length was used. Mobile phase contains methanol (10%), octyl sodium sulfate (0.023%) and EDTA (0.0035%) in 0.1 M phosphate buffer at pH 2.9.Operating pressure was 2000-3000psi and UV detector of 0.8 volts was used.
Water Maze Test
The effects on spatial memory were examined in this study by using water maze (WM) apparatus. The WM apparatus used in this study consisted of a rectangular tank that is made up of transparent glass. Tank was filled with room temperature clean tap water; to the depth of 18 cm. water of the tank was made opaque with the help of powdered milk so that the platform became invisible to rats. Platform (16 cm x 16 cm) was hidden 2 cm below the surface of water in a fixed location. First the rats were trained for a cut off time of 3 minutes and in this session each rat was introduced in the water maze and they were given 3 minutes to find the platform. If it failed in finding the platform in the training session, it was gently guided onto the platform. Short term memory and long term memory of rats was tested 1 h and 24 h respectively after training by recording the time taken by each rat to relocate the hidden platform 1h and 24 h after training. The cut off time for each session was 3 minutes.
Object Recognition Task
The novel object recognition test was used to monitor the recognition memory in rats. It is used to assess the rat's ability to distinguish a novel (new) object from a familiar object. Square wooden box of dimensions 45x45x45cm3 were used. The objects to be discriminated were two similar wooden square objects and a circular object (new object). Height of all object were same. The test was performed in three phases; habituation, training, and test session. Habituation was done on first day in which rats were placed in empty wooden box for 10 minutes without any objects. Training session was performed next day by placing two same objects in wooden box and placing each rat in the box for 10 minutes. Test session was performed on third day during which animal is exposed to one of the known object and novel object, for 5 minutes. In the test phase the sniffing time for the novel and familiar object was measured. Increase in time spent with the novel object was considered as increase in recognition memory.
Data are presented as mean SD. Data were analyzed by Student's t-test; p values less than 0.05 were considered significant.
Results and Discussion
Fig. 1 and 2 shows the influence of repeated MSG administration on plasma and brain TRP levels in rats. Analysis by Student's t-test revealed that MSG administration significantly decreased plasma TRP (t=4.45; pless than 0.01) and brain TRP (t=2.58; pless than 0.05) as compared to controls.
Fig. 3 shows the influence of repeated MSG administration upon short term memory in rats as assessed by water maze test. Analysis by Student's t- test revealed a significant effect (t=5.75; pless than 0.01) on short term memory. The results indicate that MSG administration significantly impaired short term memory functions in rats. Fig. 4 shows the influence of repeated MSG administration upon long term memory in rats as assessed by water maze test. Analysis by Student's t-test revealed a significant effect (t=8.32; pless than 0.01) on long term memory. The results indicate that MSG administration significantly impaired long term memory functions in rats.
Fig. 5 shows the influence of repeated MSG administration upon recognition memory in rats as assessed by object recognition task. Analysis by Student's t-test revealed a significant effect (t=2.86; pless than 0.05) on recognition memory. The results indicate a significant decrease in time taken by MSG treated rats to sniff novel object as compared to control rats exhibiting impairment in recognition memory by MSG treated rats as compared to control rats.
Now days a significant part of our diet includes various chemical additives, which if not avoided may increase the risk of developing health problems . Hundreds of such additives are added in our diet to mimic natural flavor. One of the most used food additive is monosodium glutamate which is deeply involved in everyday life. Its intake has been known to affect dietary habits and endocrine system. MSG administration results in obesity  and may impaired learning and memory functions [18, 19]. In the present study MSG administration for 3 weeks significantly impaired short term memory, long term memory and recognition memory in rats as compared to control rats. The alteration manifested as more time taken by the MSG treated rats to reach the hidden platform in water maze and shorter time spent with the new object in object recognition task.
Abnormal behaviors such as increase in irritability and hypo activity were reported following MSG administration . Learning and memory functions were affected by the administration of MSG . MSG administration has been reported to damage the hypothalamic regulation of appetite, and thereby determines the propensity of world-wide obesity [22, 23].It has been reported previously that administration of MSG in rats impaired both place learning acquisition and retrieval abilities .In the current study impairment in memory functions following MSG administration has been related to decrease in plasma and brain TRP levels of MSG treated rats. Tryptophan depletion was reported to impaired memory processes by affecting brain TRP levels, thereby decreasing 5-HT synthesis. The present results provide a strong evidence for the association between low plasma and brain TRP levels and memory impairments in MSG treated rats as compared to controls.
Concentration of TRP in blood is known to be an important determinant of brain 5-HT. Increase in brain 5-HT depends on the increase in brain tryptophan as the enzyme tryptophan hydroxylase is half saturated with its substrate . Thus, increased concentration of brain TRP increases brain 5-HT and reduced levels of plasma TRP leads to decreased brain TRP and decreases brain 5-HT concentration . Present study reported that tryptophan uptake by brain was decreased in MSG treated rats leading to reduction in brain tryptophan levels due to which synthesis and turnover of 5-HT in brain was also decreased.
Previously anxiogenic and depressive behaviors exhibited by MSG treated rats were also related to an abnormal functioning of serotonergic system . In the present study repeated MSG administration significantly impaired memory functions in rats. Recognition memory, short term memory and long term memory of MSG treated rats were significantly impaired. Evidence shows that brain TRP and 5-HT have a significant role in learning and memory . Alterations in brain 5-HT levels can contribute to behavioral differences in mice and psychiatric disorders in humans . Previously it is reported that increasing brain 5-HT by administration of TRP enhanced memory function  while decreased brain 5-HT has also been shown to impair memory . It was also reported that alteration in serotonin levels may be closely linked to one type of memory and not affecting other type of memory .
Decrease in brain serotonin levels has been reported to affect cholinergic activity in those brain areas that play an important role in learning and memory . Memory alteration in this study following MSG administration has been related to serotonergic system. The present study suggests that decreased plasma and brain TRP levels may be responsible for the impaired memory functions observed in MSG treated rats.
The authors wish to acknowledge Federal Urdu University, Pakistan for providing the funds for this study.
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|Author:||Khaliq, Saima; Mujahid, Rubab; Haider, Saida; Anis, Lubna; Sana Mustafa,; Farooq, Umar; Khan, Ajmal|
|Publication:||Journal of the Chemical Society of Pakistan|
|Date:||Apr 30, 2015|
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